Claims:We claim the following invention disclosers from our research outcomes and the scope of the invention is defined based on the following claims,
Claim:
1. We present, a facile method for the synthesis of amino acid based block copolymer T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] followed by the design and development of football shaped buck minister fullerene type porous capsules.
a). The shape and size of the capsules is tunable from micro to nanoscale pores and its morphology can be tuned by changing external factors like temperature, solvents and sonication methods.
b). These fullerene types of amino acid based block copolymer capsules were unique for various nanotechnology and pharmaceutical applications. These football shaped capsules were fabricated without using any precursors and surfactants. Based on IC50 and MIC values of MTT assay these are evidenced and claimed as biocompatible, biodegradable and bio-safe carrier capsules.
c). The capsule size ranges from ~10 to 100 µm (in diameter) and porosity on the surface of the capsule is in the range of ~500 nm to 1 µm (in diameter).
2. As mentioned in claim 1, the surface of the football shaped or fullerene type of capsules has been functionalized with active functional groups of polymers such as acids (-COOH), amide (-NH), ester (-COOR), hydroxyl (-OH) etc.
3. As mentioned in claim 1, along with surface properties of capsules, the porosity throughout the surface of the capsule has been observed from SEM. These football shaped capsules are spherical in shape and size of the capsules can be tuned by changing the reaction conditions and factors such as temperature, concentration, solvents, sonication methods etc.
4. As mentioned in claim 1, The pore size (deep excavation) is in the range of 10 µm (in diameter) for the football shaped fullerene type capsules. Whereas, the size of the capsule is in the range of ~30 to 50 µm (figure 1). The porosity on the surface of the smaller capsule (figure 4) is in the range of 500 nm to 1 µm (in diameter).
5. As mentioned in claim 1, these pore formulation on the surface of the spherical capsules has been engineered through the mechanistic approach followed by the molecular imprinting of hydrophobic (PCL) particles on the surface of the soft spherical capsules designed and developed by using [(PNIPAM)-b-(MeO-PEG-NH)].
6. As mentioned in claim 1, such porous spherical shaped capsules are recommended as potential carrier capsules and tools for the loading and release of nanomedicines and anticancer drugs for targeted drug delivery systems. , Description:Surface Functionalized Football Shaped Porous Nanocapsules as Potential Payload for Nanomedicines and Cancer Theranostics

Field of Invention
The present invention relates to, the polymer synthesis followed by the engineering sciences to retard the design and development of football/fullerene shaped porous capsules and nanomaterials rejuvenated through the materials engineering and nanoscience and technology.
The objectives of this invention
Synthesis of tri-block copolymer (T-BCP) of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] followed by the design and development of bio-inspired nanoarchitectured football shaped/buckminister fullerene type of large scale porous capsules for various theranostics and healthcare applications.
Background of the invention
Synthesis of tri-block copolymer (T-BCP) of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] followed by the design and development of bio-inspired nanoarchitectured materials can help in minimizing the side effects and cytotoxicity. This football shaped or buckminister fullerene type of large scale porous capsules can be used as a potential drug or nanomedicine carriers due to presence of large scale porosity (C Amgoth et al., [2016], Nanotechnology, 27, 125101). Targeted drug delivery and pH sensitive response of carrier capsules can boost the drug delivery systems with various theranostics and healthcare applications. These types of bio-inspired nanoarchitectured and surface functionalized porous capsules are recommended for potential carriers for drug loading and target specific release. Core idea to synthesize T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] is to design and develop amino acid based nanocarrier systems to minimize the existing issue in drug delivery (DDSs) systems. The biodegradable, biocompatible and bio-safe polymers with thermosensitive and pH responsive nature can help in targeted drug delivery with reduce the toxic issues (A P Esser Kahn et al., [2011], Macromolecules, 44, 5539–5553). These football shaped porous capsules of T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] are further recommended for cell seeding (growth) and tissue engineering applications. The amino acid based block copolymer of T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] will enhance the adjuvant based activity on cancer cells. The capsule size ranges from ~10 to 100 µm (in diameter) and porosity on the surface of the capsule is in the range of ~500 nm to 1 µm (in diameter). Polymer nanocapsules with surface functionalized features and porosity is one of the most promising tools for drug delivery and biomedical therapeutics. The design and development of amino acid based nanocarrier capsules in association with nanotechnology and nanobiotechnology plays a vital role for advanced drug and gene delivery followed by the cancer therapy and treatment. Various polymers like MeO-PEG-NH2, PNIPAM, and PCL are biodegradable and biocompatible for molecular biotechnology because of their harmless effects over the various cancer theranostics (T Chen et al., [2012], ACS Appl. Mater. Interfaces, 4, 5766-5774). A very limited number of amino acid based porous polymer nanocapsules and particles have been reported for similar purposes. In most of the reports, polymer–drug nanoformulations were developed in the many facile and modified ways. The drug molecules are blended with a soluble polymer at the stage of formation of capsules and particles through ring opening polymerization, addition polymerization and re-precipitation during formation of polymer capsules and particles from monomers to polymers. These are believed as promising tools or carrier capsules for varieties of nanomedicines and anticancer drug molecules for subsequent drug delivery systems (R Pantani et al., [2013], Polym. Degrad. Stab. 98, 1089–96). The impregnation of such bio-inspired nanoarchitectured capsules impact direct asperities followed by the surface functionalized properties, their dimensions, size, shape, morphology, porosity, chemical nature, and regenerative medicinal activity (M Hu et al., [2012], Biomacromolecules, 13, 3552-3561). Scientists and researchers have already designed and developed various carrier capsules for drug delivery systems such as liposomes, nanocapsules, polymerosomes, quantum dots, micelles, carbon nanotubes, nanocages, solid lipid nanoparticles, and dendrimers etc. The carrier capsules and nanoparticles for drug delivery systems are usually synthesized through the emulsion polymerization, micro-precipitation, addition polymerization, emulsion polymerization, and phase separation followed by the self-assembly process through the formation of micelles and dendrimers (J Fan et al., [2012], Biomacromolecules, 13, 4126-4137). The hollow polymer nanocapsules were also designed through the layer-by-layer deposition of polymer residues on a template followed by the extraction of layers. Interestingly, the chemical structure and functionality of the polymer chains can play a crucial role in the preparation of nanoparticles and capsules with surface modified features. The surface properties, size, shape, morphology, porosity are highly rely on the presence of active functional groups present in the synthesized block copolymers (S Zamani et al., [2012], Polymer, 53, 5723-5736). Furthermore, these properties can also rely on the synthesis methods, solvents used for synthesis purpose, reaction conditions, pH of the reactions etc. Numerous block copolymers are being designed for drug delivery applications but for all these are having limitations that confine the low drug delivery efficacies. The existing nanoparticles and capsules are very unstable in physiological conditions, issue with biodegradability, non-biocompatibility, and issues concern with bio-safety followed by the unable to protect the drugs, disrupt and degrade the activity of nanomedicines and anticancer drugs while carrying them to deliver organ or tissue specifically. The problems associated with excretion of carrier capsules and nanoparticles after successful delivery of drug molecules to the tissue and tumor targeted. The prolonged remaining of carrier capsules or nanoparticles inside the body can lead to kidney damage and other health issues. The nonspecific drug loaded nanoparticles or capsules can mislead the delivery trajectory followed by the delay, destroy, and alter the standard activities of porous capsules which cause the side effects and cytotoxicity. Therefore, we designed a novel and unique bio-inspired nanoarchitectured porous polymer capsules of MeO-PEG-NH2, PNIPAM, and PCL (P Paik et al., [2009], ACS Appl. Mater. Interfaces, 1, 1834–42). The synthesis of ABC-type tri-block copolymer T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] lead to self-assembly of active functional groups and helps in formulation of spherical football shaped capsules with large scale porosity as well smooth and rough surface capsules and nanoparticles. The L-Glutamic acid-5-benzyl ester, L-Aspartic acid-4-benzyl ester, glycine in combination with PCL, and PNIPAM were used to synthesize block copolymers for the design and development of porous materials for drug delivery and other theranostic applications. The individual PNIPAM, MeO-PEG-NH2, PCL based capsules were previously reported on hollow-mesoporous, microporous and nanoporous capsules synthesis and their usage in drug delivery systems. The hydrophobic PCL with molecular imprinting on capsules or particle surface has been discussed in detail with the presence of MP-SiO2 nanoparticles and surfactant free micelle templates where we found that the removal of templates is essential for loading sufficient quantities of drugs (A Sharma et al., [2008], J. Appl. Phys., 103, 07A308). However, the SDS (sodium dodecyl sulphate) is usually used for the formulation of mesoscale porosity within the SiO2 nanoparticles followed by the heat treatment at higher temperatures such as 400 to 800 ?C. We have synthesized mesoporous SiO2 nanoparticles and their hybridization with thermosentitive PNIPAM thin films and insertion or incorporation of MP-SiO2NPs inside the nanoscale polymer capsules for selective loading of chiral drug molecules with enhanced biocompatibility. In a very recent work, we revealed the effectiveness of gold nanoparticles which are acting as self-assembling agents to randomly distributed nanopartilces into hollow core-shell capsules with pH based structural disintegration and release of drug molecules. It is believed that the amino acid based block copolymer of T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] will enhance the adjuvant based activity and regenerative medicinal activity to kill and rupture the cancer cells. These are recommended as promising tools for drug delivery systems to cure and treat various types of cancers for better healthcare and theranostics (C Amgoth et al., [2018], Materials Science & Engineering-C, 92, 790-799).
Description of Prior Art
We disclose a unique and novel outcome of football shaped or buckminister fullerene type of large scale porous nanocapsules which was designed from the tri-block copolymer of poly-N-isopropyl acryl amide (PNIPAM), a-methoxy-?-amino ethylene glycol (MeO-PEG-NH2), and poly-e-caprolactone (PCL). The final synthesized and confirmed block copolymer has been denoted as [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)]. The synthesis of tri-block copolymer (T-BCP) has been followed by the modified facile ring opening polymerization (ROP) with the combination of two solvents such as dry tetrahydrofuran (THF), extra pure isopropanol (IPA) and acetone in the 1:2:1 ratio at inert atmospheric conditions. The synthesis of T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] carried out under inert atmospheric conditions which are maintained through the continuous passage of N2 gas. Initially, ~2.5 g of PNIPAM has been dissolved in the 50 mL of dry THF solvent, ~2.5 g of MeO-PEG-NH2 with 2000 Da has been dissolved in 100 mL of in isopropanol solvent and ~2.5 g of PCL has been dissolved in 50 mL of acetone. All the three polymers are weighed in equivalent amount i.e. 2.5 g each (1:1:1) and finally added into a 250 mL round bottom (RB) flask and mixture is allowed to stir for 36 h at ~60 ?C. The final concentration of mixture of T-BCP with THF, IPA and acetone has been adjusted to 0.01 mmol. The reactions between thermo-sensitive PNIPAM, hydrophilic a-methoxy-?-amino poly ethylene glycol (2000 Da) (MeO-PEG-NH2) and hydrophobic PCL has lead to formation of cyclic anhydride ring and its opening for the addition polymerization with (MeO-PEG-NH2). Further, this AB-type di-block copolymer of [(PNIPAM)-b-(MeO-PEG-NH)] has been reacted with N-terminals of MeO-PEG-NH2 for the formation of tri-block copolymer of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)]. The addition polymerization of PCL to di-block copolymer has been followed by the modified precipitation polymerization methods with the help of moisture free (dry) tetrahydrofuran (THF) and IPA solvents. The mixture of T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] is dried under reduced pressure (~50 pa) followed by the lyophilization. The yield of the final T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] product is ~6.8/7.5 g (wt/wt %) which is equal to ~90.6%. The final T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] powder is thermosensitive and amphiphilic (hydrophobic and hydrophilic) in nature. The freeze dried (lyophilized) T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] was once again dried at room temperature by keeping in desiccators for 48 h. To check the morphology of synthesized T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)], ~500 µg mL-1 was dispersed in 1 mL of ethanol (CH3-CH2-OH) (high pure 98%). Dispersion followed by the ultra sonication for 2 min at ~25 ?C. The dispersion of 5 µL drop casted on the clean and neat piece of glass slide and it is further allowed to dry at room temperature and well dried sample has been used to various microscopic imaging such as SEM, TEM, AFM, BET, FE-SEM etc. Since, T-BCP is non-conductive in nature, the sample before SEM imaging has been sputter coated with metal (Au) nanoparticles throughout the surface of the sample to get the conductivity while SEM imaging. The SEM imaging for T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] dispersion has been carried out at 15 kV accelerating voltages. The SEM micrographs are acquired from lower to higher magnifications. The football shaped spherical capsule with large scale porosity has been captured through the SEM and micrographs were produced in the respective sections.
Summary of the invention
The main strategy behind the formation of football or buck minister fullerene shaped nanoporous capsules within the block copolymer T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] can figure out with the effect of solvents, self-assembly as well as other factors such as temperature and sonication methods. However, these capsules of block copolymer T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] are stable at body temperature (~37.4 °C). The thermosensitive PNIPAM, hydrophilic MeO-PEG-NH2 and hydrophobic PCL lead to the formulation of football shaped porous capsules with large scale holes which seems to be deep excavation on the surface of the capsule, ditch and trenches for cell culture, hollow cavities, excavation and potholes for cell growth, porous surface, and porous block copolymer capsules. Design and development of football shaped capsules of block copolymer T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] with deep excavation can help as a payload for anticancer drug molecules and nanomedicines for pharmacokinetics and dynamics in cancer therapeutics. Designing of deep excavations on the surface of the block copolymer capsules of T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] has been happened due to the molecular imprinting of hydrophobic PCL molecules. These deep excavations can help in cell growth, tissue engineering and cellular imaging for cancer theranostics.
Detailed description of the invention
Our invention discloses a novel approach for the synthesis of block copolymer followed by the preparation, fabrication, design and development of football or buck minister fullerene shaped porous capsules of T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] for drug delivery systems (DDSs). The deep excavations on the surface of spherical capsules can be filled with anticancer drugs and nanomedicines. These football shaped porous capsules are the potential tools for the nanomedicnes for targeted drug delivery. The large scale deep excavation can be used for cell seeding for better growth (culture); pharmacokinetics and pharmacodynamics followed by the cancer therapeutics and diagnostics, tissue engineering and other applications. The synthesis of block copolymer T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] and development of porous capsules with active functional groups were self-assembled into spherical shape morphology. The deep excavation is due to molecular imprinting of hydrophobic PCL moieties. The block copolymer T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] with large scale hollow cavities and potholes on the surface of the capsules were functionalized for smooth and rough surface of spherical capsules which is a unique approach in the biomedical and nanobiomedicine fields. Amino acid based block copolymer T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] is an essential composite with medicinal and adjuvant characteristic properties which can enhances usage in pharmaceutics. Block copolymers play active role in development and designing of effective and potential capsules as payload for anticancer drug molecules and nanomedcines as such as transporters of slow and safe controlled target specific release and cellular imaging studies. Furthermore, capsules of (ranges from ~10 to 100 µm (in diameter) block copolymer T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] has deep excavation or large scale porosity (pore size on the surface of the capsule is in the range of ~500 nm to 1 µm (in diameter). The deep excavation on the surface of the capsules are most important for cell seeding (cell growth and cell transporter) to cure cellular damages and tissue engineering (TE) as well as for patch therapy (PT). Further, these porous capsules are functionalized with active functional groups and smooth surface of the capsule can be changed or modified into rough surface. Design, development and fabrication of smaller particles and capsules with functionalized surfaces can import large surface area, large number of porosity, hollow dome, biocompatibility and fewer side effects. Capsules of football shaped or buck minister fullerene type morphology which has been designed and developed from the amino acid based block copolymer T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] are themselves active towards the cancerous cells due to presence of active functional groups like –NH2, -COOH, -CH3, -CH3O which improves the effectiveness of transporters for cancer diagnostics and other biomedicinal theranostic applications.

6 Claims & 5 Figures

Brief description of Drawing
In the figures which are illustrate exemplary embodiments of the invention.
Figure 1 SEM micrograph represents the football shaped buck minister fullerene type morphology of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] based capsule with deep excavations/large scale porosity on the surface of the capsule.
Figure 2 SEM micrograph of spherical shaped [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] based capsule with rough surface functionalized by the active functional groups.
Figure 3 FE-SEM micrograph of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] based capsules with non-functionalized smooth surface.
Figure 4 SEM micrograph illustrates the size, shape and morphology of capsule with surface profile and porosity distribution throughout the surface.
Figure 5 SEM micrographs (a-c) correspond to spherical capsules with rough surface and (d-f) corresponds to spherical capsules with smooth surface. The images were captured at 15 kV accelerating voltage with varied working distances. The scale bar is 10 µm.
Detailed description of the drawing
As described above, the present invention discloses about the methods, methodologies followed to synthesis of the T-BCP of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)]. Furthermore, design and development of football shaped fullerene type of porous capsules with surface functionalized properties.
The SEM micrograph in figure 1 corroborates the football shaped capsules with deep excavations on the surface of the capsules. This has been patterned through the hydrophobic (PCL) molecular imprinting on soft spherical surface of [(PNIPAM)-b-(MeO-PEG-NH)]. The captured capsules are in micron scale size i.e. 10 µm. the size of the capsules can be tuned or reduced to certain nanoscale upon impinging and controlling the number of monomer units through the polymerization systems.
The SEM micrograph in figure 2 corroborates the football shaped capsules with rough surface functionalized due to active functional groups present in the polymers. The size, shape, morphology of the designed capsules has been controlled worthwhile controlling the factors such as solvent, concentration, reaction conditions, synthesis approaches.
The figure 3 SEM micrograph of [(PNIPAM)-b-(MeO-PEG-NH)-b-(PCL)] based capsules corroborates the non-functionalized smooth surface of the capsule. The size of the capsules ranges from 1 to 10 µm and figure shows uneven distribution of capsules with highly inconsistent capsules. The different sized nanoparticles are obtained because of hydrophobic and hydrophilic nature of polymers.
The SEM micrograph in figure 4 illustrates the size, shape and morphology of capsule with surface profile and porosity distribution throughout the surface of the capsule. The exact reason and mechanism for the pore formation on the surface of the capsules is evidenced through the molecular imprinting of hydrophobic (solvent hating/repelling) particles interacted with soft surfaces of hydrophilic (solvent loving) particles.
The SEM micrographs (a, b, c) in figure 5 correspond to spherical capsules with rough surface and (d, e, f) corroborate the spherical football shaped capsules with smooth surface. The images were captured at 15 kV accelerating voltage with varied working distances. The scale bar is 10 µm.