AST-3424 for treating liver cancer

Abstract

The use of AST-3424 in the preparation of a drug for treating a patient with hepatocellular carcinoma. A pathological paraffin block or pathological section of ex-vivo liver tumor tissue from the patient with hepatocellular carcinoma is subjected to immunohistochemical staining to detect the protein expression level of an AKR1C3 enzyme, with the detection result showing an H-score greater than or equal to 200; or the pathological paraffin block or pathological section of ex-vivo liver tumor tissue from the patient with hepatocellular carcinoma is subjected to immunohistochemical staining to detect the protein expression level of an AKR1C3 enzyme, with the detection result showing that the sum of percentages of moderate-intensity staining and high-intensity staining is greater than or equal to 70%. The dosing regimen for AST-3424 is as follows: in each 21-day cycle, administration is performed on day 1 and day 8; the initial dose is 6.0 mg / m2 per administration; and the patient will be permitted to receive treatment for a maximum of 34 cycles.

Description

AST-3424 treatment of liver cancer TECHNICAL FIELD

[0001] The present application relates to a method for treating human malignant tumors, in particular a method for treating hepatocellular carcinoma (HCC) using AKR1C3-activated anticancer prodrug compound AST-3424, and the pharmaceutical use thereof, belonging to the field of tumor chemotherapy. BACKGROUND

[0002] DNA alkylating agent prodrug AST-3424 (WO2016145092, WO2017087428) with CAS number 2097713-69-2, which is targeted at overexpressed aldehyde-ketone reductase 1C3 (AKR1C3), has the following structure:

[0003] AST-3424 (also known as OBI-3424, TH-3424) is activated by AKR1C3, which is overexpressed in cancer cells, to release the metabolite AST-2660 (also known as AST-2660) inside the cancer cells. AST-3424 itself has little toxicity to cancer cells, and its pharmacological effects in animal models and in vitro pharmacological experiments are all related to the expression of AKR1C3 enzyme: the prodrug AST-3424 is metabolized to AST-2660 under the action of AKR1C3 enzyme and NADPH, and the expression of the enzyme is positively correlated with the drug efficacy (Literature 1; Literature 2; Literature 3; Literature 4).

[0004] Currently, the drug has entered phase I / II clinical trials in China and the United States respectively (NCT03592264 in the United States, indications: liver cancer, pancreatic cancer and other solid tumors, sponsor: Taiwan Hao Ding Shengzhi Company OBI Pharama Inc (4174), drug name: OBI-3424; NCT04315324 in the United States, T-ALL / T-LBL (acute T lymphoblastic leukemia / T lymphoblastic lymphoma), drug name: OBI-3424; CTR20191371 in China, indications: various solid tumors, sponsor: Shenzhen Aixin Dawei Pharmaceutical Technology Co., Ltd. Ascentawits Pharmaceuticals, LTD., drug name: AST-3424; CTR20201915, indications: acute T lymphoblastic leukemia and acute B lymphoblastic leukemia, sponsor: Shenzhen Aixin Dawei Pharmaceutical Technology Co., Ltd. Ascentawits Pharmaceuticals, LTD., drug name: AST-3424).

[0005] In these phase I / II clinical trials conducted in China, USA, one of the dosing regimens was 21 days as a treatment cycle, with dosing on days 1 and 8 for a total of two doses, and no dosing on days 2-7 and 9-21.

[0006] The information results of the phase I clinical trial completed in the USA are summarized as follows (see documents 5, 7, Abstract part of document 5 is excerpted and translated, Table 2, Figure 4, the applicant translates the abstract as follows, and the corresponding table is translated into Figure 1, Figure 2 in this application):

[0007] Trial design: OBI-3424 was administered intravenously at a dose of 1, 2, 4, 6, 8, or 12 mg / m 2 (every 21 days, days 1 and 8, regimen A) or 8, 10, 12, or 14 mg / m 2 (every 21 days, only on day 1, regimen B). Dose escalation used a “3+3” design. Patients received study treatment until disease progression, intolerable toxicity, or up to 2 years of treatment.

[0008] Results: A total of 39 adult patients were treated.

[0009] In patients receiving regimen A, the maximum tolerated dose (MTD) of OBI-3424 was determined to be 8 mg / m 2 ; dose-limiting toxicities (DLTs) were reported at the 12 mg / m 2 dose level, including thrombocytopenia (grade 3-4 in 5 of 6 patients) and anemia (grade 3-4 in 5 of 6 patients); the lowest platelet count was observed on day 15 or day 22.

[0010] In patients receiving regimen B, the MTD was not reached at the maximum dose tested (14 mg / m 2 ); grade 3 or higher anemia occurred in 3 of 6 patients receiving 14 mg / m 2 ; the phase II recommended dose (RP2D) was 12 mg / m 2 (every 21 days, day 1).

[0011] Treatment drug-related adverse events (TRAEs) occurred in 82% (32 / 39) of patients. The most common TRAEs were anemia (64%), thrombocytopenia (51%), nausea (26%), and fatigue (21%). No patients experienced fatal TRAEs, 49% of patients reported ≥ grade 3 TRAEs, including 3 patients with serious TRAEs.

[0012] OBI-3424 at 1 to 14 mg / m 2 Doses showed linear pharmacokinetic characteristics with mild accumulation after repeated dosing.

[0013] One validated retrospective immunohistochemistry (IHC) assessment indicated that 27% of patients had high AKR1C3 staining (H-Score > 135). Twenty-one patients (54%) had confirmed best response of stable disease.

[0014] Conclusion: The investigators completed the dose escalation part of the OBI-3424 clinical study. The RP2D was determined to be 12 mg / m 2 administered every 3 weeks (total dose of 12 mg / m 2 ). OBI-3424 was well tolerated. Dose-dependent non-cumulative thrombocytopenia and anemia were dose-limiting toxicities.

[0015] That is, after the phase I clinical trial of OBI-3424 conducted in the United States, it was considered that patients with solid tumors with high AKR1C3 staining, i.e., the above-mentioned IHC H-Score > 135, had a better response to OBI-3424 by immunohistochemistry (IHC) detection (using patent application PCT / CN2021 / 114774, publication number WO2022048492A1, corresponding to Chinese application CN202180031349.X, publication number CN115485560A; corresponding to US application US18 / 043610, publication number US20240142453A1; corresponding to European application EP21863557.1, publication number EP4209785A1; corresponding to Japanese application JP2023514086, publication number JP2023540283A; corresponding to Korean application KR10-2023-7011169, publication number KR10-2023-0058507A; corresponding to Australian application AU2021337711, publication number AU2021337711A1; corresponding to Canadian application CA3192258, publication number CA3192258A1; corresponding to Israeli application IL300833, publication number IL300833A; corresponding to Brazilian application BR112023003973-8, publication number BR112023003973-8A2) that AKR1C3 enzyme-selective activated alkylating agent prodrug OBI-3424 had a better effect on patients with solid tumors with AKR1C3 enzyme level H-Score > 135 detected by the above-mentioned specific IHC method, so it is recommended in documents 5, 7 that the AKR1C3 enzyme expression level of patients enrolled in phase II clinical trials be IHC H-Score > 135. SUMMARY

[0016] Applicant as the sponsor of AST-3424 China Phase I clinical trial, the I phase solid tumor clinical trial (CTR20191371) in China has been completed, and the results of the trial are disclosed and summarized by Applicant for the first time as follows.

[0017] A total of 21 subjects were enrolled in the trial. Every 21 days was a treatment cycle, and the drug was administered on the 1st and 8th day. The doses of 1.0, 2.0, 4.0, 6.0 and 8.0 mg / m 2 There were 5 dose levels for dose escalation. After discussion by the safety review committee (SRC), it was confirmed that every 21 days was a treatment cycle, and the MTD and RP2D of the drug administered on the 1st and 8th day was 6.0 mg / m 2 (12 mg / m 2 total dose of drug administered per cycle), and the regimen was determined as the administration regimen for subsequent phase II clinical trials.

[0018] The safety and tolerability of American subjects were slightly better than those of Chinese subjects (the MTD / RP2D of the same administration regimen was slightly higher than that of the Chinese population); the safety spectrum trend was consistent (most adverse events were mild; anemia, decreased platelet count, fatigue, and vomiting were the most common adverse events). Other safety, PK, and effectiveness results were generally similar to those of OBI-3424 clinical trials.

[0019] In addition to the dose exploration of the above-mentioned administration regimen of every 21 days as a treatment cycle, and the drug administered on the 1st and 8th day, the administration regimen exploration of every 21 days as a treatment cycle, and the drug administered only on the 1st day (dose: 8, 10, 12 or 14 mg / m 2 ) was also conducted in the clinical trial in the United States. After comparison, it was finally determined that the administration regimen for subsequent phase II clinical trials in the United States was every 21 days as a treatment cycle, and the drug administered once on the 1st day of each cycle, and the RP2D was 12 mg / m 2 , and it was finally determined that the AKR1C3 enzyme expression level of the patients enrolled in the phase II clinical trial was IHC H-Score≥135.

[0020] The administration regimen of the phase II clinical trial determined according to the results of the above-mentioned phase I clinical trial conducted in China was every 21 days as a treatment cycle, and the drug administered once on the 1st and 8th day of each cycle, and the RP2D was 6.0 mg / m 2 .

[0021] That is, the safety and tolerability data obtained from the phase I clinical trials of AST-3424 (OBI-3424) in China and the United States are not much different, but the finally determined recommended dose and administration regimen for phase II have great differences.

[0022] Applicant found that patients with hepatocellular carcinoma who have excellent results when treated with AST-3424 have a tumor tissue biopsy that, when tested for AKR1C3 enzyme protein expression level via immunohistochemical staining method, has an H-score score of greater than or equal to 200 or shows a sum of percentages of moderate intensity staining and high intensity staining of greater than or equal to 70%. Based on this, the following methods for treating hepatocellular carcinoma with AST-3424 and corresponding pharmaceutical uses are proposed.

[0023] A method for treating hepatocellular carcinoma with AST-3424 as a single agent, characterized in that:

[0024] The patient with hepatocellular carcinoma has a tumor tissue biopsy that, when tested for AKR1C3 enzyme protein expression level via immunohistochemical staining method, has an H-score score of greater than or equal to 200.

[0025] Or

[0026] The patient with hepatocellular carcinoma has a tumor tissue biopsy that, when tested for AKR1C3 enzyme protein expression level via immunohistochemical staining method, shows a sum of percentages of moderate intensity staining and high intensity staining of greater than or equal to 70%.

[0027] A use of AST-3424 as a single agent in the preparation of a medicament for treating a patient with hepatocellular carcinoma, characterized in that:

[0028] The patient has a tumor tissue biopsy that, when tested for AKR1C3 enzyme protein expression level via immunohistochemical staining method, has an H-score score of greater than or equal to 200.

[0029] Or

[0030] The patient has a tumor tissue biopsy that, when tested for AKR1C3 enzyme protein expression level via immunohistochemical staining method, shows a sum of percentages of moderate intensity staining and high intensity staining of greater than or equal to 70%.

[0031] The AKR1C3 enzyme protein expression level of a tumor tissue biopsy of a patient is tested via immunohistochemical staining method (IHC), and the specific IHC testing method is disclosed in patent application PCT / CN2021 / 114774, publication number WO2022048492A1, the calculation of which is as follows:

[0032] The percentage of cells stained in the area of interest, the AKR1C3 assay is evaluated on a semi-quantitative scale and the percentage of cells staining for cytoplasmic and nuclear staining is recorded at the following four levels (0, 1+, 2+ and 3+).

[0033] (3) Tumor sample scoring criteria

[0034] The degree of staining, i.e. the level of AKR1C3 enzyme expression, is scored using the H-score (H-score): % of tumor cells staining nuclear-cytoplasmic (the total value from 0 to 3+ should not exceed 100):

[0035] 0 (unstained): value between 0 and 100

[0036] Tumor cell nuclear-cytoplasmic 1+ (weak staining): value between 0 and 100

[0037] Tumor cell nuclear-cytoplasmic 2+ (moderate staining, medium): value between 0 and 100

[0038] Tumor cell nuclear-cytoplasmic 3+ (high degree of staining, strong): value between 0 and 100

[0039] Total % of nuclear-cytoplasmic positive staining: value between 0 and 100

[0040] The total H-score will be calculated from the proportion of tumors scored for each intensity. The final H-score is calculated as follows: H-score = (% weak [1+] x 1) + (% medium [2+] x 2) + (% high [3+] x 3), with the final H-score value ranging from 0-300.

[0041] Obviously, the H-score or H-score score result 200 described above, with the sum of the percentage of moderate intensity staining and high intensity staining greater than or equal to 70%, the two are not completely equivalent in theory, the latter does not take into account the case of weak staining, but the applicant found in the ongoing phase II clinical trial that the patients with advanced hepatocellular carcinoma whose detection results of the sum of the percentage of moderate intensity staining and high intensity staining were greater than or equal to 70% had H-score score results greater than or equal to 200!

[0042] AST-3424 refers to a medicament containing AST-3424, i.e. a pharmaceutical preparation or drug product (as opposed to an API) that can be administered directly to a human patient.

[0043] By drug as described herein is meant a drug product or preparation, the resulting drug product comprising a specific dose range of the active ingredient AST-3424, the resulting drug being a specific dosage form, administered in a specific manner.

[0044] The prepared drug, medicine, preparation can also contain pharmaceutically acceptable adjuvants or excipients. The medicine can be in any dosage form for clinical administration, such as tablets, suppositories, dispersible tablets, enteric-coated tablets, chewable tablets, oral disintegrating tablets, capsules, sugar-coated tablets, granules, dry powder, oral solutions, small needles for injection, lyophilized powder needles for injection, or large infusions. Depending on the specific dosage form and administration method, the pharmaceutically acceptable adjuvants or excipients in the medicine can include one or more of the following: diluents, solubilizers, disintegrants, suspending agents, lubricants, binders, fillers, flavorings, sweeteners, antioxidants, surfactants, preservatives, coating agents, and pigments, etc.

[0045] The currently available dosage form is an injection, i.e., AST-3424 concentrated solution for injection disclosed in patent application PCT / CN2020 / 101870, publication number WO2021008520 (corresponding to Chinese application CN202080001484.5, publication number CN112469394A), which is diluted before use and administered by intravenous injection, preferably by intravenous infusion or injection pump bolus. However, other administration dosage forms can be developed later.

[0046] AST-3424 is prepared as AST-3424 concentrated solution for injection, which has a specification of containing 10 mg of AST-3424 raw drug per 1 mL.

[0047] Preferably, AST-3424 is prepared as AST-3424 concentrated solution for injection, which has a specification of containing 10 mg of AST-3424 raw drug per 1 mL, and is labeled as containing 0.75 ml of ethanol, 0.25 ml of propylene glycol, and 10 mg of AST-3424 raw drug.

[0048] Preferably, the above-mentioned AST-3424 concentrated solution for injection is provided in the following two specifications:

[0049] 0.5 ml containing 5 mg of AST-3424 raw drug, labeled as containing 0.375 ml of ethanol, 0.125 ml of propylene glycol, and 5 mg of AST-3424 raw drug, packaged in a 1 ml or 2 ml specification brown vial;

[0050] 1.0 ml containing 10 mg of AST-3424 raw drug, labeled as containing 0.75 ml of ethanol, 0.25 ml of propylene glycol, and 10 mg of AST-3424 raw drug, packaged in a 2 ml or 5 ml specification brown vial.

[0051] Obviously, the above-mentioned injection concentrated solution cannot be directly administered and needs to be diluted and prepared:

[0052] Before administration, 0.1ml of 5% sodium bicarbonate injection is added to 100ml of sterile 5% glucose injection to adjust the pH value in the intravenous infusion bag without di(2-ethylhexyl) phthalate;

[0053] The required number of milliliters of AST-3424 injection concentrate is added to the 5% glucose injection bag after adjusting the pH value, accurate to 0.01ml, to prepare AST-3424 injection for intravenous infusion administration for intravenous injection administration.

[0054] If the patient is not suitable for injection of glucose, use normal saline instead:

[0055] Before administration, 0.1ml of 5% sodium bicarbonate injection is added to 100ml of sterile 0.9% normal saline for injection to adjust the pH value in the intravenous infusion bag without di(2-ethylhexyl) phthalate;

[0056] The required number of milliliters of AST-3424 injection concentrate is added to the 5% glucose injection bag after adjusting the pH value, accurate to 0.01ml, to prepare AST-3424 injection for intravenous infusion administration for intravenous injection administration.

[0057] AST-3424 can be used as a single drug to treat hepatocellular carcinoma, or in combination with other drugs to treat hepatocellular carcinoma.

[0058] Single drug, i.e. single drug treatment, also requires that the drug contains only AST-3424 as the active ingredient, and does not contain other active ingredients, that is, the drug is not a compound drug. Single drug treatment refers to the use of only one anticancer drug in a course of treatment.

[0059] Combination, i.e. combination therapy. Combination therapy refers to the simultaneous or sequential use of two or more anticancer drugs in a course of treatment.

[0060] The drug used in combination with AST-3424 can be other anticancer drugs, and there are documents showing that combination with abiraterone, prednisolone, 5-fluorouracil, sunitinib (WO2022178821), immune checkpoint inhibitors such as PD-1 / L1 (WO2022231580), nelarabine (WO2019062919), oxaliplatin / 5-fluorouracil (document 6) has obvious combination effect.

[0061] The administration scheme when using AST-3424 for treatment is as follows:

[0062] Each cycle of 21 days, each dose is administered once on the 1st and 8th day, the first dose is 6.0mg / m 2 The longest will be allowed to receive 34 cycles of treatment.

[0063] The recommended dose of 6.0 mg / m 2 The recommended dose is based on the results of a phase I clinical trial of solid tumors in China (CTR20191371), and the medical professional can adjust the dose according to the patient's condition and other factors.

[0064] AST-3424 belongs to targeted chemotherapy, and the final metabolite AST-2660 belongs to a cytotoxic drug. When administered, the specific dose is calculated according to the body surface area of the patient. The calculation dose is not fixed, and the dose should be reduced according to the adverse reactions of the patient during treatment, or even at the beginning of treatment, the dose should be adjusted according to the patient's age, concomitant diseases, etc. to ensure the safe and effective treatment. The efficacy of cytotoxic drugs is linearly related to the dose, so under the premise of patient tolerance, sufficient dose should be given as much as possible to ensure efficacy, and dose reduction should follow a unified and strict plan: if the patient is intolerant, the dose should be reduced to 4.5, 3.0, 1.5 mg / m 2 based on the recommended dose of 6.0 mg / m 2 .

[0065] Body surface area (BSA), according to the effective dose described above, for an ordinary patient (height 175 cm, weight 75 kg), the corresponding equivalent body surface area BSA (m 2 ) = ([height (cm) x weight (kg)] / 3600) 1 / 2 = 1.90 (calculated using the Mosteller formula), the corresponding dose is 11.4 mg! Correspondingly, using the above 1 ml: 10 mg AST-3424 injection concentrate, then 1.14 ml of the above injection concentrate should be taken during dilution.

[0066] The prepared intravenous AST-3424 injection solution should be injected within 8 hours, because through research, the AST-3424 solution diluted with glucose or the AST-3424 solution diluted with normal saline is stable at room temperature (25°C) for 8 hours. As a preferred, it is better to be injected within 25-35 minutes.

[0067] In the China II phase clinical trial disclosed in this application, all enrolled patients were advanced refractory hepatocellular carcinoma patients who had undergone multiple treatments, including:

[0068] hepatectomy, TACE (transcatheter arterial chemoembolization), hepatic arterial infusion chemotherapy, single drug chemotherapy and multi-drug combination chemotherapy for systemic administration, antibody drug immunotherapy (including single antibody drugs such as PD-1 / PD-L1 / CTAL-4 or double antibody drugs, multi-antibody drugs), kinase inhibitor drug targeted therapy (i.e. molecular targeted therapy such as tyrosine kinase inhibitors, etc.), combination therapy of antibody drugs and kinase inhibitor drugs (targeted immunotherapy), radiotherapy, cell therapy, and traditional Chinese medicine treatment.

[0069] According to the statistical classification, the 30 patients enrolled in the above-mentioned phase II clinical trial have undergone antibody drug immunotherapy and molecular targeted therapy, or antibody drug immunotherapy and molecular targeted therapy combination therapy (targeted immunotherapy). The preliminary results of the clinical trial show that AST-3424 monotherapy is still effective for patients with advanced refractory hepatocellular carcinoma who have experienced disease progression (PD) after the above-mentioned treatment.

[0070] Further p53 gene mutation detection of the above-mentioned 30 patients can know that patients with negative p53 gene mutation detection or positive p53 gene detection results but can be judged as non-fatal mutations without affecting the normal expression of p53 protein have a significantly more excellent response to AST-3424, that is, compared with patients with positive p53 gene mutation detection and affecting the normal expression of p53 protein, the above-mentioned patients have better therapeutic effect after receiving AST-3424 treatment. That is, for patients whose AKR1C3 enzyme protein expression level is detected by immunohistochemical staining method, the H-score score is greater than or equal to 200, and the sum of the percentage of moderate intensity staining and high intensity staining is greater than or equal to 70%, if further p53 gene mutation detection result is negative or p53 gene detection result is positive but can be judged as non-fatal mutation without affecting the normal expression of p53 protein, receiving AST-3424 treatment will obtain better benefit.

[0071] p53 gene, also known as tp53 gene, is tumor protein p53 gene (Tumor protein p53).

[0072] In this application, p53 (protein) and p53 (gene) are not distinguished, and both are used.

[0073] In this application, p53 gene mutation negative includes the case of not detecting mutation and although detecting mutation but not reaching the designated positive degree.

[0074] Currently, there are related detection kits approved for commercial use, which can be directly purchased and used for detection, such as:

[0075] ​tp53 Six Mutations Detection Kit,

[0076] FISH detection Kit for the p53 gene produced by Sino-Technologies Biotech Co., Ltd. in Henan, China,

[0077] VariantPlex produced by Integrated DNA Technologies, Inc. in the United States VariantPlex TM p53 kit for

[0078] p53 protein expression is normal, including normal expression and high expression, overexpression. The normal value is a value artificially drawn according to the statistics of clinical practice.

[0079] p53 protein normal expression, that is, it can be determined by comparing typical WB experiments, and it can also be determined by IHC in medical clinics. The amount of p53 protein in the test sample is directly determined by these methods, and then compared with the normal value or the set threshold to determine whether it is normal expression or high expression.

[0080] It can also be screened by gene detection. Generally, if pathogenic gene mutations are detected, it can be determined that p53 protein is not normally expressed or overexpressed. If no gene mutation is detected or although a gene mutation is detected, it is not a pathogenic mutation, then it is most likely that p53 protein is normally expressed. Therefore, whether p53 protein is normally expressed / overexpressed can be determined by detecting whether p53 gene has pathogenic gene mutations.

[0081] The I-phase and II-phase clinical trials of AST-3424 (OBI-3424) disclosed in the above invention content in China and the United States have been approved by the drug regulatory departments of the places, and the related clinical trials have been completed by clinical research institutions. The related clinical trial schemes have been approved by the ethics committees, and the clinical trial processes comply with the relevant medical ethics requirements and legal and regulatory requirements of the places where the clinical trials are located. BRIEF DESCRIPTION OF DRAWINGS

[0082] Figure 1 is the TEAE statistics of patients enrolled in the I-phase solid tumor clinical trial of OBI-3424 (extracted from Table 2 in document 5 and translated into Chinese).

[0083] Figure 2 is a preliminary treatment effect statistics of enrolled patients in the phase I clinical trial of OBI-3424 (extracted from figure 4 in document 5 and translated into Chinese). Examples

[0084] The English abbreviations appearing in the examples are subject to the explanations and descriptions in medical textbooks or relevant medical societies, if not otherwise stated.

[0085] Example 1 AST-3424 phase I clinical trial conducted in China

[0086] The clinical trial has a clinical registration number of CTR20191371.

[0087] The trial has been approved by the ethics committee of the medical institutions participating in the clinical trial, conducted in accordance with the principles of the Helsinki Declaration, and has obtained the informed consent of all subjects.

[0088] Inclusion criteria

[0089] 1. Male or female, aged 18-70 years.

[0090] 2. Pathohistologically and / or cytologically confirmed malignant solid tumor (including but not limited to hepatocellular carcinoma, intrahepatic cholangiocarcinoma, gastric cancer, esophageal cancer, colorectal cancer, pancreatic cancer, renal cell carcinoma, non-small cell lung cancer, and castration-resistant prostate cancer), and metastatic or unresectable advanced cases, and standard treatment failure, or no standard treatment, or not suitable for standard treatment at this stage.

[0091] 3. Once the MTD is confirmed, the dose expansion group (MTD group) expands the enrollment of subjects who need at least one measurable lesion meeting the RECIST 1.1 criteria. The lesion that has been irradiated before cannot be used as a measurable lesion unless it has clear imaging progression after radiotherapy.

[0092] 4. Eastern Cooperative Oncology Group (ECOG) performance status score is 0 or 1.

[0093] 5. Life expectancy ≥ 12 weeks.

[0094] 6. All toxicities (except alopecia, fatigue, or peripheral neuropathy) of previous anticancer therapy must have recovered to grade 1 or baseline level (NCI CTCAE version 5) before starting the use of the study drug.

[0095] 7. Male cardiac QTcF interval is ≤ 450 milliseconds, and female is ≤ 470 milliseconds.

[0096] 8. Laboratory tests must meet the following criteria. Within 14 days prior to the screening period, the indicators cannot be corrected by blood transfusions or hematopoietic stimulating factors to meet the inclusion criteria: a. Hemoglobin ≥ 90 g / L; b. Platelet count ≥ 100 × 10⁻⁶. 9 c. Absolute neutrophil count (ANC) ≥ 1.5 × 10⁹ / L; 9 / L; d. Total bilirubin ≤1.5×ULN; e. ALT and AST ≤3.0×ULN; ≤5.0×ULN in the presence of liver tumors; f. Creatinine clearance >50 mL / min as determined by the Cockcroft-Gault equation.

[0097] 9. No history of alcoholism, drug abuse, or substance abuse within the past year.

[0098] 10. Female patients of childbearing age should have a negative pregnancy test result within 5 days prior to the start of treatment and should not be breastfeeding (a positive urine pregnancy test result needs to be confirmed by a serum pregnancy test).

[0099] 11. Female and male participants of childbearing age must agree to use effective contraception with their partners from the start of the study (e.g., surgical sterilization or condoms or diaphragm contraception combined with spermicide gel or intrauterine device [IUD], etc.) until 6 months after the last dose.

[0100] 12. Participants must voluntarily participate in this study, fully understand the relevant risks, demonstrate good compliance, and sign an informed consent form.

[0101] Exclusion criteria:

[0102] 1. Untreated active central nervous system (CNS) metastases or leptomeningeal disease. Subjects with adequately treated CNS metastases are eligible to participate in the study if their CNS metastases are confirmed to be stable for at least 4 weeks by clinical examination and brain imaging (MRI or CT) during the screening period.

[0103] 2. Underwent major surgery (excluding diagnostic surgery) within 4 weeks prior to the first dose.

[0104] 3. Patients who have received radiotherapy, surgery, chemotherapy, immunotherapy, cancer-specific biotherapy, targeted therapy, or hormone therapy within 4 weeks prior to the first dose (nitrosourea or mitomycin C treatment requires a 6-week washout period; oral fluorouracil drugs require a 2-week washout period; small molecule targeted therapy requires a 2-week washout period)

[0105] 4. Participated in a study of the investigational drug (diagnostic or therapeutic) or device within 4 weeks prior to the first dose.

[0106] 5. During the study, a potent CYP3A4 inhibitor or inducer must be used in combination.

[0107] 6. Uncontrolled, systemic therapy-required active bacterial, viral, or fungal infection.

[0108] 7. Known infection with human immunodeficiency virus (HIV) or positive for syphilis.

[0109] 8. Women who are pregnant, lactating, or planning to become pregnant.

[0110] 9. Concomitant illness or symptoms that can interfere with study conduct, or physical abnormalities that the investigator considers to pose an undue risk to the patient, including but not limited to active peptic ulcer or gastritis, changes in mental status or psychiatric abnormalities that can interfere with the patient’s understanding of the informed consent.

[0111] 10. Prior hypersensitivity to ethanol, propylene glycol.

[0112] 11. Subjects who are unwilling or unable to comply with the study protocol for any reason.

[0113] Study drug:

[0114] AST-3424 Injection Concentrate Solution: Manufactured by a pharmaceutical company commissioned by Shenzhen Aixinda Wei Medicine Technology Co., Ltd., specification 1 mL: 10 mg; contains 0.75 ml of ethanol, 0.25 ml of propylene glycol, and 10 mg of AST-3424.

[0115] Dosing regimen:

[0116] Each cycle of 21 days, administered on days 1 and 8, in the order of 1.0, 2.0, 4.0, 6.0, and 8.0 mg / m 2 A total of 5 dose levels of dose escalation, continuous administration until disease progression or occurrence of intolerable toxicity, up to 2 years.

[0117] Specific administration operation:

[0118] Before administration, 0.1 ml of 5% sodium bicarbonate injection is added to 100 ml of commercially available sterile 5% glucose injection for water (D5W) in a DEHP-free (di(2-ethylhexyl) phthalate) intravenous infusion bag. The required milliliters of AST-3424 Injection Concentrate Solution calculated to 0.01 ml are added to the D5W bag after adjusting the pH value to prepare AST-3424 injection for intravenous infusion administration.

[0119] The solute of this intravenous injection aqueous solution is composed of AST-3424 drug substance, glucose, ethanol, propylene glycol, and pH adjuster sodium bicarbonate, wherein the concentration of AST-3424 drug substance is 0.004-0.94 mg / ml, the pH is 7.4, the content of glucose is 4.5-5.0% by mass, and it is an isotonic solution.

[0120] If the patient is not suitable for injection of glucose, use normal saline instead:

[0121] Before use, add 0.1 ml of 5% sodium bicarbonate injection to 100 ml of commercially available sterile 0.9% normal saline for injection without DEHP (di-(2-ethylhexyl) phthalate). Add the calculated number of milliliters of AST-3424 concentrated solution for injection (to the nearest 0.01 ml) to the normal saline bag after adjusting the pH value to prepare AST-3424 injection for intravenous infusion administration.

[0122] The intravenous injection aqueous solution solute consists of AST-3424 bulk drug, sodium chloride, ethanol, propylene glycol, and pH adjuster sodium bicarbonate, wherein the concentration of AST-3424 bulk drug is 0.004-0.94 mg / ml, the pH is 7.4, the content of sodium chloride is 0.81-0.90% by mass, and it is an isotonic solution.

[0123] The accurate calculation method of the required number of milliliters of AST-3424 concentrated solution for injection is as follows:

[0124] For a patient with a height of 175 cm and a weight of 75 kg, the corresponding equivalent body surface area BSA (m 2 ) = ([height (cm) x weight (kg)] / 3600) 1 / 2 = 1.90, the corresponding dose is 1.90 x 6.0 = 11.40 mg, so 1.14 ml of the above specification AST-3424 concentrated injection should be extracted.

[0125] The prepared intravenous AST-3424 injection should be injected within 8 hours. In actual operation, an injection pump is used for intravenous bolus injection, and the injection is completed within 25-35 minutes.

[0126] Test results

[0127] A total of 21 subjects were enrolled in this trial. Each 21-day cycle, on day 1 and day 8, 1.0, 2.0, 4.0, 6.0 and 8.0 mg / m 2 There are 5 dose levels of dose escalation. After discussion by the Safety Review Committee (SRC), it is confirmed that each 21-day cycle, on day 1 and day 8, the MTD and RP2D of AST-3424 are 6.0 mg / m 2 . 2 ).

[0128] The efficacy analysis showed that among the 15 subjects who completed the evaluation, the best efficacy was stable disease (SD) in 10 cases, including 1 case of submandibular gland cancer, 1 case of prostate cancer, 1 case of breast cancer, 1 case of pancreatic cancer, 3 cases of hepatocellular carcinoma, and 3 cases of colorectal cancer.

[0129] Specifically, the enrolled hepatocellular carcinoma patients had a 100% SD (stable disease) rate, with doses distributed at 4.0, 6.0, and 8.0 mg / m². 2 The efficacy of the treatment in the dosage group for patients with hepatocellular carcinoma enrolled in stage I is shown in the table below (one patient was not evaluated):

[0130] Safety data showed that 4 cases of DLT were observed among the 21 subjects. Among them, 6.0 mg / m²... 2 One of the six subjects in the dosage group reported DLT, a grade 4 decrease in platelet count; 8.0 mg / m² 2 Of the nine subjects in the dosage group, three reported drug-related adverse events (DLT), including two cases of grade 4 decreased platelet count and one case of grade 2 elevated γ-GT (gamma-glutamyl transferase). The reported drug-related adverse events were mainly anemia, decreased platelet count, and fatigue, as well as gastrointestinal disorders (such as vomiting and nausea), most of which were grade 1-2 in severity. During the study, the laboratory indicators, vital signs, physical examination, electrocardiogram, and weight of the vast majority of subjects remained stable relative to baseline.

[0131] Specifically:

[0132] During the trial, a total of 147 adverse events (AEs) occurred in 21 subjects (N=21), with an AE incidence rate of 100.0%; among them

[0133] 1.0 mg / m 2 In group N=1, one subject experienced one adverse event (AE).

[0134] 2.0 mg / m 2 In group N=1, one subject experienced 12 adverse events (AEs).

[0135] 4.0 mg / m 2 In group N=4, 4 subjects experienced 27 adverse events;

[0136] 6.0 mg / m 2 In group N=6, 6 subjects experienced 37 adverse events;

[0137] 8.0 mg / m 2 In group N=9, 9 subjects experienced 70 adverse events.

[0138] A total of 21 subjects (N=21) experienced 145 TEAEs during the trial, with a TEAE incidence of 100.0%; of which

[0139] 1.0 mg / m 2 One subject (N=1) in the 1.0 mg / m

[0140] 2.0 mg / m 2 One subject (N=1) in the 2.0 mg / m

[0141] 4.0 mg / m 2 Four subjects (N=4) in the 4.0 mg / m

[0142] 6.0 mg / m 2 Six subjects (N=6) in the 6.0 mg / m

[0143] 8.0 mg / m 2 Nine subjects (N=9) in the 8.0 mg / m

[0144] A total of 10 subjects (N=21) experienced 20 Grade 3-5 TEAEs during the trial, with a Grade 3-5 TEAE incidence of 47.6%; of which

[0145] 4.0 mg / m 2 One subject (N=4) in the 4.0 mg / m

[0146] 6.0 mg / m 2 One subject (N=6) in the 6.0 mg / m

[0147] 8.0 mg / m 2 Eight subjects (N=9) in the 8.0 mg / m

[0148] 1.0 mg / m 2 No Grade 3-5 TEAEs occurred in the 1.0 mg / m 2 2.0 mg / m

[0149] A total of 4 subjects (N=21) experienced 4 DLTs during the trial, with a DLT incidence of 19.0%; of which

[0150] 6.0 mg / m 2 One subject (N=6) in the 6.0 mg / m

[0151] 8.0 mg / m2 Three DLTs occurred in 3 subjects in Cohort N=9, with an incidence of 33.3%, including 2 cases of Grade 4 reduction in platelet count and 1 case of Grade 2 increase in γ-GT (γ-glutamyl transpeptidase) (resulting in a delay of more than 14 days in the administration of the drug and the determination of DLT), 1.0 mg / m2 2 No DLTs occurred in Cohort N=1, 2.0 mg / m2 2 No DLTs occurred in Cohort N=1, 2.0 mg / m2 2 No DLTs occurred in Cohort N=4.

[0152] No adverse reactions leading to death occurred in the five groups during the trial,

[0153] During the study period, the laboratory indicators, vital signs, physical examination, electrocardiogram, body weight, etc. of most subjects remained stable compared with baseline.

[0154] Example Two: AST-3424 Phase II Clinical Trial in China

[0155] The clinical trial has the clinical registration number CTR20191399.

[0156] The trial has been approved by the ethics committee of the medical institution participating in the clinical trial, conducted in accordance with the principles of the Helsinki Declaration, and has obtained the informed consent of all subjects.

[0157] Inclusion criteria

[0158] 1. Male or female, aged ≥ 18 years.

[0159] 2. Pathologically histologically confirmed advanced HCC that cannot be controlled by surgical resection or local treatment.

[0160] 3. Previously received standard systemic therapy, including but not limited to sorafenib and / or oxaliplatin-containing systemic chemotherapy, lenvatinib, regorafenib and / or nivolumab, and developed disease progression, toxicity intolerance or refused to continue receiving these drugs.

[0161] 4. At least one measurable lesion meeting the RECIST 1.1 standard. Lesions previously treated with radiotherapy cannot be used as measurable lesions unless they have clear imaging progression after radiotherapy.

[0162] 5. Pathological paraffin blocks or sections (including archived pathological paraffin blocks and sections) for AKR1C3 expression analysis can be provided, and it is confirmed that the liver tumor tissue AKR1C3 expression is strongly positive (the center laboratory immunohistochemical result confirms that the proportion of tumor cells with AKR1C3 staining intensity of 2+ and / or 3+ is ≥ 70%).

[0163] 6. Eastern Cooperative Oncology Group (ECOG) performance status score of 0 or 1.

[0164] 7. Life expectancy > 12 weeks.

[0165] 8. With or without HBV or HCV infection. a. Subjects with HBV infection must have HBV-DNA less than 2,000 IU / ml, and be on antiviral treatment with Entecavir, Tenofovir disoproxil fumarate, or Prodrug of Tenofovir fumarate according to the National Guidelines for the Prevention and Treatment of Chronic Hepatitis B, and need to maintain treatment during the study and continue until 6 months after the last dose. b. Subjects with HCV infection (presence of detectable HCV-RNA or anti-HCV antibodies) can be treated according to medical practice.

[0166] 9. Child-Pugh score ≤ 6 points.

[0167] 10. No history of hepatic encephalopathy.

[0168] 11. All toxicities of prior anticancer therapy (except alopecia, fatigue, or peripheral neuropathy) must have recovered to Grade 1 or baseline levels (NCI CTCAE version 5) prior to starting study drug.

[0169] 12. Laboratory tests must meet the following criteria. Within 14 days prior to screening laboratory tests, the indicators cannot be corrected by blood transfusion or hematopoietic stimulating factors, input of albumin to meet the inclusion criteria. a. Hemoglobin ≥ 90 g / L; b. Platelet count ≥ 80 x 10 9 / L; c. Absolute neutrophil count (ANC) ≥ 1.5 x 10 9 / L; d. Serum total bilirubin ≤ 3 mg / dL; e. ALT and AST ≤ 5.0 x ULN; f. International normalized ratio (INR) ≤ 2.3 or prothrombin time prolongation ≤ 6 seconds; g. Albumin ≥ 29 g / L; h. Creatinine clearance > 50 mL / min according to Cockcroft-Gault equation.

[0170] 13. No history of alcoholism, drug use, or drug abuse within the last year.

[0171] 14. Female patients of childbearing potential must be in a non-lactating period and have a negative pregnancy test result within 5 days prior to treatment initiation (a positive result of a urine pregnancy test requires confirmation by a serum pregnancy test).

[0172] 15. Female and male subjects of childbearing potential must agree to use effective means of contraception (e.g., surgical sterilization or condom or diaphragm contraceptive measures combined with spermicidal gel or intrauterine device [IUD], etc.) with their partner from the start of participation in the study until 6 months after the last dose.

[0173] 16. Willing to participate in the study, fully understand the risks, be compliant, and sign the informed consent form. Subjects can also sign the Future Biomedical Research (FBR) consent form. However, subjects who do not participate in FBR can also participate in the main trial.

[0174] Exclusion Criteria:

[0175] 1. Untreated active central nervous system (CNS) metastases or leptomeningeal disease. Subjects with CNS metastases who have been adequately treated and are confirmed to be stable for at least 4 weeks by clinical examination and brain imaging (MRI or CT) during the screening period can participate in the study.

[0176] 2. History of other malignancy within 2 years, except for adequately treated basal cell carcinoma, carcinoma in situ of another site, or other neoplasm whose natural history or treatment will not interfere with the safety or efficacy assessments of the current study.

[0177] 3. Major surgery other than diagnostic surgery within 4 weeks prior to first dose.

[0178] 4. Received radiotherapy, surgical therapy, chemotherapy, immunotherapy, biological therapy directed against cancer, targeted therapy, or hormonal therapy within 4 weeks prior to first dose (6-week washout period for nitrosoureas or mytomycin C therapy; 2-week washout period for oral fluorouracil; 2-week washout period for small molecule targeted therapy).

[0179] 5. Participation in a study drug (diagnostic or therapeutic) or device study within 4 weeks prior to first dose.

[0180] 6. Concomitant use of strong CYP3A4 inhibitors or inducers during the study.

[0181] 7. Uncontrolled, active bacterial, viral, or fungal infection requiring systemic treatment.

[0182] 8. Known positive infection with human immunodeficiency virus (HIV) or syphilis.

[0183] 9. Clinically significant ascites, defined as found by physical examination and requiring control by paracentesis or increased medical intervention to maintain symptoms (patients with ascites found only by imaging can be enrolled).

[0184] 10. Women who are pregnant, breastfeeding, or planning to become pregnant.

[0185] 11. Concomitant illness or symptoms that can interfere with study conduct or that the investigator considers to pose an excessive risk to the patient. This includes, but is not limited to, a history of gastrointestinal bleeding or a higher risk of bleeding within three months, active peptic ulcer or gastritis, changes in mental status or psychiatric abnormalities that can interfere with the patient's understanding of the informed consent form.

[0186] 12. Previous hypersensitivity to ethanol, propylene glycol.

[0187] 13. Subjects unwilling or unable to comply with the study protocol for any reason.

[0188] Study drug:

[0189] AST-3424 injection concentrated solution: AST-3424 injection concentrated solution is manufactured by a pharmaceutical company entrusted by Shenzhen Aishinda Wei Medicine Technology Co., Ltd. The specification is 1 mL: 10 mg; containing 0.75 ml of ethanol, 0.25 ml of propylene glycol and 10 mg of AST-3424.

[0190] Dosing regimen:

[0191] Every 21 days for one cycle, each dose is given once on the first day and the eighth day, with a dose of 6 mg / m 2 The longest will be allowed to receive 34 cycles of treatment.

[0192] The specific operation of administration is the same as in Example 1.

[0193] Clinical evaluation

[0194] The effectiveness evaluation includes clinical efficacy evaluation.

[0195] The clinical efficacy evaluation standard adopts the RECIST 1.1 standard for evaluating the efficacy of solid tumors. The measurement method uses MRI / CT to evaluate the lesions, and the same evaluation method should be used for the same lesions during the study. The subjects must have measurable tumor lesions at baseline.

[0196] The efficacy evaluation indicators include complete remission (complete response, CR), partial remission (partial response, PR), stable disease (stable disease, SD) and progressive disease (progressive disease, PD).

[0197] Complete remission (CR): all target lesions disappear, and the short diameter of all pathological lymph nodes (including target nodes and non-target nodes) must be reduced to <10 mm.

[0198] Partial remission (PR): the sum of the diameters of the target lesions is reduced by at least 30% compared with the baseline level.

[0199] Progressive disease (PD): the minimum value of the sum of the diameters of all measured target lesions throughout the experimental study is taken as the reference, and the diameter and relative increase is at least 20% (if the baseline measurement value is the minimum, the baseline value is taken as the reference); in addition, the absolute value of the diameter and must increase by at least 5 mm (the appearance of one or more new lesions is also considered as disease progression).

[0200] Stable Disease (SD): The extent of the decrease in the target lesions is not enough to meet the PR, and the extent of the increase is also not enough to meet the PD level, between the two, the minimum sum of the diameters can be used as a reference during the study.

[0201] Study Endpoints

[0202] The efficacy of AST-3424 monotherapy in the treatment of malignant tumors such as HCC was preliminarily evaluated according to the objective response rate (ORR) of the subjects, disease control rate (DCR), duration of remission (DOR), and progression-free survival (PFS).

[0203] Objective Response Rate (ORR): Refers to the percentage of cases that achieve complete remission (CR) and partial remission (PR) after treatment relative to the total number of evaluable cases.

[0204] Disease Control Rate (DCR): Refers to the percentage of cases that achieve complete remission (CR), partial remission (PR), and stable disease (SD) in patients who can evaluate the efficacy.

[0205] Trial Results

[0206] As of March 11, 2024, 30 subjects have been enrolled: 5 are in the group treatment, 25 have been out of the group (1 has withdrawn informed consent, 10 have died, and 14 are in survival follow-up), and the current longest PFS is > 11.5 months; the longest OS is > 17.6 months. Among the subjects enrolled before the end of January 2023, there are 11 cases with OS greater than 12 months.

[0207] Investigator Assessment:

[0208] 26 cases have completed the first investigator tumor evaluation, 2 cases have achieved partial remission (PR), 15 cases have stable disease (SD, including 1 case of unplanned tumor evaluation after C2D1), and 9 cases have progressive disease (PD). The disease control rate (DCR) is 60.7% (17 / 28), and the objective response rate (ORR) is 7.1% (2 / 28) (2 cases not meeting the tumor evaluation date are not included).

[0209] Among them, 03003 patients have completed 6 investigator tumor evaluations, all of which are PR, and the target lesion size decreases by 41.5%, 44.1%, 58.4%, 62.2%, 60.3%, and 64.9%, respectively, showing a shrinking trend. The tumor also gradually decreased, with AFP being > 1210 (baseline), 436.1, 80.31, 21.2, 6.09, 3.41, and 2.46 ng / ml, respectively.

[0210] 06010The patient has completed 3 investigator tumor assessments, the first two tumor assessments were PR, the target lesions were reduced by 50%, 53.8% successively; the third tumor assessment was PD (the target lesions were reduced by 57.7% compared with baseline, and PD was judged due to new lesions in the right parietal lobe, retroperitoneal space, and diaphragm), and AFP was 2630.30 (baseline), 58.52, 174.29, and 1377.57 ng / ml successively.

[0211] The details are shown in Table 6 as follows.

[0212] Table 6: Clinical data of AST-3424-liver cancer efficacy and gene mutation relationship as of March 11, 2024

[0213] Among the 26 enrolled subjects with efficacy evaluation results, 20 had p53 gene mutation detection results, and the efficacy was observed according to the p53 detection results:

[0214] Wild type without mutation (WT), i.e. the p53 gene mutation detection result is negative (-);

[0215] Mutation of unknown significance (VUS), i.e. the p53 gene mutation detection result is positive (+), and it is not clear whether it affects protein expression;

[0216] Mutation that may affect protein function (MUT), i.e. the p53 gene mutation detection result is positive (+), which may affect protein expression, and the three groups are classified and counted, and the results are shown in Tables 7, 8, and 9.

[0217] Table 7: Efficacy data of 8 patients with P53 mutation positive (MUT)

[0218] The statistical results show that among the 8 cases, 2 have died, 6 are alive and under follow-up, the current longest PFS is 1.6 months, the longest OS is > 14.0 months, the average PFS is 1.4 months, and the average OS is greater than 9.3 months. Among the 8 cases with p53 gene mutation positive (+), which may affect protein expression, 0 cases had PR, 2 cases had SD, and 7 cases had PD, i.e. the DCR of this subgroup of cases was 25% (2 / 8), and the ORR was 0% (0 / 5). The disease control rate (DCR) of the whole group of 28 patients was 60.7% (17 / 28), and the objective response rate (ORR) was 7.1% (2 / 28). The difference is obvious: the disease control rate (DCR) and the objective response rate (ORR) of the subgroup with p53 gene mutation detection result positive (+) which may affect protein expression are significantly lower than those of the whole group.

[0219] Table 8: Efficacy data of 9 patients with P53 mutation negative (WT)

[0220] Statistical results show that 4 of the 9 cases have died, 5 cases are alive in follow-up, the current longest PFS is more than 11.5 months, the longest OS is > 13.8 months, the average PFS is more than 4.1 months, and the average OS is more than 7.7 months.

[0221] In the 9 cases of p53 gene mutation negative (-), 2 cases were PR, 5 cases were SD, and 2 cases were PD, that is, the DCR of this subgroup of case population was 77.8% (7 / 9), and the ORR was 22.2% (2 / 9). The data is significantly different from the overall disease control rate (DCR) of 60.7% (17 / 28) and the objective response rate (ORR) of 7.1% (2 / 28) of the overall 28 patients. The disease control rate (DCR) and objective response rate (ORR) of the p53 gene mutation negative (-) subgroup are significantly higher than those of the overall group.

[0222] Table 9: 3 cases of P53 mutation positive, not sure if the protein expression (VUS) is affected

[0223] Therefore, for the current clinical results, those skilled in the art have reason to believe that AST-3424 has better therapeutic effect on p53 gene mutation or defect negative (-) cancer and tumor patients than p53 gene mutation or defect positive (+) patients. Therefore, the applicant speculates that AST-3424 will have better therapeutic effect on p53 gene mutation or defect negative (-) tumor and cancer patients, that is, p53 gene mutation or defect negative (-) tumor and cancer patients receiving AST-3424 treatment will have more obvious clinical benefit.

[0224] Patient's previous treatment

[0225] UK, unknown, unknown.

[0226] By comparing the previous treatment of the above-mentioned 28 enrolled patients and the efficacy evaluation, it can be seen that AST-3424 drug still has therapeutic effect on patients with progressive disease after targeted therapy and immunotherapy, or patients with progressive disease after targeted and immunotherapy combined treatment.

[0227] The patent applications cited in this application are incorporated in their entirety into this specification.

[0228] The non-patent literatures (academic journal papers, academic conference papers, etc.) cited in this application are:

[0229] Document 1, Meng F, Li WF, Jung D, et al. A novel selective AKR1C3-activated prodrug A ST-3424 / OBI-3424 exhibits broad anti-tumor activity. Am J Cancer Res. 2021; 11(7): 3645-3659;

[0230] Document 2, Evans K, Duan J, Pritchard T, et al. OBI-3424, a Novel AKR1C3-Activated Prodrug, Exhibits Potent Efficacy against Preclinical Models of T-ALL. Clin Cancer Res. 2019; 25(14): 4493-4503. doi: 10.1158 / 1078-0432.CCR-19-0551;

[0231] Document 3, Wang Y, Liu Y, Zhou C, et al. An AKR1C3-specific prodrug with potent anti-tumor activities against T-ALL. Leuk Lymphoma. 2020; 61(7): 1660-1668. doi: 10.1080 / 10428194.2020.1728746;

[0232] Document 4, He P, Wang C, Wang Y, et al. A Novel AKR1C3 Specific Prodrug TH3424 With Potent Antitumor Activity in Liver Cancer [retracted in: Clin Pharmacol Ther. 2021 Jul; 110(1): 262]. Clin Pharmacol Ther. 2021, 110(1): 229-237. doi: 10.1002 / cpt.2171;

[0233] Document 5, Tsimberidou, Apostolia & Verschraegen, Claire & Hsu, Pei & Pearce, Tillman. (2022). Safety, pharmacokinetics, and clinical activity of OBI-3424, an AKR1C3-activated prodrug, in patients with advanced or metastatic solid tumors: A phase 1 dose-escalation study. Journal of Clinical Oncology. 40. 3030-3030. 10.1200 / JCO.2022.40.16_suppl.3030, poster available for download at OBI Pharma Inc website at https: / / www.obipharma.com / news / news-2022 / poster-presentations-at-the-2022-asco-annual-meeting-for-adagloxad-simolenin-obi-999-and-obi-3424 / ;

[0234] Document 6: Zhang Y, Qin S, Chao J, Luo Y, Sun Y and Duan J (2022) The In-Vitro Antitumor Effects of AST-3424 Monotherapy and Combination Therapy With Oxaliplatin or 5-Fluorouracil in Primary Liver Cancer. Front. Oncol. 12:885139. doi: 10.3389 / fonc.2022.885139;

[0235] Document 7: Tsimberidou, A.M., Verschraegen, C.F., Wesolowski, R. et al. Phase 1 dose-escalation study evaluating the safety, pharmacokinetics, and clinical activity of OBI-3424 in patients with advanced or metastatic solid tumors. Br J Cancer 129, 266-274 (2023). https: / / doi.org / 10.1038 / s41416-023-02280-4.

[0236] Various English abbreviations in this application are subject to the interpretation in the pharmacological and medical textbooks unless otherwise stated.

Claims

1. A method for treating hepatocellular carcinoma by AST-3424, characterized in that: the pathological paraffin block or pathological section of the liver tumor tissue of the hepatocellular carcinoma patient is detected for AKR1C3 enzyme protein expression level by immunohistochemical staining method, and the detection result H-score score is greater than or equal to 200; or the pathological paraffin block or pathological section of the liver tumor tissue of the hepatocellular carcinoma patient is detected for AKR1C3 enzyme protein expression level by immunohistochemical staining method, and the detection result shows that the sum of the percentages of moderate intensity staining and high intensity staining is greater than or equal to 70%. 2.The use of AST-3424 in the preparation of a drug for treating hepatocellular carcinoma patients, characterized in that: the pathological paraffin block or pathological section of the liver tumor tissue of the hepatocellular carcinoma patient is detected for AKR1C3 enzyme protein expression level by immunohistochemical staining method, and the detection result H-score score is greater than or equal to 200; or the pathological paraffin block or pathological section of the liver tumor tissue of the hepatocellular carcinoma patient is detected for AKR1C3 enzyme protein expression level by immunohistochemical staining method, and the detection result shows that the sum of the percentages of moderate intensity staining and high intensity staining is greater than or equal to 70%.

3. Use or method according to claim 1 or 2, characterized in that The dosage regimen of AST-3424 is: Each 21 -day cycle, on days 1 and 8, with a first dose of 6.0 mg / m 2 The longest treatment will be allowed to receive 34 cycles.

4. Use or method according to claim 3, characterized in that The dosing regimen for AST-3424 is 6.0 mg / m 2 at the first dose, and if the patient is intolerant, then the dose is reduced by 4.5, 3.0, 1.5 mg / m 2 , in that order. 5.The use or method according to claim 1 or 2, wherein the AST-3424 is prepared as a concentrated solution for injection of AST-3424, and the specification is 10 mg of AST-3424 raw material per 1 mL. 6.The use or method according to claim 5, wherein the AST-3424 is prepared as a concentrated solution for injection of AST-3424, and the specification is 10 mg of AST-3424 raw material per 1 mL, which is marked as containing 0.75 ml of ethanol, 0.25 ml of propylene glycol and 10 mg of AST-3424 raw material. 7.The use or method according to claim 6, characterized in that, before administration, 0.1 ml of 5% sodium bicarbonate injection is added to 100 ml of sterile 5% glucose injection to adjust the pH value in a bag containing no di(2-ethylhexyl) phthalate; the calculated milliliter number of AST-3424 concentrated solution for injection is added to the 5% glucose injection bag after pH adjustment, accurate to 0.01 ml, to prepare AST-3424 injection for intravenous infusion administration for intravenous injection administration; if the patient is not suitable for injection of glucose, use normal saline instead: before administration, 0.1 ml of 5% sodium bicarbonate injection is added to 100 ml of sterile 0.9% injection of normal saline to adjust the pH value in a bag containing no di(2-ethylhexyl) phthalate; the calculated milliliter number of AST-3424 concentrated solution for injection is added to the normal saline bag after pH adjustment, accurate to 0.01 ml, to prepare AST-3424 injection for intravenous infusion administration for intravenous injection administration.

8. Use or method according to claim 7, characterized in that, The prepared intravenous AST-3424 injection solution should be injected within 8 hours, preferably within 25-35 minutes.

9. Use or method according to claim 1 or 2, characterized in that, Treatment is carried out using AST-3424 as a single agent.

10. Use or method according to claim 1 or 2, characterized in that, The hepatocellular carcinoma patient is one who has experienced Progressive Disease after molecular targeted therapy and antibody immunotherapy, or one who has experienced Progressive Disease after combined molecular targeted-antibody immunotherapy.

11. Use or method according to claim 1 or 2, characterized in that, The hepatocellular carcinoma patient is one who has a negative p53 gene mutation test or normal p53 protein expression.

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