A pharmaceutical composition for treating pancreatic ductal adenocarcinoma

Abstract

This invention provides a pharmaceutical composition for treating pancreatic ductal adenocarcinoma, wherein an ADAM10 inhibitor and an immune checkpoint inhibitor work synergistically to fundamentally solve the "cold tumor" problem faced by pancreatic cancer immunotherapy. Compared with existing strategies such as chemotherapy combined with immunotherapy and radiotherapy combined with immunotherapy, this invention reshapes the tumor immune microenvironment from the source, and has the characteristics of clear mechanism, innovative target, and significant synergistic effect, providing a brand-new immunotherapy strategy for pancreatic cancer patients.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceuticals, specifically relating to pharmaceutical compositions for treating pancreatic ductal adenocarcinoma. Background Technology

[0002] Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest malignant tumors of the digestive system, with a 5-year survival rate of less than 12%. Surgical resection is currently the only potentially curative treatment, but even with radical surgery, most patients experience recurrence and metastasis shortly after surgery. Chemotherapy regimens based on gemcitabine and nab-paclitaxel offer limited survival benefits, necessitating novel treatment strategies. Immune checkpoint inhibitors (ICIs), represented by PD-1 / PD-L1 antibodies, have shown significant efficacy in various solid tumors such as melanoma and non-small cell lung cancer, but their monotherapy efficacy in pancreatic cancer is extremely limited. Multiple clinical trials have demonstrated that the objective response rate of anti-PD-1 / PD-L1 monotherapy for PDAC is almost zero, and current attempts at chemotherapy combined with immunotherapy and radiotherapy combined with immunotherapy have limited benefits in pancreatic cancer, failing to fundamentally address the lack of effector T cells in the tumor microenvironment. The core reason for pancreatic cancer's resistance to immunotherapy lies in its unique immunosuppressive tumor microenvironment (TME): the lack of pre-existing effector T cell infiltration within the tumor, a characteristic known as "immunocold tumor." Without tumor-reactive CD8+ T cells, simply blocking the PD-1 / PD-L1 pathway cannot exert an anti-tumor effect. Therefore, how to transform pancreatic cancer from a "cold tumor" to a "hot tumor" and increase the infiltration of effector T cells within the tumor is a key scientific problem in overcoming immunotherapy resistance. ADAM10 (A Disintegrin and Metalloproteinase 10) is a transmembrane metalloproteinase involved in the cleavage and processing of various membrane proteins. However, the specific role of ADAM10 in the remodeling of the pancreatic cancer immune microenvironment, its potential as a therapeutic target, and the synergistic anti-tumor effect of combining ADAM10 inhibitors with immune checkpoint inhibitors have not been reported; existing technologies have also not found an effective means to systematically increase intratumoral CD8+ T cells by targeting tumor cell-derived immunosuppressive mechanisms (such as sIL-2R-mediated IL-2 signaling pathway blockade). + The infiltration and killing functions of T cells create the preconditions for immune checkpoint inhibitors to exert their effects. Summary of the Invention

[0003] This invention provides a pharmaceutical composition for treating pancreatic ductal adenocarcinoma, which combines ADAM10 inhibitors and immune checkpoint inhibitors for synergistic therapy, fundamentally solving the "cold tumor" problem faced by pancreatic cancer immunotherapy. It has significant clinical translational value and broad application prospects.

[0004] The present invention provides a pharmaceutical composition for treating pancreatic ductal adenocarcinoma, the composition comprising an ADAM10 inhibitor and an immune checkpoint inhibitor.

[0005] Furthermore, the aforementioned ADAM10 inhibitors include one or more of GI254023X, INCB007839, INCB8765, and GW280264X.

[0006] In this invention, INCB007839 is Aderbasib.

[0007] Furthermore, the aforementioned immune checkpoint inhibitors include one or more of PD-L1 antibodies, CTLA-4 antibodies, and PD-1 antibodies.

[0008] Furthermore, the aforementioned ADAM10 inhibitors restore IL-2 signaling by inhibiting the cleavage of IL-2Rα on the surface of immune cells by ADAM10, thereby enhancing the infiltration and killing functions of immune cells and upregulating MHC-I expression.

[0009] In this invention, the ADAM10 inhibitor is not limited to the compounds described above, but should include compounds with equivalent ADAM10 enzyme activity inhibitory function.

[0010] In this invention, ADAM10 is highly expressed in pancreatic cancer cells (upregulated by KRAS mutations), and its protease activity cleaves IL-2Rα on the surface of immune cells to produce sIL-2R. sIL-2R, acting as a decoy receptor for IL-2, competitively binds to IL-2, inhibiting the activation and proliferation of effector T cells, reducing IFN-γ secretion, and subsequently downregulating the expression of MHC-I molecules in tumor cells, creating an immunosuppressive microenvironment. ADAM10 inhibitors, by blocking this pathway, restore IL-2 signaling, enhance the infiltration and killing function of CD8+ T cells, upregulate MHC-I expression and antigen presentation capacity, transforming "cold tumors" into "hot tumors," creating the preconditions for the action of immune checkpoint inhibitors. Based on this, immune checkpoint inhibitors further relieve the inhibition of activated T cells by the pathway, and the two work synergistically to achieve highly efficient immune killing of pancreatic cancer.

[0011] In this invention, ADAM10 inhibitors and immune checkpoint inhibitors have significant anti-tumor effects, with tumor growth inhibition effects significantly superior to single-drug administration, and can significantly prolong survival time, demonstrating good safety and clinical translation potential.

[0012] In this invention, the pharmaceutical composition can successfully transform pancreatic cancer from a "cold tumor" to a "hot tumor," fundamentally solving the key bottleneck of the ineffectiveness of immune checkpoint inhibitors in pancreatic cancer.

[0013] This invention is based on a complete immune escape chain of "ADAM10→sIL-2R→IL-2 signaling blockade→T cell function inhibition→IFN-γ reduction→MHC-I downregulation", with a clear mechanism, well-defined target, and is not an empirical drug combination.

[0014] Compared with existing strategies such as chemotherapy combined with immunotherapy and radiotherapy combined with immunotherapy, this invention reshapes the tumor immune microenvironment from the source. It has the characteristics of clear mechanism, innovative target and significant synergistic effect, and provides a brand-new immunotherapy strategy for pancreatic cancer patients. Attached Figure Description

[0015] Figure 1 (A) Schematic diagram of drug administration in animal experiments; (B) Gross photographs of in situ tumors in each treatment group; (C) Comparison of tumor weight among groups; (D) Kaplan-Meier survival curves for each group; (E) Intratumoral IFN-γ levels (ELISA) for each group; (F) Intratumoral CD8 levels for each group. + T cell infiltration rate; (G)CD8 + IFN-γ in T cells + Ratio; (H)CD8 + Granzyme B in T cells + Ratio; (I) CD8 + TNF-α in T cells + Ratio; (J)CD8 + PD-1 in T cells + Tim-3 + Depletion ratio;

[0016] Figure 2 (A) Curves of weight change in mice in each treatment group; (B) Comparison of serum biochemical indicators (ALT, AST, BUN, CRE) in each group; (C) Representative images of H&E staining of major organs (heart, liver, spleen, lung, kidney) in each group. Detailed Implementation

[0017] To enable those skilled in the art to better understand the present application, the technical solutions in the embodiments of the present application will be clearly and completely described below with reference to examples. Obviously, the described embodiments are only some embodiments of the present application, and not all embodiments. Based on the embodiments in the present application, all other embodiments obtained in the art without creative effort should fall within the scope of protection of the present invention.

[0018] Unless otherwise specified, the experimental methods used in the following examples are conventional methods, performed according to the techniques or conditions described in the literature in this field or according to the product instructions. Unless otherwise specified, the materials and reagents used in the following examples are commercially available.

[0019] Example 1: Experiment testing tumor suppression effect

[0020] 1. The KPC mouse pancreatic cancer cell line was derived from spontaneous pancreatic tumors in KrasG12D / +; Trp53R172H / +; Pdx1-Cre (KPC) transgenic mice. KPC cells in logarithmic growth phase were harvested and cultured at a concentration of 5 × 10⁻⁶ cells / year. 5 Orthotopic tumor models were established by in situ injection of 1 / mouse into the pancreas of 6–8 week old female C57BL / 6 mice. All animal experimental protocols were approved by the laboratory animal ethics committee of our institution.

[0021] 2. Grouping and administration: Tumor-bearing mice were randomly divided into 4 groups: control group (solvent), Aderbasib monotherapy group (30 mg / kg / d intraperitoneal injection), anti-PD-1 monotherapy group (200 μg / mouse / 3 days intraperitoneal injection), and combination therapy group (Aderbasib + anti-PD-1) (the administration method was the same as that of each monotherapy group).

[0022] 3. Experimental methods:

[0023] 3.1 Evaluation of tumor suppression effect

[0024] Mice body weight was recorded every 3 days during the administration period. After administration, mice were sacrificed, the orthotopic pancreatic tumor was completely dissected, and the tumor weight was measured and photographed. A parallel group was set up for survival analysis; mice in this group were not sacrificed, and their survival time was continuously observed and recorded. Survival curves were plotted using the Kaplan-Meier method.

[0025] 3.2 Flow cytometry analysis

[0026] Fresh tumor tissue was collected, mechanically minced, and digested with collagenase / DNase to prepare a single-cell suspension. After filtration, erythrocytes were lysed. Cell surface markers (CD45, CD3, CD8, PD-1, Tim-3) and tumor cell markers (H2-K / MHC-I) were stained with fluorescently labeled antibodies. For intracellular cytokine detection, the cells were stimulated with PMA / Ionomycin and Brefeldin A for 4–6 hours. After fixation and membrane perforation, intracellular staining for IFN-γ, TNF-α, and Granzyme B was performed using fluorescently labeled antibodies. Data were acquired using flow cytometry and analyzed using FlowJo software.

[0027] 3.3 ELISA detection

[0028] Tumor tissues from each group were collected, homogenized with lysis buffer, and centrifuged to obtain the supernatant. Intratumoral IFN-γ protein levels were detected using a mouse IFN-γ ELISA kit, following the manufacturer's instructions.

[0029] 4. Experimental Results: Comparison of tumor growth inhibition effects among the groups showed that: there was no significant difference in tumor growth between the anti-PD-1 monotherapy group and the control group, confirming that PD-1 antibody monotherapy was ineffective against KPC in situ tumors; the tumor weight in the Aderbasib monotherapy group was significantly reduced compared to the control group; the combination therapy group showed significantly better tumor growth inhibition effects than each monotherapy group (see appendix). Figure 1 AC). Kaplan-Meier survival analysis showed that the survival of tumor-bearing mice in the combination therapy group was significantly longer than that in each single-drug group and the control group (see appendix). Figure 1 D). ELISA testing showed that intratumoral IFN-γ levels were significantly higher in the Aderbasib monotherapy group than in the control group, and further higher in the combination therapy group than in the Aderbasib monotherapy group (see appendix). Figure 1 J). Flow cytometry analysis showed that the combination therapy group had CD8+ in the tumor. + The proportion of T cell infiltration was significantly higher than that in each single-drug group and the control group; CD8 + IFN-γ in T cells + TNF-α + Granzyme B + The proportion of effector cells was further increased in the combination therapy group, while PD-1 + Tim-3 + Exhaustion-type CD8 + The proportion of T cells did not increase significantly in the combination therapy group (see appendix). Figure 1 EI). The above results indicate that Aderbasib increases intratumoral CD8 by restoring IL-2 signaling. + T cell infiltration and effector function provide the necessary immune cell basis for anti-PD-1 antibodies to exert their effects; on this basis, anti-PD-1 antibodies maintain the effector function of newly infiltrating T cells without increasing their depletion, and the two work together to achieve highly efficient immune killing of pancreatic cancer.

[0030] Example 2 Safety Evaluation

[0031] 1. Experimental Method:

[0032] Mice from each treatment group in Example 1 were sacrificed at the experimental endpoint, and the following samples were collected for safety evaluation:

[0033] 1.1 Weight monitoring

[0034] The weight of mice in each group was measured and recorded every 3 days during the drug administration period. Weight change curves were plotted to assess the effect of the drug on the general condition of the mice.

[0035] 1.2 Serum biochemical marker detection

[0036] Blood was drawn from the orbit or heart before sacrifice, and serum was separated. The following routine organ function indicators were measured using a fully automated biochemical analyzer: alanine aminotransferase (ALT) and aspartate aminotransferase (AST) (liver function indicators); blood urea nitrogen (BUN) and creatinine (CRE) (kidney function indicators).

[0037] 1.3 Pathological examination of major organs

[0038] Major organs such as heart, liver, spleen, lung, and kidney of mice in each group were taken, fixed in 10% neutral formalin, embedded in paraffin, sectioned, stained with hematoxylin and eosin (H&E), and observed under an optical microscope for pathological changes.

[0039] 2. Experimental Results:

[0040] There were no significant differences in the body weight change curves among the groups during the drug administration period, and the body weight of mice in the combined treatment group did not decrease significantly (see attached). Figure 2 A). Serum ALT, AST, BUN, and CRE levels in mice from all groups were within the normal range, and there were no significant differences between the treatment groups and the control group (see appendix). Figure 2 B). H&E staining showed no obvious pathological changes in the major organs of mice in all groups, including the heart, liver, spleen, lungs, and kidneys (see appendix). Figure 2 C).

[0041] The above results indicate that the combination therapy of Aderbasib and anti-PD-1 antibody has a good safety profile, without producing significant systemic toxicity or organ damage, supporting the clinical translational application of this drug combination.

Claims

1. A pharmaceutical composition for treating pancreatic ductal adenocarcinoma, characterized in that: The composition includes an ADAM10 inhibitor and an immune checkpoint inhibitor.

2. The pharmaceutical composition according to claim 1, characterized in that: The ADAM10 inhibitors include one or more of GI254023X, INCB007839, INCB8765, and GW280264X.

3. The pharmaceutical composition according to claim 1, characterized in that: The immune checkpoint inhibitors include one or more of PD-L1 antibodies, CTLA-4 antibodies, and PD-1 antibodies.

4. The pharmaceutical composition according to any one of claims 1-3, characterized in that: The ADAM10 inhibitor restores IL-2 signaling by inhibiting the cleavage of IL-2Rα on the surface of immune cells by ADAM10, thereby enhancing the infiltration and killing functions of immune cells and upregulating MHC-I expression.

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