DESC:FORM 2
THE PATENTS ACT, 1970
(39 of 1970)
&
THE PATENTS RULES, 2003
COMPLETE SPECIFICATION
(Section 10 and Rule 13)

1. TITLE OF THE INVENTION:
SYNTHETIC DELIVERY SYSTEMS AND CONJUGATES THEREOF

2. APPLICANT:
MICRO CRISPR Pvt. Ltd., an Indian Company, of the address Survey No: 1574, Muktanand Marg, Chala, Vapi-396191, Gujarat, India

The following specification particularly describes the invention and the manner in which it is to be performed:

FIELD OF INVENTION
[1] The present disclosure relates to a delivery system. More specifically, the present disclosure relates to a synthetic delivery system and conjugates thereof.
BACKGROUND OF INVENTION
[2] There are numerous ways to deliver a therapeutic agent to a targeted organ inside a body. Usually, the therapeutic agent is a pharmaceutical composition for example, nucleic acid. With age, nucleic acids are directly delivered to the targeted organ for therapeutic effects.
[3] One of the most common therapeutic agents being ribonucleic acid (RNA) and structures thereof. Conventionally, for efficient delivery of the RNA structure to the targeted organ, lipid nanoparticles (an exemplary delivery system) are used. The LNPs are manufactured by encapsulating the RNA structures inside the LNPs. Thereafter, the LNPs are administered inside the body. Basis the surface structure and properties of the lipid nanoparticles, the LNPs are directed towards a targeted organ.
[4] However, conventional method of delivering the therapeutic agents suffers from a number of drawbacks such as off-target delivery, tissue/organ specificity, lack of widespread tissue/organ delivery, undesirable immune response, long-term stability, and safety concerns.
[5] Therefore, there is a need for a delivery system which overcomes the aforementioned challenges associated with the conventional systems.
SUMMARY OF INVENTION
[6] Particular embodiments of the present disclosure are described herein below with reference to the accompanying drawings, however, it is to be understood that the disclosed embodiments are mere examples of the disclosure, which may be embodied in various forms. Well-known functions or constructions are not described in detail to avoid obscuring the present disclosure in unnecessary detail. Therefore, specific structural and functional details disclosed herein are not to be interpreted as limiting, but merely as a basis for the claims and as a representative basis for teaching one skilled in the art to variously employ the present disclosure in virtually any appropriately detailed structure.
[7] In an exemplary embodiment, the present disclosure relates to a synthetic delivery system for targeted delivery of an agent including a tetravalent atom/moiety, at least one first spacer, at least one linker, one or more second spacers, and a plurality of ligands. The first spacer is coupled to the tetravalent atom/moiety. The linker is coupled to the tetravalent atom/moiety via the at least one first spacer. The second spacers are coupled to the tetravalent atom/moiety. The ligands are coupled to the tetravalent atom/moiety via the at least one second spacer. The linker is conjugated to an agent. The linker includes at least one of 4-(hydroxymethyl)-4-methylpiperidin-3-ol having a formula V, 2-(hydroxymethyl)piperidin-4-ol having a formula VI, 5-(hydroxymethyl)-5-methylpyrrolidin-3-ol HCl salt having a formula VII, 2-[4-(hydroxymethyl)piperidin-4-yl]propan-2-ol having a formula VIII, 3-aminopropane-1,2-diol having a formula IX, 4-(hydroxymethyl)piperidin-4-ol having a formula X, 3-aminobutane-1,2-diol having a formula XI, 5-(1-hydroxyethyl)pyrrolidin-3-ol having a formula XII, 2-aminobutane-1,3-diol having a formula XIII, 4-amino-1-[4-hydroxy-2-(O-dimethoxy tritylmethyl)pyrrolidin-1-yl]butan-1-one having a formula XIV, 8-((2R,4R)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-hydroxypiperidin-1-yl)-8-oxooctanoic acid having a formula XV, 4-((2R,4R)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-hydroxypiperidin-1-yl)-4-oxobutanoic acid having a formula XVI, 8-((4-(bis(4-methoxyphenyl)(phenyl)methoxy)-3-hydroxybutan-2-yl)amino)-8-oxooctanoic acid having a formula XVII, and 3-(3-(4-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-3-hydroxy-4-methylpiperidin-1-yl)-3-oxopropoxy)propanoic acid having a formula XVIII.
[8] In another exemplary embodiment, the present disclosure relates to a method to synthesize a synthetic delivery system for targeted delivery of an agent. The method commences by preparing a first mixture to obtain a viscous solid having a formula XXXVIII from galactosamine pentaacetate having a formula XXXVII. A second mixture is prepared to obtain a compound having a formula XXXX from the viscous solid having the formula XXXVIII. A first suspension is prepared to obtain 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having a formula XXXXI from the compound having the formula XXXX. A third mixture is prepared to obtain a compound having a formula XXXXIV from 3,3'-((2-(((Benzyloxy)carbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy))dipropionic acid (Tris-base) having a formula XXXXII. A fourth mixture is prepared to obtain a compound having a formula XXXXV from the compound having the formula XXXXIV. A fifth mixture is prepared to obtain a compound having a formula XXXXVI from the compound having the formula XXXXV and 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having the formula XXXXI. A sixth mixture is prepared to obtain a compound having a formula LXV from the compound having the formula XXXXVI. A seventh mixture is prepared to obtain a compound having either one of a formula LXVI, LXVII, LXVIII, or LXIX from the compound having the formula LXV along with at least one linker (130) having a formula XV, XVI, XVII, or XVIII. An eighth mixture is prepared to obtain a compound having either one of a formula LXX, LXXI, LXXII, or LXXIII from the compound having either one of the formula LXVI, LXVII, LXVIII, or LXIX. A ninth mixture is prepared to obtain a synthetic delivery system having either one of a formula LXXIV, LXXV, LXXVI, or LXXVII from the compound having either one of the formula LXX, LXXI, LXXII, or LXXIII.
BRIEF DESCRIPTION OF DRAWINGS
[9] The summary above, as well as the following detailed description of illustrative embodiments, is better understood when read in conjunction with the apportioned drawings. For the purpose of illustrating the present disclosure, exemplary constructions of the disclosure are shown in the drawings. However, the disclosure is not limited to specific methods and instrumentality disclosed herein. Moreover, those in the art will understand that the drawings are not to scale.
[10] Fig. 1 depicts a delivery system 100 coupled to a therapeutic agent 200 in accordance with one or more embodiments of the present disclosure.
[11] Fig. 2 depicts a method 300 to synthesize the delivery system 100 in accordance with one or more embodiments of the present disclosure.
[12] Figs. 3-13a depict the experimental data associated with the delivery system 100 in accordance with an exemplary embodiment of the present disclosure.
DETAILED DESCRIPTION OF THE DRAWINGS
[13] Prior to describing the invention in detail, definitions of certain words or phrases used throughout this patent document will be defined: the terms "include" and "comprise", as well as derivatives thereof, mean inclusion without limitation; the term "or" is inclusive, meaning and/or; the phrases "coupled with" and "associated therewith", as well as derivatives thereof, may mean to include, be included within, interconnect with, contain, be contained within, connect to or with, couple to or with, be communicable with, cooperate with, interleave, juxtapose, be proximate to, be bound to or with, have a property of, or the like. Definitions of certain words and phrases are provided throughout this patent document, and those of ordinary skill in the art will understand that such definitions apply in many, if not most, instances to prior as well as future uses of such defined words and phrases.
[14] Reference throughout this specification to “one embodiment,” “an embodiment,” or similar language means that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one embodiment. Thus, appearances of the phrases “in one embodiment,” “in an embodiment,” and similar language throughout this specification may, but do not necessarily, all refer to the same embodiment, but mean “one or more but not all embodiments” unless expressly specified otherwise. The terms “including,” “comprising,” “having,” and variations thereof mean “including but not limited to” unless expressly specified otherwise. An enumerated listing of items does not imply that any or all of the items are mutually exclusive and/or mutually inclusive, unless expressly specified otherwise. The terms “a,” “an,” and “the” also refer to “one or more” unless expressly specified otherwise.
[15] Although the method steps of exemplary embodiments of the disclosed method may be described in a particular, sequential order for convenient presentation, it should be understood that the disclosed embodiments can encompass an order of method steps other than the particular, sequential order disclosed. For example, method steps described sequentially may in some cases be rearranged or performed concurrently. Further, descriptions and disclosures provided in association with one particular embodiment are not limited to that embodiment, and may be applied to any embodiment disclosed herein. Moreover, for the sake of simplicity, the attached figures may not show the various ways in which the disclosed system, method, and apparatus can be used in combination with other systems, methods, and apparatuses.
[16] Furthermore, the described features, advantages, and characteristics of the embodiments may be combined in any suitable manner. One skilled in the relevant art will recognize that the embodiments may be practiced without one or more of the specific features or advantages of a particular embodiment. In other instances, additional features and advantages may be recognized in certain embodiments that may not be present in all embodiments. These features and advantages of the embodiments will become more fully apparent from the following description and apportioned claims, or may be learned by the practice of embodiments as set forth hereinafter.
[17] The present disclosure discloses a synthetic delivery system (or delivery system) and conjugates thereof. The delivery system may have a pre-defined affinity towards a specific tissue/organ. In an exemplary embodiment, the delivery system is hepatocyte-specific and has affinity towards asialoglycoprotein receptor (ASGPR) which is highly expressed on cell surfaces of the liver.
[18] Although the delivery system of the present disclosure is described with the example of liver and tissues/cells thereof, the delivery system may be used to target other and more than one organs/tissue and the same is within the scope of the teachings of the present disclosure. For example, the delivery system may have affinity towards lungs, central nervous system (CNS), eye, kidney, etc.
[19] One or more therapeutic agents (or agents) may be conjugated to the delivery system. In an exemplary embodiment, a small interfering ribonucleic acid (siRNA) is conjugated to the delivery system of the present disclosure. The siRNA is a double stranded oligonucleotide that silences a pre-determined gene via RNA interference (RNAi) mechanism.
[20] Although the agent in the present disclosure is described with the example of siRNA, other agents may be used to impart therapeutic effect and the same is within the scope of the teachings of the present disclosure. For example, the agent may be one of miRNA, antisense oligonucleotides (ASO), aptamers, etc.
[21] The delivery system of the present disclosure increases the circulation time of the agent conjugated to the delivery system by preventing degradation of the agent thus, providing stability. The delivery system also enhances specificity, improves uptake and reduces off targets.
[22] Now referring to the figures, Fig. 1 illustrates an exemplary synthetic delivery system (or delivery system) 100 conjugated to an agent 200. The delivery system 100 includes a carbon atom 110, at least one first spacer 120, at least one linker 130, one or more second spacers 140, a plurality of ligands 150, etc. The delivery system 100 may have a pre-defined structure including at least one of Tris-GalNAc-NHCbz (peracetylated, Cbz), GaLNAc Functionalized with alkynes, amines, azides, carboxylic acids, N-succinimidyl esters, monovalent, divalent, etc.
[23] The carbon atom 110 helps to couple the first spacer 120 and the second spacers 140 together via covalent bonds. The carbon atom 110 may be replaced with other tetravalent atom/moiety and the same is within the scope of the teachings of the present disclosure.
[24] The first spacer 120 couples the carbon atom 110 to the linker 130. The first spacer 120 includes at least one of N-Hydroxysuccinimide (NHS) ester, polyethylene glycol (PEG), amino alcohols, allyl amine, ethers, etc. The first spacer 120 makes a covalent bond with the linker 130. The first spacer 120 includes at least one of bis(2,5-dioxopyrrolidin-1-yl) pentanedioate having a formula I, 7-[(2,5-dioxopyrrolidin-1-yl)oxy]-7-oxoheptanoic acid having a formula II, bis(2,5-dioxopyrrolidin-1-yl) 3,3'-oxydipropanoate having a formula III, 2,5-dioxopyrrolidin-1-yl hex-5-ynoate having a formula IV, etc. In an exemplary embodiment, the delivery system 100 includes only one first spacer 120.

(Formula I) (Formula II) (Formula III) (Formula IV)

[25] The linker 130 couples the first spacer 120 to the agent 200. The linker 130 includes at least one of cleavable linkers, cyclic and acyclic amino alcohols, piperdine (substituted and unsubstituted) alcohols, amino alcohols, etc. The linker 130 includes at least one of 4-(hydroxymethyl)-4-methylpiperidin-3-ol having a formula V, 2-(hydroxymethyl)piperidin-4-ol having a formula VI, 5-(hydroxymethyl)-5-methylpyrrolidin-3-ol HCl salt having a formula VII, 2-[4-(hydroxymethyl)piperidin-4-yl]propan-2-ol having a formula VIII, 3-aminopropane-1,2-diol having a formula IX, 4-(hydroxymethyl)piperidin-4-ol having a formula X, 3-aminobutane-1,2-diol having a formula XI, 5-(1-hydroxyethyl)pyrrolidin-3-ol having a formula XII, 2-aminobutane-1,3-diol having a formula XIII, 4-amino-1-[4-hydroxy-2-(O-dimethoxy tritylmethyl)pyrrolidin-1-yl]butan-1-one having a formula XIV, 8-((2R,4R)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-hydroxypiperidin-1-yl)-8-oxooctanoic acid having a formula XV, 4-((2R,4R)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-hydroxypiperidin-1-yl)-4-oxobutanoic acid having a formula XVI, 8-((4-(bis(4-methoxyphenyl)(phenyl)methoxy)-3-hydroxybutan-2-yl)amino)-8-oxooctanoic acid having a formula XVII, 3-(3-(4-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-3-hydroxy-4-methylpiperidin-1-yl)-3-oxopropoxy)propanoic acid having a formula XVIII, etc. In an exemplary embodiment, the delivery system 100 includes only one linker 130.

(Formula V) (Formula VI) (Formula VII) (Formula VIII)

(Formula IX) (Formula X) (Formula XI) (Formula XII)

(Formula XIII) (Formula XIV)
(Formula XV)
(Formula XVI)
(Formula XVII)
(Formula XVIII)

[26] In an exemplary embodiment, a hydroxyl group of the linker 130 makes a phosphodiester bond with a hydroxyl group of the agent 200.
[27] In an exemplary embodiment, as shown in Fig. 1, the agent 200 includes a small interfering ribonucleic acid (siRNA) having a sense strand 210 and an antisense strand 220. Each of the sense strand 210 and the antisense strand 220 have respective 3’-ends and 5’-ends. A hydroxyl group at the 3’-end of the sense strand 210 makes a phosphodiester bond with one of the hydroxyl groups of the linker 130.
[28] The second spacers 140 couple the carbon atom 110 to at least one ligand 150 each. The second spacer 140 makes a covalent bond with the ligand 150. The second spacer 140 includes at least one of N-Hydroxysuccinimide (NHS) ester, polyethylene glycol (PEG), alkynes, azides, ethers, etc. The second spacer 140 includes at least one of2-((tert-butoxycarbonyl)amino)-2-((hex-5-ynoyloxy)methyl)propane-1,3-diyl bis(hex-5-ynoate) having a formula XIX, 2-Azidopropyl-3,4,6-tri-O-acetyl-2-(acetylamino)-2-deoxy-D-galactopyranosyl or any azidoalkyl-Ac-GalNAc having a formula XX, or a combination thereof. In an exemplary embodiment, the delivery system 100 includes three second spacers 140.

(Formula XIX) (Formula XX)

[29] The ligand 150 includes N-Acetylgalactosamine (GalNAc) or derivatives thereof. The ligand 150 includes at least one of 5-[[3,4,6-tri-O-acetyl-2-(acetylamino)-2-deoxy-D-galactopyranosyl]oxy]-pentanoic acid having a formula XXI, 5-[[3,4,6-tri-O-acetyl-2-(acetylamino)-2-deoxy-D-galactopyranosyl]oxy]-pentanoate N-Hydroxysuccinimide (NHS) ester having a formula XXII, etc. In an exemplary embodiment, one ligand 150 is coupled to each of the second spacers 140. The ligand 150 may have an affinity to bind with a pre-defined cell receptor(s) or the like. In an exemplary embodiment, the ligand 150 of the delivery system 100 is hepatocyte-specific and has affinity towards asialoglycoprotein receptor (ASGPR) which is highly expressed on cell surfaces of the liver.

(Formula XXI) (Formula XXII)

[30] In an exemplary embodiment, the delivery system 100 includes 4-((1-(24-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-9,9-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-7,14,20-trioxo-4,11-dioxa-8,15,19-triazatetracosan-1-oyl)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)piperidin-4-yl)oxy)-4-oxo-succinoyl-CPG having a formula XXIII.

(Formula XXIII)

[31] In an exemplary embodiment, the delivery system 100 includes 4-((1-(24-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-9,9-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-7,14,20-trioxo-4,11-dioxa-8,15,19-triazatetracosan-1-oyl)-4-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-methylpiperidin-3-yl)oxy)-4-oxo-succinoyl-CPG having a formula XXIV.

(Formula XXIV)

[32] In an exemplary embodiment, the delivery system 100 includes 33-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-18,18-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-6-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-7-methyl-4,9,16,23,29-pentaoxo-5,20-dioxa-8,17,24,28-tetraazatritriacontan-1-oyl-CPG(Tris-ß-GalNAc-C6-CPG) having a formula XXV.

(Formula XXV)

[33] In an exemplary embodiment, the delivery system 100 includes 33-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-18,18-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-7-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-6-methyl-4,9,16,23,29-pentaoxo-5,20-dioxa-8,17,24,28-tetraazatritriacontan-1-oyl-CPG(Tris-ß-GalNAc-C6-CPG) having a formula XXVI.

(Formula XXVI)

[34] In an exemplary embodiment, the delivery system 100 includes 4-((4-(28-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-13,13-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-4-methylene-8,11,18,24-tetraoxo-2,15-dioxa-7,12,19,23-tetraazaoctacosan-1-oyl)-3-((bis(4-methoxy phenyl)(phenyl)methoxy)methyl)cyclohexyl)oxy)-4-oxo-succinoyl-CPG(Tris-ß-GalNAc-Allylamine-cyclohexyl)oxy)-4-oxo-succinoyl-CPG) having a formula XXVII.

(Formula XXVII)

[35] In an exemplary embodiment, the delivery system 100 includes 4-((2-(28-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-13,13-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-4-methylene-8,11,18,24-tetraoxo-2,15-dioxa-7,12,19,23-tetraazaoctacosan-1-oyl)-5-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-5-methylcyclohexyl)oxy)-4-oxo-succinoyl-CPG(Tris-ß-GalNAc-Allylamine-methylcyclohexyl)oxy)-4-oxo-succinoyl-CPG) having a formula XXVIII.

(Formula XXVIII)

[36] In an exemplary embodiment, the delivery system 100 includes 36-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxy methyl)tetrahydro-2H-pyran-2-yl)oxy)-21,21-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-6-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-7-methyl-12-methylene-4,9,16,19,26,32-hexaoxo-5,10,23-trioxa-8,15,20,27,31-pentaazahexatriacontan-1-oyl-CPG(Tris-ß-GalNAc-Allylamine-6-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-7-methyl-12-methylene-4,9,16,19,26,32-hexaoxo-5,10,23-trioxa-8,15,20,27,31-pentaazahexatriacontan-1-oyl-CPG) having a formula XXIX.

(Formula XXIX)

[37] In an exemplary embodiment, the delivery system 100 includes 36-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-21,21-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-7-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-6-methyl-12-methylene-4,9,16,19,26,32-hexaoxo-5,10,23-trioxa-8,15,20,27,31-pentaazahexatriacontan-1-oyl-CPG(Tris-ß-GalNAc-Allylamine-7-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-6-methyl-12-methylene-4,9,16,19,26,32-hexaoxo-5,10,23-trioxa-8,15,20,27,31-pentaazahexatriacontan-1-oyl-CPG) having a formula XXX.

(Formula XXX)

[38] In an exemplary embodiment, the delivery system 100 includes 4-((1-(1-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-16,16-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-5,11,18-trioxo-14-oxa-6,10,17-triazapentacosan-25-oyl)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)piperidin-4-yl)oxy)-4-oxo-succinoyl-CPG(Tris-ß-GalNAc-C6-piperidin-4-yl)oxy)-4-oxo-succinoyl-CPG) having a formula XXXI.

(Formula XXXI)

[39] In an exemplary embodiment, the delivery system 100 includes 4-((1-(1-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-16,16-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-5,11,18-trioxo-14-oxa-6,10,7-triazapentacosan-25-oyl)-3-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-methylpiperidin-4-yl)oxy)-4-oxo-succinoyl-CPG(Tris-ß-GalNAc-C6-4-methylpiperidin-4-yl)oxy)-4-oxo-succinoyl-CPG) having a formula XXXII.

(Formula XXXII)

[40] In an exemplary embodiment, the delivery system 100 includes 4-((1-(4-((1,25-bis(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacet oxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-13-((3-((6-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)hexyl)amino)-3-oxopropoxy)methyl)-8,18-dioxo-11,15-dioxa-7,19-diazapentacosan-13-yl)amino)-4-oxobutanoyl)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)-piperidin-4-yl)oxy)-4-oxo-succinoyl-CPG(Tris-ß-GalNAc-C2-4-methoxyphenyl)(phenyl)methoxy)piperidin-4-yl)oxy)-4-oxo-succinoyl-CPG) having a formula XXXIII.

(Formula XXXIII)

[41] In an exemplary embodiment, the delivery system 100 includes 4-((1-(4-((1,25-bis(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-13-((3-((6-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)hexyl)amino)-3-oxopropoxy)methyl)-8,18-dioxo-11,15-dioxa-7,19-diazapentacosan-13-yl)amino)-4-oxobutanoyl)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)piperidin-4-yl)oxy)-4-oxobutanoic acid (Tris-ß-GalNAc-C2-4-methoxyphenyl)(phenyl)methoxy)methyl)piperidin-4-yl)oxy)-4-oxo-succinoyl-CPG) having a formula XXXIV.

(Formula XXXIV)

[42] In an exemplary embodiment, the delivery system 100 includes 4-((1-(4-((1,25-bis(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-13-((3-((6-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)hexyl)amino)-3-oxopropoxy)methyl)-8,18-dioxo-11,15-dioxa-7,19-diazapentacosan-13-yl)amino)-4-oxobutanoyl)-4-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-methylpiperidin-3-yl)oxy)-4-oxo-succinoyl-CPG(Tris-ß-GalNAc-C2-4-methoxyphenyl)(phenyl)methoxy)methyl)-4-methylpiperidin-3-yl)oxy)-4-oxo-succinoyl-CPG) having a formula XXXV.

(Formula XXXV)

[43] In an exemplary embodiment, the delivery system 100 includes 26-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxy methyl)tetrahydro-2H-pyran-2-yl)oxy)-14,14-bis((3-((6-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)hexyl)amino)-3-oxopropoxy)methyl)-6-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-7-methyl-4,9,12,19-tetraoxo-5,16-dioxa-8,13,20-triazahexacosan-1-oyl-CPG(Tris-ß-GalNAc-C2-4-methoxyphenyl)(phenyl)methoxy)methyl)-7-methyl-4,9,12,19-tetraoxo-5,16-dioxa-8,13,20-triazahexacosan-1-oyl-CPG) having a formula XXXVI.

(Formula XXXVI)

[44] The delivery system 100 of the present disclosure may be made via one or more biological fermentation and/or by chemical synthesis. In an exemplary embodiment, the delivery system 100 is chemically synthesized by sequentially adding different moieties of the delivery system 100 over a solid support, for example over long chain alkylamine controlled pore glass (LCAA-CPG). Other functionally equivalent synthesis method to prepare the delivery system 100 of the present disclosure is within the scope of the teachings of the present disclosure.
[45] The delivery system 100 of the present disclosure is synthesized by an exemplary method 300 as shown in Fig. 2. Unless stated otherwise, the steps of the method 300 are carried out in dry conditions under an inert environment created using, for example, nitrogen and/or argon gas. The method 300 commences at step 301 by dissolving a pre-defined amount of galactosamine pentaacetate having a formula XXXVII in at least one solvent to obtain a first mixture. A pre-defined amount of trimethylsilyl trifluoromethanesulfonate (TMSOTf) or any other functionally equivalent Lewis acid is added to the first mixture. TMSOTf has high selectivity, efficiency (in activating amidites), and compatibility with sensitive substrates. The pre-defined amount of galactosamine pentaacetate having the formula XXXVII in the first mixture ranges from 12 mM to 13 mM. The solvent includes, but are not limited to, dichloroethane (DCE), dichloromethane (DCM), chloroform, etc. The pre-defined amount of TMSOTf in the first mixture ranges from 15 mM to 16 mM. In an exemplary embodiment, 5g of galactosamine pentaacetate is dissolved in 25mL of DCE at room temperature to obtain the first mixture. Thereafter, 3.5g of TMSOTf is added to the first mixture.
[46] At step 301a, the first mixture obtained from step 301 is maintained at a pre-defined temperature for a pre-defined time period while optionally being subjected to stirring or the like. The pre-defined temperature ranges from 45 °C to 55 °C. The pre-defined time period ranges from 75 minutes to 90 minutes. In an exemplary embodiment, the first mixture is stirred at 50 °C for 90 minutes to maintain reaction homogeneity and efficient reaction conversion.
[47] At an optional step 301b, the first mixture obtained from step 301a is cooled to room temperature while being subjected to stirring or the like for a pre-defined time period. The pre-defined time period ranges from 10 hours to 12 hours. In an exemplary embodiment, the first mixture is cooled to room temperature and stirred for 12 hours for better reaction conversion.
[48] At step 301c, the first mixture obtained from either step 301a or 301b is added to an ice-cold (to prevent thermal degradation of the first mixture) sodium bicarbonate (NaHCO3) solution or any other functionally equivalent weak base for mild neutralization of the first mixture, subjected to solvent extraction with dichloromethane (DCM), and then washed with water and dried over sodium sulphate (Na2SO4) or magnesium sulphate (Mg2SO4) to remove moisture. Instead of DCM used for solvent extraction, other functionally equivalent solvents like chloroform, ethyl acetate (EtOAc), toluene, hexane, methyl tert-butyl ether (MTBE), acetone, tetrahydrofuran (THF), or diethyl ether may be used depending upon polarity, density, boiling point, reactivity, and safety of the solvent.
[49] At step 301d, the first mixture obtained from step 301c is filtered to remove the solvent(s). A residue obtained from filtering the first mixture is dried overnight under high vacuum to obtain a viscous solid having a formula XXXVIII.

(Formula XXXVII) (Formula XXXVIII)

[50] At step 303, a pre-defined amount of the viscous solid having the formula XXXVIII obtained from step 301d, a pre-defined amount of at least one benzyl ester, and a pre-defined amount of at least one molecular-sieve are dissolved in at least one solvent to obtain a second mixture. In an exemplary embodiment, the benzyl ester has a formula XXXIX to block carbolic acids (or groups thereof). In an exemplary embodiment, the second mixture is prepared by stirring the second mixture for 30 minutes. The pre-defined amount of the viscous solid having the formula XXXVIII in the second mixture ranges from 11.50 mM to 12 mM. The pre-defined amount of benzyl ester having the formula XXXIX in the second mixture ranges from 17.54 mM to 18 mM. The pre-defined amount of molecular-sieve in the second mixture ranges from 1 gm to 2 gm. The molecular-sieve include type A zeolites or the like including, but not limited to, one of molecular sieve 3A, molecular sieve 4A, molecular sieve 5A, etc. In an exemplary embodiment, the molecular-sieve includes molecular sieve 4A having a pore size of 4 ?. The solvent includes, but are not limited to, dichloromethane (DCM), chloroform, etc. In an exemplary embodiment, 3.9 g of the viscous solid having the formula XXXVIII, 3.6 g of the benzyl ester having the formula XXXIX, and 1 g of molecular-sieve having pore size of 4 ? are dissolved in 25 mL of DCM to obtain the second mixture.
[51] At step 303a, a pre-defined amount of trimethylsilyl trifluoromethanesulfonate (TMSOTf) or any other functionally equivalent Lewis acid is added to the second mixture obtained from step 303. TMSOTf has high selectivity, efficiency (in activating amidites), and compatibility with sensitive substrates. The second mixture is subjected to stirring or the like for a pre-defined amount of time period. The pre-defined time period ranges from 10 hours to 12 hours. The pre-defined amount of TMSOTf in the second mixture ranges from 4.5 mM to 5 mM. In an exemplary embodiment, 1 g of TMSOTf is added to the second mixture and stirred for 12 hours.
[52] At step 303b, the second mixture obtained from step 303a is added to an ice-cold (to prevent thermal degradation of the second mixture) sodium bicarbonate (NaHCO3) solution or any other functionally equivalent weak base for mild neutralization of the second mixture, subjected to solvent extraction with dichloromethane (DCM), and then washed with water and dried over sodium sulphate (Na2SO4) or magnesium sulphate (Mg2SO4) to remove moisture at 40 °C and 100-120 mbar pressure. Instead of DCM used for solvent extraction, other functionally equivalent solvents like chloroform, ethyl acetate (EtOAc), toluene, hexane, methyl tert-butyl ether (MTBE), acetone, tetrahydrofuran (THF), or diethyl ether may be used depending upon polarity, density, boiling point, reactivity, and safety of the solvent.
[53] At step 303c, the second mixture obtained from step 303b is filtered or decanted to remove the solvent(s). A residue obtained from filtering the second mixture is further subjected to purification by column chromatography to yield a compound having a formula XXXX. Instead of column chromatography, flash chromatography, crystallization techniques or the like may be used. In an exemplary embodiment, the residue obtained from filtering the second mixture is eluted with 20% to 100% ethyl acetate in hexanes through a column containing silica gel.

(Formula XXXVIII) (Formula XXIX) (Formula XXXX)

[54] At step 305, a pre-defined amount of the compound having the formula XXXX obtained from step 303c is suspended in at least one solvent along with a pre-defined amount of 10% (w/w) palladium/carbon (Pd/C) mixture (used for hydrogenation) to obtain a first suspension. Instead of the Pd/C mixture for hydrogenation, other functionally equivalent catalyst like platinum/carbon (Pt/C), Raney nickel, homogenous palladium, or the like may be used depending upon reaction type, selectivity, cost, and environmental impact. The pre-defined amount of the compound having the formula XXXX in the first suspension ranges from 8.37 mM to 4 mM. The solvent is at least one of methyl alcohol, ethyl acetate (EtOAc), dichloromethane (DCM), or a combination thereof. The pre-defined amount of the Pd/C mixture in the first suspension ranges from 8 % (w/w) to 10 % (w/w). In an exemplary embodiment, 4.5 g of the compound having the formula XXXX and 0.45 g of the Pd/C mixture are suspended in 2mL of methyl alcohol and 20mL of EtOAc to obtain the first suspension.
[55] At step 305a, the first suspension obtained from step 305 is flushed with hydrogen gas for a pre-defined time period to cleave the benzyl chloroformate (CBz) protecting group from the compound having the formula XXXX. The pre-defined time period ranges from 12 hours to 16 hours. In an exemplary embodiment, the first suspension is flushed with hydrogen gas and then hydrogenated under balloon pressure for 16 hours.
[56] At step 305b, the first suspension obtained from step 305a is subjected to filtration or the like. In an exemplary embodiment, the first suspension is filtered through a celite column and then the filter bed of the celite column is washed with methanol. The filtrate obtained therefrom is combined and concentrated under reduced pressure to yield 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having a formula XXXXI, i.e., the ligand 150.

(Formula XXXX) (Formula XXXXI)

[57] At step 307, a pre-defined amount of 3,3'-((2-(((Benzyloxy)carbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy))dipropionic acid (Tris-base) having a formula XXXXII, a pre-defined amount of mono-Boc protected 1,3- diaminopropane having a formula XXXXIII or any other functionally equivalent amine protecting groups like fluorenylmethoxycarbonyl (Fmoc), benzyl chloroformate (CBz), and/or acyl groups, a pre-defined amount of Hexafluorophosphate Benzotriazole Tetramethyl Uronium (HBTU) or any other functionally equivalent coupling reagent or the like to impede racemization and a pre-defined amount of N,N-diisopropylethylamine (DIEA) or any other functionally equivalent mild, non-nucleophilic base for selective protonation/base-mediated activation (without nucleophilic side reactions) are dissolved in at least one solvent to obtain a third mixture. The pre-defined amount of 3,3'-((2-(((Benzyloxy)carbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy))dipropionic acid (Tris-base) having the formula XXXXII in the third mixture ranges from 11.70 mM to 12 mM. The pre-defined amount of mono-Boc protected 1,3-diaminopropane having the formula XXXXIII in the third mixture ranges from 30 mM to 40 mM. The pre-defined amount of HBTU in the third mixture ranges from 39.55 mM to 40 mM. The pre-defined amount of DIEA in the third mixture ranges from 5 mL to 10 mL. The solvent is at least one of dimethylformamide (DMF), dimethylsulfoxide (DMSO), etc. In an exemplary embodiment, 5 g of 3,3'-((2-(((Benzyloxy)carbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy))dipropionic acid (Tris-base) having the formula XXXXII, 3.5 mol equivalent mono-Boc protected 1,3- diaminopropane having the formula XXXXIII, 10 g of HBTU, and 5 mL of DIEA are added to 30 mL of DMF to obtain the third mixture.
[58] At an optional step 307a, the third mixture obtained from step 307 is subjected to stirring or the like for a pre-defined time period. In an exemplary embodiment, the third mixture is stirred overnight.
[59] At step 307b, the third mixture either obtained from step 307 or 307a is added to ice-cold water (to prevent thermal degradation of the third mixture), subjected to solvent extraction with dichloromethane (DCM), and then washed with saturated sodium bicarbonate (NaHCO3) solution or any other functionally equivalent weak base for mild neutralization of the third mixture and/or brine to break emulsions and dry organic layers by reducing water solubility, and dried over sodium sulphate (Na2SO4) or magnesium sulphate (Mg2SO4) to remove moisture. Instead of DCM used for solvent extraction, other functionally equivalent solvents like chloroform, ethyl acetate (EtOAc), toluene, hexane, methyl tert-butyl ether (MTBE), acetone, tetrahydrofuran (THF), or diethyl ether may be used depending upon polarity, density, boiling point, reactivity, and safety of the solvent.
[60] At step 307c, the third mixture obtained from step 307b is filtered to remove the solvent(s). The filtrate is combined and concentrated under reduced pressure and a residue obtained therefrom is further subjected to purification by column chromatography or flash chromatography to yield a compound having a formula XXXXIV. In an exemplary embodiment, the residue obtained from filtering the third mixture is eluted with ethyl acetate followed by 2% to 10% methanol in dichloromethane (DCM) through a column containing silica gel.

(Formula XXXXII) (Formula XXXXIII) (Formula XXXXIV)

[61] At step 309, a pre-defined amount of the compound having the formula XXXXIV obtained from step 307c is dissolved in a pre-defined amount of Trifluoroacetic acid (TFA) (or methanesulfonic acid) to obtain a fourth mixture. TFA helps to remove tert-butoxycarbonyl (Boc) protecting group from the compound having the formula XXXXIV. The pre-defined amount of compound having the formula XXXXIV in the fourth mixture ranges from 3.81 mM to 4 mM. In an exemplary embodiment, 3.58 g the compound having the formula XXXXIV is dissolved in 15 mL of TFA to obtain the fourth mixture.
[62] At an optional step 309a, the fourth mixture is subjected to stirring or the like for a pre-defined time period. In an exemplary embodiment, the fourth mixture is subjected to stirring at room temperature for 35 minutes.
[63] At step 309b, the fourth mixture obtained from either step 309 or 309a is diluted with a pre-defined amount of at least one solvent and concentrated under reduced pressure to yield a compound having a formula XXXXV. The solvent may be at least one of toluene, hexane, heptane, xylene etc. In an exemplary embodiment, the fourth mixture is diluted with 75-100 mL of toluene and concentrated (dried) under reduced pressure using a high vacuum pump to yield TFA salt of the compound having the formula XXXXV.

(Formula XXXXIV) (Formula XXXXV)

[64] At step 311, a pre-defined amount of 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having the formula XXXXI as obtained from step 305b, a pre-defined amount of Hexafluorophosphate Benzotriazole Tetramethyl Uronium (HBTU) or any other functionally equivalent coupling reagent or the like to impede racemization, and a pre-defined amount of N,N-diisopropylethylamine (DIEA) or any other functionally equivalent mild, non-nucleophilic base for selective protonation/base-mediated activation (without nucleophilic side reactions) are dissolved in at least one solvent to obtain a fifth mixture. The pre-defined amount of 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having the formula XXXXI in the fifth mixture ranges from 7.40 mM to 8 mM. The pre-defined amount of HBTU in the fifth mixture ranges from 16.40 mM to 17.00 mM. The pre-defined amount of DIEA in the fifth mixture ranges from 4 mL to 8 mL. The solvent is at least one of dimethylformamide (DMF), dimethylsulfoxide (DMSO), etc. In an exemplary embodiment, 3.31 g of 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having the formula XXXXI, 3.31 g of HBTU, and 4 mL of DIEA are added to 15 mL DMF to obtain the fifth mixture.
[65] At an optional step 311a, the fifth mixture obtained from step 311 is subjected to stirring or the like for a pre-defined time period. In an exemplary embodiment, the fifth mixture is stirred for 5 minutes.
[66] At step 311b, a pre-defined amount of the compound having the formula XXXXV obtained from step 309b and dissolved in at least one solvent is added to the fifth mixture obtained either from step 311 or 311a. The pre-defined amount of the compound having the formula XXXXV ranges from 3.96 mM to 4 mM. The solvent is at least one of dimethylformamide (DMF), dimethylsulfoxide (DMSO), etc. In an exemplary embodiment, 2.5 g of the compound having the formula XXXXV is dissolved in 25 mL of DMF and added to the fifth mixture.
[67] At an optional step 311c, the fifth mixture obtained from step 311b is subjected to stirring or the like for a pre-defined time period. The pre-defined time period ranges from 15 hours to 18 hours. In an exemplary embodiment, the fifth mixture is stirred at room temperature for 18 hours.
[68] At step 311d, the fifth mixture obtained either from step 311b or 311c is subjected to reduced pressure to remove the solvent(s). In an exemplary embodiment, a residue obtained after removing the solvents from the fifth mixture is dissolved in 50mL of dichloromethane (DCM), washed with 50 mL of saturated aqueous sodium bicarbonate (NaHCO3) solution or any other functionally equivalent weak base for mild neutralization of the residue, 50 mL of water and 50 mL of brine to break emulsions and dry organic layers by reducing water solubility, and dried over sodium sulphate (Na2SO4) or magnesium sulphate (Mg2SO4) to remove moisture. Instead of DCM used for solvent extraction, other functionally equivalent solvents like chloroform may be used.
[69] At step 311e, the fifth mixture obtained from step 311d is filtered and subjected to reduced pressure to remove the solvent(s). A residue obtained therefrom is further subjected to purification by column chromatography (or flash chromatography) to yield a compound having a formula XXXXVI. In an exemplary embodiment, the residue obtained from filtering the fifth mixture is eluted with ethyl acetate followed by gradient elution from 5% to 25% methanol in dichloromethane (DCM) through a column containing silica gel.

(Formula XXXXV) (Formula XXXXI) (Formula XXXXVI)

[70] At step 313, the linker 130 is prepared. The linker 130 is at least one of the formula XV, XVI, XVII, and XVIII.
[71] In an exemplary embodiment, the linker 130 having the formula XV and the formula XVI are prepared by dissolving 50 g of a compound having a formula XXXXVII in 1.5 L of dry Tetrahydrofuran (THF) or any other functionally equivalent polar aprotic solvent to obtain a first reaction mixture. The first reaction mixture is cooled in an ice bath at approximately 5 °C. Thereafter, 36.18 g of Borane-dimethylsulfide (BH3·Me2S) or any other functionally equivalent reducing agent is added drop-wise to the first reaction mixture over a period of 20-25 minutes. After addition of BH3·Me2S in the first reaction mixture, the first reaction mixture is heated to 60 °C for 4-5 hours. After heating the first reaction mixture, the first reaction mixture is quenched in an ice bath by adding 50 mL of water until no more gas is released from the first reaction mixture. After quenching the first reaction mixture, the first reaction mixture is neutralized by adding 50 mL of 2N sodium hydroxide (NaOH) while maintaining the first reaction mixture at 0 °C. The first reaction mixture is concentrated by reducing the volume of the first reaction mixture to half its initial volume and subjected to solvent extraction using 200 mL of chloroform to obtain an organic layer. Instead of chloroform used for solvent extraction, other functionally equivalent solvents like dichloromethane (DCM) may be used. The organic layer is washed with 250 mL of aqueous saturated sodium bicarbonate (NaHCO3) solution or any other functionally equivalent weak base for mild neutralization of the organic layer, and 250 mL of brine to break emulsions and dry organic layers by reducing water solubility, dried over magnesium sulphate (Mg2SO4), filtered and evaporated. A residue obtained therefrom is suspended in ethyl acetate (EtOAc) and filtered to obtain a compound having a formula XXXXVIII. 35 g of the compound having the formula XXXXVIII in anhydrous pyridine is gradually added to 36.91 g of 4,4’-dimethoxytrityl chloride (DMTrCl) or any other functionally equivalent protecting group over a period of 15 minute at room temperature to obtain a second reaction mixture. The second reaction mixture is stirred for 4-5 hours at room temperature. Thereafter, the second reaction mixture is concentrated by reducing the volume of the second reaction mixture to half its initial volume under reduced pressure and then diluted with 200 mL of ethyl acetate (EtOAc) or any other functionally equivalent polar aprotic solvent. The second reaction mixture is then subjected to solvent extraction using 200 mL of chloroform to obtain an organic layer. Instead of chloroform used for solvent extraction, other functionally equivalent solvents like dichloromethane (DCM) may be used. The organic layer is washed with 2 x 200 mL of aqueous saturated sodium bicarbonate (NaHCO3) solution or any other functionally equivalent weak base for mild neutralization of the organic layer, and 200 mL of saturated brine to break emulsions and dry organic layers by reducing water solubility, dried over sodium sulphate (Na2SO4) or magnesium sulphate (Mg2SO4) to remove moisture, filtered and evaporated to yield a compound having a formula XXXXIX. 40 g of the compound having the formula XXXXIX is dissolved in 250 mL of Dimethylformamide (DMF) and 25 mL piperidine to cleave the fluorenylmethoxycarbonyl (Fmoc) group from the compound having the formula XXXXIX and obtain a third reaction mixture. The third reaction mixture is stirred for 3-4 hours at room temperature. The third reaction mixture is then subjected to solvent extraction using 250 mL of chloroform to obtain an organic layer. Instead of chloroform used for solvent extraction, other functionally equivalent solvents like dichloromethane (DCM) may be used. The organic layer is washed with 2 x 250 mL of aqueous saturated sodium bicarbonate (NaHCO3) solution or any other functionally equivalent weak base for mild neutralization of the organic layer, and 250 mL of brine to break emulsions and dry organic layers by reducing water solubility, dried over sodium sulphate (Na2SO4) or magnesium sulphate (Mg2SO4) to remove moisture, filtered and evaporated under reduced pressure, and purified by column chromatography (silica gel column, eluted with 0% to 5% methanol in dichloromethane) to yield a compound having a formula L.

(Formula XXXXVII) (Formula XXXXVIII) (Formula XXXXIX) (Formula L)

[72] To prepare the linker 130 having the formula XV, 5 g of 8-methoxy-8-oxooctanoic acid having a formula LI, 10 g or 10mL of Hexafluorophosphate Benzotriazole Tetramethyl Uronium (HBTU) or any other functionally equivalent coupling reagent or the like to impede racemization, and 10 mL of N, N-diisopropylethylamine (DIEA) or any other functionally equivalent mild, non-nucleophilic base for selective protonation/base-mediated activation (without nucleophilic side reactions) are dissolved in 75 mL of dimethylformamide (DMF) to obtain a fourth reaction mixture. The fourth reaction is stirred for 5 minutes at room temperature. 11 g of the compound having the formula L in 50 mL of anhydrous DMF is added in the fourth reaction mixture and stirred for 18 hours at room temperature. Thereafter, the reaction mixture is added to ice water and subjected to solvent extraction using 300 mL of dichloromethane (DCM) to obtain an organic layer. Instead of DCM used for solvent extraction, other functionally equivalent solvents like chloroform may be used. The organic layer is washed with 250 mL of aqueous saturated sodium bicarbonate (NaHCO3) solution or any other functionally equivalent weak base for mild neutralization of the organic layer, and 150 mL of brine to break emulsions and dry organic layers by reducing water solubility, dried over sodium sulphate (Na2SO4) or magnesium sulphate (Mg2SO4) to remove moisture, filtered and evaporated under reduced pressure, and purified by column chromatography (silica gel column, eluted with 20% to 100% ethyl acetate in hexane) to yield a compound having a formula LII. 2 g of the compound having the formula LII, and 0.41 g lithium hydroxide (LiOH) or any other strong base used in organic transformations are dissolved in 25 mL of dioxane (or a mixture of 10 mL methanol, 5 mL Tetrahydrofuran (THF), 5.5 mL water) to obtain a fifth reaction mixture. The fifth reaction mixture is stirred for 1-2 hours at room temperature. The solvent(s) is removed from the fifth reaction mixture under reduced pressure and then diluted by adding 10 mL of dichloromethane to obtain a slurry. The slurry is purified with column chromatography (silica gel column, eluted with 0% to 5% methanol in dichloromethane) to yield 8-((2R,4R)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-hydroxypiperidin-1-yl)-8-oxooctanoic acid (linker 130) having the formula XV.

(Formula L) (Formula LI) (Formula LII) (Formula XV)

[73] To prepare the linker 130 having fthe ormula XVI, 5 g of 4-methoxy-4-oxobutanoic acid having a formula LIII, 1.2 mol equivalent of Hexafluorophosphate Benzotriazole Tetramethyl Uronium (HBTU) or any other functionally equivalent coupling reagent or the like to impede racemization, and 10 mL of N, N-diisopropylethylamine (DIEA) or any other functionally equivalent mild, non-nucleophilic base for selective protonation/base-mediated activation (without nucleophilic side reactions) are dissolved in 75 mL of Dimethylformamide (DMF) to obtain a fourth reaction mixture. The fourth reaction is stirred for 5 minutes at room temperature. 10 g of the compound having the formula L in 50 mL of anhydrous DMF is added in the fourth reaction mixture and stirred for 18 hours at room temperature. Thereafter, the reaction mixture is added to ice water and subjected to solvent extraction using 300-350 mL of dichloromethane (DCM) to obtain an organic layer. Instead of DCM used for solvent extraction, other functionally equivalent solvents like chloroform may be used. The organic layer is washed with 150 mL of aqueous saturated sodium bicarbonate (NaHCO3) solution or any other functionally equivalent weak base for mild neutralization of the organic layer, and 150 mL of brine to break emulsions and dry organic layers by reducing water solubility, dried over sodium sulphate (Na2SO4) or magnesium sulphate (Mg2SO4) to remove moisture, filtered and evaporated under reduced pressure, and purified by column chromatography (silica gel column, eluted with 20% to 100% ethyl acetate in hexane) to yield a compound having a formula LIV. 2.5 g of the compound having the formula LIV, and 0.5 g lithium hydroxide (LiOH) or any other strong base used in organic transformations are dissolved in 25 mL dioxane (or a mixture of 10 mL methanol, 5 mL Tetrahydrofuran (THF), 5.5 mL water) to obtain a fifth reaction mixture. The fifth reaction mixture is stirred for 1-2 hours at room temperature. The solvent(s) is removed from the fifth reaction mixture under reduced pressure and then diluted by adding 10 mL of dichloromethane to obtain a slurry. The slurry is purified with column chromatography (silica gel column, eluted with 0% to 5% methanol in dichloromethane) to yield 4-((2R,4R)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-hydroxypiperidin-1-yl)-4-oxobutanoic acid (linker 130) having the formula XVI.

(Formula L) (Formula LIII) (Formula LIV) (Formula XVI)

[74] In an exemplary embodiment, the linker 130 having the formula XVII is prepared by dissolving 10 g of tert-butyl (3,4-dihydroxybutan-2-yl)carbamate having a formula LV is dissolved in 100 mL anhydrous pyridine to obtain a sixth reaction mixture. 16.51 g of 4,4’-dimethoxytrityl chloride (DMTrCl) or any other functionally equivalent protecting group is gradually added to the sixth reaction mixture over a period of 5-10 minutes at room temperature. Thereafter, the sixth reaction mixture is stirred for 4-5 hours at room temperature and concentrated by reducing the volume of the sixth reaction mixture to half its initial volume under reduced pressure. The sixth reaction mixture is then diluted with 100 mL ethyl acetate (EtOAc) and 100 mL water. The sixth reaction mixture is then subjected to solvent extraction using 250 mL of ethyl acetate (EtOAc) and 100 mL water to obtain an organic layer. The organic layer is washed twice with 150 mL of aqueous saturated sodium bicarbonate (NaHCO3) solution or any other functionally equivalent weak base for mild neutralization of the organic layer, and once with 100 mL of saturated brine to break emulsions and dry organic layers by reducing water solubility, dried over sodium sulphate (Na2SO4) or magnesium sulphate (Mg2SO4) to remove moisture, filtered and evaporated under reduced pressure to yield a compound having a formula LVI. The compound having the formula LVI is treated with 1.2 mol equivalent of trifluoroacetic acid (TFA) diluted with 100 mL of Toluene to deprotect the tert-butyloxycarbonyl (Boc) group and yield a compound having a formula LVII.

(Formula LV) (Formula LVI) (Formula LVII)

[75] 5.46 g of 9-methoxy-9-oxononanoic acid having a formula LVIII is dissolved in 100 mL of dimethylformamide (DMF) to obtain a seventh reaction mixture. 1.2 mol equivalent of Hexafluorophosphate Benzotriazole Tetramethyl Uronium (HBTU) or any other functionally equivalent coupling reagent or the like to impede racemization and 10 mL of N,N-diisopropylethylamine (DIEA) or any other functionally equivalent mild, non-nucleophilic base for selective protonation/base-mediated activation (without nucleophilic side reactions) is added to the seventh reaction mixture and stirred for 5 minutes at room temperature. 10 g of the compound having the formula LVII is dissolved in 15 mL of anhydrous DMF and added to the seventh reaction mixture. The seventh reaction mixture is stirred for overnight at room temperature and poured in ice-cold water. The seventh reaction mixture is then subjected to solvent extraction using 250 mL of dichloromethane (DCM) to obtain an organic layer. Instead of DCM used for solvent extraction, other functionally equivalent solvents like chloroform may be used. The organic layer is washed with 150 mL of aqueous saturated sodium bicarbonate (NaHCO3) solution or any other functionally equivalent weak base for mild neutralization of the organic layer, and 100 mL of brine to break emulsions and dry organic layers by reducing water solubility, dried over sodium sulphate (Na2SO4) or magnesium sulphate (Mg2SO4) to remove moisture, filtered and evaporated under reduced pressure, and purified by column chromatography (silica gel column, eluted with 10% to 100% ethyl acetate in hexane) to yield a compound having a formula LIX. 5 g of the compound having the formula LIX, and 9 g lithium hydroxide (LiOH) or any other strong base used in organic transformations are dissolved in a mixture of 25 mL methanol, 10 mL tetrahydrofuran (THF), 10 mL water to obtain an eighth reaction mixture. The eighth reaction mixture is stirred for 1-2 hours at room temperature. The solvent(s) is removed from the eighth reaction mixture under reduced pressure and then diluted by adding 50 mL of dichloromethane (DCM) to obtain a slurry. The slurry is purified with column chromatography (silica gel column, eluted with 0% to 10% methanol in dichloromethane) to yield 8-((4-(bis(4-methoxyphenyl)(phenyl)methoxy)-3-hydroxybutan-2-yl)amino)-8-oxooctanoic acid (linker 130) having the formula XVII.

(Formula LVII) (Formula LVIII) (Formula LIX) (Formula XVII)

[76] In an exemplary embodiment, the linker 130 having the formula XVIII is prepared by dissolving 10 g of a compound having a formula LX in 100 mL of anhydrous pyridine to obtain a ninth reaction mixture. 15.19 g is gradually added to the ninth reaction mixture over a period of 10-15 minutes at room temperature. Thereafter, the ninth reaction mixture is stirred for 4-5 hours at room temperature and concentrated by reducing the volume of the ninth reaction mixture to half of its initial volume under reduced pressure. The ninth reaction mixture is then diluted with 150 mL of ethyl acetate (EtOAc) and 150 mL of water. The ninth reaction mixture is then subjected to solvent extraction using 200 mL ethyl acetate (EtOAc) to obtain an organic layer. The organic layer is washed twice with 150 mL of aqueous saturated sodium bicarbonate (NaHCO3) solution or any other functionally equivalent weak base for mild neutralization of the organic layer, and once with 150 mL of saturated brine to break emulsions and dry organic layers by reducing water solubility, dried over sodium sulphate (Na2SO4) or magnesium sulphate (Mg2SO4) to remove moisture, filtered and evaporated to yield a compound having a formula LXI. The compound having the formula LXI is treated with 1.5 mol equivalent of trifluoroacetic acid (TFA) diluted with 75 mL of toluene to deprotect the tert-butyloxycarbonyl (Boc) group and yield a compound having a formula LXII.

(Formula LX) (Formula LXI) (Formula LXII)

[77] 4.72 g of 3-(3-methoxy-3-oxopropoxy)propanoic acid having a formula LXIII is dissolved in 75 mL of dimethylformamide (DMF) to obtain a tenth reaction mixture. 1.2 mol equivalent of Hexafluorophosphate Benzotriazole Tetramethyl Uronium (HBTU) or any other functionally equivalent coupling reagent or the like to impede racemization and 10 mL of N,N-diisopropylethylamine (DIEA) or any other functionally equivalent mild, non-nucleophilic base for selective protonation/base-mediated activation (without nucleophilic side reactions) is added to the seventh reaction mixture and stirred for 5 minutes at room temperature. 10 g of the compound having the formula LXII is dissolved in 15 mL of anhydrous DMF and added to the tenth reaction mixture. The tenth reaction mixture is stirred for overnight at room temperature and poured in ice-cold water. The tenth reaction mixture is then subjected to solvent extraction using 350-400 mL of dichloromethane (DCM) to obtain an organic layer. Instead of DCM used for solvent extraction, other functionally equivalent solvents like chloroform may be used. The organic layer is washed with 200 mL of aqueous saturated sodium bicarbonate (NaHCO3) solution or any other functionally equivalent weak base for mild neutralization of the organic layer, and 100 mL of brine to break emulsions and dry organic layers by reducing water solubility, dried over sodium sulphate (Na2SO4) or magnesium sulphate (Mg2SO4) to remove moisture, filtered and evaporated under reduced pressure, and purified by column chromatography (silica gel column, eluted with 10% to 100% ethyl acetate in hexane) to yield a compound having a formula LXIV. 5 g of the compound having the formula LXIV, and 1.78 g lithium hydroxide (LiOH) or any other strong base used in organic transformations are dissolved in 25 mL dioxane (or a mixture of 20 mL methanol, 5 mL tetrahydrofuran (THF), 10 mL water) to obtain an eleventh reaction mixture. The eleventh reaction mixture is stirred for 1-2 hours at room temperature. The solvent(s) is removed from the eleventh reaction mixture under reduced pressure and then diluted by adding 50 mL of dichloromethane (DCM) to obtain a slurry. The slurry is purified with column chromatography (silica gel column, eluted with 0% to 10% methanol in dichloromethane) to yield 3-(3-(4-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-3-hydroxy-4-methylpiperidin-1-yl)-3-oxopropoxy)propanoic acid (linker 130) having the formula XVIII.

(Formula LXII) (Formula LXIII) (Formula LXIV) (Formula XVIII)

[78] At step 315, a pre-defined amount of the compound having the formula XXXXVI obtained from step 311e is dissolved in at least one solvent along with a pre-defined amount of 10% (w/w) palladium/carbon (Pd/C) mixture (used for hydrogenation) to obtain a sixth mixture. Instead of the Pd/C mixture for hydrogenation, other functionally equivalent catalyst like platinum/carbon (Pt/C), Raney nickel, homogenous palladium, or the like may be used depending upon reaction type, selectivity, cost, and environmental impact. The pre-defined amount of the compound having the formula XXXXVI in the sixth mixture ranges from 2.65 mM to 3.00 mM. The solvent is at least one of methanol, ethanol, or a combination thereof. The pre-defined amount of the Pd/C mixture in the sixth mixture ranges from 8% (w/w) to 10% (w/w). In an exemplary embodiment, 5 g of the compound having the formula XXXXVI is dissolved in 30 mL of methanol to obtain the sixth mixture. 10% (w/w) of the Pd/C mixture is then added to the sixth mixture.
[79] At step 315a, the sixth mixture obtained from step 315 is flushed with hydrogen gas for a pre-defined time period to cleave the benzyl chloroformate (CBz) protecting group from the compound having the formula XXXXVI. The pre-defined time period ranges from 10 hours to 12 hours. In an exemplary embodiment, the sixth mixture is flushed with hydrogen gas and then hydrogenated under balloon pressure for 12 hours.
[80] At step 315b, the sixth mixture obtained from step 315a is subjected to filtration or the like. In an exemplary embodiment, the sixth mixture is filtered through a celite column and then the filter bed of the celite column is washed with methanol.
[81] At step 315c, a pre-defined amount of trifluoroacetic acid (TFA) (or methanesulfonic acid) is added to the sixth mixture obtained from step 315b. TFA helps to remove tert-butoxycarbonyl (Boc) protecting group from the compound having the formula XXXXVI. The pre-defined amount of TFA in the sixth mixture ranges from 0.5 ml to 1 ml. In an exemplary embodiment, 0.5 mL of TFA is added to the sixth mixture.
[82] At step 315d, the solvent(s) from the sixth mixture is removed and a residue obtained therefrom is evaporated (optionally co-evaporated with a solvent) and dried for a pre-defined time period to yield a compound having a formula LXV. In an exemplary embodiment, the solvents of the sixth mixture are removed under reduced pressure. The residue obtained therefrom is co-evaporated 2-3 times with toluene and dried under high vacuum for overnight to yield a TFA salt of the compound having the formula LXV.

(Formula XXXXVI)
(Formula LXV)

[83] At step 317, a pre-defined amount of the linker 130 having either of the formula XV, XVI, XVII, or XVIII as obtained from step 313 is treated with a pre-defined amount of Hexafluorophosphate Benzotriazole Tetramethyl Uronium (HBTU) or any other functionally equivalent coupling reagent or the like to impede racemization dissolved in at least one solvent to obtain a seventh mixture. A pre-defined amount of N,N-diisopropylethylamine (DIEA) or any other functionally equivalent mild, non-nucleophilic base for selective protonation/base-mediated activation (without nucleophilic side reactions) to the seventh reaction mixture and stirred or the like for a pre-defined time period. The pre-defined time period ranges from 10 minutes to 15 minutes. The solvent is at least one of dimethylformamide (DMF), dimethylsulfoxide (DMSO), etc. The pre-defined amount of linker 130 in the seventh mixture ranges from 0.5 mM to 1 mM. The pre-defined amount of HBTU in the seventh mixture ranges from 1.5 mM to 3 mM. The pre-defined amount of DIEA in the seventh mixture ranges from 2.5 mM to 3.5 mM. In an exemplary embodiment, 1 mol equivalent of the linker 130 is treated with 1.5 mol equivalent HBTU in 25 mL of anhydrous DMF. 3 mol equivalent of DIEA is added to the seventh mixture and stirred for 10-15 minutes.
[84] At step 317a, a pre-defined amount of the compound having the formula LXV as obtained from step 315d dissolved in at least one solvent is added to the seventh mixture and stirred or the like for a pre-defined time period. The solvent is at least one of dimethylformamide (DMF), dimethylsulfoxide (DMSO), etc. In an exemplary embodiment, 1 mol equivalent of the compound having the formula LXV dissolved in 30 mL of DMF is added to the seventh mixture and stirred at room temperature for overnight.
[85] At step 317b, the seventh mixture obtained from step 317a is subjected to solvent extraction, filtration, etc. to yield a compound having either one of a formula LXVI, LXVII, LXVIII, or LXIX corresponding to the linker 130 having the formula XV, XVI, XVII, or XVIII used in step 317. In an exemplary embodiment, the seventh mixture is subjected to solvent extraction using water and 500 mL of dichloromethane (DCM), dried over sodium sulphate (Na2SO4) or magnesium sulphate (Mg2SO4) to remove moisture, filtered and concentrated under reduced pressure. A residue obtained therefrom is purified by column chromatography (silica gel column, eluted with 0% to 25% methanol in dichloromethane) to yield a compound having either one of the formula LXVI, LXVII, LXVIII, or LXIX. Instead of DCM used for solvent extraction, other functionally equivalent solvents like chloroform, ethyl acetate (EtOAc), toluene, hexane, methyl tert-butyl ether (MTBE), acetone, tetrahydrofuran (THF), or diethyl ether may be used depending upon polarity, density, boiling point, reactivity, and safety of the solvent.

(Formula LXV)
(Formula LXVI)
(Formula LXV)
(Formula LXVII)
(Formula LXV)
(Formula LXVIII)
(Formula LXV)
(Formula LXIX)

[86] At step 319, a pre-defined amount of the compound having either one of the formula LXVI, LXVII, LXVIII, or LXIX as obtained from step 317b is treated with a pre-defined amount of succinic anhydride and 4-dimethylaminopyridine (DMAP) dissolved in at least one solvent to obtain an eighth mixture. Succinic anhydride reacts with hydroxyl or amine groups to form succinylated derivatives through acylation reaction using DMAP as a catalyst. The eight mixture is subjected to stirring or the like for a pre-defined time period. The solvent is at least one of dichloromethane (DCM), dichloroethane (DCE), etc. The pre-defined amount of the compound having either one of the formula LXVI, LXVII, LXVIII, or LXIX in the eighth mixture ranges from 1 mM to 2 mM. The pre-defined amount of succinic anhydride in the eighth mixture ranges from 2 mM to 3 mM. The pre-defined amount of DMAP in the eighth mixture ranges from 2.5 mM to 3.5 mM. In an exemplary embodiment, 2 g (1 mol equivalent) of the compound having either one of the formula LXVI, LXVII, LXVIII, or LXIX is treated with 2 mol equivalent succinic anhydride and 3 mol equivalent DMAP dissolved in 50 mL DCM to obtain the eighth mixture. The eighth mixture is stirred at room temperature for overnight.
[87] At step 319a, the eighth mixture as obtained from step 319 is diluted with at least one solvent, washed, dried, and filtered to yield a compound having either one of a formula LXX, LXXI, LXXII, or LXXIII corresponding to the compound having either one of the formula LXVI, LXVII, LXVIII, or LXIX used in step 319. The solvent is at least one of dichloromethane (DCM), dichloroethane (DCE), etc. In an exemplary embodiment, the eighth mixture is diluted with 150 mL DCM, washed with 250 mL water and 250 mL of cold 10% citric acid solution (as a neutralizer). An organic layer obtained therefrom is dried over sodium sulphate (Na2SO4) or magnesium sulphate (Mg2SO4) to remove moisture, concentrated under reduced pressure, and filtered through a small pad of silica gel to yield succinate derivative of a compound having either one of the formula LXX, LXXI, LXXII, or LXXIII. Instead of neutralizing the eight mixture with 10% citric acid solution, acetic acid, phosphoric acid, and/or ethylenediaminetetraacetic acid (EDTA) may be used depending upon buffering, chelation, or acid strength requirements.
(Formula LXVI)
(Formula LXX)
(Formula LXVII)
(Formula LXXI)
(Formula LXVIII)
(Formula LXXII)
(Formula LXIX)
(Formula LXXIII)

[88] At step 321, a pre-defined amount of the compound having either one of the formula LXX, LXXI, LXXII, or LXXIII as obtained from step 319a, is dissolved in anhydrous acetonitrile along with a pre-defined amount of Hexafluorophosphate Benzotriazole Tetramethyl Uronium (HBTU) or any other functionally equivalent coupling reagent or the like to impede racemization to obtain a ninth mixture. Instead of anhydrous acetonitrile, solvents like dimethylformamide (DMF) or dimethyl sulfoxide (DMSO) having high boiling point, protic solvents like methanol or ethanol, or less polar solvents like tetrahydrofuran (THF) may be used. The ninth mixture is subjected to stirring or the like under an inert environment for a pre-defined time period. The pre-defined time period ranges from 15 minutes to 20 minutes. The pre-defined amount of the compound having either one of the formula LXX, LXXI, LXXII, or LXXIII in the ninth mixture ranges from 1 mM to 1.5 mM. The pre-defined amount of HBTU in the ninth mixture ranges from 1.5 mM to 2.5 mM. In an exemplary embodiment, the compound having either one of the formula LXX, LXXI, LXXII, or LXXIII is dissolved in 25 mL of anhydrous acetonitrile along with 1.2 mol equivalent HBTU to obtain the ninth mixture. The ninth mixture is stirred for 15 minutes at room temperature under either argon or nitrogen protection.
[89] At step 321a, a pre-defined amount of controlled pore glass (CPG) (as a solid support) having a pre-defined particle size is added to the ninth mixture as obtained from step 321. The particle size of the CPG ranges from 500 Å to 3000 Å. The pre-defined amount of CPG in the ninth mixture ranges from 100 µM to 150 µM. In an exemplary embodiment, 0.050 g of CPG having particle size of 1000 Å is added to the ninth mixture. Instead of CPG, polystyrene (PS) resins, polyethylene glycol (PEG)-based supports, or silica-based materials may be used based on specific needs like higher loading, aqueous compatibility, or cost.
[90] At step 321b, the ninth mixture as obtained from step 321a is subjected to gentle shaking or the like for a pre-defined time period in an inert environment. The pre-defined time period ranges from 30 minutes to 60 minutes. In an exemplary embodiment, the ninth mixture is gently shaken for 1 hour under either argon or nitrogen protection.
[91] At step 321c, the ninth mixture as obtained from step 321b is subjected to filtration or the like, washed and dried to obtain the delivery system 100 having either one of a formula LXXIV, LXXV, LXXVI, or LXXVII of the present disclosure. In an exemplary embodiment, the ninth mixture is filtered, washed 5 times with 20 mL acetonitrile and dried under vacuum at 25 °C for 48 hours to obtain the delivery system 100 having either one of the formula LXXIV, LXXV, LXXVI, or LXXVII of the present disclosure. Instead of anhydrous acetonitrile, solvents like dimethylformamide (DMF) or dimethyl sulfoxide (DMSO) having high boiling point, protic solvents like methanol or ethanol, or less polar solvents like tetrahydrofuran (THF) may be used.

(Formula LXX) (Formula LXXIV)

(Formula LXXI) (Formula LXXV)

(Formula LXXII) (Formula LXXVI)

(Formula LXXIII) (Formula LXXVII)

[92] At an optional step 323, at least one agent 200 is conjugated to the delivery system 100 having either one of the formula LXXIV, LXXV, LXXVI, or LXXVII as obtained from step 321c.
[93] Although the steps of the method 300 has been described in an exemplary order, at least few of the steps of the method 300 may be executed either simultaneously, randomly, and/or in a different order and the same is within the scope of the teachings of the present disclosure. For example, the step 313 of the method 300 may be executed before commencing the method 300.
[94] Now the delivery system 100 of the present disclosure will be explained with the help of the following examples:
[95] Example 1: Preparing the delivery system 100 having the formula LXXIV of the present disclosure
[96] 5g of galactosamine pentaacetate was dissolved in 25mL of DCE at room temperature to obtain the first mixture. Thereafter, 3.5g of TMSOTf was added to the first mixture and stirred at 50 °C for 90 minutes. The first mixture was cooled to room temperature and stirred for 12 hours. The first mixture was then added to an ice-cold NaHCO3 solution, subjected to solvent extraction with DCM, and then washed with water and dried over Na2SO4. The first mixture was then filtered to remove the solvent(s). A residue obtained from filtering the first mixture was dried overnight under high vacuum to obtain a viscous solid having the formula XXXVIII.
[97] 3.9 g of the viscous solid having the formula XXXVIII, 3.6 g of the benzyl ester having the formula XXXIX, and 1 g of molecular-sieve having pore size of 4 ? were dissolved in 25 mL of DCM to obtain the second mixture. 1 g of TMSOTf was added to the second mixture and stirred for 12 hours. The second mixture was added to an ice-cold NaHCO3 solution, subjected to solvent extraction with DCM, and then washed with water and dried over Na2SO4. The second mixture was filtered and a residue obtained from filtering the second mixture was eluted with 20% to 100% ethyl acetate in hexanes through a column containing silica gel to yield a compound having the formula XXXX.
[98] 4.5 g of the compound having the formula XXXX and 0.45 g of the Pd/C mixture were suspended in 2mL of methyl alcohol and 20mL of EtOAc to obtain the first suspension. The first suspension was flushed with hydrogen gas and then hydrogenated under balloon pressure for 16 hours. The first suspension was filtered through a celite column and then the filter bed of the celite column was washed with methanol. The filtrate obtained therefrom was combined and concentrated under reduced pressure to yield 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having the formula XXXXI, i.e., the ligand 150.
[99] 5 g of 3,3'-((2-(((Benzyloxy)carbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy))dipropionic acid (Tris-base) having the formula XXXXII, 3.5 mol equivalent mono-Boc protected 1,3- diaminopropane having the formula XXXXIII, 10 g of HBTU, and 5 mL of DIEA were added to 30 mL of DMF to obtain the third mixture. The third mixture was stirred overnight. The third mixture was added to ice-cold water, subjected to solvent extraction with DCM, and then washed with saturated NaHCO3 solution and brine, and dried over Na2SO4. The third mixture was filtered. A residue obtained from filtering the third mixture was eluted with ethyl acetate followed by 2% to 10% methanol in DCM through a column containing silica gel to yield a compound having the formula XXXXIV.
[100] 3.58 g the compound having the formula XXXXIV is dissolved in 15 mL of TFA to obtain the fourth mixture. The fourth mixture was stirred at room temperature for 35 minutes. The fourth mixture was diluted with 75-100 mL of toluene and concentrated (dried) under reduced pressure using a high vacuum pump to yield TFA salt of the compound having the formula XXXXV.
[101] 3.31 g of 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having the formula XXXXI, 3.31 g of HBTU, and 4 mL of DIEA were added to 15 mL DMF to obtain the fifth mixture. The fifth mixture was stirred for 5 minutes. 2.5 g of the compound having the formula XXXXV was dissolved in 25 mL of DMF and added to the fifth mixture. The fifth mixture was stirred at room temperature for 18 hours. The fifth mixture was subjected to reduced pressure to remove the solvent(s). A residue obtained after removing the solvents from the fifth mixture was dissolved in 50mL of DCM, washed with 50 mL of saturated aqueous NaHCO3 solution, 50 mL of water and 50 mL of brine, and dried over Na2SO4. The fifth mixture was filtered and subjected to reduced pressure to remove the solvent(s). A residue obtained therefrom was eluted with ethyl acetate followed by gradient elution from 5% to 25% methanol in DCM through a column containing silica gel to yield a compound having the formula XXXXVI.
[102] 5 g of the compound having the formula XXXXVI was dissolved in 30 mL of methanol to obtain the sixth mixture. 10% (w/w) of the Pd/C mixture was then added to the sixth mixture. The sixth mixture was flushed with hydrogen gas and then hydrogenated under balloon pressure for 12 hours. The sixth mixture was filtered through a celite column and then the filter bed of the celite column was washed with methanol. 0.5 mL of TFA was added to the sixth mixture. The solvents of the sixth mixture were removed under reduced pressure. The residue obtained therefrom was co-evaporated 2-3 times with toluene and dried under high vacuum for overnight to yield a TFA salt of the compound having the formula LXV.
[103] 1 mol equivalent of the linker 130 having the formula XV was treated with 1.5 mol equivalent HBTU in 25 mL of anhydrous DMF to obtain the seventh mixture. 3 mol equivalent of DIEA was added to the seventh mixture and stirred for 10-15 minutes. 1 mol equivalent of the compound having the formula LXV dissolved in 30 mL of DMF was added to the seventh mixture and stirred at room temperature for overnight. The seventh mixture was subjected to solvent extraction using water and 500 mL of dichloromethane (DCM), dried over sodium sulphate (Na2SO4), filtered and concentrated under reduced pressure. A residue obtained therefrom was purified by column chromatography (silica gel column, eluted with 0% to 25% methanol in dichloromethane) to yield a compound having the formula LXVI.
[104] 2 g (1 mol equivalent) of the compound having the formula LXVI was treated with 2 mol equivalent succinic anhydride and 3 mol equivalent DMAP dissolved in 50 mL DCM to obtain the eighth mixture. The eighth mixture was stirred at room temperature for overnight. The eighth mixture was diluted with 150 mL DCM, washed with 250 mL water and 250 mL of cold 10% citric acid solution. An organic layer obtained therefrom was dried over sodium sulphate (Na2SO4), concentrated under reduced pressure, and filtered through a small pad of silica gel to yield succinate derivative of a compound having the formula LXX.
[105] The compound having the formula LXX was dissolved in 25 mL of anhydrous acetonitrile along with 1.2 mol equivalent HBTU to obtain the ninth mixture. The ninth mixture was stirred for 15 minutes at room temperature under argon protection. 0.050 g of CPG having particle size of 1000 Å was added to the ninth mixture. The ninth mixture was gently shaken for 1 hour under argon protection. The ninth mixture was filtered, washed 5 times with 20 mL acetonitrile and dried under vacuum at 25 °C for 48 hours to obtain the delivery system 100 having the formula LXXIV of the present disclosure.
[106] The delivery system 100 having the formula LXXIV had a white solid powder-like appearance. The loading capacity of the delivery system 100 having the formula LXXIV on CPG support was determined using a standard trityl release assay. The loading capacity was found to be 24.5 µmol/g.
[107] Example 2: Preparing the delivery system 100 having the formula LXXV of the present disclosure
[108] 5g of galactosamine pentaacetate was dissolved in 25mL of DCE at room temperature to obtain the first mixture. Thereafter, 3.5g of TMSOTf was added to the first mixture and stirred at 50 °C for 90 minutes. The first mixture was cooled to room temperature and stirred for 12 hours. The first mixture was then added to an ice-cold NaHCO3 solution, subjected to solvent extraction with DCM, and then washed with water and dried over Na2SO4. The first mixture was then filtered to remove the solvent(s). A residue obtained from filtering the first mixture was dried overnight under high vacuum to obtain a viscous solid having the formula XXXVIII.
[109] 3.9 g of the viscous solid having the formula XXXVIII, 3.6 g of the benzyl ester having the formula XXXIX, and 1 g of molecular-sieve having pore size of 4 ? were dissolved in 25 mL of DCM to obtain the second mixture. 1 g of TMSOTf was added to the second mixture and stirred for 12 hours. The second mixture was added to an ice-cold NaHCO3 solution, subjected to solvent extraction with DCM, and then washed with water and dried over Na2SO4. The second mixture was filtered and a residue obtained from filtering the second mixture was eluted with 20% to 100% ethyl acetate in hexanes through a column containing silica gel to yield a compound having the formula XXXX.
[110] 4.5 g of the compound having the formula XXXX and 0.45 g of the Pd/C mixture were suspended in 2mL of methyl alcohol and 20mL of EtOAc to obtain the first suspension. The first suspension was flushed with hydrogen gas and then hydrogenated under balloon pressure for 16 hours. The first suspension was filtered through a celite column and then the filter bed of the celite column was washed with methanol. The filtrate obtained therefrom was combined and concentrated under reduced pressure to yield 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having the formula XXXXI, i.e., the ligand 150.
[111] 5 g of 3,3'-((2-(((Benzyloxy)carbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy))dipropionic acid (Tris-base) having the formula XXXXII, 3.5 mol equivalent mono-Boc protected 1,3- diaminopropane having the formula XXXXIII, 10 g of HBTU, and 5 mL of DIEA were added to 30 mL of DMF to obtain the third mixture. The third mixture was stirred overnight. The third mixture was added to ice-cold water, subjected to solvent extraction with DCM, and then washed with saturated NaHCO3 solution and brine, and dried over Na2SO4. The third mixture was filtered. A residue obtained from filtering the third mixture was eluted with ethyl acetate followed by 2% to 10% methanol in DCM through a column containing silica gel to yield a compound having the formula XXXXIV.
[112] 3.58 g the compound having the formula XXXXIV is dissolved in 15 mL of TFA to obtain the fourth mixture. The fourth mixture was stirred at room temperature for 35 minutes. The fourth mixture was diluted with 75-100 mL of toluene and concentrated (dried) under reduced pressure using a high vacuum pump to yield TFA salt of the compound having the formula XXXXV.
[113] 3.31 g of 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having the formula XXXXI, 3.31 g of HBTU, and 4 mL of DIEA were added to 15 mL DMF to obtain the fifth mixture. The fifth mixture was stirred for 5 minutes. 2.5 g of the compound having the formula XXXXV was dissolved in 25 mL of DMF and added to the fifth mixture. The fifth mixture was stirred at room temperature for 18 hours. The fifth mixture was subjected to reduced pressure to remove the solvent(s). A residue obtained after removing the solvents from the fifth mixture was dissolved in 50mL of DCM, washed with 50 mL of saturated aqueous NaHCO3 solution, 50 mL of water and 50 mL of brine, and dried over Na2SO4. The fifth mixture was filtered and subjected to reduced pressure to remove the solvent(s). A residue obtained therefrom was eluted with ethyl acetate followed by gradient elution from 5% to 25% methanol in DCM through a column containing silica gel to yield a compound having the formula XXXXVI.
[114] 5 g of the compound having the formula XXXXVI was dissolved in 30 mL of methanol to obtain the sixth mixture. 10% (w/w) of the Pd/C mixture was then added to the sixth mixture. The sixth mixture was flushed with hydrogen gas and then hydrogenated under balloon pressure for 12 hours. The sixth mixture was filtered through a celite column and then the filter bed of the celite column was washed with methanol. 0.5 mL of TFA was added to the sixth mixture. The solvents of the sixth mixture were removed under reduced pressure. The residue obtained therefrom was co-evaporated 2-3 times with toluene and dried under high vacuum for overnight to yield a TFA salt of the compound having the formula LXV.
[115] 1 mol equivalent of the linker 130 having the formula XVI was treated with 1.5 mol equivalent HBTU in 25 mL of anhydrous DMF to obtain the seventh mixture. 3 mol equivalent of DIEA was added to the seventh mixture and stirred for 10-15 minutes. 1 mol equivalent of the compound having the formula LXV dissolved in 30 mL of DMF was added to the seventh mixture and stirred at room temperature for overnight. The seventh mixture was subjected to solvent extraction using water and 500 mL of dichloromethane (DCM), dried over sodium sulphate (Na2SO4), filtered and concentrated under reduced pressure. A residue obtained therefrom was purified by column chromatography (silica gel column, eluted with 0% to 25% methanol in dichloromethane) to yield a compound having the formula LXVII.
[116] 2 g (1 mol equivalent) of the compound having the formula LXVII was treated with 2 mol equivalent succinic anhydride and 3 mol equivalent DMAP dissolved in 50 mL DCM to obtain the eighth mixture. The eighth mixture was stirred at room temperature for overnight. The eighth mixture was diluted with 150 mL DCM, washed with 250 mL water and 250 mL of cold 10% citric acid solution. An organic layer obtained therefrom was dried over sodium sulphate (Na2SO4), concentrated under reduced pressure, and filtered through a small pad of silica gel to yield succinate derivative of a compound having the formula LXXI.
[117] The compound having the formula LXXI was dissolved in 25 mL of anhydrous acetonitrile along with 1.2 mol equivalent HBTU to obtain the ninth mixture. The ninth mixture was stirred for 15 minutes at room temperature under argon protection. 0.050 g of CPG having particle size of 1000 Å was added to the ninth mixture. The ninth mixture was gently shaken for 1 hour under argon protection. The ninth mixture was filtered, washed 5 times with 20 mL acetonitrile and dried under vacuum at 25 °C for 48 hours to obtain the delivery system 100 having the formula LXXV of the present disclosure.
[118] The delivery system 100 having the formula LXXV had a white solid powder-like appearance. The loading capacity of the delivery system 100 having the formula LXXV on CPG support was determined using a standard trityl release assay. The loading capacity was found to be 19.2 µmol/g.
[119] Example 3: Preparing the delivery system 100 having the formula LXXVI of the present disclosure
[120] 5g of galactosamine pentaacetate was dissolved in 25mL of DCE at room temperature to obtain the first mixture. Thereafter, 3.5g of TMSOTf was added to the first mixture and stirred at 50 °C for 90 minutes. The first mixture was cooled to room temperature and stirred for 12 hours. The first mixture was then added to an ice-cold NaHCO3 solution, subjected to solvent extraction with DCM, and then washed with water and dried over Na2SO4. The first mixture was then filtered to remove the solvent(s). A residue obtained from filtering the first mixture was dried overnight under high vacuum to obtain a viscous solid having the formula XXXVIII.
[121] 3.9 g of the viscous solid having the formula XXXVIII, 3.6 g of the benzyl ester having the formula XXXIX, and 1 g of molecular-sieve having pore size of 4 ? were dissolved in 25 mL of DCM to obtain the second mixture. 1 g of TMSOTf was added to the second mixture and stirred for 12 hours. The second mixture was added to an ice-cold NaHCO3 solution, subjected to solvent extraction with DCM, and then washed with water and dried over Na2SO4. The second mixture was filtered and a residue obtained from filtering the second mixture was eluted with 20% to 100% ethyl acetate in hexanes through a column containing silica gel to yield a compound having the formula XXXX.
[122] 4.5 g of the compound having the formula XXXX and 0.45 g of the Pd/C mixture were suspended in 2mL of methyl alcohol and 20mL of EtOAc to obtain the first suspension. The first suspension was flushed with hydrogen gas and then hydrogenated under balloon pressure for 16 hours. The first suspension was filtered through a celite column and then the filter bed of the celite column was washed with methanol. The filtrate obtained therefrom was combined and concentrated under reduced pressure to yield 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having the formula XXXXI, i.e., the ligand 150.
[123] 5 g of 3,3'-((2-(((Benzyloxy)carbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy))dipropionic acid (Tris-base) having the formula XXXXII, 3.5 mol equivalent mono-Boc protected 1,3- diaminopropane having the formula XXXXIII, 10 g of HBTU, and 5 mL of DIEA were added to 30 mL of DMF to obtain the third mixture. The third mixture was stirred overnight. The third mixture was added to ice-cold water, subjected to solvent extraction with DCM, and then washed with saturated NaHCO3 solution and brine, and dried over Na2SO4. The third mixture was filtered. A residue obtained from filtering the third mixture was eluted with ethyl acetate followed by 2% to 10% methanol in DCM through a column containing silica gel to yield a compound having the formula XXXXIV.
[124] 3.58 g the compound having the formula XXXXIV is dissolved in 15 mL of TFA to obtain the fourth mixture. The fourth mixture was stirred at room temperature for 35 minutes. The fourth mixture was diluted with 75-100 mL of toluene and concentrated (dried) under reduced pressure using a high vacuum pump to yield TFA salt of the compound having the formula XXXXV.
[125] 3.31 g of 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having the formula XXXXI, 3.31 g of HBTU, and 4 mL of DIEA were added to 15 mL DMF to obtain the fifth mixture. The fifth mixture was stirred for 5 minutes. 2.5 g of the compound having the formula XXXXV was dissolved in 25 mL of DMF and added to the fifth mixture. The fifth mixture was stirred at room temperature for 18 hours. The fifth mixture was subjected to reduced pressure to remove the solvent(s). A residue obtained after removing the solvents from the fifth mixture was dissolved in 50mL of DCM, washed with 50 mL of saturated aqueous NaHCO3 solution, 50 mL of water and 50 mL of brine, and dried over Na2SO4. The fifth mixture was filtered and subjected to reduced pressure to remove the solvent(s). A residue obtained therefrom was eluted with ethyl acetate followed by gradient elution from 5% to 25% methanol in DCM through a column containing silica gel to yield a compound having the formula XXXXVI.
[126] 5 g of the compound having the formula XXXXVI was dissolved in 30 mL of methanol to obtain the sixth mixture. 10% (w/w) of the Pd/C mixture was then added to the sixth mixture. The sixth mixture was flushed with hydrogen gas and then hydrogenated under balloon pressure for 12 hours. The sixth mixture was filtered through a celite column and then the filter bed of the celite column was washed with methanol. 0.5 mL of TFA was added to the sixth mixture. The solvents of the sixth mixture were removed under reduced pressure. The residue obtained therefrom was co-evaporated 2-3 times with toluene and dried under high vacuum for overnight to yield a TFA salt of the compound having the formula LXV.
[127] 1 mol equivalent of the linker 130 having the formula XVII was treated with 1.5 mol equivalent HBTU in 25 mL of anhydrous DMF to obtain the seventh mixture. 3 mol equivalent of DIEA was added to the seventh mixture and stirred for 10-15 minutes. 1 mol equivalent of the compound having the formula LXV dissolved in 30 mL of DMF was added to the seventh mixture and stirred at room temperature for overnight. The seventh mixture was subjected to solvent extraction using water and 500 mL of dichloromethane (DCM), dried over sodium sulphate (Na2SO4), filtered and concentrated under reduced pressure. A residue obtained therefrom was purified by column chromatography (silica gel column, eluted with 0% to 25% methanol in dichloromethane) to yield a compound having the formula LXVIII.
[128] 2 g (1 mol equivalent) of the compound having the formula LXVIII was treated with 2 mol equivalent succinic anhydride and 3 mol equivalent DMAP dissolved in 50 mL DCM to obtain the eighth mixture. The eighth mixture was stirred at room temperature for overnight. The eighth mixture was diluted with 150 mL DCM, washed with 250 mL water and 250 mL of cold 10% citric acid solution. An organic layer obtained therefrom was dried over sodium sulphate (Na2SO4), concentrated under reduced pressure, and filtered through a small pad of silica gel to yield succinate derivative of a compound having the formula LXXII.
[129] The compound having the formula LXXII was dissolved in 25 mL of anhydrous acetonitrile along with 1.2 mol equivalent HBTU to obtain the ninth mixture. The ninth mixture was stirred for 15 minutes at room temperature under argon protection. 0.050 g of CPG having particle size of 1000 Å was added to the ninth mixture. The ninth mixture was gently shaken for 1 hour under argon protection. The ninth mixture was filtered, washed 5 times with 20 mL acetonitrile and dried under vacuum at 25 °C for 48 hours to obtain the delivery system 100 having the formula LXXVI of the present disclosure.
[130] The delivery system 100 having the formula LXXVI had a white solid powder-like appearance. The loading capacity of the delivery system 100 having the formula LXXVI on CPG support was determined using a standard trityl release assay. The loading capacity was found to be 23.2 µmol/g.
[131] Example 4: Preparing the delivery system 100 having the formula LXXVII of the present disclosure
[132] 5g of galactosamine pentaacetate was dissolved in 25mL of DCE at room temperature to obtain the first mixture. Thereafter, 3.5g of TMSOTf was added to the first mixture and stirred at 50 °C for 90 minutes. The first mixture was cooled to room temperature and stirred for 12 hours. The first mixture was then added to an ice-cold NaHCO3 solution, subjected to solvent extraction with DCM, and then washed with water and dried over Na2SO4. The first mixture was then filtered to remove the solvent(s). A residue obtained from filtering the first mixture was dried overnight under high vacuum to obtain a viscous solid having the formula XXXVIII.
[133] 3.9 g of the viscous solid having the formula XXXVIII, 3.6 g of the benzyl ester having the formula XXXIX, and 1 g of molecular-sieve having pore size of 4 ? were dissolved in 25 mL of DCM to obtain the second mixture. 1 g of TMSOTf was added to the second mixture and stirred for 12 hours. The second mixture was added to an ice-cold NaHCO3 solution, subjected to solvent extraction with DCM, and then washed with water and dried over Na2SO4. The second mixture was filtered and a residue obtained from filtering the second mixture was eluted with 20% to 100% ethyl acetate in hexanes through a column containing silica gel to yield a compound having the formula XXXX.
[134] 4.5 g of the compound having the formula XXXX and 0.45 g of the Pd/C mixture were suspended in 2mL of methyl alcohol and 20mL of EtOAc to obtain the first suspension. The first suspension was flushed with hydrogen gas and then hydrogenated under balloon pressure for 16 hours. The first suspension was filtered through a celite column and then the filter bed of the celite column was washed with methanol. The filtrate obtained therefrom was combined and concentrated under reduced pressure to yield 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having the formula XXXXI, i.e., the ligand 150.
[135] 5 g of 3,3'-((2-(((Benzyloxy)carbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy))dipropionic acid (Tris-base) having the formula XXXXII, 3.5 mol equivalent mono-Boc protected 1,3- diaminopropane having the formula XXXXIII, 10 g of HBTU, and 5 mL of DIEA were added to 30 mL of DMF to obtain the third mixture. The third mixture was stirred overnight. The third mixture was added to ice-cold water, subjected to solvent extraction with DCM, and then washed with saturated NaHCO3 solution and brine, and dried over Na2SO4. The third mixture was filtered. A residue obtained from filtering the third mixture was eluted with ethyl acetate followed by 2% to 10% methanol in DCM through a column containing silica gel to yield a compound having the formula XXXXIV.
[136] 3.58 g the compound having the formula XXXXIV is dissolved in 15 mL of TFA to obtain the fourth mixture. The fourth mixture was stirred at room temperature for 35 minutes. The fourth mixture was diluted with 75-100 mL of toluene and concentrated (dried) under reduced pressure using a high vacuum pump to yield TFA salt of the compound having the formula XXXXV.
[137] 3.31 g of 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having the formula XXXXI, 3.31 g of HBTU, and 4 mL of DIEA were added to 15 mL DMF to obtain the fifth mixture. The fifth mixture was stirred for 5 minutes. 2.5 g of the compound having the formula XXXXV was dissolved in 25 mL of DMF and added to the fifth mixture. The fifth mixture was stirred at room temperature for 18 hours. The fifth mixture was subjected to reduced pressure to remove the solvent(s). A residue obtained after removing the solvents from the fifth mixture was dissolved in 50mL of DCM, washed with 50 mL of saturated aqueous NaHCO3 solution, 50 mL of water and 50 mL of brine, and dried over Na2SO4. The fifth mixture was filtered and subjected to reduced pressure to remove the solvent(s). A residue obtained therefrom was eluted with ethyl acetate followed by gradient elution from 5% to 25% methanol in DCM through a column containing silica gel to yield a compound having the formula XXXXVI.
[138] 5 g of the compound having the formula XXXXVI was dissolved in 30 mL of methanol to obtain the sixth mixture. 10% (w/w) of the Pd/C mixture was then added to the sixth mixture. The sixth mixture was flushed with hydrogen gas and then hydrogenated under balloon pressure for 12 hours. The sixth mixture was filtered through a celite column and then the filter bed of the celite column was washed with methanol. 0.5 mL of TFA was added to the sixth mixture. The solvents of the sixth mixture were removed under reduced pressure. The residue obtained therefrom was co-evaporated 2-3 times with toluene and dried under high vacuum for overnight to yield a TFA salt of the compound having the formula LXV.
[139] 1 mol equivalent of the linker 130 having the formula XVIII was treated with 1.5 mol equivalent HBTU in 25 mL of anhydrous DMF to obtain the seventh mixture. 3 mol equivalent of DIEA was added to the seventh mixture and stirred for 10-15 minutes. 1 mol equivalent of the compound having the formula LXV dissolved in 30 mL of DMF was added to the seventh mixture and stirred at room temperature for overnight. The seventh mixture was subjected to solvent extraction using water and 500 mL of dichloromethane (DCM), dried over sodium sulphate (Na2SO4), filtered and concentrated under reduced pressure. A residue obtained therefrom was purified by column chromatography (silica gel column, eluted with 0% to 25% methanol in dichloromethane) to yield a compound having the formula LXIX.
[140] 2 g (1 mol equivalent) of the compound having the formula LXIX was treated with 2 mol equivalent succinic anhydride and 3 mol equivalent DMAP dissolved in 50 mL DCM to obtain the eighth mixture. The eighth mixture was stirred at room temperature for overnight. The eighth mixture was diluted with 150 mL DCM, washed with 250 mL water and 250 mL of cold 10% citric acid solution. An organic layer obtained therefrom was dried over sodium sulphate (Na2SO4), concentrated under reduced pressure, and filtered through a small pad of silica gel to yield succinate derivative of a compound having the formula LXXIII.
[141] The compound having the formula LXXIII was dissolved in 25 mL of anhydrous acetonitrile along with 1.2 mol equivalent HBTU to obtain the ninth mixture. The ninth mixture was stirred for 15 minutes at room temperature under argon protection. 0.050 g of CPG having particle size of 1000 Å was added to the ninth mixture. The ninth mixture was gently shaken for 1 hour under argon protection. The ninth mixture was filtered, washed 5 times with 20 mL acetonitrile and dried under vacuum at 25 °C for 48 hours to obtain the delivery system 100 having the formula LXXVII of the present disclosure.
[142] The delivery system 100 having the formula LXXVII had a white solid powder-like appearance. The loading capacity of the delivery system 100 having the formula LXXVII on CPG support was determined using a standard trityl release assay. The loading capacity was found to be 22.7 µmol/g.
[143] Example 5: Conjugating agent 200 with the delivery system 100 of the present disclosure
[144] A plurality of sense strands (SS) were synthesized on Mermade 12 oligosynthsizer machine using commercially available 5'-O-(4,4'-dimethoxytrityl)-2'-deoxy-2'-fluoro-, 5'-O-(4,4'-dimethoxytrityl)- 2'-O-(tert-butyldimethylsilyl)-, and 5'-O-(4,4'-dimethoxytrityl)-2'-O-methyl- 3'-O-(2-cyanoethyl N,N-diisopropyl) phosphoramidite monomers of uridine, 4-N-acetylcytidine, 6-N benzoyladenosine, and 2-N-isobutyrylguanosine using standard solid-phase oligonucleotide synthesis and deprotection protocols.
[145] The delivery system 100 having the formula LXXIV, LXXV, LXXVI, and LXXVII as obtained from example 1-4 were conjugated with 3’-end of the sense strand of SEQ ID No. 1 and SEQ ID No. 2 oligonucleotides (Table 1) separately followed by deprotection. The respective conjugates obtained was checked for purity using Ion-Pair Reversed-Phase (IP RP) High Pressure Liquid Chromatography (HPLC). The plots obtained therefrom were plotted as depicted in Figs. 3-6.
SEQ ID No. Sequence
1 5’ – C U A G A C C U G U U U U G C U U U U G U - 3’
2 5’ – Um- Gm- Gm Gm Am Um Uf Um Cf Af Uf Gm Um Am Am Cm Cm Am Am Gm Am – 3’

[146] The IP RP-HPLC plot of SEQ ID NO. 1/SEQ ID No. 2 conjugated to the delivery system 100 having the formula LXXIV and LXXV gave very low yield, i.e., less than 5.0%. It was our assumption that the first or second base of the SEQ ID NO. 1/SEQ ID No. 2 was not connected to the delivery system 100, as a result the peaks in the plots appeared very early (i.e., less than 1 minute) to be visually significant.
[147] The IP RP-HPLC plot of SEQ ID NO. 1 conjugated to the delivery system 100 having the formula LXXVI is depicted in Fig. 3. The purity was calculated to be 93.6%. The IP RP-HPLC plot of SEQ ID NO. 2 conjugated to the delivery system 100 having the formula LXXVI is depicted in Fig. 3a. The purity was calculated to be 88%. It was observed that the main peak mass of the actual synthesized conjugates was consistent with the theoretical value.
[148] The IP RP-HPLC plot of SEQ ID NO. 1 conjugated to the delivery system 100 having the formula LXXVII is depicted in Fig. 4. The purity was calculated to be 85.4%. The IP RP-PLC plot of SEQ ID NO. 2 conjugated to the delivery system 100 having the formula LXXVII is depicted in Fig. 4a. The purity was calculated to be 82.3%. It was observed that the main peak mass of the actual synthesized conjugates was consistent with the theoretical value.
[149] Example 5: Spectral analysis of the delivery system 100 having the formula XXXI (with n=5) synthesized using the linker 130 having the formula XV of the present disclosure
[150] The delivery system 100 having formula XXXI (with n=5) synthesized using linker 130 having formula XV has a chemical formula C118H173N11O45 having structure as follows:

(Formula LXXIX)

[151] The delivery system 100 having a formula LXXIX was found to have a molecular mass 2448.1638 g/mol. The 1H-NMR (nuclear magnetic resonance) spectral data obtained was as follows (plotted in Fig. 5):
1H-NMR (DMSO-d6, 400MHz, ppm): 7.90-7.79 (m, 9H), 7.34-7.21 (m, 9H), 7.01 (s, 1H), 6.89–6.87 (m, 4H), 5.22-5.21 (d, 3H), 4.98–4.96 (dd, 3H), 4.52-4.49 (dd, 3H), 4.04-4.03 (m, 9H, 3XCH-NHCO &3 XCH2-OCOCH3 of GalNAc moiety), 3.91-3.86 (m, 4H), 3.74-3.69 (m , 11H), 3.56-3.53 (m, 16H), 3.44-3.37 (m, 14H), 2.46-2.44 (m, 5H), 2.30-2.27 (m, 7H), 2.10 (s, 9H), 2.08–2.04 (m, 7H), 2.00(s, 10H), 1.89 (m, 10H), 1.78-1.77 (m,9H), 1.53–1.48 (m, 20H), 1.46-1.43 (m,10H).
[152] The ESI-MS (electrospray ionization mass spectrometry) spectral data obtained was as follows (plotted in Fig. 5a):
ESI-MS (m/z): Calc 2464.2587, found 2466.1618 (M+2H)+, 2488.1452 (M+H+Na)+, 2567.1452 (M+2H+TEA)+, 1255.2559 (M+2Na)2+.
[153] Example 6: Spectral analysis of the delivery system 100 having the formula XXXI (with n=2) synthesized using the linker 130 having the formula XVI of the present disclosure
[154] The delivery system 100 having the formula XXXI (with n=2) synthesized using linker 130 having the formula XVI having structure as follows:

(Formula LXXX)

[155] The delivery system 100 having a formula LXXX was found to have a molecular mass 2237.0212 g/mol. The 1H-NMR (nuclear magnetic resonance) spectral data obtained was as follows (plotted in Fig. 6):
1H-NMR (400MHz, CDCl3): H-NMR (DMSO-d6, 400MHz, ppm): 7.84-7.80 (t, 5H), 7.34-7.31 (m, 4H), 7.29-7.20 (dd, 4H), 7.12 (s, 1H), 6.90-6.83 (d, 4H), 5.33-5.21 (d 3H), 4.99-4.91 (dd, 3H), 4.50-4.42 (d, 3H), 4.05-4.01 (d, 9H), 3.91-3.84 (q, 4H), 3.74 (s, 6H), 3.69-3.67 (m, 2H), 3.59-3.52 (q, 13H), 3.04-3.01 (q, 6H), 2.99-2.86 (d, 6H), 2.47-2.43 (d, 4H), 2.30-2.27 (d, 6H), 2.10 (s, 9H), 1.99 (s, 9H), 1.89 (s, 9H), 1.77 (s, 9H), 1.46-1.34 (m, 24H).
[156] The ESI-MS (electrospray ionization mass spectrometry) spectral data obtained was as follows (plotted in Fig. 6a):
ESI-MS (m/z): Calc 2237.0317, found 2237.0212 (M)+, 1135.9982 (M+NH4++NH3)2+.
[157] Example 7: Spectral analysis of the delivery system 100 having the formula XXV (with n=5) synthesized using the linker 130 having the formula XVII of the present disclosure
[158] The delivery system 100 having the formula XXV (with n=5) synthesized using linker 130 having the formula XVII has a chemical formula C116H171N11O45 having structure as follows:

(Formula LXXXI)

[159] The delivery system 100 having a formula LXXXI was found to have a molecular mass 2438.1431 g/mol. The 1H-NMR (nuclear magnetic resonance) spectral data obtained was as follows (plotted in Fig. 7):
1H-NMR (CDCl3, 400MHz, ppm): 7.41-7.36 (m, 5H), 7.30-7.26 (m, 6H), 7.06-7.04 (s, 4H), 6.84-6.81 (m, 6H), 5.36-5.34(m, 3H), 5.21-5.18 (m, 3H), 5.03-5.01(m,1H), 4.63-4.61 (m, 3H), 4.16- 4.06 (s, 11H), 3.95-3.90 (m, 8H), 3.79-3.78 (m, 15H), 3.69-3.67 (m, 6H), 3.53-3.50 (m, 17H), 3.28- 3.23 (m, 6H), 2.45-2.42 (m, 9H), 2.15 (m, 9H), 2.04-2.03 (m, 7H), 2.01-1.99(m, 9H), 1.95(m, 8H), 1.75-1.65 (m, 23H), 1,38-1.46 (m, 5H).
[160] The ESI-MS (electrospray ionization mass spectrometry) spectral data obtained was as follows (plotted in Fig. 7a):
ESI-MS (m/z): Calc 2438.1431, found 2438.1301 (Z = 1).
[161] Example 8: Spectral analysis of the delivery system 100 having the formula XXIV synthesized using the linker 130 having the formula XVIII of the present disclosure
[162] The delivery system 100 having the formula XXIV synthesized using linker 130 having formula XVIII has a chemical the formula C117H171N11O46 having structure as follows:

(Formula LXXXII)

[163] The delivery system 100 having a formula LXXXII was found to have a molecular mass 2466.1380 g/mol. The 1H-NMR (nuclear magnetic resonance) spectral data obtained was as follows (plotted in Fig. 8):
1H-NMR (CDCl3, 400MHz, ppm): 7.37-7.36 (m, 7H), 7.04-7.00 (m, 4H), 6.84-6.80 (m, 9H), 5.35-5.34(d, 3H), 5.21-5.18 (m, 3H), 4.63-4.61 (m, 3H), 4.19- 4.09 (m, 27H), 4.06-3.89 (m, 13H), 3.79-3.68 (m, 21H), 3.67-3.26 (m, 17H), 3.53-3.50 (m, 17H), 2.15 (m, 8H), 2.04-2.03 (m, 11H), 2.01-1.99(m, 18H), 1.94 (m, 8H), 1.76-1.59 (m, 24H).
[164] The ESI-MS (electrospray ionization mass spectrometry) spectral data obtained was as follows (plotted in Fig. 8a):
ESI-MS (m/z): Calc 2466.1380, found 2466.1342 (M)+.
[165] Example 9: Spectral analysis of the linker 130 having the formula XV of the present disclosure
[166] The linker 130 having the formula XV has a structure as follows:

(Formula XV)

[167] The 1H-NMR (nuclear magnetic resonance) spectral data obtained was as follows (plotted in Fig. 9):
1H-NMR (400MHz, CDCl3, ppm): 7.31-7.21 (m, 9H), 7.89-7.86 (dd, 4H), 4.96 (d, 1H,OH proton), 3.74 (s, 6H), 3.49-3.25 (m, 2H), 3.21-3.17 (t, 1H), 2.99 (m, 2H), 2.47-2.42 (dd, 2H), 2.19-2.14 (td, 2H), 1.78-1.76 (m,1H),1.70-1.66 (d, 1H),1.48 (s, 4H), 1.28-1.17 (m, 4H).
[168] The ESI-MS (electrospray ionization mass spectrometry) spectral data obtained was as follows (plotted in Fig. 9a):
ESI-MS (m/z): Calc 589.3040, found 590.3110 (M+H)+, 612.2927(M+Na)+.
[169] Example 10: Spectral analysis of the linker 130 having the formula XVI of the present disclosure
[170] The linker 130 having the formula XVI has a structure as follows:

(Formula XVI)

[171] The 1H-NMR (nuclear magnetic resonance) spectral data obtained was as follows (plotted in Fig. 10):
1H-NMR (400MHz, CDCl3, ppm): 7.33-7.28 (d, 4H), 7.22-7.19 (dd, 5H), 6.90-6.87 (m, 4H), 4.35-4.30 (m, 1H), 4.05-4.00 (q, 2H), 3.74(s, 6H), 3.23-3.17 (m, 1H), 2.98-2.96 (m, 1H), 2.80 (m, 1H), 2.72 (s, 3H),2.51 (m,2H), 2,42 (s, 3H),1.90-1.78 (m, 2H).
[172] The ESI-MS (electrospray ionization mass spectrometry) spectral data obtained was as follows (plotted in Fig. 10a):
ESI-MS (m/z): Calc 533.2412, found 534.2491 (M+H)+, 556.2308 (M+Na)+.
[173] Example 11: Subjecting liver tissue to the delivery system 100 of the present disclosure
[174] A group of cells from male hPCSK9-UTR mice were injected once as described below:
[175] A control group of cells (G1) from the liver tissue were exposed to phosphate buffer saline (PBS) buffer. A group of cells (G2) were exposed to the delivery system 100 having the formula LXXVI at 3 mg/Kg. A group of cells (G3) were exposed to the delivery system 100 having the formula LXXVI at 10 mg/Kg. A group of cells (G4) were exposed to the delivery system 100 having the formula LXXVII at 3 mg/Kg. A group of cells (G5) were exposed to the delivery system 100 having the formula LXXVI at 10 mg/Kg. Each group of cells were analyzed.
[176] Example 12: Hematoxylin and eosin (H&E) staining of liver tissue
[177] The group of cells as prepared in Example 11 were subjected to H&E staining and viewed under a microscope (as shown in Fig. 11). In group of cells (G1), it was observed that there was diffuse mild swelling of hepatocytes accompanied by slight fatty degeneration, along with a focal area of hepatocellular necrosis. In group of cells (G2), mild swelling was noted in some hepatocytes, and three areas of spotty necrosis were present, while the overall liver tissue structure appears nearly normal. In group of cells (G3), two areas of spotty necrosis were observed in the liver tissue, which remains close to a normal structural state. In group of cells (G4), one area of spotty necrosis was noted, with the liver tissue structure also appearing nearly normal. In group of cells (G5), mild swelling was seen in some hepatocytes, along with one area of hepatocellular necrosis, while the remaining liver tissue structure shows no significant abnormalities.
[178] Example 13: Oil Red O (ORO) staining of liver tissue
[179] The group of cells as prepared in Example 11 were subjected to ORO staining and viewed under a microscope (as shown in Fig. 12). The proportion of ORO stain per unit area is plotted in Fig. 12a. It was observed that the liver tissue sections compared to those treated with PBS as a control (G1), the group of cells G2-G5 exhibited a dose-dependent decrease in lipid droplet accumulation. This was indicative of the fact that effective functional delivery of the siRNA molecule (i.e., the agent 200) took place to target pathways involved in reducing lipid accumulation.
[180] Example 14: hPCSK9 immunohistochemistry in liver tissue
[181] The group of cells as prepared in Example 11 were observed under a microscope (Fig. 13). It was observed that in comparison to control group G1, the group of cells G2-G5 demonstrated a significant dose-dependent reduction in hPCSK9 levels (Fig.13a).
[182] The scope of the invention is only limited by the appended patent claims. More generally, those skilled in the art will readily appreciate that all parameters, dimensions, materials, and configurations described herein are meant to be exemplary and that the actual parameters, dimensions, materials, and/or configurations will depend upon the specific application or applications for which the teachings of the present invention is/are used. ,CLAIMS:WE CLAIM
1. A synthetic delivery system (100) for targeted delivery of an agent (200) comprising:
a. a tetravalent atom/moiety;
b. at least one first spacer (120) coupled to the tetravalent atom/moiety;
c. at least one linker (130) coupled to the tetravalent atom/moiety via the at least one first spacer (120), the linker (130) includes at least one of 4-(hydroxymethyl)-4-methylpiperidin-3-ol having a formula V, 2-(hydroxymethyl)piperidin-4-ol having a formula VI, 5-(hydroxymethyl)-5-methylpyrrolidin-3-ol HCl salt having a formula VII, 2-[4-(hydroxymethyl)piperidin-4-yl]propan-2-ol having a formula VIII, 3-aminopropane-1,2-diol having a formula IX, 4-(hydroxymethyl)piperidin-4-ol having formula X, 3-aminobutane-1,2-diol having a formula XI, 5-(1-hydroxyethyl)pyrrolidin-3-ol having a formula XII, 2-aminobutane-1,3-diol having a formula XIII, 4-amino-1-[4-hydroxy-2-(O-dimethoxy tritylmethyl)pyrrolidin-1-yl]butan-1-one having a formula XIV, 8-((2R,4R)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-hydroxypiperidin-1-yl)-8-oxooctanoic acid having a formula XV, 4-((2R,4R)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-hydroxypiperidin-1-yl)-4-oxobutanoic acid having a formula XVI, 8-((4-(bis(4-methoxyphenyl)(phenyl)methoxy)-3-hydroxybutan-2-yl)amino)-8-oxooctanoic acid having a formula XVII, and 3-(3-(4-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-3-hydroxy-4-methylpiperidin-1-yl)-3-oxopropoxy)propanoic acid a having formula XVIII;
d. one or more second spacers (140) coupled to the tetravalent atom/moiety;
e. a plurality of ligands (150) coupled to the tetravalent atom/moiety via the at least one second spacer (140);
wherein, the linker (130) is conjugated to an agent (200).
2. The synthetic delivery system (100) as claimed in claim 1, wherein the tetravalent atom/moiety includes a carbon atom (110).
3. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) has a pre-defined structure including at least one of Tris-GalNAc-NHCbz (peracetylated, Cbz), GaLNAc Functionalized with alkynes, amines, azides, carboxylic acids, N-succinimidyl esters, monovalent, and divalent.
4. The synthetic delivery system (100) as claimed in claim 1, wherein the first spacer (120) includes at least one of N-Hydroxysuccinimide (NHS) ester, polyethylene glycol (PEG), amino alcohols, allyl amine, and ethers.
5. The synthetic delivery system (100) as claimed in claim 1, wherein the first spacer (120) includes at least one of bis(2,5-dioxopyrrolidin-1-yl) pentanedioate having a formula I, 7-[(2,5-dioxopyrrolidin-1-yl)oxy]-7-oxoheptanoic acid having a formula II, bis(2,5-dioxopyrrolidin-1-yl) 3,3'-oxydipropanoate having a formula III, and 2,5-dioxopyrrolidin-1-yl hex-5-ynoate having a formula IV.
6. The synthetic delivery system (100) as claimed in claim 1, wherein the agent (200) includes a small interfering ribonucleic acid (siRNA) having a sense strand (210) and an antisense strand (220).
7. The synthetic delivery system (100) as claimed in claim 1, wherein a hydroxyl group at the 3’-end of a sense strand (210) of the agent (200) makes a phosphodiester bond with one of the hydroxyl groups of the linker (130).
8. The synthetic delivery system (100) as claimed in claim 1, wherein the second spacer (140) includes at least one of N-Hydroxysuccinimide (NHS) ester, polyethylene glycol (PEG), alkynes, azides, and ethers.
9. The synthetic delivery system (100) as claimed in claim 1, wherein the second spacer (140) includes at least one of 2-((tert-butoxycarbonyl)amino)-2-((hex-5-ynoyloxy)methyl)propane-1,3-diyl bis(hex-5-ynoate) having a formula XIX, 2-Azidopropyl-3,4,6-tri-O-acetyl-2-(acetylamino)-2-deoxy-D-galactopyranosyl and any azidoalkyl-Ac-GalNAc having a formula XX.
10. The synthetic delivery system (100) as claimed in claim 1, wherein the ligand (150) includes N-Acetylgalactosamine (GalNAc) or derivatives thereof having affinity towards asialoglycoprotein receptor (ASGPR).
11. The synthetic delivery system (100) as claimed in claim 1, wherein the ligand (150) includes at least one of 5-[[3,4,6-tri-O-acetyl-2-(acetylamino)-2-deoxy-D-galactopyranosyl]oxy]-pentanoic acid having a formula XXI, and 5-[[3,4,6-tri-O-acetyl-2-(acetylamino)-2-deoxy-D-galactopyranosyl]oxy]-pentanoate N-Hydroxysuccinimide (NHS) ester having a formula XXII.
12. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) includes one linker (130) coupled to the tetravalent atom/moiety via one first spacer (120), and three ligands (150) coupled to the tetravalent atom/moiety via respective second spacers (140).
13. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) includes 4-((1-(24-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-9,9-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-7,14,20-trioxo-4,11-dioxa-8,15,19-triazatetracosan-1-oyl)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)piperidin-4-yl)oxy)-4-oxo-succinoyl-CPG having a formula XXIII.
14. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) includes 4-((1-(24-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-9,9-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-7,14,20-trioxo-4,11-dioxa-8,15,19-triazatetracosan-1-oyl)-4-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-methylpiperidin-3-yl)oxy)-4-oxo-succinoyl-CPG having a formula XXIV.
15. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) includes 33-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-18,18-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-6-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-7-methyl-4,9,16,23,29-pentaoxo-5,20-dioxa-8,17,24,28-tetraazatritriacontan-1-oyl-CPG(Tris-ß-GalNAc-C6-CPG) having a formula XXV.
16. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) includes 33-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-18,18-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-7-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-6-methyl-4,9,16,23,29-pentaoxo-5,20-dioxa-8,17,24,28-tetraazatritriacontan-1-oyl-CPG(Tris-ß-GalNAc-C6-CPG) having a formula XXVI.
17. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) includes 4-((4-(28-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-13,13-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-4-methylene-8,11,18,24-tetraoxo-2,15-dioxa-7,12,19,23-tetraazaoctacosan-1-oyl)-3-((bis(4-methoxy phenyl)(phenyl)methoxy)methyl)cyclohexyl)oxy)-4-oxo-succinoyl-CPG(Tris-ß-GalNAc-Allylamine-cyclohexyl)oxy)-4-oxo-succinoyl-CPG) having a formula XXVII.
18. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) includes 4-((2-(28-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-13,13-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-4-methylene-8,11,18,24-tetraoxo-2,15-dioxa-7,12,19,23-tetraazaoctacosan-1-oyl)-5-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-5-methylcyclohexyl)oxy)-4-oxo-succinoyl-CPG(Tris-ß-GalNAc-Allylamine-methylcyclohexyl)oxy)-4-oxo-succinoyl-CPG) having a formula XXVIII.
19. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) includes 36-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxy methyl)tetrahydro-2H-pyran-2-yl)oxy)-21,21-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-6-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-7-methyl-12-methylene-4,9,16,19,26,32-hexaoxo-5,10,23-trioxa-8,15,20,27,31-pentaazahexatriacontan-1-oyl-CPG(Tris-ß-GalNAc-Allylamine-6-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-7-methyl-12-methylene-4,9,16,19,26,32-hexaoxo-5,10,23-trioxa-8,15,20,27,31-pentaazahexatriacontan-1-oyl-CPG) having a formula XXIX.
20. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) includes 36-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-21,21-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-7-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-6-methyl-12-methylene-4,9,16,19,26,32-hexaoxo-5,10,23-trioxa-8,15,20,27,31-pentaazahexatriacontan-1-oyl-CPG(Tris-ß-GalNAc-Allylamine-7-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-6-methyl-12-methylene-4,9,16,19,26,32-hexaoxo-5,10,23-trioxa-8,15,20,27,31-pentaazahexatriacontan-1-oyl-CPG) having a formula XXX.
21. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) includes 4-((1-(1-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-16,16-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-5,11,18-trioxo-14-oxa-6,10,17-triazapentacosan-25-oyl)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)piperidin-4-yl)oxy)-4-oxo-succinoyl-CPG(Tris-ß-GalNAc-C6-piperidin-4-yl)oxy)-4-oxo-succinoyl-CPG) having a formula XXXI.
22. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) includes 4-((1-(1-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-16,16-bis((3-((3-(5-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)pentanamido)propyl)amino)-3-oxopropoxy)methyl)-5,11,18-trioxo-14-oxa-6,10,7-triazapentacosan-25-oyl)-3-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-methylpiperidin-4-yl)oxy)-4-oxo-succinoyl-CPG(Tris-ß-GalNAc-C6-4-methylpiperidin-4-yl)oxy)-4-oxo-succinoyl-CPG) having a formula XXXII.
23. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) includes 4-((1-(4-((1,25-bis(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacet oxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-13-((3-((6-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)hexyl)amino)-3-oxopropoxy)methyl)-8,18-dioxo-11,15-dioxa-7,19-diazapentacosan-13-yl)amino)-4-oxobutanoyl)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)-piperidin-4-yl)oxy)-4-oxo-succinoyl-CPG(Tris-ß-GalNAc-C2-4-methoxyphenyl)(phenyl)methoxy)piperidin-4-yl)oxy)-4-oxo-succinoyl-CPG) having a formula XXXIII.
24. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) includes 4-((1-(4-((1,25-bis(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-13-((3-((6-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)hexyl)amino)-3-oxopropoxy)methyl)-8,18-dioxo-11,15-dioxa-7,19-diazapentacosan-13-yl)amino)-4-oxobutanoyl)-2-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)piperidin-4-yl)oxy)-4-oxobutanoic acid (Tris-ß-GalNAc-C2-4-methoxyphenyl)(phenyl)methoxy)methyl)piperidin-4-yl)oxy)-4-oxo-succinoyl-CPG) having a formula XXXIV.
25. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) includes 4-((1-(4-((1,25-bis(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-13-((3-((6-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)hexyl)amino)-3-oxopropoxy)methyl)-8,18-dioxo-11,15-dioxa-7,19-diazapentacosan-13-yl)amino)-4-oxobutanoyl)-4-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-methylpiperidin-3-yl)oxy)-4-oxo-succinoyl-CPG(Tris-ß-GalNAc-C2-4-methoxyphenyl)(phenyl)methoxy)methyl)-4-methylpiperidin-3-yl)oxy)-4-oxo-succinoyl-CPG) having a formula XXXV.
26. The synthetic delivery system (100) as claimed in claim 1, wherein the synthetic delivery system (100) includes 26-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxy methyl)tetrahydro-2H-pyran-2-yl)oxy)-14,14-bis((3-((6-(((2S,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)hexyl)amino)-3-oxopropoxy)methyl)-6-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-7-methyl-4,9,12,19-tetraoxo-5,16-dioxa-8,13,20-triazahexacosan-1-oyl-CPG(Tris-ß-GalNAc-C2-4-methoxyphenyl)(phenyl)methoxy)methyl)-7-methyl-4,9,12,19-tetraoxo-5,16-dioxa-8,13,20-triazahexacosan-1-oyl-CPG) having a formula XXXVI.
27. A method (300) to synthesize a synthetic delivery system (100) for targeted delivery of an agent (200) comprising:
a. preparing a first mixture to obtain a viscous solid having a formula XXXVIII from galactosamine pentaacetate having a formula XXXVII;
b. preparing a second mixture to obtain a compound having a formula XXXX from the viscous solid having the formula XXXVIII obtained from step a;
c. preparing a first suspension to obtain 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having a formula XXXXI from the compound having the formula XXXX obtained from step b;
d. preparing a third mixture to obtain a compound having a formula XXXXIV from 3,3'-((2-(((Benzyloxy)carbonyl)amino)-2-((2-carboxyethoxy)methyl)propane-1,3-diyl)bis(oxy))dipropionic acid (Tris-base) having a formula XXXXII;
e. preparing a fourth mixture to obtain a compound having a formula XXXXV from the compound having the formula XXXXIV obtained from step d;
f. preparing a fifth mixture to obtain a compound having a formula XXXXVI from the compound having the formula XXXXV obtained from step e and 5-(((2R,3R,4R,5R,6R)-3-acetamido-4,5-diacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy) pentanoic acid having the formula XXXXI obtained from step c;
g. preparing a sixth mixture to obtain a compound having a formula LXV from the compound having the formula XXXXVI obtained from step f;
h. preparing a seventh mixture to obtain a compound having either one of a formula LXVI, LXVII, LXVIII, or LXIX from the compound having the formula LXV obtained from step g along with at least one linker (130) having a formula XV, XVI, XVII, or XVIII;
i. preparing an eighth mixture to obtain a compound having either one of a formula LXX, LXXI, LXXII, or LXXIII from the compound having either one of the formula LXVI, LXVII, LXVIII, or LXIX obtained from step h; and
j. preparing a ninth mixture to obtain a synthetic delivery system (100) having either one of a formula LXXIV, LXXV, LXXVI, or LXXVII from the compound having either one of the formula LXX, LXXI, LXXII, or LXXIII obtained from step i.
28. The method (300) as claimed in claim 27, wherein the method (300) includes conjugating at least one agent (200) to the synthetic delivery system (100) having either one of a formula LXXIV, LXXV, LXXVI, or LXXVII.