Pharmaceutical kit for treating colorectal cancer and application thereof

Abstract

The present disclosure generally relates to a pharmaceutical kit for treating colorectal cancer; the pharmaceutical kit comprises (E)-N-hydroxyl-3-(1-(benzenesulfonyl)-indoline-5-yl)-acrylamide, and an anti-PD-1 antibody or an anti-PD-L1 antibody, and is characterized in that the pharmaceutical kit comprises the (E)-N-hydroxyl-3-(1-(benzenesulfonyl)-indoline-5-yl)-acrylamide and the Furthermore, the present disclosure sets forth a method for treating colorectal cancer in a subject comprising administering to the subject the above pharmaceutical kit wherein the (E)-N-hydroxy-3-(1-(benzenesulfonyl)-indolin-5-yl)-acrylamide is administered to the subject at a dose of 10 [mu] g / kg to 1 male g / kg, and a method for treating colorectal cancer in a subject comprising administering to the subject the (E)-N-hydroxy-3-(1-(benzenesulfonyl)-indolin-5-yl)-acrylamide to the subject at a dose of 10 [mu] g / kg to 1 male g / kg, and the anti-PD-1 or anti-PD-L1 antibody is administered to the individual at a dose of 1 [mu] g / kg to 100 [mu] g / kg.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to U.S. Patent No. 63 / 444,242, filed February 8, 2023. The contents of that application are incorporated herein by reference in their entirety. Background of the Invention 1. Technical Field

[0002] The present disclosure generally relates to the technical field of cancer treatment. More specifically, the present disclosure relates to a pharmaceutical kit comprising (E)-N-hydroxy-3-(1-(phenylsulfonyl)-indolin-5-yl)-acrylamide (ABT-301) and an anti-PD-1 or anti-PD-L1 antibody for treating colorectal cancer. 2. Background Technology

[0003] Colorectal cancer is a tumor that develops in the colon or rectum, both of which are part of the large intestine. Colorectal cancer has a progressive course that may last for years. It typically originates from benign polyps in the lining of the colon or rectum. Over time, these polyps may develop into cancer cells, leading to colorectal cancer. There are various types of colorectal cancer, with adenocarcinoma accounting for the majority of cases. According to the World Health Organization (WHO), colorectal cancer ranks second in cancer-related deaths worldwide. In 2020 alone, it is estimated that there will be over 1.9 million new cases of colorectal cancer worldwide, resulting in over 930,000 deaths. Colorectal cancer incidence is highest in Europe, Australia, and New Zealand, while mortality is highest in Eastern Europe. By 2040, colorectal cancer cases are projected to increase to 3.2 million new cases annually (a 63% increase) and 1.6 million deaths annually (a 73% increase). This serious cancer imposes a significant health and economic burden on society.

[0004] In terms of medical intervention, contemporary approaches to treating colorectal cancer typically encompass a multifaceted strategy that combines surgery, radiation therapy, chemotherapy, and, in some cases, targeted therapies and immunotherapy. Different treatment strategies are used for the management of early-stage and advanced-stage disease. For early-stage disease (i.e., tumors confined to the intestine or regional lymph nodes and not yet metastasized to distant organs), surgery is typically used to remove the tumor and nearby lymph nodes. In some cases, a temporary or permanent colostomy or ileostomy may be necessary to create a drainage channel for waste. In early-stage colorectal cancer, patients may be advised to receive adjuvant chemotherapy to kill any residual cancer cells that cannot be seen or removed during surgery, or neoadjuvant chemotherapy before surgery to shrink the tumor. Radiation therapy may be administered to rectal tumors to reduce their size. For advanced-stage disease, systemic therapy (e.g., chemotherapy) is the mainstay of treatment for metastatic colorectal cancer. For patients with specific gene mutations (e.g., KRAS or BRAF mutations), targeted therapy can be combined with chemotherapy. For patients whose tumors have specific genetic signatures (e.g., microsatellite instability-high (MSI-H) or mismatch repair deficiency (dMMR)), immunotherapy can be considered.

[0005] Although modern medical interventions have advanced and improved the prognosis for many patients, current treatments still have some limitations. For example, surgical intervention is an invasive treatment method that causes direct harm to the patient; and the performance of a temporary or permanent colostomy or ileostomy can significantly affect the patient's quality of life. Chemotherapy can cause significant side effects, such as nausea and vomiting, loss of appetite, fatigue, mouth sores and oral ulcers, and alopecia. In addition, the efficacy of chemotherapy varies from patient to patient, and the emergence of chemotherapy resistance over time poses additional challenges to the long-term management of tumors.

[0006] In view of this, the relevant field urgently needs an improved therapy for treating colorectal cancer to enhance therapeutic efficacy, reduce adverse reactions, and protect the quality of life of patients. Summary of the Invention

[0007] The following is a brief summary of the present disclosure to help readers gain a basic understanding of the present disclosure. This summary is not intended to be a comprehensive overview of the present disclosure, nor is it intended to identify key or critical components of the present disclosure, or to outline the scope of the present disclosure. Its sole purpose is to present certain concepts of the present disclosure in a simplified form as a prelude to the more detailed description that follows.

[0008] As described above, the main purpose of the present disclosure is to provide an improved method for treating colorectal cancer. As embodied and broadly described herein, one aspect of the present disclosure is a pharmaceutical kit for treating colorectal cancer in an individual; the pharmaceutical kit comprises a first container comprising (E)-N-hydroxy-3-(1-(phenylsulfonyl)-indolin-5-yl)-acrylamide; and A second container comprises an anti-PD-1 or anti-PD-L1 antibody.

[0009] According to certain embodiments of the present disclosure, the anti-PD-1 antibody is selected from the group consisting of cemiplimab, pembrolizumab, nivolumab, and RMP1-14. Furthermore, the anti-PD-L1 antibody is selected from the group consisting of atezolizumab, avelumab, and durvalumab.

[0010] Preferably, the colorectal cancer treatable by the pharmaceutical kit of the present disclosure is a microsatellite stable-type or microsatellite instable-type colorectal cancer. In addition, the colorectal cancer can be in situ colorectal cancer or metastatic colorectal cancer.

[0011] Another aspect of the present disclosure relates to a method for treating colorectal cancer in an individual by using the pharmaceutical kit of the present disclosure, the method comprising administering to the individual the pharmaceutical kit of the present disclosure, wherein The (E)-N-hydroxy-3-(1-(phenylsulfonyl)-indolin-5-yl)-acrylamide is administered to the subject in an amount of 10 μg / kg to 1 g / kg; and The anti-PD-1 or anti-PD-L1 antibody is administered to the subject in an amount of 1 μg / kg to 100 mg / kg.

[0012] More preferably, the (E)-N-hydroxy-3-(1-(phenylsulfonyl)-indolin-5-yl)-acrylamide is administered to the subject in an amount of 2.5-10 mg / kg; and the anti-PD-1 or anti-PD-L1 antibody is administered to the subject in an amount of 100 μg / kg-1 mg / kg.

[0013] Exemplary anti-PD-1 antibodies for use in the pharmaceutical kits of the present disclosure may be cemiplizumab, pembrolizumab, nivolumab, or RMP1-14. Alternatively, anti-PD-L1 antibodies for use in the pharmaceutical kits of the present disclosure may be atezolizumab, avelumab, and durvalumab.

[0014] In a preferred embodiment, the colorectal cancer that can be treated by the pharmaceutical kit of the present disclosure is a microsatellite stable or microsatellite unstable colorectal cancer. In addition, the colorectal cancer can be in situ colorectal cancer or metastatic colorectal cancer.

[0015] The subject that can be treated by the pharmaceutical kit or method of the present disclosure is a mammal, for example, humans, mice, rats, guinea pigs, hamsters, monkeys, pigs, dogs, cats, horses, sheep, goats, cows, and rabbits. Preferably, the subject is a human.

[0016] Other aspects of the disclosure include a pharmaceutical composition comprising the drug combination described herein and a pharmaceutical composition formulated with a pharmaceutically acceptable excipient; and the use of the pharmaceutical kit or pharmaceutical composition of the disclosure in the preparation of a drug for treating colorectal cancer in an individual in need.

[0017] The numerous attendant features and advantages of the present disclosure will be readily apparent after reviewing the following detailed description and accompanying drawings. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] The present disclosure and other features, aspects, and advantages will become more apparent upon review of the following detailed description, claims, and accompanying drawings, in which:

[0019] Figures 1A-1G The results are shown in Figure 3. ABT-301 and avelumab (an anti-PD-L1 antibody) were used to treat mice bearing CT26 (a microsatellite stable (MSS) subtype colorectal cancer cell line), where Figures 1A-1DThe numbers of mice showing complete response (CR) in a specific treatment group are shown (n=8 / group); Figures 1E-1F Explain that within a 28-day period ( Figure 1E ) and day 28 ( Figure 1F ) tumor volume; Figure 1G Describe the survival rate of mice after a specific treatment;

[0020] Figures 2A-2J The results of the combined treatment of ABT-301 and avelumab in mice bearing CT26 were shown in Figure 2. Figures 2A-2H Indicate the number of mice (n=10 / group) that showed progressive disease (PD), stable disease (SD), partial response (PR), or complete response (CR) in a specific treatment group; and Figures 2I-2J The mice treated with specific treatments showed Figure 2I ) and day 21 ( Figure 2J ) tumor volume;

[0021] Figures 3A-3J The results of the combined treatment of ABT-301 and RMP1-14 (an anti-mouse PD-1 antibody) with CT26-bearing mice were shown in Figure 2. Figures 3A-3H Indicate the number of mice that showed PD, SD, PR, or CR in a specific treatment group (n=10 / group); and Figures 3I-3J The mice treated with specific treatments showed Figure 3I ) and day 17 ( Figure 3J ) tumor volume;

[0022] Figures 4A-4I The results are shown in the results of the combination of ABT-301 and avelumab in mice bearing MC-38 (a colorectal cancer cell line with a high microsatellite instable-high (MSI-H) subtype), in which Figures 4A-4F The number of mice showing complete remission (CR) in a specific treatment group is indicated (n=8 / group); Figures 4G-4H Explain that within a 25-day period ( Figure 4G ) and day 25 ( Figure 4H ) of tumor volume, of which the mice in the vehicle group only showed the experimental results up to day 20 because they reached the humane endpoint; Figure 4I Describe the survival rate of mice after a specific treatment; and

[0023] Figures 5A-5KThe results are shown in Figure 2. The results are shown in Figure 2. The results are shown in Figure 2. The results are shown in Figure 2. The results are shown in Figure 2. The results are shown in Figure 2. Figures 5A-5H The tumor volume of each mouse in a specific treatment group is shown separately (n=8 or 9 / group); and Figures 5I-5K The mice treated with specific treatments showed Figure 5I ) and day 22 ( Figure 5J ) and day 31 ( Figure 5K , in which a specific treatment has been terminated). DETAILED DESCRIPTION

[0024] To provide a more detailed and complete description of the present disclosure, the following illustrative descriptions of various embodiments and examples of the present invention are provided. However, these descriptions are not intended to be the only way to implement or use the embodiments of the present invention. The detailed descriptions cover the features of various embodiments and the method steps and sequences for constructing and operating these embodiments. However, other embodiments may be used to achieve the same or equivalent functionality and step sequences.

[0025] I. Definition

[0026] For convenience, specific terms used in this specification, examples, and the appended claims are collected here. Unless otherwise defined in this specification, the scientific and technical terms used herein have the same meaning as those understood and customarily used by those having ordinary skill in the art to which the invention pertains. Furthermore, singular terms used in this specification encompass the plural form of such terms, and plural terms encompass the singular form of such terms, unless inconsistent with the context. Specifically, in this specification and the claims, the singular forms "a," "an," and "the" include plural references unless the context indicates otherwise. Furthermore, in this specification and the claims, expressions such as "atleast one" and "one or more" have the same meaning, both representing one, two, three, or more. Unless otherwise indicated, the practice of this disclosure will employ conventional techniques in the fields of molecular biology, microbiology, recombinant DNA, and immunology, all of which are within the skill of the art. Such techniques are described in detail in the published literature.

[0027] Although the numerical ranges and parameters used to define the broader scope of the present invention are approximate, the numerical values ​​in the specific examples have been presented herein as precisely as possible. However, any numerical value inherently contains standard deviations resulting from individual testing methods. As used herein, "about" generally means that the actual value is within plus or minus 10%, 5%, 1%, or 0.5% of a particular value or range. Alternatively, the term "about" means that the actual value falls within an acceptable standard error of the mean, as determined by one of ordinary skill in the art. Except in the examples, or unless otherwise expressly indicated, all ranges, amounts, values, and percentages used herein (e.g., to describe material amounts, time periods, temperatures, operating conditions, quantitative ratios, etc.) are to be understood as modified by the word "about." Accordingly, unless otherwise indicated, the numerical parameters disclosed in this specification and the accompanying claims are approximate and are subject to change as needed. At a minimum, these numerical parameters should be understood to include the number of significant digits indicated and to apply normal rounding.

[0028] Colorectal cancer can be classified according to traditional histological subtypes defined by the World Health Organization. The most common subtype is adenocarcinoma not otherwise specified, which accounts for 85% of colorectal cancer cases worldwide. The second most common subtype is mucinous carcinoma, characterized by mucinous lakes in at least 50% of the tumor area, accounting for 5-20% of colorectal cancer cases worldwide. Medullary carcinoma, characterized by solid tumor growth accompanied by inflammation, accounts for an estimated 4% of colorectal cancer cases worldwide. Signet-ring cell carcinoma accounts for less than 2% of colorectal cancer cases worldwide. Alternatively, colorectal cancer can be categorized by molecular classification, with either DNA mismatch repair-proficient (pMMR) / microsatellite stable (MSS) or DNA mismatch repair-deficient / microsatellite instability-high (dMMR / MSI-H) subtypes serving as key criteria for colorectal cancer classification. Colorectal cancers of the MSI-H (or dMMR / MSI-H) subtype account for only 8-10% of all colorectal cancer cases, while 90% of colorectal cancer patients belong to the pMMR / MSS subtype. Microsatellite instability (MSI) and the corresponding microsatellite stability (MSS) subtype can be confirmed by pentaplex PCR using the single-nucleotide MSI markers BAT-25, BAT-26, NR-21, NR-22, and NR-24. MSI-positive is defined when three or more of these five markers show allele size variation.

[0029] As used herein, the term "treatment" (treatment and treating) may refer to a curative or palliative measure. The term "treatment" encompasses partial or complete prevention, improvement, alleviation, and / or management of symptoms, secondary diseases, or conditions associated with colorectal cancer. As used herein, the term "treatment" refers to the application or administration of the pharmaceutical kit and / or method of the present disclosure to an individual suffering from colorectal cancer, or suffering from a symptom or disease associated with colorectal cancer, or suffering from a secondary disease or condition secondary to colorectal cancer, in order to partially or completely alleviate, ameliorate, relieve, delay onset, inhibit progression, reduce severity, and / or reduce the incidence of one or more symptoms, secondary diseases, or conditions associated with colorectal cancer. Symptoms, secondary diseases, and / or conditions associated with colorectal cancer include, but are not limited to, fever, weakness, fatigue, weight loss, pain, cough, bleeding, skin changes, diarrhea or constipation, nausea, vomiting, and loss of appetite. Individuals who exhibit early signs of such symptoms, diseases, and / or conditions may be treated in order to reduce the risk of developing symptoms, secondary diseases, and / or conditions associated with colorectal cancer. As defined herein, a treatment is generally considered "effective" if one or more symptoms or clinical manifestations or markers are reduced. Alternatively, a treatment is "effective" if the progression of a symptom, disease, or condition is reduced or stopped.

[0030] The terms "administered," "administering," or "administration" are used interchangeably herein to refer to the direct administration of a pharmaceutical kit of the present disclosure comprising (E)-N-hydroxy-3-(1-(phenylsulfonyl)-indolin-5-yl)-acrylamide and an anti-PD-1 or anti-PD-L1 antibody.

[0031] As used herein, the term "effective amount" refers to an amount of a component sufficient to produce the desired therapeutic result. For therapeutic purposes, an effective amount may also refer to an amount in which any toxic or deleterious effects of a component are outweighed by the therapeutically beneficial effects. The specific effective or sufficient amount will vary depending on a variety of factors, such as the specific condition being treated, the patient's physical condition (e.g., the patient's weight, age, or sex), the species of mammal or animal being treated, the duration of treatment, the nature of concurrent therapy (if any), and the specific formulation and structure of the compound or its derivative being used. The effective amount can be expressed, for example, as the total mass of active ingredient (e.g., in grams, milligrams, or micrograms), or as the ratio of the mass of active ingredient to body weight (e.g., in milligrams per kilogram (mg / kg)). Alternatively, the effective amount can be expressed as the concentration of the active ingredient (e.g., the drug ABT-301 of the present disclosure), such as molar concentration, mass concentration, volume concentration, molality, mole fraction, mass fraction, and mixing ratio. Specifically, the term "therapeutically effective amount" in relation to the drugs described herein refers to an amount of the drug sufficient to alleviate or improve colorectal cancer-related symptoms in the individual. Those skilled in the art can calculate the human equivalent dose (HED) of the drug (e.g., the drug ABT-301 of the present disclosure) based on the dose determined in the animal model. For example, the HED can be the maximum safe dose for human subjects estimated in accordance with the industry guideline "Estimating the Maximum Safe Starting Dose in Initial Clinical Trials for Therapeutics in Adult Healthy Volunteers" issued by the U.S. Food and Drug Administration (FDA).

[0032] The term "subject" or "patient" refers to an animal, including human species, that can be treated using the pharmaceutical kits and / or methods of the present disclosure. Unless a gender is explicitly indicated, the term "subject" or "patient" is intended to refer to both males and females. Accordingly, the term "subject" or "patient" includes any mammal that may benefit from treatment for colorectal cancer. Exemplary "subjects" or "patients" include, but are not limited to, humans, rats, mice, guinea pigs, monkeys, pigs, goats, cows, horses, dogs, cats, birds, and poultry. In an exemplary embodiment, the subject is a human.

[0033] As described herein, tumor response was assessed using the Response Evaluation Criteria in Solid Tumors (RECIST). A complete response (CR) was defined as the complete disappearance of measurable or evaluable tumor lesions; a partial response (PR) was defined as a decrease in the size of a measurable tumor lesion by at least 30% (≧30%) relative to the initial tumor size; no change (SD) was defined as a decrease in the size of a measurable tumor lesion by less than 30% (<30%) relative to the initial tumor size or an increase in the size of the measurable tumor lesion by less than 20% (<20%) relative to the initial tumor size; and progressive disease (PD) was defined as an increase in tumor volume by more than 30% (>30%) relative to the initial tumor size or the appearance of new lesions.

[0034] II. Detailed Description of the Invention

[0035] The present disclosure is based, at least in part, on the discovery that the compound (E)-N-hydroxy-3-(1-(phenylsulfonyl)-indolin-5-yl)-acrylamide (ABT-301) has therapeutic efficacy against colorectal cancer. In addition, it is noteworthy that ABT-301 enhances the therapeutic effect of immunotherapy on colorectal cancer, not only for the treatment of MSI-H subtype colorectal cancer, but also for the treatment of challenging refractory MSS subtype colorectal cancer. As mentioned above, ABT-301, when used in combination with immunotherapy, particularly when used with immune checkpoint inhibitors (e.g., anti-PD-1, anti-PD-L1 antibodies, etc.), exhibits a synergistic effect in the treatment of colorectal cancer.

[0036] 1. Pharmaceutical kits

[0037] Accordingly, a first aspect of the present disclosure relates to a pharmaceutical kit for treating colorectal cancer in an individual in need thereof. According to an embodiment of the present disclosure, the pharmaceutical kit comprises: a first container comprising (E)-N-hydroxy-3-(1-(phenylsulfonyl)-indolin-5-yl)-acrylamide; and A second container comprises an anti-PD-1 or anti-PD-L1 antibody.

[0038] Fundamentally, the disclosed drug, ABT-301, exhibits selective inhibition of class I and class IIb histone deacetylases (HDACs), including HDAC1, HDAC2, and HDAC3, which belong to class I HDACs, and HDAC6 and HDAC10, which belong to class IIb HDACs. In the treatment of colorectal cancer, ABT-301 not only imparts apoptosis to tumor cells but also enhances the therapeutic effects of immunotherapy, particularly for the refractory MSS subtype of colorectal cancer.

[0039] Generally speaking, the anti-PD-1 antibody suitable for the pharmaceutical kit of the present disclosure can be cemiplizumab, pembrolizumab, nivolumab, or RMP1-14; preferably, the anti-PD-1 antibody suitable for the pharmaceutical kit of the present disclosure is RMP1-14. Alternatively, the pharmaceutical kit of the present disclosure can also include an anti-PD-L1 antibody, which can be any one of atezolizumab, avelumab, or durvalumab; preferably, the anti-PD-L1 antibody suitable for the pharmaceutical kit of the present disclosure is avelumab.

[0040] Without being bound by theory, the pharmaceutical kits of the present disclosure also encompass the option of using other immune checkpoint inhibitors; these immune checkpoint inhibitors may be anti-CTLA4 antibodies, such as ipilimumab, tremelimumab, zalifrelimab, and the like.

[0041] Optionally or additionally, the pharmaceutical kit of the present disclosure may further include other agents for immunotherapy. The immunotherapy agent may be selected from CC motif chemokine ligand 3 (CCL3), CC motif chemokine ligand 26 (CCL26), CXC motif chemokine ligand 7 (CXC motif chemokine ligand 7, CXCL7); granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), interferon-α (IFN-α), interferon-β (IFN-β), interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α); interleukin-4 (IL-4), interleukin-5 (IL-5), interleukin-7 (IL-7), interleukin-10 (IL-10), interleukin-12 (IL-12), interleukin-13 (IL-13), interleukin-15 (IL-15); apremilast st), imiquimod, lenalidomide, pomalidomide, sipuleucel-T, thalidomide; a group consisting of anti-CD2 antibody, anti-CD3 antibody, anti-CD4 antibody, anti-CD11a antibody, anti-CD20 antibody, anti-CD25 antibody, anti-CD52 antibody, anti-EGFR antibody, anti-HER2 antibody, anti-PCDP1 antibody, anti-SLAMF7 antibody, and anti-Trop-2 antibody.

[0042] For the purpose of treating colorectal cancer, the pharmaceutical kit of the present disclosure may further include a chemotherapeutic agent or a targeted therapy agent. Exemplary chemotherapeutic agents include, but are not limited to, actinomycin D, aminoglutethimide, amsacrin, anastrozol, anthracycline, bexaroten, bleomycin, buselerin, busulfan, camptothecin derivatives,derivates), capecitabine, carboplatin, carmustine, chlorambucil, cisplatin, cladribin, cyclophosphamide, cytarabin, cytosinarabinoside, dacarbacin, dactinomycin, daunorubicin, docetaxel axel, doxorubicin, epirubicin, estramustine, etoposide, exemestane, fludarabin, fluorouracil, formestan, gemcitabin, goselerin, hycamtin, idarubicin, ifosfamid, imatinib, irinotecan rinotecan), letrozol, leuprorelin, lomustin, melphalan, mercaptopurine, methotrexate, miltefosin, mitomycine, mitoxantron, nimustine, oxaliplatin, paclitaxel, pentostatin, procarbazine acin, temozolomid, teniposid, testolacton, thiotepa, thioguanine, topotecan, treosulfan, tretinoin, triptorelin, trofosfamide, vinblastine, vincristine, vindesine, and vinorelbine.

[0043] According to an embodiment of the present disclosure, the targeted therapeutic agent can be bortezomib, dasatinib, erlotinib, gefitinib, lapatinib, nilotinib, sorafenib, sunitinib, tofacitinib, crizotinib, venetoclax, obatoclax, navitoclax, gossypol, olaparib, rucaparib, niraparib, talazoparib, ), perifosine, apatinib, vemurafenib, dabrafenib, trametinib, vismodegib, sonidegib, salinomycin, vintafolide, temsirolimus, everolimus, rituximab, trastuzumab, alemtuzumab, cetuximab, panitumumab, bevacizumab, trichostatin A A), phenylbutyrate, sodium butyrate, valproic acid, suberoylanilide hydroxamic acid, belinostat, citarinostat, entinostat, mocetinostat, nanatinostat, panobinostat, pracinostat, romidepsin, tucidinostat, and vorinostat.

[0044] The container suitable for containing the agent can be made of a variety of different materials, such as glass or plastic. The first container can contain the drug ABT-301 of the present disclosure or a pharmaceutical formulation thereof, in a dosage sufficient to treat colorectal cancer. The second container can contain an anti-PD-1 or anti-PD-L1 antibody or a pharmaceutical formulation thereof, in a dosage sufficient to treat colorectal cancer. Alternatively or in addition, the pharmaceutical kit can further include a third container containing a pharmaceutically acceptable excipient (e.g., a pharmaceutically acceptable buffer), such as phosphate-buffered saline (PBS), Ringer's solution, or dextrose solution. Other commercially or user-required materials may also be included, including other buffers, diluents, filters, needles, and syringes. The pharmaceutical kit can further include a label or package insert along with the container. The label or package insert is typically written instructions on the label or package insert (e.g., a piece of paper included in the pharmaceutical kit), but can also be machine-readable instructions (e.g., instructions recorded on a magnetic or optical storage disk). The label or leaflet indicates the use of ABT-301 and the anti-PD-1 or anti-PD-L1 antibody contained in the first and second containers, respectively, for treating colorectal cancer. Similarly, the label or leaflet may include instructions for administering ABT-301 and the anti-PD-1 or anti-PD-L1 antibody. The pharmaceutical kit of the present disclosure is provided in suitable packaging. Suitable packaging includes, but is not limited to, vials, bottles, jars, flexible packaging (e.g., sealed Mylar or plastic bags), boxes, etc.

[0045] 2. Methods for treating colorectal cancer

[0046] In another aspect of the present disclosure, the present invention relates to a method of using the pharmaceutical kit of the present disclosure to treat a subject in need thereof (e.g., a subject suffering from or suspected of suffering from colorectal cancer). The method comprises the steps of administering the pharmaceutical kit of the present disclosure to the subject, wherein The (E)-N-hydroxy-3-(1-(phenylsulfonyl)-indolin-5-yl)-acrylamide is administered to the subject in an amount of 10 μg / kg to 1 g / kg; and The anti-PD-1 or anti-PD-L1 antibody is administered to the subject in an amount of 1 μg / kg to 100 mg / kg.

[0047] Basically, the disclosed drug ABT-301 is used as an HDAC inhibitor to induce apoptosis in colorectal cancer cells while enhancing the therapeutic efficacy of immunotherapy. According to certain embodiments, the individual is a mouse, wherein the dosage of ABT-301 is 0.1 micrograms to 1 kilogram per kilogram of body weight per day (i.e., 0.1 micrograms-1 kilogram / kilogram of body weight / day). Preferably, the dosage of the drug is 1 microgram-100 grams / kilogram of body weight / day. More preferably, the dosage of the drug is 10 micrograms-10 grams / kilogram of body weight / day. Even more preferably, the dosage of the drug is 0.1 milligram-10 grams / kilogram of body weight / day. Even more preferably, the dosage of the drug is 0.1 milligram-10 grams / kilogram of body weight / day. Even more preferably, the dosage of the drug is 0.1 milligram-1 gram / kilogram of body weight / day. Even more preferably, the dosage of the drug is 1 milligram-1 gram / kilogram of body weight / day. According to certain working examples of the present disclosure, administration of 25-100 mg / kg body weight / day (e.g., 25, 50, or 100 mg / kg body weight / day) of ABT-301 is sufficient to induce apoptosis in colorectal cancer cells and enhance the therapeutic efficacy of immunotherapy.

[0048] Those skilled in the art can calculate the human equivalent dose (HED) of the drug ABT-301 of the present disclosure based on the dose determined in the animal model. Accordingly, the dosage of the drug ABT-301 for human body is 10 ng to 100 g per kilogram of body weight per day (e.g., 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 310, 320, 330, 340, 350, 360, 370, 380, 390, 400, 410, 420, 430, 440, 450, 460, 470, 480, 490, 500, 510, 520, 530, 540, 550, 560, 570, 580, 590, 600, 610, 620, 630, 640, 650, 660, 670, 680, 690, 700, 710, 720, 730, 740, 750, 760, 770, 780, 790, 800, 810, 820, 830, 840, 850, 860, 870, 880, 890, 900, 910, 920, 930, 940, 950, 960, 970, 980, or 990 90 ng / kg bw / day; or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 310, 320, 330, 340, 350, 360, 370, 380, 390, 400, 410, 420, 430, 440, 450, 460, 470, 480, 490, 500, 510, 520, 530, 540, 550, 560, 570, 580, 590, 600, 610, 620, 630, 640, 650, 660, 670, 680, 690, 700, 710, 720, 730, 740, 750, 760, 770, 780, 790, 800, 810, 820, 830, 840, 850, 860, 870, 880, 890, 900, 910, 920, 930, 940, 950, 960, 970, 980, or 990 micrograms per kilogram of body weight per day;or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 310, 320, 330, 340, 350, 360, 370, 380, 390, 400, 410, 420, 430, 440, 450, 460, 470, 480, 490, 500, 510, 520, 530, 540, 550, 560, 570, 70, 80, 90, or 100 g / kg body weight per day). Preferably, the dosage of the drug ABT-301 administered to a human body is 100 ng to 10 g / kg body weight / day. More preferably, the dosage of the drug ABT-301 administered to a human body is 1 μg to 1 g / kg body weight / day. Even more preferably, the dosage of the drug ABT-301 administered to a human body is 10 μg to 1 g / kg body weight / day. Even more preferably, the dosage of the drug ABT-301 administered to a human body is 10 μg to 100 mg / kg body weight / day. Even more preferably, the dosage of the drug ABT-301 administered to a human body is 0.1 mg to 100 mg / kg body weight / day. According to certain operating embodiments of the present disclosure, the dosage of the drug ABT-301 administered to a human body is 2.5 to 10 mg / kg body weight / day (e.g., 2.5, 5, or 10 mg / kg body weight / day).

[0049] Depending on the desired purpose, the drug ABT-301 can be administered by any appropriate route, for example, enteral, oral, nasal, parenteral (e.g., intratumoral, intramuscular, intravenous, intraarterial, subcutaneous, intraperitoneal, intracerebral, intraventricular, or intraspinal injection), or transmucosal administration. According to one embodiment of the present disclosure, the drug ABT-301 is administered orally.

[0050] To induce apoptosis in colorectal cancer cells and / or enhance the effectiveness of immunotherapy, the drug ABT-301 may be administered to the subject one or more times. For example, the drug ABT-301 may be administered once to complete the entire course of treatment. Alternatively, the drug ABT-301 may be administered to the subject daily for at least 7 days; for example, for 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, or more days. In certain embodiments, when multiple doses are administered to an individual, the frequency of multiple doses administered to an individual is three doses per day, two doses per day, one dose per day, one dose every other day, one dose every three days, one dose per week, one dose every other Monday, one dose per month, one dose every other month, one dose per quarter, one dose every six months, or one dose per year. In certain embodiments, the frequency of multiple doses administered to an individual is one dose per day. In certain embodiments, the frequency of multiple doses administered to an individual is two doses per week. In certain embodiments, when multiple doses are administered to an individual, the duration between the first dose and the last dose of the multiple dose is one day, two days, four days, one week, two weeks, three weeks, one month, two months, three months, four months, six months, nine months, one year, two years, three years, four years, five years, seven years, ten years, fifteen years, twenty years, or the lifetime of the individual. In certain embodiments, the duration between the first dose and the last dose of the multiple dose is about 1-1.5 months.

[0051] According to the embodiments disclosed herein, immunotherapies that may benefit from co-administration with the drug ABT-301 include anti-PD-1 or anti-PD-L1 antibodies as described above. According to certain embodiments, the individual is a mouse, and the anti-PD-1 or anti-PD-L1 antibody is administered at a dose of 1 ng to 1 kg per kilogram of body weight per day (i.e., 1 ng to 1 kg per kilogram of body weight per day). Preferably, the anti-PD-1 or anti-PD-L1 antibody is administered at a dose of 10 ng to 100 g per kilogram of body weight per day. More preferably, the anti-PD-1 or anti-PD-L1 antibody is administered at a dose of 100 ng to 10 g per kilogram of body weight per day. Even more preferably, the anti-PD-1 or anti-PD-L1 antibody is administered at a dose of 1 microgram to 10 g per kilogram of body weight per day. Still more preferably, the dosage of the anti-PD-1 or anti-PD-L1 antibody is 10 μg-1 g / kg body weight / day. Still more preferably, the dosage of the anti-PD-1 or anti-PD-L1 antibody is 100 μg-100 mg / kg body weight / day. According to certain working embodiments of the present disclosure, the dosage of the anti-PD-1 or anti-PD-L1 antibody is 1-10 mg / kg body weight / day (e.g., 3.3 or 6.7 mg / kg body weight / day), which is sufficient to produce a synergistic effect with the drug ABT-301 in the treatment of colorectal cancer.

[0052] When the subject is a human, the anti-PD-1 or anti-PD-L1 antibody is administered in an amount of 0.1 ng to 100 g per kg body weight per day (e.g., 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 310, 320, 330, 340、350、360、370、380、390、400、410、420、430、440、450、460、470、480、490、500、510、520、530、540、550、560、570、580、590、600、610、620、630、640、650、660、670、680、690、700、710、720、730、740、750、760、770、780、790、800、810、820、830、840、850、860、870、880、890、900、91 0, 920, 930, 940, 950, 960, 970, 980, or 990 ng / kg bw / day; or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 310, 320, 330, 340, 350, 360, 370, 380, 390, 400, 410, 420, 430, 440, 450, 460, 470, 480, 490, 500, 510, 520, 530, 540, 550, 560, 570, 580, 590, 600, 610, 620, 630, 640, 650, 660, 670, 680, 690, 700, 710, 720, 730, 740, 750, 760, 770, 780, 790, 800, 810, 820, 830, 840, 850, 860, 870, 880, 890, 900, 910, 920, 930, 940, 950, 960, 970, 980, or 990 micrograms per kilogram of body weight per day;or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 310, 320, 330, 340, 350, 360, 370, 380, 390, 400, 410, 420, 430, 440, 450, 460, 470, 480, 490, 500, 510, 520, 530, 540, 550, 560, 570, 70, 80, 90, or 100 g / kg body weight per day). Preferably, the anti-PD-1 or anti-PD-L1 antibody is administered to a human body at a dose of 1 ng to 10 g / kg body weight / day. More preferably, the anti-PD-1 or anti-PD-L1 antibody is administered to a human body at a dose of 10 ng to 1 g / kg body weight / day. Even more preferably, the anti-PD-1 or anti-PD-L1 antibody is administered to a human body at a dose of 100 ng to 1 g / kg body weight / day. Even more preferably, the anti-PD-1 or anti-PD-L1 antibody is administered to a human body at a dose of 1 μg to 100 mg / kg body weight / day. Even more preferably, the anti-PD-1 or anti-PD-L1 antibody is administered to a human body at a dose of 10 μg to 10 mg / kg body weight / day. According to certain working embodiments of the present disclosure, the anti-PD-1 or anti-PD-L1 antibody is administered to a human at a dose of 100 μg to 1 mg / kg body weight / day (e.g., 330 or 670 μg / kg body weight / day).

[0053] Alternatively, the actual dosage of the presently disclosed drug ABT-301 and anti-PD-1 or anti-PD-L1 antibody may be determined by the attending physician based on the individual's physical and physiological factors, including, but not limited to, age, sex, weight, body surface area, disease to be treated, severity of disease, past medical history, use of other medications, route of administration, etc.

[0054] Non-limiting routes of administration include, but are not limited to, enteral, oral, nasal, parenteral, topical, or transmucosal administration, wherein parenteral administration can be any of the following, including intratumoral, intramuscular, intravenous, intraarterial, subcutaneous, intraperitoneal, intracerebral, intracerebroventricular, or intraspinal injection. In one embodiment, the anti-PD-1 or anti-PD-L1 antibody is administered to the subject via the intraperitoneal route.

[0055] It will be appreciated that, under the guidance of a healthcare professional, the subject may be administered ABT-301, an agent of the present disclosure, before, simultaneously with, or after administration of an anti-PD-1 or anti-PD-L1 antibody. For the purpose of treating colorectal cancer, the anti-PD-1 or anti-PD-L1 antibody may be administered to the subject one or more times. For example, the anti-PD-1 or anti-PD-L1 antibody may be administered once to complete the entire course of treatment. Alternatively, the anti-PD-1 or anti-PD-L1 antibody can be administered to the subject daily for at least 7 days; for example, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, or more days. In certain embodiments, when multiple doses are administered to an individual, the frequency of multiple doses administered to an individual is three doses per day, two doses per day, one dose per day, one dose every other day, one dose every three days, one dose per week, one dose every other Monday, one dose per month, one dose every other month, one dose per quarter, one dose every six months, or one dose per year. In certain embodiments, the frequency of multiple doses administered to an individual is one dose per day. In certain embodiments, the frequency of multiple doses administered to an individual is two doses per week. In certain embodiments, when multiple doses are administered to an individual, the duration between the first dose and the last dose of the multiple dose is one day, two days, four days, one week, two weeks, three weeks, one month, two months, three months, four months, six months, nine months, one year, two years, three years, four years, five years, seven years, ten years, fifteen years, twenty years, or the lifetime of the individual. In certain embodiments, the duration between the first dose and the last dose of the multiple dose is about 1-1.5 months.

[0056] The colorectal cancer that can be treated by the pharmaceutical kit and / or method of the present disclosure is an MSS type or MSI type colorectal cancer. In addition, the colorectal cancer can be in situ colorectal cancer or metastatic colorectal cancer. Optionally, the colorectal cancer may be resistant to chemotherapy (e.g., 5-fluorouracil (5-fluorouracil, 5-FU)), radiotherapy (e.g., ultraviolet (UV) irradiation), or immunotherapy (e.g., adaptive immune cell therapy (AIT)). Accordingly, the pharmaceutical kit and / or method of the present disclosure provides a possible way to treat colorectal cancer patients who are resistant to cancer treatment.

[0057] Optionally or in addition, the method of the present disclosure further comprises administering an additional anti-cancer therapy to the individual before, simultaneously with, or after administering the pharmaceutical kit of the present disclosure to the individual, wherein the additional anti-cancer therapy is at least one of surgery, radiotherapy, hyperthermia, immunotherapy, chemotherapy, or targeted therapy. Exemplary drugs used in immunotherapy, chemotherapy, or targeted therapy are described above; for the sake of brevity, they are not further described here.

[0058] Alternatively, ABT-301, the drug of the present disclosure, can be administered individually to patients with colorectal cancer without concomitant treatment (e.g., co-treatment or pre-treatment) with an anti-PD-1 or anti-PD-L1 antibody, and it is still possible to achieve a measurable therapeutic effect in eliminating colorectal cancer; and vice versa.

[0059] Essentially, the subject suitable for treatment by the pharmaceutical kits and / or methods of the present disclosure is a mammal; preferably, the subject is a human.

[0060] The following examples illustrate certain aspects of the present invention to facilitate practice of the present invention by those skilled in the art. These examples should not be construed as limiting the scope of the present invention. It is believed that those skilled in the art, after reading the description provided herein, will be able to fully utilize and practice the present invention without undue interpretation. All publications cited herein are incorporated herein in their entirety as a part of this specification. Example

[0061] Materials and Methods

[0062] 1. HADC enzyme activity detection

[0063] To assess the HDAC enzyme activity of ABT-301, a biochemical HDAC enzyme activity assay service platform provided by Reaction Biology Corp. (Malvern, PA, USA) was used to perform HDAC enzyme activity assays. The HDAC enzyme activity assay was performed using a synthetic HDAC substrate, the structure of which is shown in Formula (I). The moiety used for signal generation is attached to the carboxyl group of acetyllysine (K(Ac)), which is the target of HDAC deacetylation. After HDAC deacetylation, the amount of the fluorescent signal moiety 7-amino-4-methyl coumarin (AMC) generated can be quantified as an indicator of HDAC enzyme activity.

[0064] The fluorogenic peptides used to measure HDAC enzyme activity are listed in Table 1 below.

[0065] Table 1. Fluorescent substrate peptides for HDAC enzyme activity detection

[0066] Histone deacetylase activity is detected by the release of AMC fluorescence from the aforementioned fluorescent substrate peptide under the catalysis of the deacetylase. The half-maximal inhibitory concentration (IC50) is the drug concentration that causes a 50% decrease in HDAC activity observed in the control wells during the compound reaction in a 10-dose IC50 format. Control group: No inhibitor control group (DMSO vehicle only), and for each target, a control group compound that is specific for a specific target was tested in a 10-dose IC50 format. HDAC reference compounds: (1) trichostatin A (TSA), a control group compound that is specific for HDAC1, 2, 3, 6, 8, and 11 targets; (2) TMP269, a control group compound that is specific for HDAC4, 5, 7, and 9 targets; (3) quisinostat, a control group compound that is specific for HDAC10 targets.

[0067] 2. Cell Culture

[0068] CT26.WT cells (ATCC accession number: CRL-2638 TM) are mouse colorectal cancer cells, generally classified as microsatellite stable, and cultured in RPMI medium supplemented with 10% fetal bovine serum, 100 units / ml penicillin, 100 μg / ml streptomycin, 10 mM HEPES, and 1 mM sodium pyruvate. MC-38 cells (purchased from Kerafast, Inc.) are derived from C57BL / 6 mouse colorectal adenocarcinoma cells, generally classified as microsatellite unstable, and cultured in DMEM medium supplemented with 10% fetal bovine serum, 100 units / ml penicillin, 100 μg / ml streptomycin, 10 mM HEPES, and 1× non-essential amino acids. All cells were cultured at 37°C in a humidified atmosphere containing 5% CO2.

[0069] 3. Animal Experiments

[0070] Female BALB / c mice (BALB / cByJNarl), aged 5-7 weeks (for the CT26 subcutaneous tumor model); female C57BL / 6 mice (C57BL / 6JNarl), aged 5-7 weeks (for the MC-38 subcutaneous tumor model). These mice were housed in standard cages in the animal facility of the Industrial Technology Research Institute (ITRI). A week prior to the start of the study was used for acclimatization, and the health of the mice was monitored daily. The animals were housed in an animal room with a temperature of 22-26°C, a humidity of 40-70%, positive pressure, 60% air circulation, a ventilation rate of 15-20 times per hour, and a controlled light-dark cycle (12-12 hours).

[0071] 4.CT26 subcutaneous tumor model

[0072] CT26 cells (2×10 5 Tumors were suspended in 100 μl of PBS and then subcutaneously inoculated into the right flank of female BALB / c mice. Tumors were measured using a caliper and the tumor size was calculated as follows: tumor volume (V) = (L × S 2) / 2(L, longest diameter (mm); S, shortest diameter (mm)). Tumor size and mouse body weight were monitored and recorded two to three times a week. The antitumor activity of a specific treatment was expressed as percentages of tumor growth inhibition (%TGI), which was calculated as follows: [1-(final tumor volume of the treatment group-initial tumor volume) / (final tumor volume of the control group-initial tumor volume)]×100. The body weight of each mouse was also compared with the first day of treatment (i.e., day 0) and expressed as a percentage of the value obtained on day 0. All animal experimental procedures were performed in accordance with the Guide for the Care and Use of Laboratory Animals (NRC, USA) and were approved by the Institutional Animal Care and Use Committee (IACUC) of ITRI (IACUC approval numbers: ITRI-IACUC-2022-031, ITRI-IACUC-2023-031, and ITRI-IACUC-2023-010).

[0073] 4-1. ABT-301 single dose, with or without avelumab

[0074] Treatment was started five days after inoculation of CT26 cells, with the treatment start day designated as day 0. When the average tumor volume reached 50-100 mm3 (mm 3 ), the mice were divided into four groups according to tumor size in an S-shaped group. The four groups were: (1) vehicle group; (2) avelumab group (100 μg, intraperitoneal injection (IP), twice a week (BIW); (3) ABT-301 group (100 mg / kg, oral (per os, PO), once a day (quaque die, QD)); and (4) ABT-301 (100 mg / kg, PO, QD) + avelumab (100 μg, IP, BIW) group. Eight mice were used in each group. Treatment lasted for 43 days. ABT-301 was formulated in 0.5% carboxymethylcellulose + 0.1% Tween 80 (in 5% glucose). Avelumab was formulated in PBS. The treatment schedule is summarized in Table 2.

[0075] Table 2. Treatment Schedule for ABT-301 Single Dose, with or without Avelumab *Vehicle control: 0.5% carboxymethylcellulose + 0.1% Tween 80 (in 5% glucose).

[0076] 4-2. ABT-301 at multiple doses, with or without avelumab

[0077] Treatment began when the mean tumor volume reached approximately 80-100 cubic millimeters, with the start of treatment designated day 0. Mice were divided into eight groups, including: (1) vehicle group; (2) avelumab group (200 μg, IP, BIW); (3) ABT-301 group (25 mg / kg, PO, QD); (4) ABT-301 (25 mg / kg, PO, QD) + avelumab (200 μg, IP, BIW) group; (5) ABT-301 group (50 mg / kg, PO, QD); (6) ABT-301 (50 mg / kg, PO, QD) + avelumab (200 μg, IP, BIW) group; (7) ABT-301 group (100 mg / kg, PO, QD); and (8) ABT-301 (100 mg / kg, PO, QD) + avelumab (200 μg, IP, BIW) group. Treatment was continued for a specific time (24 days) or until humane endpoints were reached. Ten mice were used per group. ABT-301 was formulated in 0.5% carboxymethylcellulose + 0.1% Tween 80 (in 5% glucose). Avelumab was formulated in PBS. The treatment schedule is summarized in Table 3.

[0078] Table 3. Treatment Plans for Various Doses of ABT-301 Combined with Avelumab *Vehicle control: 0.5% carboxymethylcellulose + 0.1% Tween 80 (in 5% glucose). #Avelumab was administered on days 0, 3, 7, and 10.

[0079] 4-3. ABT-301 at various doses, with or without RMP1-14

[0080] Treatment began when the mean tumor volume reached approximately 80-100 cubic millimeters, with the start of treatment designated day 0. The mice were divided into eight groups, including: (1) vehicle group; (2) RMP1-14 group (200 μg, IP, BIW); (3) ABT-301 group (25 mg / kg, PO, QD); (4) ABT-301 (25 mg / kg, PO, QD) + RMP1-14 (200 μg, IP, BIW) group; (5) ABT-301 group (50 mg / kg, PO, QD); (6) ABT-301 (50 mg / kg, PO, QD) + RMP1-14 (200 μg, IP, BIW) group; (7) ABT-301 group (100 mg / kg, PO, QD); and (8) ABT-301 (100 mg / kg, PO, QD) + RMP1-14 (200 μg, IP, BIW) group. Treatment was continued for a specified time (27 days) or until a humane endpoint was reached. Ten mice were used per group. ABT-301 was formulated in 0.5% carboxymethylcellulose + 0.1% Tween 80 (in 5% glucose). RMP1-14 was formulated in PBS. The treatment schedule is summarized in Table 4.

[0081] Table 4. Treatment plan for various doses of ABT-301 combined with RMP1-14 *Vehicle control: 0.5% carboxymethylcellulose + 0.1% Tween 80 (in 5% glucose).

[0082] 4-4. HDAC inhibitors, with or without RMP1-14

[0083] This study used HDAC inhibitors, including ABT-301, voriconazole, and cetaxel. Treatment began when the mean tumor volume reached approximately 80-100 cubic millimeters, with the start of treatment designated day 0. Mice were divided into eight groups, including: (1) vehicle group; (2) RMP1-14 group (200 μg, IP, BIW); (3) ABT-301 group (100 mg / kg, PO, QD); (4) ABT-301 (100 mg / kg, PO, QD) + RMP1-14 (200 μg, IP, BIW) group; (5) vorinostat group (100 mg / kg, PO, QD); (6) vorinostat (100 mg / kg, PO, QD) + RMP1-14 (200 μg, IP, BIW) group; (7) sitastat group (50 mg / kg, IP, QD); and (8) sitastat (50 mg / kg, IP, QD) + RMP1-14 (200 μg, IP, BIW) group. Treatment was performed for a specific time (31 days) or until a humane endpoint was reached. Eight or nine mice were used per group. ABT-301 and voriconazole were formulated in 0.5% carboxymethylcellulose + 0.1% Tween 80 (in 5% glucose). Sitacotestat was formulated in 10% DMSO, 40% PEG300, 5% Tween-80, and 45% normal saline. RMP1-14 was formulated in PBS. The treatment plan is summarized in Table 5.

[0084] Table 5. Treatment plans for different HDAC inhibitors combined with RMP1-14 *Vehicle control: 0.5% carboxymethylcellulose + 0.1% Tween 80 (in 5% glucose).

[0085] 5. MC-38 subcutaneous tumor model

[0086] MC-38 cells (2×10 5 Tumors were suspended in 100 μl of PBS and then subcutaneously inoculated into the right flank of female C57BL / 6 mice. Tumors were measured using a caliper and the tumor size was calculated as follows: tumor volume (V) = (L × S 2) / 2(L, longest diameter (mm); S, shortest diameter (mm)). Tumor size and mouse weight were monitored and recorded two to three times a week. The antitumor activity of a specific treatment was expressed as tumor growth inhibition rate (%TGI), which was calculated as follows: [1-(final tumor volume of the treatment group-initial tumor volume) / (final tumor volume of the control group-initial tumor volume)]×100. The body weight of each mouse was also compared with the first day of treatment (i.e., day 0) and expressed as a percentage of the value obtained on day 0. All animal experimental procedures were performed in accordance with the Guide for the Care and Use of Laboratory Animals (NRC, USA) and approved by the IACUC of ITRI (IACUC approval number: ITRI-IACUC-2023-003).

[0087] 5-1. ABT-301 single dose, with or without avelumab or RMP1-14

[0088] Treatment was initiated 6 days after inoculation of MC-38 cells, with the treatment start date designated as day 0. When the mean tumor volume reached 50-100 mm3, mice were divided into six groups according to tumor size using an S-shaped grouping. Mice were divided into six groups, including: (1) vehicle group; (2) ABT-301 group (100 mg / kg, PO, QD); (3) avelumab group (200 μg, IP, BIW); (4) ABT-301 (100 mg / kg, PO, QD) + avelumab (200 μg, IP, BIW) group; (5) RMP1-14 group (200 μg, IP, BIW); and (6) ABT-301 (100 mg / kg, PO, QD) + RMP1-14 (200 μg, IP, BIW) group. Treatment was continued for a specific time (29 or 36 days) or until humane endpoints were reached. Eight mice were used per group. ABT-301 was formulated in 0.5% carboxymethylcellulose + 0.1% Tween 80 (in 5% glucose). Avelumab and RMP1-14 were formulated in PBS. The treatment schedule is summarized in Table 6.

[0089] Table 6. Treatment Schedule for ABT-301 Single Dose, with or without Avelumab or RMP1-14 *Vehicle control: 0.5% carboxymethylcellulose + 0.1% Tween 80 (in 5% glucose).

[0090] 6. Statistical Analysis

[0091] Graphs and data were analyzed using Excel and GraphPad Prism. Statistical analysis was performed using GraphPad Prism. Statistical significance was determined using Student's t-test or one-way ANOVA. A P value of less than 0.05 indicated a statistically significant difference.

[0092] Example 1 Confirmation of HDAC Inhibitory Activity of ABT-301

[0093] In this embodiment, the main goal is to understand the overview of the HDAC enzymatic activity displayed by ABT-301. The steps for evaluating HDAC enzymatic activity are as described in the "Materials and Methods" section, and the experimental results are presented in Table 7. Based on the results of the study, ABT-301 clearly demonstrates HDAC enzymatic activity to HDAC1, HDAC2, and HDAC3 (belonging to Class I HDAC), as well as HDAC6 and HDAC10 (belonging to Class IIb HDAC). However, ABT-301 does not show activity to HDAC4, HDAC5, HDAC7, and HDAC9 (belonging to Class IIa HDAC), or HDAC11 (belonging to Class IV HDAC). These experimental results clearly show that ABT-301 is an inhibitor of Class I and Class IIb HDAC.

[0094] Table 7. HDAC enzyme activity (IC 50 (nanomolar concentration, nM)

[0095] In the HDAC enzyme study, in addition to ABT-301, vorinostat and toxistat were also included in the parallel comparative analysis; these two compounds are class I and / or class IIb HDAC inhibitors. Despite this, ABT-301 demonstrated superior inhibitory activity against class I and / or class IIb HDACs overall compared to the other compounds.

[0096] Example 2: Efficacy of ABT-301 in Inhibiting MSS Subtype Colorectal Cancer

[0097] 2.1 Preliminary Evaluation of Combination Therapy with ABT-301 and Avelumab

[0098] In this example, the primary objective was to evaluate the anticancer efficacy of ABT-301 against MSS subtype colorectal cancer, as well as its ability to enhance the efficacy of immunotherapy against MSS subtype colorectal cancer, which accounts for the majority (approximately 90%) of colorectal cancer patients.

[0099] To achieve this goal, a preliminary study was conducted according to the procedures described in the "Materials and Methods" section, "4-1. ABT-301 single dose, with or without avelumab". In this study, animals were injected with avelumab for immunotherapy, and the experimental results were as follows: Figures 1A-1G The tumor growth results of mice bearing CT26 are shown in Figures 1A-1D The focus of the study was on demonstrating the complete remission (CR) efficacy of specific treatments on tumor growth in mice. According to the experimental results, compared to the vehicle group, administering either avelumab or ABT-301 alone to mice failed to significantly increase the number of mice that achieved a CR to their tumors (i.e., the CR rates in the vehicle, avelumab, and ABT-301 groups were 0 / 8, 1 / 8, and 2 / 8, respectively). However, the combined administration of avelumab and ABT-301 significantly increased the CR rate in treated mice (the CR rate in the combined treatment group was 7 / 8).

[0100] Results for the 28-day period and tumor volume on day 28 are presented in Figures 1E-1F As shown in Table 8, ABT-301 and avelumab each exhibited varying degrees of antitumor activity, but combined treatment demonstrated the greatest inhibitory effect on tumor growth. No statistically significant differences in mouse body weight (%) were observed between each treatment group and the vehicle group on day 28 (data not shown).

[0101] Table 8. Tumor size and tumor growth inhibition rate on day 28 after ABT-301 and avelumab treatment in mice bearing CT26 allograft tumors Experimental data are presented as mean ± standard error of the mean (SEM). Data from mice that died or reached humane endpoints were excluded. Vehicle group, n = 6; ABT-301 group, n = 6; avelumab group, n = 4; and ABT-301 + avelumab group, n = 8. *P < 0.05, compared with the avelumab group (by t-test).

[0102] In addition, the combined treatment significantly improved the survival rate of treated mice compared with the vehicle-, avelumab-, and ABT-301-treated mice ( Figure 1G (See Table 9). In summary, while ABT-301 and avelumab exhibited anticancer activity when administered alone, combined treatment with ABT-301 and avelumab significantly enhanced the antitumor growth effect of avelumab against CT26 tumors, as reflected by a reduction in tumor growth and inhibition of tumor volume. Furthermore, combined treatment with ABT-301 and avelumab prolonged the survival of CT26-bearing mice compared to treatment with either ABT-301 or avelumab alone.

[0103] Table 9. Median survival of mice bearing CT26 homograft tumors treated with ABT-301 and avelumab throughout the study period (Days) Vector group Avelumab ABT-301 ABT-301 + Avelumab Median survival 30.5 28 33.5 >86

[0104] 2.2 Further evaluation of the combination of ABT-301 and avelumab

[0105] Based on the knowledge gained from Example 2.1, this study further expanded to explore the ability of ABT-301 to inhibit cancer and the ability of ABT-301 to enhance the treatment of colorectal cancer with avelumab, and further extended to the use of different doses of ABT-301, with or without avelumab. The experimental procedures are described in the "Materials and Methods" and "4-2. Various doses of ABT-301, with or without avelumab", and the experimental results are presented in Figures 2A-2J In. Based on Figures 2A-2H , compared with the vehicle group ( Figure 2A ), administration of avelumab alone could produce a certain degree of anticancer activity, as shown by the complete remission (CR) rate (i.e., 7 out of 10 mice in the avelumab group showed CR; Figure 2B However, increasing the ABT-301 dose from 25 mg / kg to 100 mg / kg resulted in only a slight enhancement of the therapeutic effect (i.e., from PD = 9 and PR = 1 in the ABT-301(25) group to PD = 8 and SD = 2 in the ABT-301(100) group; Figure 2C 、 2E , and 2G). However, the combination of ABT-301 and avelumab significantly enhanced CR to tumors in treated mice, and this was also evident even at the lowest dose of 25 mg / kg of ABT-301 ( Figure 2C contrast Figure 2D , Figure 2E contrast Figure 2F ,as well as Figure 2G contrast Figure 2H In addition, the combined treatment still showed a dose-dependent therapeutic efficacy with increasing ABT-301 dose (i.e., from PD=1 and CR=9 in the ABT-301(25)+avelumab group to CR=10 in the ABT-301(100)+avelumab group; Figure 2D 、 2F , and 2H).

[0106] The results for tumor volume during the 24-day period and on day 21 are presented in Figures 2I-2JAs shown in Table 10, ABT-301 and avelumab each exhibited varying degrees of antitumor activity, but combined treatment further inhibited tumor growth. No statistically significant differences in mouse body weight (%) were observed between each treatment group and the vehicle group on day 24 (data not shown).

[0107] Table 10. Tumor size on day 24 after ABT-301 and Avelumab treatment in mice bearing CT26 homograft tumors. Experimental data are presented as mean ± standard error of the mean (SEM). Data from experiments in mice that reached humane endpoints were excluded. n = 8 for the vehicle and ABT-301 (25 mg / kg) groups; n = 10 for other groups. *P < 0.05, **P < 0.01, compared to the vehicle group (by t-test).

[0108] In summary, ABT-301 and avelumab jointly slowed the growth of CT26 tumors. Combination therapy with ABT-301 and avelumab significantly enhanced the anticancer effect against CT26 tumors, with ABT-301 strongly enhancing the anti-tumor growth efficacy of avelumab against CT26 tumors. This was demonstrated by increased CR rates and inhibition of tumor volume, with efficacy observed even at the lowest ABT-301 dose of 25 mg / kg. These findings suggest a synergistic therapeutic effect, with a reduced ABT-301 dose required to achieve potent anticancer effects.

[0109] 2.3 Evaluation of Combination Therapy with ABT-301 and RMP1-14

[0110] In this example, the study exploring the potential of ABT-301 in enhancing immunotherapy for the treatment of colorectal cancer was expanded to include the use of different doses of ABT-301 in combination with another immune checkpoint inhibitor, specifically the anti-PD-1 antibody RMP1-14, as an example. The experimental procedures are described in the "Materials and Methods" and "4-3. Various doses of ABT-301, with or without RMP1-14" sections, and the experimental results are presented in Figures 3A-3J In. Based on Figures 3A-3H , similar to Figures 2A-2H The results in the vehicle group were compared with those in the control group ( Figure 3A ), administration of RMP1-14 alone could produce a certain degree of anticancer activity, as shown by the complete remission (CR) rate (i.e., 4 out of 10 mice in the RMP1-14 group showed CR; Figure 3BHowever, increasing the ABT-301 dose from 25 mg / kg to 100 mg / kg resulted in only a slight enhancement of the therapeutic effect (i.e., from PD=8, SD=1, PR=1 in the ABT-301(25) group to PD=7, PR=2, CR=2 in the ABT-301(100) group; Figure 3C 、 3E , and 3G). However, combined treatment with ABT-301 and RMP1-14 significantly enhanced CR to tumors in treated mice, and this was also evident even at the lowest dose of 25 mg / kg of ABT-301 ( Figure 3C contrast Figure 3D , Figure 3E contrast Figure 3F ,as well as Figure 3G contrast Figure 3H In addition, the combined treatment still showed a dose-dependent therapeutic efficacy with increasing ABT-301 dose (i.e., from PD=2, PR=1, and CR=7 in the ABT-301(25)+RMP1-14 group to CR=10 in the ABT-301(100)+RMP1-14 group; Figure 3D 、 3F , and 3H).

[0111] After 27 days of monitoring and the tumor volume results on day 17, see Figures 3I-3J , and are summarized in Table 11. ABT-301 and RMP1-14 individually exhibited varying degrees of antitumor activity. However, combined treatment demonstrated significant inhibition of tumor growth. Furthermore, no statistically significant differences in mouse body weight (%) were observed between each treatment group and the vehicle group on day 27 (data not shown).

[0112] Table 11. Tumor size on day 27 after ABT-301 and RMP1-14 treatment of mice bearing CT26 homograft tumors. Experimental data are presented as mean ± standard error of the mean (SEM). Data from mice that died or reached humane endpoints were excluded. Vehicle group, n = 7; other groups, n = 10. *P < 0.05, **P < 0.01, ***P < 0.001, compared with the vehicle group (one-way ANOVA followed by Tukey's multiple comparisons test (t-test)).

[0113] In summary, ABT-301 and RMP1-14 jointly slowed the growth of CT26 tumors. The combined treatment of ABT-301 and RMP1-14 significantly enhanced the anti-cancer effect on CT26 tumors, and ABT-301 continued to strongly enhance the anti-tumor growth efficacy of RMP1-14 on CT26 tumors. This can be confirmed by increasing the CR rate of tumor growth and inhibiting tumor volume, and it is effective even at the lowest ABT-301 dose of 25 mg / kg. These findings indicate that a synergistic therapeutic effect is produced when ABT-301 is used in combination with RMP1-14, that is, a potent anti-cancer effect can be achieved using a smaller amount of ABT-301.

[0114] Example 3: Efficacy of ABT-301 in inhibiting MSI-H subtype colorectal cancer

[0115] 3.1 Evaluation of Combination Therapy with ABT-301 and Avelumab or RMP1-14

[0116] The following experiments were conducted to investigate the efficacy of ABT-301 in treating MSI-H subtype colorectal cancer and its effectiveness in enhancing the effects of immunotherapy against MSI-H subtype colorectal cancer.

[0117] The study was conducted according to the procedures described in the section "Materials and Methods", "5-1. ABT-301 single dose, with or without avelumab or RMP1-14". Immunotherapy was performed by administering avelumab or RMP1-14 to mice, and the results are presented in Figures 4A-4I The results of tumor growth in mice with MC-38 are presented in Figures 4A-4F The focus of the study was on demonstrating the efficacy of specific treatments in producing complete remission (CR) of tumor growth in treated mice. The results showed that administering ABT-301 alone did not significantly increase the number of mice that achieved complete remission (CR) compared to the vehicle group (i.e., the CR rates for both the vehicle and ABT-301 groups were 0 / 8; Figures 4A-4B ). According to the literature, immune checkpoint inhibitors show higher response rates and survival rates in the treatment of MSI-H subtype colorectal cancer compared with MSS subtype tumors, resulting in higher survival rates. Similarly, it was observed that the administration of either avelumab or RMP1-14 alone achieved a certain degree of anti-cancer effect (i.e., the CR rates in the avelumab group and the RMP1-14 group were 4 / 8 and 5 / 8, respectively; Figure 4C and 4EHowever, in this case, when used in combination with avelumab or RMP1-14, ABT-301 was still able to further enhance the efficacy of avelumab or RMP1-14 in treating MSI-H subtype colorectal cancer; the combined administration of ABT-301 with avelumab or RMP1-14 significantly increased the CR rate of treated mice (i.e., the CR rates of the ABT-301+avelumab group and the ABT-301+RMP1-14 group were 6 / 8 and 8 / 8, respectively; Figure 4D and 4F ).

[0118] After 25 days of recording and the tumor volume results on day 25, see Figures 4G-4H , and are summarized in Table 12. ABT-301, avelumab, and RMP1-14 individually exhibited varying degrees of antitumor activity. However, combined treatment demonstrated a more potent inhibitory effect on tumor growth. Due to reaching a humane endpoint, mice in the vehicle group were sacrificed on day 20. In addition, no statistically significant differences in the body weight (%) of mice in each treatment group and the vehicle group were observed on day 25 (data not shown).

[0119] Table 12. Tumor size on day 25 after treatment with ABT-301 and avelumab or RMP1-14 in mice bearing MC-38 homograft tumors. Experimental data are presented as mean ± standard error of the mean (SEM). Data from mice that died or reached humane endpoints were excluded. Vehicle group, n = 1; ABT-301 group, n = 8; avelumab group, n = 8; ABT-301 + avelumab group, n = 8; RMP1-14 group, n = 7; ABT-301 + RMP1-14 group, n = 8.

[0120] In addition, although mice treated with avelumab or RMP1-14 showed a significantly higher survival rate than mice in the vehicle group, combined treatment (either ABT-301 + avelumab or ABT-301 + RMP1-14) significantly improved the survival rate of treated mice ( Figure 4I and Table 13). In summary, although avelumab or RMP1-14 can exhibit anticancer activity when used alone, the combination of ABT-301 and avelumab or RMP1-14 can more strongly enhance the anti-tumor growth effect of avelumab or RMP1-14 on MC-38 tumors, as demonstrated by CR of tumor growth and inhibition of tumor volume. In addition, compared to treatment with ABT-301, avelumab, or RMP1-14 alone, the combined treatment of ABT-301 and avelumab or avelumab can prolong the survival of MC-38-bearing mice.

[0121] Table 13. Median survival of mice bearing MC-38 homograft tumors treated with ABT-301 and avelumab or RMP1-14 over the entire study period.

[0122] Example 4 Effect of Combination of HDAC Inhibitor and RMP1-14 on the Growth of Colorectal Cancer

[0123] This embodiment aims to evaluate the efficacy of combined treatment of different HDAC inhibitors with RMP1-14 for colorectal cancer. To this end, this study co-administered different HDAC inhibitors, including ABT-301 (full-spectrum HDAC (pan-HDAC) inhibitor), voriconstat (full-spectrum HDAC inhibitor), and cetaxel (specific HDAC inhibitor for HDAC6) with RMP1-14 to the test animals for combined treatment. The experimental steps are as described in the section "4-4. HDAC inhibitors, with or without RMP1-14" of this specification, and the experimental results are presented in Figures 5A-5K The results of tumor growth in mice bearing CT26 are presented in Figures 5A-5H Compared with the vehicle group, the administration of RMP1-14 alone or the use of HDAC inhibitors (i.e., ABT-301, voriconazole, or cetaxel) alone did not induce a significant response in tumor growth in mice ( Figures 5A-5C , 5E, and 5G). However, co-administration of HDAC inhibitors with RMP1-14 induced varying degrees of anti-cancer responses in mice, particularly inhibition of colorectal cancer volume. The results showed that co-administration of ABT-301 and RMP1-14 exhibited a better tumor suppression response than co-administration of vorisstat and RMP1-14, or co-administration of cetaxel and RMP1-14. Figure 5D 、 5F , and 5H).

[0124] In another batch of experiments, it was also found that compared with the vehicle group, the administration of RMP1-14 or other HDAC inhibitors (i.e., ABT-301, PCI-34051 (a specific HDAC inhibitor targeting HDAC8), or elevenostat (a specific HDAC inhibitor targeting HDAC11) alone did not induce a significant response in tumor growth in mice. In contrast, co-administration of ABT-301 and RMP1-14 showed a better response to tumor suppression than co-administration of PCI-34051 and RMP1-14 or co-administration of elevenostat and RMP1-14 (data not shown).

[0125] After 49 days of recording and the results of tumor volume changes on days 22 and 31, see Figures 5I-5K and Table 14. Mice receiving combined treatment with ABT-301 and RMP1-14 had the smallest tumor volume among all experimental groups ( Figure 5I On day 22, individual administration of ABT-301, voriconazole, cetaxel, or RMP1-14 showed varying degrees of antitumor activity. However, combined treatment showed a more potent tumor growth inhibitory effect, with the combination of ABT-301 and RMP1-14 showing the most significant inhibitory effect ( Figure 5J and Table 14). Mice in the vehicle, ABT-301, voriconazole, and cetaxel groups were sacrificed due to reaching the humane endpoint. On day 31, after treatment termination, mice receiving the combination of ABT-301 and RMP1-14 still showed the smallest tumor volume among all experimental groups. This finding indicates that the combination treatment (ABT-301 and RMP1-14) can continuously and effectively inhibit tumor growth, in contrast to other combination treatments (which showed tumor regrowth). Figure 5K In addition, no statistically significant differences in the body weight (%) of mice between each treatment group and the vehicle group were observed throughout the experimental period (data not shown).

[0126] Table 14. Tumor growth inhibition rate on day 22 after HDAC inhibitor and RMP1-14 treatment in mice bearing CT26 homograft tumors

[0127] In summary, these data suggest that the combination of a pan-HDAC inhibitor and an anti-PD-1 antibody (e.g., RMP1-14) exhibits superior anti-cancer activity compared to the combination of a specific HDAC inhibitor (e.g., voriconazole, PCI-34051, or elestat) and an anti-PD-1 antibody. However, among all combinations of pan-HDAC inhibitors and anti-PD-1 antibodies, the combination of ABT-301 and RMP1-14 enhanced anti-cancer activity compared to other combinations of pan-HDAC inhibitors (e.g., voriconazole) and RMP1-14.

[0128] In summary, the present invention provides an improved treatment method for colorectal cancer, including MSS and MSI-H subtypes, by combining the disclosed drug ABT-301 with an anti-PD-1 or anti-PD-L1 antibody. This combination therapy exhibits enhanced therapeutic efficacy, thereby reducing the burden on patients suffering from the disease.

[0129] It should be understood that the description of the above embodiments is only for illustrative purposes, and that persons skilled in the art may make various changes and modifications thereto. The above description, examples, and experimental data provide a complete description of the structure and use of the present disclosure as illustrative embodiments. Although various embodiments of the present disclosure have been described above with certain characteristics, or with reference to one or more individual embodiments, persons skilled in the art may make numerous modifications to the disclosed embodiments without departing from the spirit and scope of the present disclosure.

Claims

1. A pharmaceutical kit for treating colorectal cancer in an individual, comprising A first container comprising (E)-N-hydroxy-3-(1-(phenylsulfonyl)-indolin-5-yl)-acrylamide; and A second container comprises an anti-PD-1 or anti-PD-L1 antibody.

2. The pharmaceutical kit according to claim 1, wherein the anti-PD-1 antibody is selected from the group consisting of cemiplimab, pembrolizumab, nivolumab, and RMP 1-14.

3. The pharmaceutical kit of claim 1, wherein the anti-PD-L1 antibody is selected from the group consisting of atezolizumab, avelumab, and durvalumab. 4 . The pharmaceutical kit according to claim 1 , wherein the colorectal cancer is a microsatellite stable-type or microsatellite instable-type colorectal cancer.

5. A method for treating colorectal cancer in an individual, comprising administering to the individual the pharmaceutical kit of claim 1, wherein The (E)-N-hydroxy-3-(1-(phenylsulfonyl)-indolin-5-yl)-acrylamide is administered to the subject in an amount of 10 μg / kg to 1 g / kg; and The anti-PD-1 or anti-PD-L 1 antibody is administered to the subject in an amount of 1 μg / kg to 100 mg / kg. 6 . The method of claim 5 , wherein the anti-PD-1 antibody is selected from the group consisting of cemiplizumab, pembrolizumab, nivolumab, and RMP 1-14.

7. The method of claim 5, wherein the anti-PD-L1 antibody is selected from the group consisting of atezolizumab, avelumab, and durvalumab.

8. The method of claim 5, wherein the colorectal cancer is a microsatellite stable or microsatellite unstable colorectal cancer.

9. The method of claim 5, wherein said (E)-N-Hydroxy-3-(1-(phenylsulfonyl)-indolin-5-yl)-acrylamide is administered to the subject in an amount of 2.5-10 mg / kg.

10. The method of claim 5, wherein the anti-PD-1 or anti-PD-L 1 antibody is administered to the subject in an amount of 100 μg / kg to 1 mg / kg.

11. The method of claim 5, wherein the individual is a human.

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