A therapeutic agent that is a heat shock protein (HSP) inhibitor and its use in cancer treatment

Abstract

The invention relates to the use of the compound ES3707 as a therapeutic agent for the treatment of cancer diseases and for alleviating symptoms caused by such diseases, specifically by inhibiting heat shock proteins involved in cell division and by preventing resistance induced by clinical drugs. The therapeutic agent subject to the invention is characterized by its enhanced efficacy when used in combination with photodynamic therapy.

Description

[0001] A THERAPEUTIC AGENT THAT IS A HEAT SHOCK PROTEIN (HSP) INHIBITOR AND ITS USE IN CANCER TREATMENT

[0002] TECHNICAL FIELD

[0003] The present invention relates to the use of the compound ES3707 as a therapeutic agent for the treatment of cancer and for alleviating symptoms associated with cancer. The compound functions through the inhibition of heat shock proteins (HSPs), which play a role in cell division, and through the prevention of resistance caused by clinical drugs. The therapeutic agent of the invention is characterized by its enhanced efficacy when used in combination with photodynamic therapy. It is an effective agent particularly for the treatment of triple-negative breast cancer.

[0004] BACKGROUND

[0005] Cancers are typically characterized by continuously and rapidly growing tumors. Cancerous cells undergo uncontrolled and unlimited division. In contrast, healthy cells in the human body grow and develop in a regulated manner while performing their physiological functions. Cancer cells, however, exhibit uncontrolled proliferation, variations in shape and size, lack regular morphology, and have the ability to invade different tissues — a process known as metastasis. In a rapidly proliferating healthy human cell with a total cell cycle duration of approximately 24 hours, the G1 phase lasts about 11 hours, the S phase 8 hours, the G2 phase 4 hours, and the M phase 1 hour. When a cell is not dividing or preparing to divide, it enters a resting state known as the GO phase. Cancer cells, however, tend to avoid entering the GO phase during mitosis and progress through this phase much more rapidly than normal cells.

[0006] Heat shock proteins (HSPs) are molecular structures in cells that assist in the proper folding of substrate proteins. Proteins must fold into their native conformations to become functionally active. HSPs facilitate this folding process. Proteins that fail to attain their native structure lose functionality and are subsequently degraded by the proteasome. Due to their high metabolic activity and rapid cell division, cancer cells require an increased amount of proteins to sustain their functions. Therefore, they exhibit a heightened need for HSPs to ensure proper protein folding. As a result, the expression levels of HSPs are significantly elevated in cancer cells. More than 100 different HSP proteins and isoforms have been identified. Among them, HSP70 and HSP90 play particularly critical roles; they function in a coordinated and cooperative manner with various co-chaperones (auxiliary HSPs), forming distinct complexes with diverse cellular functions. Owing to these functional properties, HSPs have become important targets in ongoing research and development efforts for cancer therapy.

[0007] Breast cancer is one of the most common types of cancer in women. In current treatment protocols for breast cancer, various chemotherapeutic agents are employed. These include anthracyclines such as doxorubicin and epirubicin; taxanes such as paclitaxel and docetaxel; antimetabolites such as 5-fluorouracil and capecitabine; and alkylating agents such as cyclophosphamide and carboplatin.

[0008] In cancer treatment, a combination of drugs, often referred to as a "chemotherapy cocktail," is typically administered to inactivate and kill cancer cells. However, effective drugs specifically targeting resistance mechanisms, such as resistance to targeted therapies or doxorubicin, are currently lacking. Heat shock proteins (HSPs) function complementarily; when HSP70 is inhibited, HSP90 compensates by becoming active. This compensatory mechanism undermines the effectiveness of therapies targeting a single HSP, thereby limiting the development of effective treatments against cancer.

[0009] As a result, in addition to the known drug molecules in the art, the development of new therapeutic agents has become imperative to provide additional technical solutions and advantages in the relevant technical field.

[0010] SUMMARY OF THE INVENTION

[0011] It is known that in the relevant technical field, certain drugs and drug combinations are used for the treatment of cancer patients and the alleviation of symptoms arising from the disease. The active ingredients of these drugs function by inhibiting heat shock proteins (HSPs), which play a role in cell proliferation. However, these active ingredients do not allow for the simultaneous inhibition of both HSP70 and HSP90 proteins. Consequently, effective cancer treatment is not achieved. The present invention primarily relates to the use of a therapeutic agent that enables the simultaneous inhibition of HSP70 and HSP90 proteins.

[0012] Doxorubicin belongs to a class of drugs known as cytotoxic agents used in cancer treatment. This drug inhibits or slows down the growth of actively proliferating cells, such as cancer cells, thereby increasing the likelihood of their death. However, there are currently no drugs available that prevent the development of resistance in cancer cells to doxorubicin. The present invention relates to the use of a therapeutic agent that enhances the efficacy of treatment when administered in combination with doxorubicin.

[0013] BRIEF DESCRIPTION OF THE DRAWINGS

[0014] Figure 1 shows the HR-MS spectrum of the compound Ethyl 3-(3-benzoyl-1-phenyl- 1 H-pyrazol-5-yl)-3-oxopropanoate (Compound D).

[0015] Figure 2 shows the FT-IR spectrum of Compound D.

[0016] Figure 3 shows the 1 H NMR spectrum of Compound D.

[0017] Figure 4 shows the 13C NMR spectrum of Compound D.

[0018] Figure 5 shows the HR-MS spectrum of 3-(3-Benzoyl-1-phenyl-1 H-pyrazole-5- carbonyl)-2H-chromen-2-one (ES3707).

[0019] Figure 6 shows the FT-IR spectrum of ES3707.

[0020] Figure 7 shows the 1 H NMR spectrum of ES3707.

[0021] Figure 8 illustrates the binding sites of ES3707 (a) and ATP molecule (b) with the Hsp90 alpha complex in YASARA Structure software.

[0022] Figure 9 shows the best interactions between ES3707 and the Hsp90 alpha protein.

[0023] Figure 10 shows the best interactions between ATP and the Hsp90 alpha protein.

[0024] Figure 11 shows the effects of varying doses of ES3707 (100-3.125 pM) on cell viability in MCF-7 cells after incubation periods of 24 (a), 48 (b), and 72 (c) hours; and the effects of varying doses of paclitaxel (100-3.125 nM) on cell viability in MCF-7 cells after incubation periods of 24 (d), 48 (e), and 72 (f) hours.

[0025] Figure 12a shows the percentage viability analysis of ES3707-treated MCF-10A cells at 24 hours.

[0026] Figure 12b shows the percentage viability analysis of ES3707-treated MCF-10A cells at 48 hours.

[0027] Figure 12c shows the percentage viability analysis of ES3707-treated MCF-10A cells at 72 hours.

[0028] Figure 13a shows Annexin V-stained NT (control) MCF-7 cells after 48 hours treatment with IC50 concentrations of ES3707 and paclitaxel.

[0029] Figure 13b shows MCF-7 cells treated with 20 nM paclitaxel at IC50 concentration for 48 hours.

[0030] Figure 13c shows MCF-7 cells treated with 3.14 pM ES3707 at IC50 concentration for 48 hours. Figure 13d shows MCF-7 cells treated with both ES3707 and paclitaxel at IC50 concentrations for 48 hours, stained with Annexin V.

[0031] Figure 13e shows the statistical comparison of cell death rates in MCF-7 cells treated with paclitaxel followed by ES3707 after 24 hours.

[0032] Figure 14 shows PCR array results demonstrating the effect of ES3707 on the cell cycle; flow cytometry experiments indicate that ES3707 arrests cells in the G1 / G0 phase, while paclitaxel arrests cells in the G2 / M phase.

[0033] Figure 15 shows changes in gene expression in signaling pathways of MCF-7 cell lines in the presence of HSP inhibitors.

[0034] Figure 16 shows changes in gene expression in signaling pathways of MCF-7 cell lines in the presence of paclitaxel.

[0035] Figure 17 shows the anticancer effects of ES3707 on immune subtypes of triplenegative breast cancer.

[0036] Figure 18 shows the effects of ES3707 on another triple-negative breast cancer cell line (MDAMB453).

[0037] Figure 19 shows further effects of ES3707 on the triple-negative breast cancer cell line (MDAMB453).

[0038] Figure 20 shows findings in the 4T1 group: (A) tumor mass, (B) liver metastases (arrows), (C) large metastatic mass in the lung (arrow), HE staining, scale bars = 50 pm.

[0039] Figure 21 shows findings in the 4T1 + 10th day drug group: (A) tumor mass, (B) no liver metastases, (C) a metastatic mass in the lung (arrow), HE staining, scale bars = 50 pm.

[0040] Figure 22 shows findings in the 4T1 + 15th day drug group: (A) tumor mass, (B) liver metastases (arrows), (C) a metastatic mass in the lung (arrow), HE staining, scale bars = 50 pm.

[0041] Figure 23 shows findings in the 4T1 + DOXO group: (A) tumor mass, (B) liver metastases (arrows), (C) a metastatic mass in the lung (arrow), HE staining, scale bars = 50 pm.

[0042] Figure 24 shows findings in the 4T1 + drug + DOXO group: (A) tumor mass, (B) liver metastases (arrows), (C) large metastatic mass in the lung (arrow), HE staining, scale bars = 50 pm.

[0043] Figure 25. Findings of the 4T1 + DMSO group: (A) tumor mass, (B) liver metastases (arrows), (C) a metastatic mass in the lung (arrow), HE staining, scale bars = 50 pm. DETAILED DESCRIPTION OF THE INVENTION

[0044] The present detailed description relates to a therapeutic agent.

[0045] The therapeutic agent of the present invention is the compound 3-(3-Benzoyl- 1 -phenyl-1 H-pyrazole-5-carbonyl)-2H-chromen-2-one (ES3707).

[0046] The therapeutic agent of the present invention is suitable for use in the treatment of cancer diseases and the alleviation of symptoms arising from these diseases.

[0047] The therapeutic agent is primarily used to inhibit heat shock proteins 15 (HSP15), which are involved in cell division.

[0048] The therapeutic agent is employed to overcome resistance caused by clinical drugs.

[0049] It has been determined that the therapeutic agent increases its efficacy when used in combination with photodynamic therapy.

[0050] The synthesis of the therapeutic agent involves the following procedural steps: Synthesis of (E)-3-(dimethylamino)-1-phenylprop-2-en-1-one Compound (B):

[0051] 1 mmol of acetophenone and 1 .2 mmol of dimethylformamide dimethyl acetal (DMF-DMA) reagent are placed into a 100 mL round-bottom flask. To this mixture, 20 mL of xylene is added, and the reaction is refluxed under a condenser for 24 hours. Completion of the reaction is monitored by Thin Layer Chromatography (TLC). At the end of the reaction, the reaction flask is allowed to cool in a refrigerator for one day. The precipitated solid in the reaction flask is filtered under vacuum. The yellow crude product (B) is then dried over P2O5 in a vacuum desiccator and subsequently used in further reactions.

[0052] Synthesis of (E)-3-oxo-3-phenyl-2-(2-phenylhydrazinylidene)propanal Compound (C):

[0053] 1 mmol of aniline is converted into its amine salt under acidic conditions (HCI) and then reacted with an aqueous solution of sodium nitrite at 0°C to form the diazonium salt. This diazonium salt solution is added dropwise to a solution of (E)-3- (dimethylamino)-1-arylprop-2-en-1-one (1 mmol, B) and sodium acetate (1 mmol) in ethanol at a constant temperature of 0°C. After five minutes, the yellow diazo compound (C) precipitated in the reaction medium is filtered under vacuum. The crude product is purified by washing with ethanol and dried over P2O5 in a vacuum desiccator. Synthesis of Ethyl 3-(3-benzoyl-1-phenyl-1H-pyrazol-5-yl)-3-oxopropanoate Compound (D):

[0054] Compound C (1.00 mmol) is dissolved in 40 mL of acetone. To this solution, 1 .20 mmol of ethyl 4-chloro-3-oxobutanoate and 1 .00 mmol of potassium carbonate are added. The reaction mixture is refluxed under a condenser with magnetic stirring for 24 hours. Completion of the reaction is monitored by Thin Layer Chromatography (TLC). At the end of the reaction, the insoluble materials are removed by filtration. The solvent of the filtrate is evaporated under reduced pressure. 1 -Propanol is added to the remaining crude product, and the mixture is stirred at room temperature. The precipitated product is filtered under vacuum and recrystallized using 1 -propanol. Figure 1 shows the HR-MS spectrum of Ethyl 3-(3-benzoyl-1 -phenyl-1 H-pyrazol-5-yl)- 3-oxopropanoate (Compound D). Figure 2 shows the FT-IR spectrum of Compound D. Figure 3 shows theA1 H NMR spectrum, and Figure 4 shows theA13C NMR spectrum of Compound D.

[0055] Synthesis of 3-(3-Benzoyl-1 -phenyl-1 H-pyrazol-5-carbonyl)-2H-chromen-2-one (abbreviated as ES3707):

[0056] In a 100 mL reaction flask, Compound D (1 mmol) and an equimolar amount of salicylaldehyde derivative (1 mmol) are dissolved in 30 mL of methanol or chloroform (CHCI_3). Catalytic amount (~5%) of piperazine is added to the mixture. The reaction mixture is refluxed under a condenser for 24 hours. The yellow precipitate formed in the reaction mixture is collected by suction filtration. The crude product is washed sequentially with chloroform, acetonitrile, and diethyl ether. It is then recrystallized and purified using a DMF-water (20:1 ) solvent mixture. The purified product is dried over P2O5 in a vacuum desiccator.

[0057] Figure 5 shows the HR-MS spectrum of 3-(3-Benzoyl-1 -phenyl-1 H-pyrazol-5- carbonyl)-2H-chromen-2-one (ES3707). Figure 6 shows the FT-IR spectrum of ES3707, and Figure 7 shows theA1 H NMR spectrum of ES3707.

[0058] Evaluation of Molecular Docking Studies

[0059] To understand the mechanism by which the ES3707 molecule inhibits Hsp90 (Heat Shock Protein 90), molecular docking was performed between the human Hsp90a crystal structure and ES3707. Through this interaction, the binding energy and inhibition constant (Ki) of the ES3707 inhibitor to Hsp90a were calculated. As a control, the same interactions between Hsp90a and ATP (adenosine triphosphate) were also simulated using YASARA Structure software. The results of the molecular docking are shown in Figure 8, illustrating the binding sites of ES3707 (a) and ATP (b) with the Hsp90 alpha complex within the YASARA Structure software.

[0060] According to the molecular docking results, the ES3707 inhibitor binds with high affinity to the ATP binding site of Hsp90a. The binding energy of the ES3707 inhibitor was calculated as -10.20 kcal / mol with an inhibition constant (Ki) of 86.88 nM, whereas ATP exhibited a binding energy of -7.50 kcal / mol and a Ki of 3.14 pM. These results indicate that ES3707 interacts with Hsp90a with higher affinity compared to ATP. Based on these data, ES3707 is expected to block ATP binding to the relevant site of the HSP90 protein, thereby inhibiting its chaperone function.

[0061] ES3707 inhibitor interacts at the ATP binding site of Hsp90a with amino acid residues Leu48, Asn51 , Ser52, Asp54, Ala55, Lys58, Asp93, Met98, Asn106, Leu 107, lle110, Ala111 , Lys112, Ser113, Gly135, Val136, Phe138, Tyr139, Thr184, and Val186. Consequently, the inhibitor disrupts the folding processes of oncoproteins, preventing their proper folding and thereby inhibiting tumorigenesis.

[0062] The best interactions of the protein-ligand complexes shown in Figure 8 (a) and (b) are presented in Figures 9 and 10, respectively. The key amino acids contributing to binding were identified as Lys58, Phe138, and Tyr139.

[0063] Evaluation of Cell Culture Studies

[0064] ES3707 was compared with paclitaxel, a clinically widely used microtubule function inhibitor, and was found to operate via a different mechanism, particularly evident from cell cycle experiments.

[0065] Evaluation of the Anticancer Activity of the Inhibitor and the Clinical Drug

[0066] For the XTT viability assays, MCF-7 and MCF-10A cells were seeded into 96- well plates at a density of 5,000 cells per well. All experiments were performed in five replicates. Cells were treated with ES3707 at concentrations ranging from 100 to 3.125 pM, and paclitaxel (PX) at concentrations ranging from 100 to 3.125 nM, and cytotoxicity assays were conducted at 24, 48, and 72 hours. Evaluation of Anticancer Activity of ES3707 and Paclitaxel in the MCF-7 Cell Line

[0067] The IC50 values of ES3707 in the MCF-7 cell line were determined as 3.924 pM (p<0.001 ) at 24 hours, 4.376 pM (p=0.002) at 48 hours, and 4.150 pM (p<0.001 ) at 72 hours. These data are presented in Figure 11 (a-c). The IC50 values of PX in MCF-7 cells were 7.722 nM (p<0.001 ) at 24 hours, 3.143 nM (p<0.0001 ) at 48 hours, and 3.668 nM (p<0.0001 ) at 72 hours, as shown in Figure 11 (d-f).

[0068] Although paclitaxel exerted effects at lower concentrations compared to ES3707, ES3707 was determined to be a highly effective anticancer agent. Additionally, toxicity was evaluated in healthy cells using the MCF-10A cell line.

[0069] Evaluation of the Effect on Healthy Breast Cells

[0070] Assessment of Anticancer Activity of the Hsp Inhibitor in MCF-10A (Normal Breast Cells) Cell Line

[0071] Dose-dependent changes in cell proliferation of MCF-10A cells treated with ES3707 at 24, 48, and 72 hours are shown in Figure 12 (a-c). The IC50 values detected in MCF-7 cells did not reduce the viability of MCF-10A cells, indicating low toxicity towards healthy cells.

[0072] Evaluation of Apoptotic Effect in MCF-7 Cell Line

[0073] Following viability assays by XTT, apoptosis and necrosis rates induced by ES3707 and PX were measured by Annexin V-FITC assay using flow cytometry. The apoptotic effects on MCF-7 cells are shown in Figures 13 (a-e) and Table 1. After 48 hours of treatment with either ES3707 or PX, the total early and late apoptotic rates were found as 16.50% (p<0.001 ) and 30.86% (p<0.0005), respectively.

[0074] When ES3707 and PX were administered simultaneously to MCF-7 cells, apoptosis reached 35% (p=0.005), while sequential treatment with PX followed by ES3707 after 24 hours induced approximately 40% apoptosis. Table 1 . Total Apoptotic and Necrotic Cell Percentages Determined After 48 Hours of Treatment

[0075] Q1 : Necrotic cells, Q2: Late apoptotic cells, Q3: Viable cells, Q4: Early apoptotic cells

[0076] When considering the sum of Q2 and Q4 populations, it can be observed that the ES3707 molecule induces apoptosis in breast cancer cells to a degree comparable to the clinical drug paclitaxel.

[0077] Evaluation of Cell Cycle Results in MCF-7 Cell Line

[0078] MCF-7 cells incubated with the IC50 concentrations of ES3707 and paclitaxel for 48 hours were stained using a propidium iodide (PI) staining procedure and analyzed for cell cycle distribution by flow cytometry. Representative flow cytometry data for the treated MCF-7 cells are shown in Figure 14. For each treatment group, 10,000 cells were analyzed by the flow cytometer, and cell cycle phases were determined using CytExpert software.

[0079] It was observed that ES3707 and paclitaxel arrest cancer cells at different cell cycle phases (ES3707 at G0 / G1 , paclitaxel at G2 / M). However, both agents trigger immune system activation and intrinsic apoptotic pathways through similar mechanisms.

[0080] Array Studies and Gene Enrichment Analysis

[0081] Figures 15 and 16 demonstrate that ES3707 induces the immune system via the ERK / MAPK signaling pathway (including MEF2A, ELK1 , MAPK1) in MCF-7 cancer cells. This induction involves activation of MAP kinases. Additionally, toll-like receptors, cytokine and interferon signaling pathways, as well as growth hormone receptors, are affected. This signaling is transcriptionally regulated via the SMAD2:SMAD3 / SMAD4 complex. While apoptosis occurs through the intrinsic pathway, genes associated with pyroptosis and senescence are activated. Furthermore, expression of cancer stem cell markers POU5F1 and SOX2 is down regulated. Gene enrichment analysis indicates that despite ES3707 and paclitaxel arresting cancer cells at different stages, they inhibit cancer progression through similar molecular mechanisms. Table 2. ES3707 Cancer Pathway PCR Array Results

[0082] Molecular Effect of ES3707 on PI3K / AKT Pathway

[0083] Investigation of the molecular mechanism of ES3707 revealed that the compound utilizes the PI3K / AKT signaling pathway (involving ERBB2-4, KIT, PDGFRA-B, ESR1-2). The inhibitory effect of ES3707 on cancer signaling pathways supports its potential as a significant drug candidate molecule (see Table 2). Anticancer Effects of ES3707 on Triple-Negative Breast Cancer Immune Subtypes

[0084] The effect of ES3707 on MDAMB231 triple-negative breast cancer cells was determined to be 10 pM on day 3. This IC50 value decreased to 4.66 pM within the first 24 hours when combined with photodynamic therapy (laser application). ES3707 demonstrates activity against immune subtypes of triple-negative breast cancer, with a reduced IC50 upon laser treatment, as shown in Figure 20.

[0085] In contrast, ES3707’s effect on another triple-negative breast cancer cell line, MDAMB453, remained relatively high at 15 pM on day 3, as illustrated in Figure 21.

[0086] Resistance Studies

[0087] ES3707 inhibited doxorubicin-resistant cells at a low concentration (7.40 pM) after 48 hours. However, the photodynamic therapy effect (laser application) was not observed in resistant cells. Cells were incubated with various concentrations of ES3707 for 24 hours, followed by addition of 2.5 pM methylene blue and incubation at 37°C for 1 hour. Subsequently, 660 nm diode laser was applied for 1 minute, and cell viability was assessed by MTT assay 24 hours post-treatment. ES3707 and its combination with doxorubicin showed high efficacy on doxorubicin-resistant ER+ breast cancer cells, with no enhancement from laser treatment.

[0088] Animal Studies

[0089] Female Balb / c mice were orthotopically inoculated with 4T1 breast cancer cells into the mammary fat pad. Tumor growth to a size of 2-3 mm was observed with 90% success by day 10 post-inoculation (in a pilot study, 56 mice were inoculated, with tumors developing in 50). The present study was conducted on these 50 animals. Euthanasia was performed on day 39 post-inoculation (29 days after tumor development at day 10). Preliminary histopathological examinations of the collected samples have been completed, and results are summarized below.

[0090] 4T1 mouse breast cancer cells (ATCC; #CRL2539) were cultured under standard conditions in RPMI medium supplemented with 10% fetal bovine serum (FBS), 1% penicillin / streptomycin (PS), and HEPES buffer at 37°C with 5% CO2 in a sterile incubator. Cells were expanded and 1 x 1OA5 cells were orthotopically injected into the mammary tissue of female Balb / c mice aged 8-10 weeks. Mice weights and tumor progression were monitored starting from day 7 postinjection and subsequently every three days. Tumor masses became palpable in 50 mice from day 10 onwards. Experimental Groups and Treatment Protocols

[0091] Group 1 : 4T1 cells injected; no treatment applied. Group 2: Treatment initiated on day 10 post-inoculation, continued every 3 days until study end. Group 3: Treatment initiated on day 15 post-inoculation, continued every 3 days until study end. Group 4: Positive control group treated with doxorubicin (DOXO) starting on day 10, repeated every 7 days. Group 5: Combined treatment with ES3707 and DOXO. Group 6: Vehicle control group treated with DMSO.

[0092] The detailed protocols are presented in Table 3.

[0093] Tumor sizes were measured with calipers every 3 days along with body weight monitoring. On day 39, all animals were euthanized; tumor tissues, lungs, and liver samples were collected. Primary tumor masses were carefully excised, weighed with a precision scale, and documented. Tumor-bearing mammary tissues were fixed in 10% formalin solution and subjected to histopathological analysis.

[0094] Table 3. Experimental Groups

[0095] Histopathological Findings

[0096] The largest tumor sizes and highest incidence of metastases were observed in Group 1 (4T1 tumor-bearing untreated controls). All mice in this group developed metastases (Figure 20). The group that started drug treatment on day 10 showed the smallest and least metastases, with some mice lacking liver metastases entirely (Figure 21).

[0097] In the group where treatment started on day 15, tumors and metastases were larger and more numerous compared to the day 10 treatment group (Figure 22).

[0098] In the positive control group treated with DOXO starting on day 10, metastases were observed in both lungs and liver. The number and size of metastatic masses resembled those of the day 15 treatment group (Figure 23).

[0099] Interestingly, in the group treated with both drug and DOXO starting on day 7, the combination therapy increased tumor size and metastasis (Figure 24).

[0100] The DMSO-treated control group showed no therapeutic effect, displaying tumor and metastasis characteristics similar to the untreated tumor group (Figure 25).

[0101] Summary

[0102] These findings indicate that the designed drug candidate slows tumor growth in the 4T1 breast cancer model, with earlier treatment initiation producing more effective results. No synergistic or additive therapeutic effect was observed with DOXO combination therapy. DMSO treatment showed no therapeutic benefit.

[0103] Gene enrichment analyses were also performed on tissue samples collected from these experiments.

[0104] Table 4. PCR Array Results of Cancer Pathways in Mouse Breast Tissue Treated with ES3707

[0105]

[0106] ES3707 enhances oxidative phosphorylation in breast cancer tissue (increased C0X5A, decreased SLC2A1 / GLUT1), induces apoptosis (CASP2, CASP7, CFLAR, APAF1), and reduces oxidative stress (SOD1). These effects were found to be more effective than those of the clinically used drug doxorubicin. The clinical agent does not exhibit the same effect on oxidative phosphorylation (C0X5A decreases, while SLC2A1 / GLUT1 increases).

[0107] Moreover, ES3707 exerts an anti-cancer effect by increasing the expression of the angiogenesis inhibitor SERPINF1 and decreasing the expression of SNAI2, a key gene in epithelial-mesenchymal transition (see Table 4).

[0108] The scope of protection of the invention is defined in the claims appended hereto and shall in no way be limited to the examples described in this detailed disclosure. It is evident that a person skilled in the art may, in light of the above description and without departing from the core concept of the invention, develop similar embodiments.

Claims

CLAIMS1. The invention relates to the use of the compound ES3707 as a therapeutic agent in the treatment of cancer diseases and alleviation of symptoms caused by such diseases, specifically by inhibiting heat shock proteins involved in cell division and preventing resistance induced by clinical drugs.

2. The use according to claim 1, characterized in that the inhibition concentration in cancer cells is between 3.9 pM and 4.3 pM.

3. The use according to any of the preceding claims, characterized in that it is applied in the treatment of breast cancer.

4. The use according to claim 1 , characterized in that it is suitable for use in combination with photodynamic therapy.

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