Description:FIELD OF THE DISCLOSURE
The present disclosure generally relates to anti-cancer compounds. In particular, the present disclosure relates to biologically active compounds that inhibit breast cancer cell growth. The present disclosure also relates to greener synthesis of such biologically active compounds.
BACKGROUND
One of the leading causes of cancer death among women is breast cancer, which is one of the most frequently diagnosed as a very life-threatening cancer found in women. Breast cancer accounts for ~25% of all women's cancer cases and ~12% of all gender-specific cancer cases. Cancers of this type originate from the breast tissue and are mostly found in the inner lining of milk ducts and lobules. Approximately 10.4% of all cancer cases among women worldwide are breast cancer, making it the second most common non-skin cancer after lung cancer. Women are 100 times more likely than men to develop breast cancer as per one of the research studies. Worldwide, 2.3 million women were diagnosed with breast cancer in 2020, and 6,85,000 women died from it. Breast cancer was diagnosed in 7.8 million women in the past 5 years by the end of 2020, making it the most prevalent cancer in the world. 70,000 cases and 90,408 deaths were recorded in India according to the global cancer observatory (maintained by WHO) in 2020, representing the highest percentage (13.5%) of cancer cases out of all other cancers recorded in India.
The Indian subcontinent is diverse in terms of ethnicities, cultures, lifestyles, and economics. Several areas of the Indian healthcare program have not yet reached their full potential and are highly heterogeneous. Often, women do not seek medical care early because of literacy issues and economic concerns. About 60-75% of all breast cancers are invasive ductal carcinomas of no particular type. The special types of breast cancer account for up to 25% of all cases and there are now at least 17 such entities recognized by the WHO.
A number of therapeutic approaches are available to control breast cancer, including chemotherapy, radiotherapy, surgery (mastectomy), and combination therapy. Each of such approaches has their own side effects, well known in the art. Recent new breast cancer drugs as approved by the U.S. Food and Drug Administration bear high cost which becomes a financial burden for low-income and middle-class families. Thus, developing countries are unable to afford such expensive medicines. identify some synthesized anti-breast cancer compounds with fewer side effects and low cost, higher efficacy, lower toxicity than the existing drugs.
Therefore, there exists a need for synthesizing anti-breast cancer compounds which with minimal side effects, higher efficacy, lower toxicity along with low cost.
OBJECTS OF THE EMBODIMENT
One object of the present disclosure is to provide biologically active compounds that are potent anti-cancer agents.
Another object of the present disclosure is to provide the biologically active compounds that specifically target breast cancer cells.
Another object of the present disclosure is to provide the biologically active compounds that inhibits cancer cell proliferation with nano- to micro-molar range affinity against breast cancer cell lines MCF-7 and MDA-MB-231.
Another object of the present disclosure is to provide low dosage administration of the biologically active compounds to reduce breast cancer significantly.
Another object of the present disclosure is to provide the anti-breast cancer compounds that do not cross blood brain barrier to avoid neurodegenerative damage.
Another object of the present disclosure is to provide the anti-breast cancer compounds/agents that are specific, non-toxic and highly effective compounds useful to prevent and manage cancer.
Another object of the present disclosure is to provide the anti-breast cancer compounds/agents that have reduction in side effects including such as but not limited to cytotoxicity and non-specificity.
Another object of the present disclosure is to provide the anti-breast cancer compounds/agents that bear low cost.
Another object of the present disclosure is to provide a green process to provide the biologically active compounds that specifically target breast cancer cells.
In this respect, before explaining at least one embodiment of the present disclosure in detail, it is to be understood that the disclosure is not limited to in its application to the details of processing and to the arrangements of the components set forth in the following description or illustrated in the drawings. The disclosure is capable of embodiments in addition to those described and of being practised and carried out in various ways. Also, it is to be understood that the phraseology terminology employed herein, as well as the abstract, are for the purpose of description and should not be regarded as limiting.
SUMMARY
In an embodiment, the present disclosure discloses a biologically active anti-cancer compound (7) and derivative thereof. The compound (7) is an anti-cancer agent to inhibit breast cancer cells proliferation. The breast cancer cells are selected from one or more of MCF-7, HEK-293, and MDA-MB-231.
In another embodiment, an anti-breast cancer formulation comprising a biologically active compound is disclosed. The compound (7) includes benzothiazole-based dihydropyrimidine in the range of 0.1 µg/µL - 38.8µg/µL and excipients.
In yet another embodiment, a green process (900) for synthesizing biologically active anti-cancer compound (7) is disclosed. The process (900) includes preparing Fe-dopped Ce oxide nanoparticles, followed by treating ethyl 4-chloroacetoacetate (1) with propargyl alcohol in the presence of NaH and in THF. The process (900) involves converting ethyl 4-chloroacetoacetate to alkyne derivative thereof, the alkyne derivative is ethyl 3-oxo-4-(prop-2-yn-1-yloxy) butanoate and dissolving propargylated ethyl 4-chloroacetoacetate (2), 3-chlorobenzaldehyde (3) and 2-aminobenzothiazole (5) in C2H5OH to prepare the compound (7), followed by adding Fe-doped Ce oxide nanoparticles and refluxed thereto for 8 hours to get starting materials via a one-pot cascade reaction.
BRIEF DESCRIPTION OF THE DRAWINGS
Other objects, features, and advantages of the embodiment will be apparent from the following description when read with reference to the accompanying drawings. In the drawings, wherein like reference numerals denote corresponding parts throughout the several views:
Referring to Figure 1A, shows a chemical structure of a biologically active compound 7 as a potent anti-breast cancer agent, in accordance with an illustrative embodiment of a present disclosure;
Referring to Figure 1B, shows a reaction scheme depicting green synthesis of the biologically active compound 7, in accordance with another illustrative embodiment of the present disclosure;
Referring to Figure 2, shows cell viability evaluation of MCF-7 cells treated with different compounds including the compound 7, in accordance with the illustrative embodiment of the present disclosure;
Referring to Figure 3, shows cell viability evaluation of MDA-MB-231 cells treated with different compounds including the compound 7, in accordance with the illustrative embodiment of the present disclosure;
Referring to Figure 4, shows Cell viability evaluation of HEK-293 cells treated with different compounds including the compound 7, in accordance with the illustrative embodiment of the present disclosure;
Referring to Figures 5A-5C, show microscopic images of cells before any treatment: MCF-7 (breast-cancer cell line) (Figure 5A), MDA-MB-231 (another breast-cancer cell line) (Figure 5B); and HEK-293 (Human embryonic kidney cells, control: non-cancerous cell line) (Figure 5C), in accordance with the illustrative embodiment of the present disclosure;
Referring to Figures 6A-6D, show microscopic images of MCF-7 cells after treatment with the compound 7 (Figure 6A), PBS (solvent control) (Figure 6B), DMSO (vehicle control) (Figure 6C), and Tamoxifen (positive control) (Figure 6D), in accordance with the illustrative embodiment of the present disclosure;
Referring to Figures 7A-7D shows microscopic images of MDA-MB-231 cells after treatment with the compound 7 (Figure 7A), PBS (solvent control) (Figure 7B), DMSO (vehicle control) (Figure 7C), and Tamoxifen (positive control) (Figure 7D), in accordance with the illustrative embodiment of the present disclosure;
Referring to Figures 8A and 8B, show dose response curves of the compound (7) in MCF-7 and MDA-MB-231 cells for calculating IC50 respectively, in accordance with the illustrative embodiment of the present disclosure; and
Referring to Figure 9, discloses a flowchart depicting various steps involved in a process (900) for synthesis of the compound 7, in accordance with the illustrative embodiment of the present disclosure.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
The embodiments herein and the various features and advantageous details thereof are explained more fully with reference to the non-limiting embodiments that are illustrated in the accompanying drawings and detailed in the following description. Descriptions of well-known components and processing techniques are omitted so as to not unnecessarily obscure the embodiments herein. The examples used herein are intended merely to facilitate an understanding of ways in which the embodiments herein may be practiced and to further enable those of skill in the art to practice the embodiments herein. Accordingly, the examples should not be construed as limiting the scope of the embodiments herein.
Many modifications will be apparent to those skilled in the art without departing from the scope of the present invention as hereinbefore described with reference to the accompanying drawings.
Throughout this specification and the claims which follow, unless the context requires otherwise, the word “comprise”, and variations such as “comprises” and “comprising”, will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not the exclusion of any other integer or step or group of integers or steps.
As used herein, the singular forms “a”, “an”, “the” include plural referents unless the context clearly dictates otherwise. Further, the terms “like”, “as such”, “for example”, “including” are meant to introduce examples which further clarify more general subject matter, and should be contemplated for the persons skilled in the art to understand the subject matter.
Figure 1A shows a schematic view of a biologically active chemical compound (7) and derivative(s) thereof. The anti-cancer agent compound (7) is benzothiazole-based dihydropyrimidine. The compound (7) is an anti-cancer agent that inhibits breast cancer cells proliferation. The breast cancer cells may include such as but not limited to MCF-7, HEK-293, and MDA-MB-231. The compound (7) may be added to many additives or substituents or excipients. The additives or substituents or excipients may be polar and/or non-polar solvents. The excipients include such as but not limited to 0.01%-3% (v/v) ethanol, 0.01%- 1% (v/v) DMSO, 100% of Saline, and so on. Other additives may include such as but not limited to permissible edible colour during formulation as tablets, if any, microcrystalline cellulose, microcrystalline cellulose, pharmaceutical polymer coatings, croscarmellose sodium, colloidal silicon dioxide, lactose monohydrate, povidone, hypromellose, sodium starch glycolate etc. The compound (7) when added to various additives or substituents or excipients, constitutes a chemical formulation to target specifically breast cancer cells, may be in the range of 0.1 µg/µL - 38.8µg/µL. The derivatives of the compound (7) may be added to the additives or substituents or excipients in the range of 0.1 µg/µL - 59.64 µg/µL.
The formulation may be administered to the patient in nano-molar concentration ranging from 100 µM to 1 nM, which is very low in dosage. The formulation is administered in solid tablet or liquid syrup form.
Figure 1B shows a reaction scheme for the synthesis of the compound 7.
In step 1, iron-doped nanoparticles were prepared through processs as well known in the art. FeCl3 in ethanol was added to a solution of Ce(NO3)3.6H2O in C2H5OH and toluene. This is followed by the addition of oxalic acid solution in C2H5OH. An orange-coloured solution was formed, which was dried in a muffle furnace to get red-coloured iron-doped Ce-oxide nanoparticles (Scheme 2.1). These nanoparticles were further used for the synthesis of the compound 7 (Scheme 2.4). This is followed by the addition of oxalic acid solution in C2H5OH. An orange-coloured solution was formed, which was dried in a muffle furnace to get red-coloured iron-doped Ce-oxide nanoparticles (Scheme 2.1). These nanoparticles were further used for the synthesis of compounds 7 (Scheme 2.3).

Scheme 2.1 Preparation of Fe-dopped Ce oxide nanoparticles
In step 2, ethyl 4-chloroacetoacetate was converted to its alkyne derivative, ethyl 3-oxo-4-(prop-2-yn-1-yloxy) butanoate (2), by treating ethyl 4-chloroacetoacetate (1) with propargyl alcohol in the presence of NaH and in THF (Scheme-2.2).

Scheme 2.2 Synthesis of ethyl 3-oxo-4-(prop-2-yn-1-yloxy) butanoate.
Same product forms when rection is carried out in polar aprotic solvents like DMSO, Acetonitrile etc.
Target compound (7) was synthesised by taking (1 mmol or 38.78% w/w) propargylated ethyl 4-chloroacetoacetate (2), (1 mmol or 29.60% w/w) 3-chlorobenzaldehyde (3) and (1 mmol or 31.62% w/w) 2-aminobenzothiazole (5) dissolved in C2H5OH. To this, 0.05% (with respect to reactant 3) Fe-doped Ce oxide nanoparticles were added and refluxed it for 8 hr to get starting materials via a one-pot cascade reaction Scheme 2.3.

Scheme 2.3 Synthesis of alkyne derivatives of benzothiazole-based dihydropyrimidine which is compound 7 respectively, using Fe-doped Ce oxide nanoparticles in C2H5OH.
Same product forms when reaction is carried out in polar protic solvents like methanol, butanol, tertiary butanol etc.
Structures of derivative compounds 6, 14a-d and 15 of the compound (7) are shown in following Table 1. Structures of derivative compounds 16a-d and 17 of the compound (7) as shown in Table 2.

Table 1: Structures of derivative compounds 14a-d and 15 of claimed compound 7.

Table 2: Structures of synthesised compounds 16a-d and 17

S. No. Compound No Structure of compounds Melting Point Yield (%)
1 6 114-116°C 90
2 14a 141-143°C 88
3 7 117-119 °C 91
4 16c 150-152 °C 86
Table 3: Synthesized compound 7 and its derivatives which are found active against cancer cell lines with their IUPAC name

S. No. Compound No Structure of compounds Melting Point Yield (%)
1 14b 145-147°C 89
2 14c 148-150 °C 85
3 14d 146-148 °C 87
4 15 139-141 °C 85
5 16a 144-146 °C 87
6 16b 147-149 °C 89
7 16d 149-151 °C 88
8 17 143-145 °C 84
Table 4: Other derivatives of the compound 7, which are found less active against cancer cell lines with their IUPAC name.
Process for the synthesis of derivative compounds 14a-d and 15.
Reactants 6 (1 mmol) and isatin azides 10a-d (1 mmol) or acylated glucose azide (13) were dissolved in DMF-H2O (1:1) and stirred; sodium ascorbate (0.4 mmol) and CuSO4.5H2O (0.2 mmol) were added as catalysts and reaction contents were heated for 14-18 min at 60°C (Scheme 2.6).

Scheme-2.6 Synthesis of 1,2,3-triazole derivatives of thiazole-fused dihydropyrimidines 14a-d and 15.
Process for the synthesis of derivative compounds 16a-d and 17.
Reactants 7 (1 mmol) and isatin azides 10a-d (1 mmol) or acylated glucose azide (13) were dissolved in DMF-H2O (1:1) and stirred; sodium ascorbate (0.4 mmol) and CuSO4.5H2O (0.2 mmol) were added as catalysts and reaction contents were heated for 14-18 min at 60°C (Scheme 2.7).

Weight % of each of the anti-cancer compound (7) and derivatives thereof to be added to additives or substituents or excipients as follows:
Compound 7: 0.1 µg/µL - 38.8µg/µL
Compound 14a: 0.1 µg/µL - 59.64 µg/µL
Compound 6: 0.1 µg/µL - 35.5 µg/µL
Compound 16c: 0.1 µg/µL - 62.63 µg/µL
Experimental details
Various alternative compounds were synthesized and tested along with the compound (7) to check potential of the compound (7). The compound (7) or the corresponding formulation has a tendency to inhibit the MCF-7 and MDA-MB-231 cells growth with high specificity and efficacy.
MCF-7 (Michigan Cancer Foundation-7):
Origin: MCF-7 cells originate from human breast adenocarcinoma.
Function: MCF-7 cells are frequently employed as a model for estrogen receptor (ER)-positive breast cancer. They exhibit responsiveness to hormones and express estrogen receptors, making them instrumental in investigating the impact of estrogen and anti-estrogen compounds. Studies involving MCF-7 cells often center around breast cancer biology, endocrine therapy, and the elucidation of mechanisms related to hormone receptor signaling.
MDA-MB-231(MD Anderson - Mammalian Breast 231):
Origin: MDA-MB-231 cells are derived from human breast adenocarcinoma.
Function: MDA-MB-231 cells serve as a model for triple-negative breast cancer, signifying the absence of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) expression. Known for their invasiveness, these cells are crucial for studying metastasis, tumor cell migration, and invasion mechanisms. Researchers utilize MDA-MB-231 cells to explore potential therapeutic targets for aggressive forms of breast cancer.
HEK293 (Human Embryonic Kidney 293):
Origin: HEK293 cells are derived from human embryonic kidney cells.
Function: Unlike cancer cells, HEK293 cells are non-cancerous and commonly employed as controls in experiments. Originating from non-cancerous tissues, they play a versatile role in biotechnology and molecular biology research. HEK293 cells are utilized for producing recombinant proteins and investigating diverse cellular processes, such as signal transduction, gene expression, and protein-protein interactions. While not cancerous, HEK293 cells contribute to understand relevant cellular processes in cancer research by providing a non-cancerous reference point in experiments.
In an exemplary embodiment, the compound (7) or the corresponding formulation shows very low IC50 values of 24 nM and 55 nM against the MDA-MB-231 and MCF-7 cell lines respectively, that are comparable to or better than known drug viz., tamoxifen. The compound (7) shows insignificant cytotoxic profiles against HEK293 cell lines (non-cancerous) making it suitable lead molecules for anti-cancer drug development.
Figure 2 shows MTT cell viability assay result with % viability vs compounds/drugs upon treating the MCF-7 cells with 100 µM compounds and 100 µM of positive control tamoxifen treated cells were taken as anti-breast cancer positive controls while DMSO and PBS treated cells were treated as vehicle control and solvent control respectively. All the treatments were done in triplicates.
Figure 3 shows MTT cell viability assay result showing % viability vs compounds/drugs upon treating the MDA-MB-231cells. with 100 µM compounds and 100 µM of positive control tamoxifen treated cells were taken as anti-breast cancer positive controls while DMSO and PBS treated cells were treated as vehicle control and solvent control respectively. All the treatments were done in triplicates. Figure 4 shows MTT cell viability assay result showing % viability vs compounds/drugs upon treating the HEK-293 cells. with 100 µM compounds and 100 µM of positive control tamoxifen treated cells were taken as anti-breast cancer positive controls while DMSO and PBS treated cells were treated as vehicle control and solvent control respectively. All the treatments were done in triplicates.
All the synthesized compounds were evaluated for their anti-breast cancer activity by using MTT viability assay. MCF-7 and MDA-MB-231 cell lines were used to investigate the anti-breast cancer activity of compounds whereas, HEK293 cell line was used to investigate the non-specific effects of compounds. After compound treatment, MDA-MB-231, MCF-7, and HEK-293 cell lines were tested for cell viability assay using MTT assay. The cells appear violet after MTT treatment. MTT is reduced to formazan crystal which is purple in colour by the metabolically active cells by the action of oxidoreductase enzymes in the mitochondria. Data indicates low cell viability in case of MDA-MB-231 and MCF-7 cells in presence of 100 ?M doses of compound (7), while none of the tested compounds found to be toxic against HEK-293 as indicated by dark purple colour upon treatment with same high doses of the compounds as well as control drug tamoxifen. % cell viability was then calculated by normalizing the data taking the absorbance of cell viability due to PBS and DMSO treatment as 100%. When MCF-7 cells were treated with compounds with 100 ?M, wells with the compound (7) which could kill 48% MCF-7 cells at a dose of 100 ?M while tamoxifen killed only 40% cells.
In contrary, when MDA-MB-231 cells were treated with compounds with 100 ?M, wells with the compound (7) had killed 52% MDA-MB-231 cells at a dose of 100 ?M. while tamoxifen killed only 37% cells. Such calculated results are shown in the MCF-7 (Figure 2), MDA-MB-231 (Figure 3) and HEK-293 (Figure 4) as plotted by % cell viability versus compound names. Nevertheless, the four compounds (compound 6, 14a, 7, 16c) were showing anti-breast cancer activities, and no toxicity against the non-breast cancer cells like HEK-293 cells, and thus they were further studied against the HEK-293.
The cells were treated with different compounds for 48 h and observed under microscope (Eclipse Ni-E, Nikon) before and after compound treatment. Cells were observed under microscope without addition of any kind of compounds, drugs, or control solutions. Following the cells were treated with 100 µM concentration of synthesized compounds, 100 µM concentration of positive control drugs (tamoxifen), and 1% of vehicle control solutions (DMSO, and PBS).
Figures 5A-5C show microscopic images of cells before any treatment: MCF-7 (breast-cancer cell line), MDA-MB-231 (another breast-cancer cell line), and C HEK-293 (Human embryonic kidney cells, control: non-cancerous cell line). Figures 6A-6D show microscopic images of MCF-7 cells after treatment with the compound (7), PBS (solvent control), DMSO (vehicle control), and d Tamoxifen (positive control). All the treatments were done in triplicates. Figures 7A-7D show microscopic images of MDA-MB-231 cells after treatment with the compound (7), PBS (solvent control), DMSO (vehicle control), and Tamoxifen (positive control). All the treatments were done in triplicates.
The microscopic analysis was done before and after compound/drug treatment using Nikon microscope at 10X magnification. MCF-7 cells treated where some of the compounds for 48 h have shown significant changes in their morphology. Cellular shrinkage and dead cells were also seen after treatment in comparison to untreated cells. The untreated MCF7 cells and HEK293 were appeared healthy under the microscope whereas the cells treated with vehicle control DMSO and PBS had least dead cells and cells were growing and flourishing as normal in these wells. Whereas the cells treated with positive control tamoxifen was mostly dead as expected.
The microscopic analysis was performed before and after compound/drug treatment using Nikon microscope at 10X magnification. MCF-7 cells treated with compound (7), and tamoxifen for 48 h have shown significant changes in their morphology which can clearly be seen in photomicrographs shown in Figures 6A-6D. Cellular shrinkage and dead cells were also seen after treatment in comparison to untreated cells. The untreated MCF7 cells (Figure 5A), MDA-MB-231 (Figure 5B) and HEK-293 (Figure 5C) were appeared healthy under the microscope whereas, the compound (7) treated MCF-7 (Figures 6A-6D) cells which are mostly dead and morphologically very different too from normal MCF7 cells. The cells treated with vehicle control DMSO and solvent control PBS had no dead cells and cells were growing and flourishing as normal in these wells. Similar results can be seen in case of MDA-MB-231 (Figure 7A-7D) cells. Whereas the cells treated with positive control tamoxifen was mostly dead as expected as shown in Figures 5A-7D.
Hence, compound (7) has the highest anti-breast cancer activity out of all the compounds and have better cancer selective effect. These compounds inducing the significant cell death and morphological changes in comparison to controls.
The cells were treated with different concentration of the compound (7) for 48 h and observed under microscope (Eclipse Ni-E, Nikon) before and after compound treatment. Cells were observed under microscope without addition of any kind of compounds, drugs, or control solutions. Following the cells were treated with 100 µM concentration of synthesized compounds, 100 µM concentration of positive control drugs (tamoxifen), and 1% of vehicle control solutions (DMSO, and PBS).
In order to determine the IC50 values, further investigation was conducted on the compound (7) (Figures 8A and 8B) by treating MDA-MB-231 and MCF-7 cells with varying doses of the compound (7) was tested in the dose range of 100 µM to 1 nM due to its potency even at low doses (Figures 8A and 8B ). Based on the results presented in Figures 8A and 8B, the calculated IC50 value of the compound (7) was approximately 24 nM against MDA-MB-231. Such findings suggest that the compound (7) demonstrates the potential for anti-breast cancer efficacy. Surprisingly, similar results were observed in MCF-7 cells, with IC50 values of approximately 55 nM for the compound (7). Such results indicate that the compound (7) is the effective compound in both MCF-7 and MDA-MB-231 cell lines.
Further experimentation is necessary to elucidate the underlying mechanisms by which these molecules exert their anti-breast cancer effects. This should include investigating whether these compounds act through anti-apoptotic pathways, anti-metastatic pathways, or other anti-oncogenic pathways. These additional experiments will provide a better understanding of the potential anti-breast cancer mechanisms of these compounds.
The compound (7) has potential to serve as lead compounds for development of anti-breast cancer small molecule that are not toxic, have low effective dose and are devoid of side effects on other normal cells. The compound (7) has melting point in the range of 117 deg C and 119 deg C, and yield of about 91%.
Figure 9 discloses a flowchart depicting a process (900) for the preparation of the anti-cancer compound (7), steps thereof already discussed hereinabove. The process (900) is a green process because conventional process involves the use of Brönsted acids, Lewis acids, nanoparticles, mixed metal or metal oxide nanoparticles, and ionic liquids, which are costlier as compared to that of the present invention. Also, the conventional arts tend to cause harm to the environment when draining after the reaction and solvents like DMF, acetonitrile, THF, etc. also harm the environment. The process (900) is less costly as the reactants used therein are easily available and cheap. The nanoparticles used hereinabove are recyclable, which lowers the price of the catalyst. Such nanoparticles can be recycled for the next reaction up to 5 cycles. Ethanol, which is used as a solvent, is also cheap and a green solvent.
The foregoing descriptions of exemplary embodiments of the present disclosure have been presented for purposes of illustration and description. They are not intended to be exhaustive or to limit the disclosure to the precise forms disclosed, and obviously many modifications and variations are possible in light of the above teaching. The exemplary embodiments were chosen and described in order to best explain the principles of the disclosure and its practical application, to thereby enable others skilled in the art to best utilize the disclosure and various embodiments with various modifications as are suited to the particular use contemplated. It is understood that various omissions, substitutions of equivalents are contemplated as circumstance may suggest or render expedient but is intended to cover the application or implementation without departing from the spirit or scope of the claims of the present disclosure.
, Claims:We Claim
1. A biologically active chemical compound (7) and derivative thereof,

2. The compound (7) as claimed in claim 1, wherein the compound is an anti-cancer agent to inhibit breast cancer cells proliferation.
3. The compound (7) as claimed in claim 2, wherein the breast cancer cells are selected from one or more of MCF-7, HEK-293, and MDA-MB-231.
4. An anti-breast cancer formulation comprising a biologically active compound comprising benzothiazole-based dihydropyrimidine in the range of 0.1 µg/µL - 38.8µg/µL and excipients.
5. The anti-breast cancer formulation as claimed in claim 4, wherein dosage of administering the formulation is in nano-molar concentration ranging from 100 µM to 1 nM.
6. The anti-breast cancer formulation as claimed in claim 4, wherein the formulation is administered in oral form.
7. The anti-breast cancer formulation as claimed in claim 4, wherein the formulation is administered in forms selected from one or more of powder, solid tablet, and syrup.
8. The anti- breast cancer formulation as claimed in claim 4, wherein the compound is administered in alone or in conjugation with additives or substituents or excipients.
9. A green process (900) for synthesizing biologically active anti-cancer compound (7), the process (900) comprising:
preparing Fe-dopped Ce oxide nanoparticles;
treating ethyl 4-chloroacetoacetate (1) with propargyl alcohol in the presence of NaH and in THF;
converting ethyl 4-chloroacetoacetate to alkyne derivative thereof, the alkyne derivative is ethyl 3-oxo-4-(prop-2-yn-1-yloxy) butanoate;
dissolving propargylated ethyl 4-chloroacetoacetate (2), 3-chlorobenzaldehyde (3) and 2-aminobenzothiazole (5) in C2H5OH to prepare the compound (7);
adding Fe-doped Ce oxide nanoparticles and refluxed thereto for 8 hours to get starting materials via a one-pot cascade reaction; and
obtaining the compound (7).