Combination of antibody-drug conjugates with cancer therapy and uses thereof

Abstract

The invention relates to the field of biological medicine, in particular to combination of antibody-drug conjugates and cancer therapy (especially kinase inhibitors or chemotherapy) and application thereof.

Description

Technical Field

[0001] This article relates to the biomedical field, specifically the combination of antibody-drug conjugates with cancer therapies (especially kinase inhibitors or chemotherapy) and their applications. Background Technology

[0002] Human epidermal growth factor receptor-2 (HER2) belongs to the epidermal growth factor (EGFR) receptor tyrosine kinase family. In addition to activating molecular pathways responsible for cell proliferation and survival under stress, HER2 also directly regulates programmed cell death. HER2 overexpression or amplification is characteristic of many common cancers. Commercially available HER2-targeting cancer therapies include antibodies, tyrosine kinase inhibitors, and antibody-drug conjugates (ADCs).

[0003] HER2-targeted adjuvants (ADCs) offer promising new avenues for the treatment of various cancers, including breast cancer, gastric or gastroesophageal junction cancer, colorectal cancer, non-small cell lung cancer, and other HER2-positive cancers. To date, microtubule inhibitors, such as tubulin polymerization inhibitors (e.g., maytansine alkaloids and auristatin), account for the vast majority of developed payloads. For example, DM1 (madensine alkaloids) targets T-DM1 (trastuzumab emtansine). It is used as a payload in Genentech / Roche and several research ADCs.

[0004] However, on the one hand, T-DM1 and other commercially available ADCs, as well as most ADCs in clinical trials, are synthesized via chemical conjugation, a method that utilizes a thiosuccinimide structure (thiosuccinimide linker) to conjugate small molecule drugs to targeting antibodies or proteins. The thiosuccinimide structure is formed through the reaction of a thiol group with maleimide. The thiosuccinimide linker is unstable. In vivo, reverse Michael addition or exchange with other thiol groups leads to payload detachment from the ADC, resulting in off-target toxicity and reduced safety. On the other hand, chemical conjugation reactions are not site-specific, and the resulting ADCs exhibit high heterogeneity. We have long focused on developing stable linkers, with results including extended half-life of ADC drugs in systemic circulation, reduced premature detachment, and increased payload accumulation on the target. Our HER-2-targeting ADCs, including GQ1001 (see CN106856656B), have demonstrated improved safety.

[0005] Drug resistance is another major obstacle in cancer therapy development. Inherent resistance and acquired new resistance mutations can be observed in conventional chemotherapy, targeted therapy, and immunotherapy. Compensatory adaptation processes in oncogenic pathways have also been described as a resistance mechanism. In-depth research is underway to overcome this obstacle. The development of combination therapies often faces challenges such as limited clinical efficacy, more complex dosing regimens, pharmacological compatibility issues, and the resulting need for higher levels of risk management. Cost-effectiveness is also a factor to consider.

[0006] For the treatment of HER2-positive breast cancer, in several cases, dual HER2 blockade with trastuzumab and pertuzumab combined with chemotherapy (such as taxanes and platinum-based drugs) is the standard first-line treatment, but resistance to trastuzumab is unavoidable. Combinations containing tyrosine kinase inhibitors such as tucatinib, neratinib, or lapatinib are primarily used for second- or third-line or later-line treatment. The tyrosine kinase inhibitor pyrotinib was approved in 2018 in combination with capecitabine for the treatment of HER2-positive patients with advanced or metastatic breast cancer who have previously received anthracycline or taxane chemotherapy, and in 2022 it was approved in combination with trastuzumab and docetaxel for neoadjuvant treatment of HER2-positive early or locally advanced breast cancer (LABC). T-DM1 can be used as monotherapy for patients with metastatic breast cancer (mBC) who have received one or more chemotherapy agents, and T-DM1 can be used as adjuvant therapy for HER2-overexpressing breast cancer patients who have received adjuvant chemotherapy containing anthracycline and radiotherapy (if applicable). Although T-DM1 has been extensively studied theoretically as a foundational drug for combination therapy strategies in mBC, the side effects of T-DM1-containing therapies limit their clinical research and application. For example, thrombocytopenia has been observed in approximately 30%–55% of patients receiving T-DM1 as a single agent. The combination of T-DM1 and docetaxel has been investigated in patients with mBC or LABC, but nearly half of these patients required dose reduction due to serious adverse events (grade 3–4 neutropenia in 72% of mBC patients). In the Phase III MARIANNE study, the combination of T-DM1 and pertuzumab showed only non-inferiority to, but not superior to, taxanes (docetaxel or paclitaxel) plus trastuzumab. Currently, clinical trials of T-DM1 in combination with other agents (e.g., immune checkpoint inhibitors, CDK4 / 6 inhibitors, and TKIs) are underway, but side effects remain a major concern. The combination regimens containing T-DM1 are associated with a higher incidence of side effects, thus limiting the number of doses and treatment cycles. Furthermore, resistance to chemotherapy drugs and / or antibodies can affect the objective response rate. Overall, the benefit to patients is limited.

[0007] Furthermore, T-DM1 has shown only limited clinical activity in gastric cancer or gastroesophageal junction cancer, and failed to demonstrate superior efficacy compared to taxanes (docetaxel or paclitaxel) in the phase II / III GATSBY study.

[0008] Therefore, new, highly effective, and low-toxicity therapies are still urgently needed. Summary of the Invention

[0009] In a first aspect, a pharmaceutical combination is provided, the pharmaceutical combination comprising the following (a) and (b);

[0010] (a) One or more couplings having the structure of formula (i-1) and / or formula (i-2).

[0011]

[0012]

[0013] Preferably, the trastuzumab is connected to the remainder of the conjugate via the C-terminus of the two light chains of the trastuzumab;

[0014] (b) One or more other cancer treatment agents, said one or more other cancer treatment agents being selected from: tyrosine kinase inhibitors, chemotherapeutic agents and pyrimidine antimetabolites.

[0015] In a second aspect, a kit or pharmaceutical composition is provided that comprises the pharmaceutical composition described herein.

[0016] In a third aspect, a method for preventing, alleviating, or treating a proliferative disorder is provided, the method comprising administering to a subject requiring such treatment an effective amount of any of the pharmaceutical combinations described herein, or the kits or pharmaceutical compositions described herein. Attached Figure Description

[0017] Figure 1a The efficacy of the combination of GQ01 and tucatinib in a human breast cancer PDX model was demonstrated.

[0018] Figure 1b The efficacy of the combination of GQ01 and neratinib in a human breast cancer PDX model was demonstrated.

[0019] Figure 1c The efficacy of the combination of GQ01 and pyrotinib in a human breast cancer PDX model was demonstrated.

[0020] Figure 2a The efficacy of the combination of GQ01 and tucatinib in a human breast cancer PDX model was demonstrated.

[0021] Figure 2bThe efficacy of the combination of GQ01 and pyrotinib in a human breast cancer PDX model was demonstrated.

[0022] Figure 3a The efficacy of the combination of GQ01 and tucatinib in a human breast cancer PDX model was demonstrated.

[0023] Figure 3b The efficacy of the combination of GQ01 and pyrotinib in a human breast cancer PDX model was demonstrated.

[0024] Figure 4a The efficacy of the combination of GQ01 and tucatinib in a human gastric cancer PDX model was demonstrated.

[0025] Figure 4b The efficacy of the combination of GQ01 and pyrotinib in a human gastric cancer PDX model was demonstrated.

[0026] Figure 5a The efficacy of the combination of GQ01 and paclitaxel in a human breast cancer CDX model was demonstrated.

[0027] Figure 5b The efficacy of the combination of GQ01 and capecitabine in a human breast cancer CDX model was demonstrated.

[0028] Figure 6a The efficacy of the combination of GQ01 and paclitaxel in a human breast cancer CDX model was demonstrated.

[0029] Figure 6b The efficacy of the combination of GQ01 and capecitabine in a human breast cancer CDX model was demonstrated. Detailed Implementation

[0030] The following provides specific implementation schemes to illustrate the technical content of this document. Through the disclosure in this specification, those skilled in the art can readily understand other advantages and effects of this document. This document can also be implemented or applied through other different specific implementation schemes. Those skilled in the art can make various modifications and changes without departing from the spirit of this document.

[0031] definition

[0032] Unless otherwise defined below, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. The term "technique" as used herein refers to those techniques commonly understood in the art, including variations and equivalents that are obvious to one of ordinary skill in the art. While it is believed that the following terms will be readily understood by one of ordinary skill in the art, the following definitions are provided for the purpose of better illustrating this document. When a trade name appears herein, it refers to the corresponding product or its active ingredient. All patents, published patent applications, and publications cited herein are incorporated herein by reference.

[0033] The terminology used herein to describe outcome measures of investigational drugs or drug combinations may be referenced to RECIST v1.1 (Criteria for Evaluation of Efficacy in Solid Tumors) or have the same meaning as commonly understood by those skilled in the art. Specifically, the following terms may be used to describe response status: CR, PR, PD, and SD. Complete Response (CR): All target lesions disappear; the short axis of any pathological lymph node (whether target or non-target) must be reduced to <10 mm. Partial Response (PR): The total diameter of target lesions decreases by at least 30% relative to the baseline total diameter. Disease Progression (PD): The total diameter of target lesions increases by at least 20% relative to the minimum total diameter in the study (including the baseline total if it is the minimum in the study); in addition to a relative increase of 20%, the total diameter must also increase by at least 5 mm in absolute terms. The presence of one or more new lesions is also considered disease progression. Stable Disease (SD): The reduction in diameter does not meet the PR criteria, and the increase in diameter does not meet the PD criteria, relative to the minimum total diameter in the study. In addition, in some implementations, the terms “clinical complete response (cCR, complete response assessed using clinical methods)” and “pathological complete response (pCR or pathCR, complete response assessed using pathological methods)” may be used to describe the outcome.

[0034] The terminology used in this article to describe human diagnostic and treatment methods may be referenced to the latest guidelines issued by authoritative bodies, especially those issued by the Chinese Society of Clinical Oncology (CSCO) or the American Society of Clinical Oncology (ASCO), or have the same meaning as commonly understood by those skilled in the art.

[0035] Unless the context explicitly indicates otherwise, the singular forms of "a / an" and "the / described" include the plural forms. Expressions of "one or more" or "at least one" can mean 1, 2, 3, 4, 5, 6, 7, 8, 9, or more. Such expressions are open-ended when the term "therapy" is used in combination with other terms. For example, the term "palbociclib therapy" does not exclude the use of other therapeutic agents. Other similar expressions such as "pyrotinib therapy" should be understood in a similar manner.

[0036] When used in conjunction with numerical variables, the terms “about” and “approximately” generally mean that the value of the variable and all values ​​of the variable are within the experimental error (e.g., within the 95% confidence interval of the mean) or within ±10% of the specified value, or a wider range.

[0037] The terms “optional” or “optionally” mean that the event described thereafter may, but is not necessarily, occur, and the description includes the possibility that the event or situation described therein may or may not occur.

[0038] The expressions "comprising," "including," "containing," and "having" are open-ended and do not exclude additional unlisted elements, steps, or components. The expression "consisting of" excludes any unspecified elements, steps, or components. The expression "substantially consisting of" means that the scope is limited to the specified elements, steps, or components, as well as other optional elements, steps, or components that do not materially affect the essential and novel features of the claimed subject matter. It should be understood that the expression "comprising" encompasses the expressions "substantially consisting of" and "consisting of."

[0039] As used herein, the term "antibody" is used broadly and specifically includes complete monoclonal antibodies, polyclonal antibodies, monospecific antibodies, multispecific antibodies (e.g., bispecific antibodies), and antibody fragments, provided they possess the desired biological activity. The antibody may have any subtype (such as IgG, IgE, IgM, IgD, and IgA) or subclass and may be derived from any suitable species. In some embodiments, the antibody is derived from human or mouse. The antibody may also be a fully human antibody, humanized antibody, or chimeric antibody prepared by recombinant methods.

[0040] As used herein, the term "monoclonal antibody (mAb)" refers to an antibody derived from a substantially homogeneous population of antibodies; that is, the individual antibodies constituting this population are identical except for the possible small number of naturally occurring mutations. Monoclonal antibodies exhibit high specificity for the same antigenic site, for example, one epitope (monospecific) or two epitopes (bispecific). The modifier "monoclonal" indicates that the antibody is characterized by being derived from a substantially homogeneous population of antibodies and should not be interpreted as requiring specific methods to produce the antibody.

[0041] A complete or full-length antibody essentially comprises an antigen-binding variable region, a light chain constant region (CL), and a heavy chain constant region (CH). The heavy chain constant region may include CH1, CH2, CH3, and / or CH4, depending on the antibody subtype. The antigen-binding variable region (also known as the variable region fragment or Fv fragment) typically contains a light chain variable region (V... L ) and heavy chain variable region (V HThe constant region can be a constant region with a natural sequence (such as a constant region with a human natural sequence), or it can be a constant region with an amino acid sequence variant of said natural sequence. The variable region recognizes and interacts with the target antigen. The constant region can be recognized and interacted with by the immune system.

[0042] Antibody fragments may contain a portion of a complete antibody, preferably its antigen-binding region or variable region. Examples of antibody fragments include Fab, Fab', F(ab')2, and V... H The Fab fragment consists of an Fd fragment, an Fv fragment, a single-domain antibody (dAb) fragment, and a separated complementarity-determining region (CDR), along with the CH1 domain. Fab fragments are antibody fragments obtained by digesting full-length immunoglobulins with papain, or fragments with the same structure generated, for example, through recombinant expression. Fab fragments contain a light chain (containing V... L and CL) and another chain, wherein the other chain contains a variable structural domain (V) of the heavy chain. H The F(ab')2 fragment contains a constant region (CH1) of the heavy chain and a Fab fragment. The F(ab')2 fragment is an antibody fragment obtained by digesting immunoglobulins with pepsin at pH 4.0–4.5, or a fragment with the same structure generated, for example, through recombinant expression. The F(ab')2 fragment essentially comprises two Fab fragments, each heavy chain portion containing a small number of additional amino acids, including cysteine ​​residues that form a disulfide bond connecting the two fragments. The Fab' fragment (one heavy chain and one light chain) is a fragment containing half of the F(ab')2 fragment. Antibody fragments may comprise multiple chains linked together, for example, via disulfide bonds and / or via peptide linkers. Examples of antibody fragments also include single-chain Fv (scFv), Fv, dsFv, biantibodies, Fd and Fd' fragments, and other fragments, including modified fragments. Antibody fragments typically contain at least or about 50 amino acids, and generally contain at least or about 200 amino acids.

[0043] The antibodies described herein can be prepared using techniques well known in the art, such as recombinant techniques, phage display techniques, synthetic techniques, or other techniques known in the art. For example, genetically engineered recombinant antibodies (or antibody mimics) can be expressed using suitable culture systems (e.g., E. coli or mammalian cells). Engineering can refer to, for example, the introduction of a ligase-specific recognition sequence at its end.

[0044] The term "drug resistance" refers to the resistance of a targeted disease to drug therapy. Multiple factors can contribute to drug resistance in cancer, such as tumor burden (tumor size, or, in the case of hematologic malignancies), growth kinetics, patient-to-patient and intratumoral heterogeneity, the immunosuppressive cancer microenvironment, increased genomic instability, and selective therapeutic pressure. In the case of HER2-positive breast cancer, the causes of resistance may include upregulation of HER2 expression, HER2 mutations, aberrant activation of HER2 downstream signaling pathways, loss of tumor suppressor genes, cell cycle dysregulation, and low infiltration of tumor immune cells. Current research to overcome this resistance involves selective inhibitors of PI3K, inhibition of the IGF1R signaling pathway, and CDK inhibitors.

[0045] A payload is a substance that inhibits or prevents cell expression activity, cell function, and / or causes cell damage. Currently, payloads used in ADCs are more toxic than chemotherapy drugs. Examples of payloads include, but are not limited to, drugs targeting the following targets: microtubule cytoskeleton, DNA, RNA, kinin-mediated protein transport, and regulation of apoptosis. Drugs targeting the microtubule cytoskeleton can be, for example, microtubule stabilizers or microtubule polymerization inhibitors. Examples of microtubule stabilizers include, but are not limited to, taxanes. Examples of microtubule polymerization inhibitors include, but are not limited to, maytansinoids, auristatin, vincristine alkaloids, colchicine, and sarsaparilla toxin. DNA-targeting drugs can be, for example, drugs that directly disrupt DNA structure or topoisomerase inhibitors. Examples of drugs that directly disrupt DNA structure include, but are not limited to, DNA double-strand breakers, DNA alkylating agents, and DNA intercalating agents. DNA double-strand breakers can be, for example, enediyne antibiotics, including but not limited to danendycin, esperamycin, neomycin, uncialamycin, etc. DNA alkylating agents can be, for example, DNA dialkylating agents (i.e., DNA cross-linking agents) or DNA monoalkylating agents. Examples of DNA alkylating agents include, but are not limited to, pyrrolo[2,1-c][1,4]benzodiazepines. (PBD) dimer, 1-(chloromethyl)-2,3-dihydro-1H-benzo[e]indole (CBI) dimer, CBI-PBD heterodimer, dihydroindole benzodiazepine (IGN) dimers, pyruvic compounds, etc. Examples of topoisomerase inhibitors include, but are not limited to, eczema and its derivatives (such as DX8951f, DXd-(1) and DXd-(2), the structures of which are shown below), camptothecin, and anthracycline. RNA-targeting drugs can be, for example, drugs that inhibit splicing, examples of which include, but are not limited to, pladienolide. Drugs that target kinin-mediated protein transport can be, for example, mitotic kinin inhibitors, including, but not limited to, kinin spindle protein (KSP) inhibitors.

[0046] HER2 refers to human epidermal growth factor receptor 2, which belongs to the epidermal growth factor (EGFR) receptor tyrosine kinase family (HER family or EGFR family).

[0047] The HER family of proteins are type I transmembrane receptor tyrosine kinases (RTKs), with four members: HER1 (epidermal growth factor receptor (EGFR), ErbB1), HER2 (ErbB2), HER3 (ErbB3), and HER4 (ErbB4). The expression status of HER2 can be determined using known methods such as immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH).

[0048] Trastuzumab is an anti-HER2 IgG1κ monoclonal antibody that binds to the fourth extracellular domain (ECD4) of HER2. In addition to triggering antibody-dependent cytotoxicity, trastuzumab also inhibits HER2 activation by suppressing the cleavage of the HER2 extracellular domain, thereby preventing the formation of its truncated and active form, p95HER2. It is approved for the treatment of HER2-overexpressing breast cancer and metastatic gastric or gastroesophageal junction adenocarcinoma. In addition to its direct antitumor effects, trastuzumab has also been found to possess immune system-mediated antitumor activity.

[0049] Trastuzumab induces sustained downregulation of membrane-bound HER2. Long-term trastuzumab treatment downregulates HER2. The expression of truncated HER2 without ECD, sustained activation of the PI3K / AKT signaling pathway, and overactivation of other tyrosine kinase receptors compensating for oncogenic mechanisms may contribute to trastuzumab resistance. Gene amplification and RNA / protein overexpression results indicate that tumor cells developing acquired resistance to trastuzumab remain dependent on the HER2 oncogene.

[0050] Pertuzumab binds to the extracellular dimerization domain (second extracellular domain, ECD2) of HER2. In addition to triggering antibody-dependent cytotoxicity, pertuzumab also prevents ligand-induced heterodimerization of HER2 with other HER family members, particularly HER3. Therefore, pertuzumab is also known as a HER2 dimerization inhibitor. It is approved for use in combination with trastuzumab and docetaxel, or in combination with trastuzumab and chemotherapy, to treat HER2-positive breast cancer, or as a monotherapy for adjuvant treatment of patients with HER2-positive early breast cancer at high risk of recurrence.

[0051] HER2-targeting ADCs are ADCs that use antibodies targeting HER2 as their target molecule. Currently commercially available HER2-targeting ADCs include vidicetuzumab (RC48) and T-DM1. Trastuzumab derutecan (T-DXd) T-DM1 is used to treat HER2-positive cancers. T-DM1 contains trastuzumab, which is covalently linked to the microtubule inhibitor DM1 (a maytansine derivative) via a thiosuccinimide linker MCC (4-[N-maleimide methyl]cyclohexane 1-carboxylate). The ADC drug GQ1001, developed by GeneQuantum, contains trastuzumab, which is covalently linked to DM1 via a stable linker comprising an open-ring thiosuccinimide structure.

[0052] As used in this article, the term "antibody-drug conjugate" is also referred to as "conjugate".

[0053] The term "tyrosine kinase inhibitor (TKI)" refers to a small molecule that can inhibit the tyrosine kinase activity of a target kinase. The terms "EGFR family tyrosine kinase inhibitor," "HER family tyrosine kinase inhibitor," or "ErbB receptor tyrosine kinase inhibitor" (also referred to herein as "EGFR family inhibitor," "HER family inhibitor," or "ErbB receptor inhibitor," respectively) refer to a TKI that can inhibit the tyrosine kinase activity of one or more EGFR family (HER family) proteins. HER2 inhibitors refer to the following TKIs that can inhibit the tyrosine kinase activity of HER2, but do not exclude the possibility of inhibiting the tyrosine kinase activity of one or more other HER family proteins. HER2 selective inhibitors refer to TKIs that selectively inhibit HER2. Currently marketed HER family TKIs include: HER2 selective inhibitors, such as tucatinib and DZD1516; pan-HER inhibitors, such as afatinib, neratinib, and pyrotinib; and lapatinib as a dual EGFR / HER2 inhibitor. In monogenic tumors, resistance to tyrosine kinase inhibitors (TKIs) is often caused by gatekeeper mutations and other driver mutations that maintain dependence on oncogenes.

[0054] Types of targeted therapies include, but are not limited to: enzyme inhibitors, enzyme activators, receptor modulators, ion channel modulators, transmembrane ion transport modulators, antibodies, antibody fragments, binding proteins to a given antigen, antibody mimics, and scaffold proteins with affinity for a given target. The term "small molecule for targeted therapy" or "small molecule targeted therapy" refers to a targeted therapy using small molecule compounds, and in some embodiments, particularly refers to enzyme inhibitors, especially kinase inhibitors. The term "targeted therapy agent" or "targeting agent" refers to an agent that can be used as a targeted therapy.

[0055] Examples of small molecule targeted therapies include, but are not limited to: kinase inhibitors, such as antisecretory agents (e.g., brefidobacterium); 4-1BB agonists (e.g., PF-05082566); HSP90 inhibitors; checkpoint inhibitors; cell cycle inhibitors and differentiation inducers; cysteine ​​aspartate protease activators; tyrosine kinase inhibitors, including but not limited to ALK inhibitors (e.g., crizotinib, ceritinib, alectinib, brigatinib, lorlatinib, entrectinib, bezatinib, ripretinib, ensartinib), and c-Met inhibitors (e.g., crizotinib, cabozantinib, carmatinib, terpollinib). Inhibitors include: nifedipine, ferritinib, glibenclamide, tevantinib, voritinib (cevotinib), beritinib, nifedipine, gumetinib, and carnitineib; HER family inhibitors (e.g., gefitinib, erlotinib, icotinib, afatinib, dacomitinib, osimertinib, ametinib, lapatinib, neratinib (HKI-272), tucatinib (tocatinib, ONT-380), ZN-A-1041, DZD1516, omamotinib, avitinib, pelitinib, and vormetinib (aflutinib); FLT3 inhibitors (e.g., tandutinib, lenatatinib, pericidatinib, giglitinib, and queza). Inhibitors of VEGF / VEGFR / FGFR / PDGFR (e.g., sorafenib, lenvatinib, regorafenib, nintedanib, apatinib, anlotinib, fruquintinib, erdatinib, pemitinib, avatinib, ricatatinib, sildenafil, dovirtinib, motishanib, surufatinib, clairabinib, daritinib (lucitanib), axitinib); RET inhibitors (e.g., vandetanib, ceprtinib, pralatinib); tropomyosin receptor kinase (TRK) inhibitors (e.g., larotrectinib, entrectinib, cabozantinib, meritinib, betaine). Ritinib, cistanotinib, atetinib, seritinib); Bcr-Abl1 inhibitors (e.g., imatinib, dasatinib, bosutinib, ladotinib, nilotinib, ponatinib, aciminib, rebamitinib, barbitinib); BTK inhibitors (e.g., ibrutinib, acalutinib, zanubrutinib, tirabrutinib, slubrutinib, evobrutinib, brumbutinib, HM71224, vicatinib, LOXO-305, fenibutinib, ARQ-531); JAK inhibitors (e.g., ruxolitinib, fenzotinib, tofacitinib, baricitinib, gandoltinib, letatinib, paktinib);Serine / threonine kinase inhibitors, including but not limited to BRAF / MEK / ERK inhibitors (e.g., vemurafenib, dabrafenib, encofenib, sorafenib, trametinib, bimetinib, cobibitinib, selumetinib, lavocitinib), CDK inhibitors (e.g., palbociclib, ribociclib, abeciclib, trelaciclib (G1T28)), PI3K / AKT / mTOR inhibitors (e.g., curpannice, duverithione, apeliximab (BYL719), serexetine, imilis, datoliximab (BEZ235), GDC-0084 (RG7666), GDC-0941, gidarisoxetine, partasertitine, capipritetine, afluritetine, euprositetine, tesimolimus, saline, etc.). Parselt (TAK-228), Vetuselt (AZD2014), CC-223, BI860585, DS-3078a, ME-344, Taselil (GDC-0032), GDC-0349, OSI-027, P529); epigenetic inhibitors, including but not limited to EZH2 inhibitors (e.g., 3-deadenine A (DZNep), EI1, EPZ005687, GSK126, GSK343, UNC1999, Tazesta (EPZ6438), SHR2554, CPI-1205, DS-3201, PF-06821497, HH2853, EED226 (MAK683)). BR-001, UNC6852, GNA002), HDAC inhibitors (e.g., TSA, vorinostat (SAHA), belistat, pabistal, romidesin, tucistat (chidamide), valproic acid, phenylbutyrate, nicotinamide, EX-527, sibutanol, cambinol, entenoxetine), IDH1 / 2 inhibitors (e.g., entidiprine (AG-221), evanib (AG-120), vorasidini (AG-881)), DNMT inhibitors (e.g., azacitidine, decitabine), DOT1L methyltransferase inhibitors (e.g., pinoxastat), BET inhibitors (e.g., pilarose, molirexate, ZEN003694) PLX51107), LSD1 inhibitors (e.g., idastat, INCB059872, CC-90011); BCL-2 inhibitors (e.g., Olimerson, ABT-737, ABT-263 (navittola), venettola, S55746 (also known as BCL201 or Servier-1), S64315 (MIK665), AZD-5991520, AMG-176, WEHI-539, A-1331852); hedgehog pathway inhibitors, including but not limited to SMO inhibitors (e.g., vemodega, sonidega, gradidega), GLI inhibitors (e.g., arsenic trioxide (ATO)) and itraconazole;Proteasome inhibitors, including but not limited to bortezomib, carfilzomib, ixazomib, marizomib (Salinomycin A), opzomib, and delanzomib; PARP inhibitors, including but not limited to nicotinamide, olaparib, rucaparib, niraparib, tazozoparib, pamiparib, INO-1001, E7449, iniparib, AZD2461, ameparib, IMP4297, RBN-2397, fluzoparib, and veriparib (ABT-888).

[0056] The term "chemotherapeutic agent" refers to compounds with selective toxicity and an appropriate therapeutic index, thus suitable for treating cancer. Types of chemotherapeutic agents include, but are not limited to: alkylating agents, antimetabolites, mitochondrial dysfunction inducers, cytotoxic / antitumor antibiotics, hormones and hormone analogs, photosensitizers, and drugs targeting the microtubule cytoskeleton, DNA, RNA, kinin-mediated protein transport, and regulation of apoptosis. Drugs targeting the microtubule cytoskeleton can be, for example, microtubule stabilizers or microtubule polymerization inhibitors. Examples of microtubule stabilizers include, but are not limited to, taxanes (e.g., docetaxel, paclitaxel, albumin-bound paclitaxel). Examples of microtubule polymerization inhibitors include, but are not limited to, maytansinoids, auratestatin, vincristine alkaloids, colchicine, and sarsaparilla toxin. DNA-targeting drugs can be, for example, drugs that directly disrupt DNA structure or topoisomerase inhibitors. Examples of drugs that directly disrupt DNA structure include, but are not limited to, DNA double-strand breakers, DNA alkylating agents, and DNA intercalating agents. DNA double-strand breaking agents can be, for example, enediyne antibiotics, including but not limited to danexycin, esperamycin, neomycin, and uncialamycin. DNA alkylating agents can be, for example, DNA dialkylating agents (i.e., DNA cross-linking agents) or DNA monoalkylating agents. Examples of DNA alkylating agents include, but are not limited to, pyrrolo[2,1-c][1,4]benzodiazepines. (PBD) dimer, 1-(chloromethyl)-2,3-dihydro-1H-benzo[e]indole (CBI) dimer, CBI-PBD heterodimer, dihydroindole benzodiazepine (IGN) dimers and pyruvicides. Examples of DNA intercalators include, but are not limited to, platinum-based drugs, ruthenium-based drugs, rhodium-based drugs, and organic dual intercalators. Examples of topoisomerase inhibitors include, but are not limited to, camptothecin and anthracycline. RNA-targeting drugs can be, for example, drugs that inhibit splicing, examples of which include, but are not limited to, pladienolide. Drugs that target kinin-mediated protein transport can be, for example, mitotic kinin inhibitors, including, but not limited to, kinin spindle protein (KSP) inhibitors. Other examples of chemotherapy agents include, but are not limited to: epothilone, cobustatin A-4 phosphate, cobustatin A-4 and its derivatives, indole-sulfonamides, sea haretoxin 10 and its analogues, spongin B, eribulin, indole-3-oxoacetamide, podophyllotoxin, 7-diethylamino-3-(2'-benzoxazolyl)-coumarin (DBC), spongilin, lelimycin, mitoxantrone, mitoxantrone hydrazone, nitrogen mustard, cyclophosphamide, nitrosourea, aziridines, benzodopa, carboquinone, metoprolol, urotepa, aclarubicin, actinomycin, atrazomycin, bleomycin, actinomycin C, carabixin, erythromycin, anticancermycin, actinomycin D, daunorubicin, detoxin, doxorubicin, epirubicin, esopixin, idarubicin. Star, Methamphetamine, Mitomycin, Nopramine, Oligomycin, Pepromycin, Pofibromycin, Purulin, Ferrous Oxytocin, Rodoxine, Streptomycin, Streptozotocin, Netostatin, Zorubicin, Trichoderma, T-2 Toxin, Veraculin A, Bacitracin A, Serpentin, Ubenimex, Diazoserine, 6-Diazo-5-oxo-L-leucine, Folic Acid, Methotrexate, Pteroside, Trimethoprim, Idatraxate, Fludarabine, 6-Mercaptopurine, Thiomipril, Thioguanine, Cytarabine, Capecitabine, Ancitabine, Gemcitabine, Enoxabine, Azacitidine, 6-azauridine, Carmoflurane, Dideoxyuridine, Deoxyfluorouridine, Fluorouracil, Calotestosterone, Drotahistone Propionate, Cyclothrostanol, Medanone, Testrolide. The term "chemotherapy" refers to a therapy that uses one or more chemotherapeutic agents, particularly a therapy that uses one chemotherapeutic agent. In some implementations, "chemotherapy" may be used interchangeably with the term "chemotherapeutic agent".

[0057] The terms “endocrine therapy (ET)” and “anti-hormone therapy” refer to treatments designed to reduce hormone release or the effect of hormones on cancer cell growth. Endocrine therapy is commonly used to treat breast cancer, prostate cancer, and thyroid cancer. Examples of endocrine therapy agents (endocrine agents) include, but are not limited to: estrogen receptor modulators (e.g., tamoxifen, clomiphene, raloxifene, toremifene, lasoxifene, azoxifen, opemifene, amoxifen, azoxifen, bardoxifene), aromatase inhibitors (e.g., anastrozole, letrozole, exemestane), antiadrenergic drugs (e.g., aminoglutethimide, mitotan, triplostertan), and androgen receptor modulators (e.g., flutamide, niglutethimide, bicalutamide, enzalutamide, abiraterone, leuprorelin acetate, galemetone, goserelin).

[0058] The term "immunoagonist" refers to an agonist capable of inducing or enhancing an immune response against a tumor, such as by activating immune cells, including but not limited to dendritic cells (DCs), B cells, macrophages, NK cells, and T cells. Examples of immune agonists include, but are not limited to, TLR agonists, such as agonists of TLR7 and / or TLR8 and / or TLR9 (e.g., imiquimod, ralsimod, 852A, and VTX-2337) and STING agonists (e.g., ADU-S100 and MK-1454).

[0059] The terms “small molecule targeted therapy” and “immune agonist”, “small molecule targeted therapy” and “endocrine therapy”, and “small molecule targeted therapy” and “chemotherapeutic agent” are not mutually exclusive when used in this article.

[0060] Small molecule compounds are molecules with a size comparable to organic molecules commonly used in medicine. This term does not cover biological macromolecules (such as proteins and nucleic acids), but it does cover low molecular weight peptides or their derivatives, such as dipeptides, tripeptides, tetrapeptides, and pentapeptides. Typically, the molecular weight of small molecule compounds can be, for example, about 100 Da to about 2000 Da, about 200 Da to about 1000 Da, about 200 Da to about 900 Da, about 200 Da to about 800 Da, about 200 Da to about 700 Da, about 200 Da to about 600 Da, or about 200 Da to about 500 Da.

[0061] Cellular proliferative disorders include, but are not limited to, tumors, cancers, and necrotic tissues, including pre-malignant and non-necrotic stages, as well as psoriasis, endometriosis, polyps, and fibroadenomas.

[0062] The terms "cancer" and "cancerous" refer to a physiological condition in mammals typically characterized by unregulated cell growth / proliferation. Examples of cancer include, but are not limited to, carcinoma, lymphoma, blastoma, sarcoma, and leukemia. More specific examples of such cancers include squamous cell carcinoma, small cell lung cancer, non-small cell lung cancer, lung adenocarcinoma, lung squamous cell carcinoma, peritoneal cancer, hepatocellular carcinoma, gastrointestinal cancer (e.g., stomach cancer or gastroesophageal junction cancer), pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, liver cancer, bladder cancer, hepatocellular carcinoma, breast cancer, colon cancer, colorectal cancer, endometrial or uterine cancer, salivary gland cancer, kidney cancer, liver cancer, prostate cancer, vulvar cancer, thyroid cancer, hepatocellular carcinoma, and various types of head and neck cancer.

[0063] As used herein, the term “tumor” refers to all proliferative cell growth and proliferation, whether malignant or benign, as well as all precancerous and cancerous cells and tissues. “Cancer” or “cancer tissue” may include tumors.

[0064] The terms “cancer,” “cancerous,” “proliferative disorder,” “proliferative disease,” and “tumor” are not mutually exclusive when used in this article.

[0065] The terms "cancer therapeutic agent," "antitumor agent," and "antimitotic agent" refer to drugs with anticancer effects. As used herein, the term "anticancer effect" refers to biological effects that can manifest as a reduction in tumor burden, such as a decrease in tumor volume, a reduction in the number of tumor cells, a reduction in tumor cell proliferation, a reduction in the number of metastases, an increase in overall survival or progression-free survival, an increase in life expectancy, or an improvement in various tumor-related physiological symptoms. Anticancer effects can also refer to the prevention of cancer, such as the therapeutic effect on precancerous lesions.

[0066] Examples of precancerous lesions include, but are not limited to: actinic keratosis (solar keratosis), nevus (dysplastic nevus), actinic cheilitis (farmer's lip), cutaneous horn, Barrett's esophagus, atrophic gastritis, congenital dyskeratosis, iron deficiency dysphagia, lichen planus, oral submucosal fibrosis, actinic (solar) elastin degeneration, and cervical dysplasia.

[0067] As used herein, the term "wild-type" is as understood in the art and refers to a polypeptide or polynucleotide sequence found in a natural population without genetic modifications. Also as understood in the art, a "mutant" includes polypeptide or polynucleotide sequences that have at least one modification on an amino acid or nucleic acid compared to the corresponding amino acid or nucleic acid found in a wild-type polypeptide or polynucleotide. The term mutant includes single nucleotide polymorphisms (SNPs), where a single base pair differs in the sequence of a nucleic acid chain compared to the most commonly found (wild-type) nucleic acid chain. Cancers identified by known methods that are associated with wild-type HER2 (Uniprot accession number: P04626) or mutants, or that have ErbB2 (Genbank accession number X03363) amplification or HER2 protein overexpression.

[0068] In some implementations, the mutant form of HER2 includes one or more of the following: an insertion in exon 20 of HER2; a deletion around amino acid residues 755-759 of HER2; or one or more of the following amino acid substitutions: D277G, D277H, D277V, D277Y, G309E, G309A, S310F, S310Y, S653C, V664E, R678Q, V697L, L726F, K753E, L755S, L755P, I767M. L768S, D769H, D769Y, V773L, A775_G776insYVMA, G776delinsVC, V777L, P780_Y781insGSP, T798I, T798M, C805S, V842I, T862A, L866M, L869R, and R896C; and the same non-synonymous potential activating mutations (or insertion / deletion mutations) present in two or more independent samples as reported in the COSMIC database or prior art references. In some implementations, the mutant form of HER2 includes one or more of the following amino acid substitutions: D277G, D277H, D277V, D277Y, G309E, S310F, S310Y, S653C, R678Q, V697L, L726F, K753E, L755S, I767M, D769H, D769Y, A775 G776insYVMA, G776delinsVC, V777L, P780 Y781insGSP, T798I, T798M, V842I, L866M, and L869R. In some embodiments, the mutant form of HER2 comprises one or more of the following amino acid substitutions: S310F, S310Y, V697L, L726F, K753E, L755S, I767M, D769H, D769Y, V777L, P780_Y781insGSP, T798I, T798M, V842I, and L869R. In some embodiments, the mutant form of HER2 comprises one or more of the following amino acid substitutions: exon 20 insertion / deletion, L755S, V777L, T798I, and V842I. In some embodiments, the mutant form of HER2 comprises one or more of the following amino acid substitutions: S310F, S310Y, R678Q, D769H, D769Y, and V777L.

[0069] "HER2-mutated" cancers include, but are not limited to, cancers characterized by overexpression of the mutant HER2 protein.

[0070] The term "pharmaceutically acceptable" means that when it comes into contact with a patient's tissues within the normal range of medical judgment, it will not produce inappropriate toxicity, irritation, allergic reactions, etc.

[0071] Pharmaceutically acceptable salts of other cancer therapeutic agents include their acid addition salts and base addition salts. Methods for preparing pharmaceutically acceptable salts of the compounds described herein are known to those skilled in the art. In one embodiment, the pharmaceutically acceptable salt is a maleate salt.

[0072] The other cancer therapeutic agents may exist in specific geometric or stereoisomer forms, including cis and trans isomers, (-)-enantiomers and (+)-enantiomers, (R)-enantiomers and (S)-enantiomers, diastereomers, (D)-isomers, (L)-isomers, and racemic mixtures thereof and other mixtures, such as mixtures enriched with enantiomers or diastereomers, all of which are within the scope of this document.

[0073] A solvate of a compound refers to a form in which the compound contains a solvent as an integral part of its crystal structure, said solvent being, in particular, water, methanol, or ethanol. The amount of solvent, especially water, may be stoichiometric or non-stoichiometric.

[0074] Metabolites of a compound are substances formed in the body when the compound is administered. Such products can be generated, for example, through oxidation, reduction, hydrolysis, amidation, deamidation, esterification, enzymatic hydrolysis, etc., of the administered compound.

[0075] Prodrugs are certain derivatives of compounds that have little or no pharmacological activity on their own, but can be converted into the compounds of this article with the desired activity through, for example, hydrolysis and cleavage when administered to the human body or in vivo.

[0076] The term “polymorph” or “polymorphic material” refers to a single polymorph or a mixture of more than one polymorph in any proportion.

[0077] The term "crystal form" or "crystal" refers to any solid substance that exhibits a three-dimensional ordered structure. In contrast to amorphous solid substances, crystalline substances can produce characteristic X-ray powder diffraction patterns with clear and well-defined diffraction peaks.

[0078] The term "amorphous" refers to any solid material that lacks an ordered structure in three dimensions.

[0079] The term "effective dose" refers to the dose of these medications that, when administered, can alleviate one or more symptoms of the disease being treated to a certain extent.

[0080] As used herein, the term "subject" in relation to an individual suffering from a disease or symptom encompasses both mammals and non-mammals. Examples of mammals include, but are not limited to, any member of the class Mammalia: humans, non-human primates such as chimpanzees, and other ape and monkey species; farm animals such as cattle, horses, sheep, goats, and pigs; domesticated animals such as rabbits, dogs, and cats; and laboratory animals, including rodents such as rats, mice, and guinea pigs. Examples of non-mammals include, but are not limited to, birds and fish. In some embodiments of the methods and uses provided herein, the subject is a human.

[0081] Drug combination

[0082] In one aspect, a pharmaceutical combination is provided, the pharmaceutical combination comprising the following (a) and (b);

[0083] (a) One or more couplings having structures of formula (I-1), (I-2), (I-3) and / or (I-4).

[0084]

[0085]

[0086] Wherein, n is an integer from 1 to 100, d is an integer from 1 to 20, A is an antibody or its antigen-binding fragment, LA3 contains 1 to 100 tandem structural units selected from one or more glycine and alanine; b is independently 0 or 1, i.e., LA3 is independently present or absent, X in the ligase recognition sequence LPXT is glutamic acid (E) or any other natural / non-natural amino acid, where L is leucine, P is proline, and T is threonine; x is a -OH or -NH2 group;

[0087] (b) One or more other cancer therapeutic agents selected from tyrosine kinase inhibitors, taxanes, and pyrimidine antimetabolites (also known as pyrimidine analogs).

[0088] In one embodiment, the ligase recognition sequence LPXT has the amino acid sequence of SEQ ID No:11.

[0089] Other cancer therapeutic agents described herein include their pharmaceutically acceptable salts, stereoisomers, solvates (preferably hydrates), polymorphs, tautomers, isotopic compounds, metabolites, or prodrugs.

[0090] Specific implementation plan for drug combination

[0091] Further embodiments of the pharmaceutical combinations described herein are described below by describing both (a) and (b), specifying any component of (a) and / or (b), or specifying other components of the pharmaceutical combinations. It should be understood that any pharmaceutical combination formed by any combination of mutually non-exclusive embodiments disclosed herein is expressly disclosed herein, whether or not the pharmaceutical combination is specifically described.

[0092] (a): Coupling

[0093] In one embodiment, the conjugates of formula (I-1), (I-2), (I-3), or (I-4) are antibody-drug conjugates formed by covalently linking the following portions: a payload, a linker, and an antibody; wherein the payload is DM-1 (a maytansine derivative); and the linker is a stable linker that tends not to undergo reverse Michael addition or exchange with other thiol groups. In one embodiment, the antibody is linked to the remainder of the conjugate via one or more linkers formed by site-specific coupling catalyzed by a ligase.

[0094] In one embodiment, the coupling has a structure as shown in formula (I-1) and / or (I-2). In another embodiment, the coupling has a structure as shown in formula (I-3) and / or (I-4).

[0095] In one embodiment, n is an integer from 3 to 10, preferably 2 to 5, particularly 3. In one embodiment, d is an integer from 1 to 10, preferably 1 to 4, more preferably 1 or 2, particularly 2. In one embodiment, LA3 is GGGGS, GGGGSGGGGS, GGGGSGGGGSGGGGS, or GA; preferably, LA3 is GA. In one embodiment, b is independently 0 or 1; preferably, b is 1, i.e., LA3 is present. In one embodiment, X in the ligase recognition sequence LPXT is glutamine (Q) or glutamate (E); preferably glutamate (E). In one embodiment, x is an -NH2 group.

[0096] In one embodiment, the antibody is an anti-CD19 antibody, an anti-CD20 antibody, an anti-CD22 antibody, an anti-CD25 antibody, an anti-CD30 / TNFRSF8 antibody, an anti-CD33 antibody, an anti-CD37 antibody, an anti-CD44v6 antibody, an anti-CD56 antibody, an anti-CD70 antibody, an anti-CD71 antibody, an anti-CD74 antibody, an anti-CD79b antibody, an anti-CD117 / KITk antibody, an anti-CD123 antibody, an anti-CD138 antibody, an anti-CD142 antibody, an anti-CD174 antibody, an anti-CD227 / MUC1 antibody, an anti-CD352 antibody, an anti-CLDN18.2 antibody, an anti-DLL3 antibody, an anti-ErbB2 / HER2 antibody, an anti-CN33 antibody, an anti-GPNMB antibody, an anti-ENPP3 antibody, an anti-Nectin-4 antibody, or an anti-EGFRvⅢ antibody. Anti-SLC44A4 / AGS-5 antibody, anti-CEACAM5 antibody, anti-PSMA antibody, anti-TIM1 antibody, anti-LY6E antibody, anti-LIV1 antibody, anti-Nectin4 antibody, anti-SLITRK6 antibody, anti-HGFR / cMet antibody, anti-SLAMF7 / CS1 antibody, anti-EGFR antibody, anti-BCMA antibody, anti-AXL antibody, anti-NaPi2B antibody, anti-GCC antibody, anti-STEAP1 antibody, anti-MUC16 antibody, anti-mesothelin antibody, anti-ETBR antibody, anti-EphA2 antibody, anti-5T4 antibody, anti-FOLR1 antibody, anti-LAMP1 antibody, anti-cadherin 6 antibody, anti-FGFR2 antibody, anti-FGFR3 antibody, anti-CA6 antibody, anti-CanAg antibody, anti-integrin αV antibody, anti-TDGF1 antibody, anti-Ephrin A4 antibody, anti-TROP2 antibody, anti-PTK7 antibody, anti-NOTCH3 antibody, anti-C4.4A antibody, anti-FLT3 antibody, anti-B7H3 / 4 antibody, anti-tissue factor antibody, or anti-ROR1 / 2 antibody; preferably, anti-HER2 antibody, anti-FGFR3 antibody, anti-Trop2 antibody, anti-HER3 antibody, or anti-FRα antibody.

[0097] In a preferred embodiment, the antibody is an anti-HER2 antibody. In one embodiment, the antibody is an anti-human HER2 antibody. Examples of anti-human HER2 antibodies include, but are not limited to, pertuzumab and trastuzumab.

[0098] In one embodiment, the antibody is an anti-HER2 antibody or its antigen-binding fragment, the anti-HER2 antibody or its antigen-binding fragment comprising a light chain variable region (V0). L ) and heavy chain variable region (V H ), wherein V LLCDR1 comprising the amino acid sequence of SEQ ID NO:1, LCDR2 comprising the amino acid sequence of SEQ ID NO:2, and LCDR3 comprising the amino acid sequence of SEQ ID NO:3, wherein the V H The antibody comprises HCDR1 containing the amino acid sequence of SEQ ID NO:4, HCDR2 containing the amino acid sequence of SEQ ID NO:5, and HCDR3 containing the amino acid sequence of SEQ ID NO:6. In one embodiment, the CDR region is defined by KABAT. In one embodiment, the antibody is an anti-HER2 antibody or an antigen-binding fragment thereof, the anti-HER2 antibody or the antigen-binding fragment thereof comprising a light chain variable region (V... L ) and heavy chain variable region (V H ), wherein V L Having an amino acid sequence having at least about 85%, at least about 90%, or at least about 95% sequence identity with SEQ ID NO:7, and the V H It has an amino acid sequence having at least about 85%, at least about 90%, or at least about 95% sequence identity with SEQ ID NO:8. In a preferred embodiment, the anti-HER2 antibody or its antigen-binding fragment comprises a light chain variable region (V... L ) and heavy chain variable region (V H ), wherein V L Having the amino acid sequence of SEQ ID NO:7, and the V H The antibody has the amino acid sequence of SEQ ID NO:8. In one specific embodiment, the anti-HER2 antibody comprises a light chain and a heavy chain, wherein the light chain has an amino acid sequence having at least about 85%, at least about 90%, or at least about 95% sequence identity with SEQ ID NO:9, and the heavy chain has an amino acid sequence having at least about 85%, at least about 90%, or at least about 95% sequence identity with SEQ ID NO:10. In a preferred embodiment, the anti-human HER2 antibody is selected from trastuzumab and engineered anti-HER2 antibodies based on trastuzumab. In one embodiment, the antibody is trastuzumab. In a preferred embodiment, the antibody is linked to the remainder of the conjugate via the C-terminus of one or both light chains of the antibody, preferably both light chains. In one specific embodiment, A in formulas (I-1), (I-2), (I-3), and (I-4) is trastuzumab.

[0099] In one embodiment, the coupling of formula (I-1) has the structure of formula (i-1).

[0100]

[0101] In one embodiment, the coupling of formula (I-2) has the structure of formula (i-2).

[0102]

[0103] In one embodiment, the conjugate has the structure of formula (i-1) and / or formula (i-2) described above. In a preferred embodiment, the trastuzumab is connected to the remainder of the conjugate via the C-termini of the two light chains of the trastuzumab.

[0104] (b): Other cancer treatments

[0105] In one embodiment, the other cancer therapeutic agent is selected from small molecule compounds. In another embodiment, the other cancer therapeutic agent is selected from: targeted therapy agents, chemotherapeutic agents, endocrine therapy agents, and immune agonists.

[0106] In one embodiment, the other cancer therapeutic agents are selected from: vaccines, antibodies, fusion proteins, HER2-targeting ADCs, fibrinolytic agents; anti-migration agents; anti-secretion agents; HSP90 inhibitors; anti-angiogenic compounds; checkpoint inhibitors; cell cycle inhibitors and differentiation inducers; mitochondrial dysfunction inducers; cysteine-aspartic protease activators; kinase inhibitors, proteasome inhibitors, poly(ADP-ribose) polymerase (PARP) inhibitors, zeste homolog enhancer 2 (EZH2) inhibitors, histone deacetylase (HDAC) inhibitors, isocitrate dehydrogenase 1 / 2 (IDH1 / 2) inhibitors, DNA methyltransferase (DNMT) inhibitors, and DOT1-like histone lysine (DOT1L). Methyltransferase inhibitors, bromodomain and terminal extra-terminal domain (BET) protein inhibitors, lysine-specific demethylase 1 (LSD1) inhibitors, B-cell lymphoma-2 (BCL-2) inhibitors, transmembrane smoothing protein (SMO) inhibitors, GLI inhibitors, itraconazole, alkylating agents, antimetabolites, cytotoxic / antitumor antibiotics, hormones and hormone analogs, photosensitizers, microtubule stabilizers, microtubule polymerization inhibitors, DNA double-strand breaking agents, DNA alkylating agents, DNA intercalating agents, topoisomerase inhibitors, mitotic kinesin inhibitors, estrogen receptor modulators, aromatase inhibitors, antiadrenergic drugs, androgen receptor modulators, Toll-like receptor (TLR) agonists, and interferon gene stimulators (STING) agonists.

[0107] In one embodiment, the other cancer therapeutic agents are selected from: tyrosine kinase inhibitors, chemotherapeutic agents, and pyrimidine antimetabolites.

[0108] In one embodiment, the kinase inhibitor is selected from: tyrosine kinase inhibitors and serine / threonine kinase inhibitors. In one embodiment, the tyrosine kinase inhibitor is selected from: anaplastic lymphoma kinase (ALK) inhibitors, c-Met inhibitors, HER family inhibitors, FMS-like tyrosine kinase 3 (FLT3) inhibitors, vascular endothelial growth factor (VEGF) / vascular endothelial growth factor receptor (VEGFR) inhibitors, fibroblast growth factor receptor (FGFR) inhibitors, platelet-derived growth factor receptor (PDGFR) inhibitors, transfection rearrangement (RET) kinase inhibitors, tropomyosin receptor kinase (TRK) inhibitors, Bcr-Abl1 inhibitors, Bruton's tyrosine kinase (BTK) inhibitors, and Janus kinase (JAK) inhibitors. In one embodiment, the serine / threonine kinase inhibitor is selected from: BRAF inhibitors, mitogen-activated protein kinase (MEK) inhibitors, extracellular signal-regulated kinase (ERK) inhibitors, cyclin-dependent kinase (CDK) inhibitors, phosphatidylinositol 3-kinase (PI3K) inhibitors, AKT inhibitors, and mammalian target of rapamycin (mTOR) inhibitors.

[0109] In one implementation, the antibody used as another cancer therapeutic agent is selected from: trastuzumab, pertuzumab, magnituximab, bevacizumab, atelizumab, pembrolizumab, PF-05082566, tislelizumab, and their biosimilars (e.g., (Trastuzumab-pkrb)

[0110] In one embodiment, the other cancer treatment agents are selected from: brefidobacterium, PF-05082566, crizotinib, ceritinib, alectinib, brigatinib, lorlatinib, entrectinib, bezatinib, ripretinib, ensartinib, cabozantinib, carmatinib, terpoxtinib, furitinib, glibenclamide, tevantinib, voritinib (cevotinib), beritinib, nigratinib, gumetinib, carnitineib, gefitinib, erlotinib, icotinib, afatinib, dacomitinib, osimertinib, ametinib, lapatinib, lenatitinib (HKI-272), tucatinib (tocatinib, ONT-380), ZN-A-1041, DZD1516, omamotinib, avitinib, and pectinib. Ritinib, Vometinib (Aflutinib), Tandutinib, Letatinib, Percidatinib, Giglitinib, Quezatinib, Ponatinib, Cabozantinib, Sorafenib, Lenvatinib, Regorafenib, Nintedanib, Apatinib, Anlotinib, Fruquintinib, Erdatinib, Pemitinib, Avatinib, Repatinib, Sildiraib, Dovirtinib, Motishanib, Suruvatinib Nitroglycerin, Cleratinib, Deritinib, Axitinib, Vandetanib, Septinib, Pramipinib, Larotrectinib, Meritinib, Cetratinib, Atetinib, Celitinib, Imatinib, Dasatinib, Bosutinib, Ladotinib, Nilotinib, Aciminib, Rebatinib, Baffitinib, Ibrutinib, Acarutinib, Zanubrutinib, Telaralutinib, Spabutinib Evorabutinib, brumbutinib, HM71224, vecartinib, LOXO-305, fenibutinib, ARQ-531, ruxolitinib, finzotinib, tofacitinib, baricitinib, gandrolinib, paclinib, vemurafenib, dabrafenib, encofenib, trametinib, bimetinib, cobibitinib, seletinib, lavocitinib, palbociclib, ribociclib, abeciclib, triamcinolone (G1T28), cupanixine, duverithione, apeliximab (BYL719), threliximab, bismiliximab, datoliximab (BEZ235), GDC-0084 (RG7666), GDC-0941, gidarisoximab, partasertit, carpicetit, aflusetit, euprosetit, tesiromos Sapaxel (TAK-228), Vetuximab (AZD2014), CC-223, BI860585, DS-3078a, ME-344, Taselix (GDC-0032), GDC-0349, OSI-027, P529, 3-Desafenone A (DZNep), EI1, EPZ005687, GSK126, GSK343, UNC1999, Tazesta (EPZ6438), SHR2554, CPI-1205, DS-3201, PF-06821497, HH2853, EED226 (MAK683), BR-001, UNC6852, GNA002, TSA,Vorinostat (SAHA), Belistat, Papirostat, Romidesin, Tuxexistat (Chidamide), Valproic acid, Phenylebyl butyrate, Nicotinamide, EX-527, Sibutanol, Canbirol, Entenol, Ensidipine (AG-221), Evonib (AG-120), Volasidini (AG-881), Azacitidine, Decitabine, Pinostat, Bilarix, Molirex, ZEN003694, PLX51107, Adalat, INCB059872, CC-90011, Olimerson, ABT-737, ABT-263 (Navetola), Vernetola, S55746 (also known as BCL201 or Servier-1), S64315 (MIK665), AZD-5991520, AMG-176, WEHI -539, A-1331852, Vimodil, Sonidil, Gladil, Arsenic Trioxide (ATO), Itraconazole, Bortezomib, Carfilzomib, Ixazomib, Marizomib (Salinomycin A), Opzomib, Delanzomib, Olaparib, Lucaparib, Niraparib, Tazozoparib, Pamiparib, INO-1001, E7449, Iniparib, AZD2461, A Meparib, IMP4297, RBN-2397, Fluzoparib, Veriparib (ABT-888), Docetaxel, Paclitaxel, Albumin-bound Paclitaxel, Maytanine Alkaloids, Auristatin, Vincristine Alkaloids, Colchicine, Sea Haretoxin, Danonemycin, Esperamycin, Neocarcinogen, Uncialamycin, Pyrrolo[2,1-c][1,4]benzodiazepines (PBD) dimer, 1-(chloromethyl)-2,3-dihydro-1H-benzo[e]indole (CBI) dimer, CBI-PBD heterodimer, dihydroindole benzodiazepine (IGN) dimer, pyruvicides, platinum-based drugs, ruthenium-based drugs, rhodium-based drugs, organic dual intercalation agents, camptothecin, anthracycline, praldidine, epothilone, cobustatin A-4 phosphate, cobustatin A-4 and its derivatives, indole-sulfonamides, sea haretoxin 10 and its analogues, spongin B, eribulin, indole-3-oxoacetamide, podophyllotoxin, 7-diethylamino-3-(2'-benzoxazolyl)-coumarin (DBC), spongin lactone, lelimycin, mitoxantrone Mitoguanidine hydrazone, nitrogen mustard, cyclophosphamide, nitrosourea, aziridines, benzodopa, carboquinone, metoprolol, ureotepa, aclarubicin, actinomycin, atrazomycin, bleomycin, actinomycin C, carabicin, erythromycin, anticancer mycotoxin, actinomycin D, daunorubicin, detoxin, doxorubicin, epirubicin, isorubicin, idarubicin, mesorubicin, mitomycin, nopramine, olivomycin, pepromycin, pofibromycin, puromycin, doxorubicin, rodorubicin, streptomycin, streptozotocin, net Saturidine, Zorubicin, Trichothecene, T-2 toxin, Veraculin A, Bacitracin A, Serpentin, Ubenimex, Diazoserine, 6-Diazo-5-oxo-L-leucine, Folic acid, Methotrexate, Pteroxate, Trimethoprim, Idatraxate, Fludarabine, 6-Mercaptopurine, Thiomipril, Thioguanine, Cytarabine, Capecitabine, Ancitabine, Gemcitabine, Enoxabine, Azacitidine, 6-azauridine, Carmoflurane, Dideoxyuridine, Deoxyfluorouridine, Fluorouracil, Calotestosterone, Drotathione Ketopropionate, Cyclothothermide, Medanone, Testrolide, Tamoxifen, Clomiphene, Raloxifene, Toremiphene, Lasoxifene, Azoxifen, Olemiphene, Amoxifen, Azoxifen, Bardoxifene, Anastrozole, Letrozole, Exemestane, Aminoglutide, Mitotan, Tralostan, Flutamide, Nilumet, Bicalutamide, Enzalutamide, Abiraterone, Leuprolide Acetate, Galetolone, Goserelin, Imiquimod, Resimod, 852A, VTX-2337, ADU-S100, and MK-1454.

[0111] In one embodiment, the other cancer therapeutic agent is selected from: kinase inhibitors and chemotherapeutic agents. In another embodiment, the other cancer therapeutic agent is selected from: tyrosine kinase inhibitors, taxanes, and pyrimidine antimetabolites (also known as pyrimidine analogs).

[0112] In one embodiment, the other cancer therapeutic agent is selected from kinase inhibitors. In one embodiment, the kinase inhibitor is selected from HER family tyrosine kinase inhibitors. In a specific embodiment, the HER family tyrosine kinase inhibitor is selected from lapatinib, tucatinib, afatinib, neratinib, and pyrotinib. In another specific embodiment, the other cancer therapeutic agent is selected from tucatinib, neratinib, and pyrotinib; preferably tucatinib or pyrotinib. In another specific embodiment, the other cancer therapeutic agent is pyrotinib. In one embodiment, the tyrosine kinase inhibitor is in the form of a pharmaceutically acceptable salt thereof. In a specific embodiment, pyrotinib is in the form of pyrotinib maleate. In another specific embodiment, the HER family tyrosine kinase inhibitor is ZN-A-1041 or DZD1516.

[0113] In one embodiment, the other cancer therapeutic agent is selected from taxanes. In another embodiment, the other cancer therapeutic agent is selected from docetaxel and paclitaxel, preferably paclitaxel.

[0114] In one embodiment, the other cancer therapeutic agent is selected from pyrimidine antimetabolites. In one embodiment, the pyrimidine antimetabolite is selected from: cytarabine, capecitabine, ancitabine, gemcitabine, enoxatabine, carmoflu, dideoxyuridine, deoxyfluorouridine, and fluorouridine. In another specific embodiment, the other cancer therapeutic agent is capecitabine.

[0115] In one embodiment, the other cancer treatment agents are selected from: tucatinib, neratinib, pyrotinib, paclitaxel, and capecitabine.

[0116] In one embodiment, the antibody, as another cancer therapeutic agent, is an anti-CD19 antibody, an anti-CD20 antibody, an anti-CD22 antibody, an anti-CD25 antibody, an anti-CD30 / TNFRSF8 antibody, an anti-CD33 antibody, an anti-CD37 antibody, an anti-CD44v6 antibody, an anti-CD56 antibody, an anti-CD70 antibody, an anti-CD71 antibody, an anti-CD74 antibody, an anti-CD79b antibody, an anti-CD117 / KITk antibody, an anti-CD123 antibody, an anti-CD138 antibody, an anti-CD142 antibody, an anti-CD174 antibody, an anti-CD227 / MUC1 antibody, an anti-CD352 antibody, an anti-CLDN18.2 antibody, an anti-DLL3 antibody, an anti-ErbB2 / HER2 antibody, an anti-CN33 antibody, an anti-GPNMB antibody, an anti-ENPP3 antibody, an anti-Nectin-4 antibody, or an anti-EGFR antibody. Anti-vIII antibody, anti-SLC44A4 / AGS-5 antibody, anti-CEACAM5 antibody, anti-PSMA antibody, anti-TIM1 antibody, anti-LY6E antibody, anti-LIV1 antibody, anti-Nectin4 antibody, anti-SLITRK6 antibody, anti-HGFR / cMet antibody, anti-SLAMF7 / CS1 antibody, anti-EGFR antibody, anti-BCMA antibody, anti-AXL antibody, anti-NaPi2B antibody, anti-GCC antibody, anti-STEAP1 antibody, anti-MUC16 antibody, anti-mesothelin antibody, anti-ETBR antibody, anti-EphA2 antibody, anti-5T4 antibody, anti-FOLR1 antibody, anti-LAMP1 antibody, anti-cadherin 6 antibody, anti-FGFR2 antibody, anti-FGFR3 antibody, anti-CA6 antibody, anti-CanAg antibody, anti-integrin αV antibody, anti-TDGF1 antibody, anti-Ephrin A4 antibody, anti-TROP2 antibody, anti-PTK7 antibody, anti-NOTCH3 antibody, anti-C4.4A antibody, anti-FLT3 antibody, anti-B7H3 / 4 antibody, anti-tissue factor antibody, anti-ROR1 / 2 antibody; preferably, anti-HER2 antibody, anti-FGFR3 antibody, anti-Trop2 antibody, anti-HER3 antibody or anti-FRα antibody.

[0117] In one embodiment, the other cancer treatment agents are selected from: palbociclib, afatinib, poziotinib, metronidazole, PD-0332991, tucatinib, abeciclib, neratinib, curapannixi, anthracycline-based chemotherapy, pertuzumab, BYL719, DZD1516, lapatinib, pembrolizumab, vinorelbine, capecitabine, atelizumab, osimertinib, ribociclib, pyrotinib (e.g., pyrotinib maleate), venetumab, GDC-0941, trastuzumab, 4-1BB agonists (e.g., PF-05082566), BN-Brachyury, TPIV100, M7824, taxanes (e.g., docetaxel, paclitaxel, albumin-bound paclitaxel, etc.). Entenolol, vemurafenib, alectinib, ZN-A-1041, saxaglastine, carboplatin, doxorubicin, epirubicin, cyclophosphamide, taselixir, eribulin, gemcitabine, tislelizumab, celecoxib

[0118] Pharmaceutical compositions and pharmaceutical preparations

[0119] The pharmaceutical combinations described herein may also comprise one or more pharmaceutically acceptable carriers. In one embodiment, the pharmaceutical combination is provided in a unique form such as a pharmaceutical composition or mixture, comprising compounds or agents combined with the same pharmaceutically acceptable carriers for simultaneous, single, or sequential use. Therefore, this document provides a pharmaceutical composition comprising the pharmaceutical combinations described herein.

[0120] The pharmaceutical compositions described herein can be prepared by methods known per se and are suitable for intra-enteral (e.g., oral or rectal) and parenteral administration to mammals (warm-blooded animals), including humans. The compositions comprise a therapeutically effective amount of conjugates of formula (I-1), (I-2), (I-3) and / or (I-4), and at least one therapeutically effective amount of another cancer therapeutic agent, or further combined with one or more pharmaceutically acceptable carriers, particularly suitable for intra-enteral or parenteral administration.

[0121] In an alternative embodiment, the drug combination is provided separately, such as in different compartments within a kit or in different kits, wherein the drugs, conjugates of formulas (I-1), (I-2), (I-3), and / or (I-4), or other cancer therapeutic agents contained therein are independently combined with pharmaceutically acceptable carriers. The pharmaceutically acceptable carriers for each drug can be the same or different, depending on practical needs. Therefore, a kit is also provided herein comprising: (a) conjugates of formulas (I-1), (I-2), (I-3), and / or (I-4) and optionally one or more pharmaceutically acceptable carriers; (b) one or more other cancer therapeutic agents and optionally one or more pharmaceutically acceptable carriers; and (c) instructions for use of (a) and (b). Conjugates of formulas (I-1), (I-2), (I-3), and / or (I-4) and other cancer therapeutic agents are as defined above. Therefore, a kit is provided herein comprising the drug combination described herein.

[0122] In one embodiment, the conjugate contained in the kit is administered at a dose of about 4 mg / kg to about 10 mg / kg every 18 days to every 28 days; for example, every 18 days to every 28 days, at about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 7 mg / kg, about 7.2 mg / kg, about 8 mg / kg, about 8.4 mg / kg, about 9 mg / kg, or about 10 mg / kg (or a range between any two of these values); every 18 days, at about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 7 mg / kg, about 8 mg / kg, about 9 mg / kg, or about 10 mg / kg (or a range between any two of these values); or every 20 days, at about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 7 mg / kg, about 7.2 mg / kg, about 8 mg / kg, about 8.4 mg / kg, about 9 mg / kg, or about 10 mg / kg. mg / kg (or a range between any two of these values); or every 21 days, approximately 4 mg / kg, approximately 5 mg / kg, approximately 6 mg / kg, approximately 7 mg / kg, approximately 7.2 mg / kg, approximately 8 mg / kg, approximately 8.4 mg / kg, approximately 9 mg / kg, or approximately 10 mg / kg (or a range between any two of these values); or every 24 days, approximately 4 mg / kg, approximately 5 mg / kg, approximately 6 mg / kg, approximately 7 mg / kg, approximately 7.2 mg / kg, approximately 8 mg / kg, approximately 8.4 mg / kg, approximately 9 mg / kg, or approximately 10 mg / kg (or a range between any two of these values); or every 28 days, approximately 4 mg / kg, approximately 5 mg / kg, approximately 6 mg / kg, approximately 7 mg / kg, approximately 7.2 mg / kg, approximately 8 mg / kg, approximately 8.4 mg / kg, approximately 9 mg / kg, or approximately 10 mg / kg (or a range between any two of these values). In one embodiment, the conjugate contained in the kit is administered at a dose of about 5 mg / kg to about 9 mg / kg every 21 days. In another embodiment, the conjugate contained in the kit is administered at a dose of about 6 mg / kg to about 8.4 mg / kg every 21 days, for example, about 6 mg / kg, about 7.2 mg / kg, or about 8.4 mg / kg every 21 days. In a very specific embodiment, the conjugate contained in the kit is administered at a dose of about 7.2 mg / kg every 21 days.

[0123] In one embodiment, the conjugate contained in the kit is administered at a dose of about 200 mg to about 700 mg every 18 to 28 days. In another embodiment, the conjugate contained in the kit is administered at a dose of about 200 mg to about 700 mg every 18 days. In another embodiment, the conjugate contained in the kit is administered at a dose of about 200 mg to about 700 mg every 21 days. In another embodiment, the conjugate contained in the kit is administered at a dose of about 200 mg to about 700 mg every 24 days. In yet another embodiment, the conjugate contained in the kit is administered at a dose of about 200 mg to about 700 mg every 28 days. In one embodiment, the conjugate contained in the kit is administered once every 21 days in doses of approximately 200 mg, approximately 260 mg, approximately 300 mg, approximately 360 mg, approximately 400 mg, approximately 452 mg, approximately 460 mg, approximately 500 mg, approximately 504 mg, approximately 560 mg, approximately 600 mg, approximately 630 mg, approximately 660 mg, approximately 700 mg (or a range between any two of these values). In another embodiment, the conjugate contained in the kit is administered once every 21 days in doses of approximately 360 mg, approximately 452 mg, or approximately 504 mg.

[0124] In one embodiment, the coupling comprises the structure shown in formula (i-1) and / or formula (i-2). In some embodiments, the coupling consists of the structures shown in formula (i-1) and formula (i-2).

[0125] In one embodiment, the dosage of other cancer therapeutic agents included in the kit is approximately 100 mg to approximately 600 mg once daily for each treatment cycle of 18 to 30 days; for example, approximately 100 mg to approximately 500 mg, or approximately 200 mg to approximately 400 mg, or approximately 260 mg to approximately 400 mg, or approximately 300 mg to approximately 350 mg once daily for each treatment cycle of 18 days; approximately 100 mg to approximately 500 mg, or approximately 200 mg to approximately 400 mg, or approximately 260 mg to approximately 400 mg, or approximately 300 mg to approximately 350 mg once daily for each treatment cycle of 21 days; or approximately 100 mg to approximately 500 mg, or approximately 200 mg to approximately 400 mg, or approximately 260 mg to approximately 400 mg, or approximately 300 mg to approximately 350 mg once daily for each treatment cycle of 24 days. Approximately 300 mg to approximately 350 mg; once daily for each treatment cycle of 28 days, approximately 100 mg to approximately 500 mg, or approximately 200 mg to approximately 400 mg, or approximately 260 mg to approximately 400 mg, or approximately 300 mg to approximately 350 mg per dose; once daily for each treatment cycle of 30 days, approximately 100 mg to approximately 500 mg, or approximately 200 mg to approximately 400 mg, or approximately 260 mg to approximately 400 mg, or approximately 300 mg to approximately 350 mg per dose; preferably once daily for each treatment cycle of 18 to 30 days, approximately 300 mg to approximately 350 mg per dose; for example, once daily for each treatment cycle of 18 to 30 days, approximately 300 mg, approximately 310 mg, approximately 315 mg, approximately 320 mg, approximately 326 mg, approximately 330 mg, approximately 340 mg, or approximately 350 mg per dose. In one embodiment, the dosage of other cancer therapeutic agents included in the kit is approximately 320 mg once daily for each treatment cycle of 28 days.

[0126] In one embodiment, the other cancer therapeutic agent is a tyrosine kinase inhibitor, a taxane, or a pyrimidine antimetabolite. In one embodiment, the other cancer therapeutic agent is tucatinib, neratinib, pyrotinib, paclitaxel, or capecitabine.

[0127] In one embodiment, the proliferative disease is HER2-positive breast cancer; preferably HER2-positive metastatic breast cancer; and / or

[0128] Subjects requiring the kit must have received prior treatment selected from: trastuzumab, pertuzumab, pyrotinib, capecitabine, palbociclib, letrozole, or combinations thereof; and / or

[0129] The conjugate contained in the kit is administered at a dose of about 5 mg / kg to about 10 mg / kg once every 21 days; preferably at a dose of about 6 mg / kg to about 8.4 mg / kg once every 21 days, for example, about 6 mg / kg once every 21 days, about 7.2 mg / kg once every 21 days, or about 8.4 mg / kg once every 21 days; more preferably, the conjugate contained in the kit is for intravenous administration; and / or

[0130] The other cancer treatment agent is pyrotinib; preferably, the dosage of pyrotinib is about 300 mg to about 350 mg once a day for each 28-day treatment cycle, for example, about 300 mg once a day for each 28-day treatment cycle; about 310 mg once a day for each 28-day treatment cycle; about 320 mg once a day for each 28-day treatment cycle; about 330 mg once a day for each 28-day treatment cycle; about 340 mg once a day for each 28-day treatment cycle; or about 350 mg once a day for each 28-day treatment cycle; preferably, about 320 mg once a day for each 28-day treatment cycle; more preferably, pyrotinib is administered orally.

[0131] In one embodiment, a kit is also provided comprising the conjugate, the other cancer therapeutic agent, and instructions for administering the compound to a patient in need; wherein the effective amount of the conjugate is approximately 5 mg / kg to 10 mg / kg per treatment cycle, and the effective amount of the other cancer therapeutic agent is approximately 100 mg to 600 mg once daily per treatment cycle.

[0132] In one implementation, each treatment cycle of said conjugate application is independently about 1 week, about 2 weeks, about 3 weeks, about 4 weeks, about 5 weeks, about 6 weeks, about 7 weeks, or a range (including endpoints) between any two of these values ​​or any value therein.

[0133] In one implementation, each treatment cycle of the other cancer treatment agent is independently about 1 week, about 2 weeks, about 3 weeks, about 4 weeks, about 5 weeks, about 6 weeks, about 7 weeks, or a range (including endpoints) between any two of these values ​​or any value therein.

[0134] In some implementations, the patient receives multiple (e.g., 2, 3, or 4) treatment cycles. In some implementations, the patient receives treatment until the condition is relieved and treatment is no longer required. In some implementations, the patient receives multiple treatment cycles of the conjugate and the other cancer therapeutic agents.

[0135] In some embodiments, the conjugate and the other cancer therapeutic agent may have the same or different administration cycles. In some embodiments, the conjugate and the other cancer therapeutic agent are each administered in combination as independent dose units. In some embodiments, the conjugate may be administered before, after, or simultaneously with the other cancer therapeutic agent. In some embodiments, the conjugate and the other cancer therapeutic agent are simultaneously administered in combination as combined dose units.

[0136] In the drug combinations described herein, the total ratio of conjugates of formulas (I-1), (I-2), (I-3) and / or (I-4) to one or more other cancer therapeutic agents may vary, for example, to meet the needs of a subgroup of patients being treated, or the needs of an individual patient, which may depend on the specific disease, age, sex, weight, etc.

[0137] Treatment methods and uses

[0138] In another aspect, methods for treating proliferative disorders are provided, the methods comprising administering an effective amount of the pharmaceutical combination described herein, or the kit or pharmaceutical composition described herein, to a subject requiring such treatment. In one embodiment, one or more conjugates and one or more other cancer therapeutic agents are administered simultaneously, separately, or sequentially.

[0139] In another aspect, the use of the drug combination described herein in the preparation of a drug for treating proliferative disorders is provided.

[0140] In another aspect, a method for treating proliferative disorders is provided, the method comprising administering to a subject requiring such treatment an effective amount of a conjugate of formula (I-1), (I-2), (I-3), and / or (I-4), and one or more other cancer therapeutic agents. In an alternative embodiment herein, the conjugate of formula (I-1), (I-2), (I-3), and / or (I-4) is administered simultaneously, sequentially, or separately with one or more other cancer therapeutic agents.

[0141] Conjugates of formulas (I-1), (I-2), (I-3), and / or (I-4) exhibit outstanding anticancer effects and good safety profiles (WO 2015165413A). Therefore, in another aspect, a method for enhancing the efficacy of cancer therapeutics is provided, the method comprising combining conjugates of formulas (I-1), (I-2), (I-3), and / or (I-4) with a cancer therapeutic.

[0142] In another aspect, a method for treating cancers expressing HER2 (HER-2 positive cancers) is provided, the method comprising administering a mixture of conjugates of formula (i-1) and formula (i-2), and other cancer therapeutic agents; wherein the other cancer therapeutic agents are selected from: tyrosine kinase inhibitors, taxanes, and pyrimidine antimetabolites.

[0143] In one embodiment, the tyrosine kinase inhibitor is selected from: tucatinib, neratinib, and pyrotinib; preferably tucatinib or pyrotinib. In one embodiment, the taxane is selected from: docetaxel and paclitaxel, preferably paclitaxel. In one embodiment, the pyrimidine antimetabolite is selected from: cytarabine, capecitabine, ancitabine, gemcitabine, enoxatabine, carmoflu, dideoxyuridine, deoxyfluorouridine, and fluorouridine, preferably capecitabine.

[0144] In one implementation, the cancer expressing HER2 is a HER2-overexpressing cancer.

[0145] In one embodiment, the HER2-overexpressing cancer is a cancer with a HER2 expression score of 2+ in immunohistochemistry.

[0146] In one embodiment, the HER2-overexpressing cancer is a cancer with a HER2 expression score of 3+ in immunohistochemistry.

[0147] In one embodiment, the cancer is resistant to or does not respond to existing anticancer drugs, wherein the existing anticancer drugs include at least one selected from: trastuzumab emtansine, trastuzumab, pertuzumab, lapatinib, irinotecan, cisplatin, carboplatin, oxaliplatin, fluorouracil, gemcitabine, capecitabine, paclitaxel, docetaxel, doxorubicin, epirubicin, cyclophosphamide, mitomycin C, tegafur-gimeracil-oteracil combination drug, cetuximab, panitumumab, bevacizumab, ramucirumab, regorafenib, trifluridine-tipiracil combination drug, gefitinib, erlotinib, afatinib, methotrexate, pyrotinib, palbociclib, and pemetrexed.

[0148] In one implementation, the HER2-positive cancer is selected from: gastric cancer or esophageal cancer, breast cancer, vaginal cancer, salivary gland cancer, endometrial cancer, bladder cancer, non-small cell lung cancer (NSCLC), cervical cancer, ovarian cancer, colorectal cancer, and pancreatic cancer.

[0149] In one embodiment, a method for preventing, alleviating, or treating proliferative disorders, tumors, or cancer is provided, the method comprising administering to a subject in need an effective amount of the conjugate (or formulation) and other cancer therapeutic agents (or formulations); wherein the conjugate is administered at an effective amount of about 5 mg / kg to 10 mg / kg per treatment cycle, and the other cancer therapeutic agents are administered at an effective amount of about 100 mg to 600 mg once daily per treatment cycle.

[0150] In one implementation, each treatment cycle of said conjugate application is independently about 1 week, about 2 weeks, about 3 weeks, about 4 weeks, about 5 weeks, about 6 weeks, about 7 weeks, or a range (including endpoints) between any two of these values ​​or any value therein. In one embodiment, the conjugate is used to be administered at an effective dose of about 4 mg / kg to about 10 mg / kg once every 18 days to every 28 days; for example, about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 7 mg / kg, about 7.2 mg / kg, about 8 mg / kg, about 8.4 mg / kg, about 9 mg / kg, or about 10 mg / kg (or a range between any two of these values) once every 18 days to every 28 days; or about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 7 mg / kg, about 8 mg / kg, about 9 mg / kg, or about 10 mg / kg (or a range between any two of these values) once every 18 days; or about 4 mg / kg, about 5 mg / kg, about 6 mg / kg, about 7 mg / kg, about 7.2 mg / kg, about 8 mg / kg, about 8.4 mg / kg, about 9 mg / kg, or about 10 mg / kg once every 20 days. kg (or a range between any two of these values); or approximately 4 mg / kg, approximately 5 mg / kg, approximately 6 mg / kg, approximately 7 mg / kg, approximately 7.2 mg / kg, approximately 8 mg / kg, approximately 8.4 mg / kg, approximately 9 mg / kg, or approximately 10 mg / kg (or a range between any two of these values) every 21 days; or approximately 4 mg / kg, approximately 5 mg / kg, approximately 6 mg / kg, approximately 7 mg / kg, approximately 7.2 mg / kg, approximately 8 mg / kg, approximately 8.4 mg / kg, approximately 9 mg / kg, or approximately 10 mg / kg (or a range between any two of these values) every 24 days; or approximately 4 mg / kg, approximately 5 mg / kg, approximately 6 mg / kg, approximately 7 mg / kg, approximately 7.2 mg / kg, approximately 8 mg / kg, approximately 8.4 mg / kg, approximately 9 mg / kg, or approximately 10 mg / kg (or a range between any two of these values) every 28 days.

[0151] In one embodiment, the conjugate is administered at an effective amount of about 5 mg / kg to about 9 mg / kg every 21 days. In another embodiment, the conjugate is administered at an effective amount of about 6 mg / kg to about 8.4 mg / kg every 21 days, for example, about 6 mg / kg, about 7.2 mg / kg, or about 8.4 mg / kg every 21 days. In a very specific embodiment, the conjugate is administered at an effective amount of about 7.2 mg / kg every 21 days.

[0152] In one embodiment, the conjugate is administered at an effective dose of about 200 mg to about 700 mg every 18 to 28 days. In another embodiment, the conjugate is administered at an effective dose of about 200 mg to about 700 mg every 18 days. In another embodiment, the conjugate is administered at an effective dose of about 200 mg to about 700 mg every 21 days. In another embodiment, the conjugate is administered at an effective dose of about 200 mg to about 700 mg every 24 days. In yet another embodiment, the conjugate is administered at an effective dose of about 200 mg to about 700 mg every 28 days. In one embodiment, the conjugate is administered once every 18 days to every 28 days, for example, once every 21 days at an effective amount of about 200 mg, about 260 mg, about 300 mg, about 360 mg, about 400 mg, about 452 mg, about 460 mg, about 500 mg, about 504 mg, about 560 mg, about 600 mg, about 630 mg, about 660 mg, about 700 mg (or a range between any two of these values). In one embodiment, the conjugate is administered once every 21 days at an effective amount of about 360 mg, about 452 mg, or about 504 mg.

[0153] In one embodiment, the coupling comprises the structure shown in formula (i-1) and / or formula (i-2). In some embodiments, the coupling consists of the structures shown in formula (i-1) and formula (i-2).

[0154] In one implementation, each treatment cycle of the other cancer treatment agent is independently about 1 week, about 2 weeks, about 3 weeks, about 4 weeks, about 5 weeks, about 6 weeks, about 7 weeks, or a range (including endpoints) between any two of these values ​​or any value therein. In one embodiment, the other cancer therapeutic agent is administered once daily at an effective dose of about 100 mg to about 600 mg for each treatment cycle of 18 to 30 days; for example, about 100 mg to about 500 mg, or about 200 mg to about 400 mg, or about 260 mg to about 400 mg, or about 300 mg to about 350 mg once daily for each treatment cycle of 18 days; about 100 mg to about 500 mg, or about 200 mg to about 400 mg, or about 260 mg to about 400 mg, or about 300 mg to about 350 mg once daily for each treatment cycle of 21 days; or about 100 mg to about 500 mg, or about 200 mg to about 400 mg, or about 260 mg to about 400 mg, or about 300 mg to about 350 mg once daily for each treatment cycle of 24 days; or about 100 mg to about 500 mg, or about 200 mg to about 400 mg, or about 260 mg to about 400 mg once daily for each treatment cycle of 30 days; or about 100 mg to about 500 mg, or about 200 mg to about 400 mg, or about 300 mg to about 350 mg once daily for each treatment cycle of 24 days. From 0 mg to about 350 mg; once daily for each 28-day treatment cycle, about 100 mg to about 500 mg, or about 200 mg to about 400 mg, or about 260 mg to about 400 mg, or about 300 mg to about 350 mg; once daily for each 30-day treatment cycle, about 100 mg to about 500 mg, or about 200 mg to about 400 mg, or about 260 mg to about 400 mg, or about 300 mg to about 350 mg; preferably once daily for each 18- to 30-day treatment cycle, about 300 mg to about 350 mg; for example, once daily for each 18- to 30-day treatment cycle, about 300 mg, about 310 mg, about 315 mg, about 320 mg, about 326 mg, about 330 mg, about 340 mg, or about 350 mg. In one embodiment, the other cancer treatment agent is administered at an effective dose of about 320 mg once daily for each 28-day treatment cycle.

[0155] In one embodiment, the other cancer therapeutic agent is a tyrosine kinase inhibitor, a taxane, or a pyrimidine antimetabolite. In one embodiment, the other cancer therapeutic agent is tucatinib, neratinib, pyrotinib, paclitaxel, or capecitabine. In one embodiment, the proliferative disease is HER2-positive breast cancer; preferably HER2-positive metastatic breast cancer; and / or

[0156] The subject had received prior treatment selected from the following: trastuzumab, pertuzumab, pyrotinib, capecitabine, palbociclib, letrozole, or a combination thereof.

[0157] In some implementations, the patient receives multiple treatment cycles (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, or 14). In some implementations, the patient receives treatment until the condition is relieved and treatment is no longer required. In some implementations, the patient receives multiple treatment cycles of the conjugate and the other cancer therapeutic agents.

[0158] In some implementations, the conjugate and the other cancer therapeutic agents may have the same or different administration cycles.

[0159] In some embodiments, the conjugate and the other cancer therapeutic agent are each administered in combination as independent dose units. In some embodiments, the conjugate may be administered before, after, or simultaneously with the other cancer therapeutic agent. In some embodiments, the conjugate and the other cancer therapeutic agent are simultaneously administered in combination as a combined dose unit.

[0160] In another aspect, the use of conjugates of formulas (I-1), (I-2), (I-3) and / or (I-4) in the preparation of medicaments for the prevention, relief or treatment of proliferative disorders of cells is provided, wherein the medicament is used in combination with one or more cancer therapeutic agents.

[0161] In one embodiment, the coupling agent is a coupling agent of formula (I-1) and / or (I-2). In another embodiment, the coupling agent is a coupling agent of formula (I-3) and / or (I-4). In one embodiment, the coupling agent is a coupling agent of formula (i-1) and / or (i-2). In one embodiment, the coupling agent is a mixture of formula (i-1) and (i-2).

[0162] In one embodiment, the proliferative disorder is a proliferative disorder associated with wild-type or mutant HER2, or a proliferative disorder with HER2 (ErbB2) amplification or HER2 protein overexpression. In one embodiment, the proliferative disorder is cancer. In one embodiment, the cancer is HER2-positive cancer; wherein, preferably, the HER2 expression level is IHC 2+ or 3+, particularly 3+. In one embodiment, the proliferative disorder is a cancer associated with wild-type or mutant HER2, or a proliferative disorder with HER2 (ErbB2) amplification or HER2 protein overexpression. In one embodiment, the proliferative disorder is characterized by increased proliferation and survival of HER2-positive tumor cells, particularly human breast cancer cells or human gastric cancer cells. In some embodiments, the cancer is primary or secondary drug-resistant.

[0163] In one embodiment, the proliferative lesion is a cancerous tumor. In one embodiment, the cancer is selected from: gastric cancer or esophageal cancer, breast cancer, vaginal cancer, salivary gland cancer, endometrial cancer, bladder cancer, non-small cell lung cancer (NSCLC), cervical cancer, ovarian cancer, colorectal cancer, and pancreatic cancer. In a preferred embodiment, the cancer is gastric cancer or esophageal cancer, breast cancer, vaginal cancer, colorectal cancer, non-small cell lung cancer, and ovarian cancer. In a specific embodiment, the cancer is gastric cancer or esophageal cancer or breast cancer. In a very specific embodiment, the cancer is breast cancer. In one embodiment, the breast cancer is a breast tumor.

[0164] In one embodiment, the proliferative disease is HER2-positive breast cancer; preferably HER2-positive metastatic breast cancer.

[0165] In one embodiment, the drug combination described herein is used as first-line treatment. In one embodiment, the drug combination described herein is used as second-line or later-line treatment. In one embodiment, the prior treatment includes antibody therapy, chemotherapy, and / or kinase inhibitor therapy; preferably antibody therapy and / or chemotherapy. In one embodiment, the prior treatment is selected from: trastuzumab, pertuzumab, tucatinib, neratinib, pyrotinib, paclitaxel, capecitabine, palbociclib, letrozole, or combinations thereof. In one embodiment, the prior treatment includes antibody therapy, particularly treatment using trastuzumab plus pertuzumab. In one embodiment, the prior treatment includes chemotherapy, particularly palbociclib therapy. In one embodiment, the prior treatment includes tyrosine kinase inhibitor therapy, particularly pyrotinib therapy. In one embodiment, the drug combination described herein is used as neoadjuvant therapy. In one embodiment, the drug combination described herein is used as adjuvant therapy. In one embodiment, the proliferative disease is cancer. In one embodiment, the cancer is breast cancer or gastric cancer, and the subject requiring such treatment has received prior treatment. In one embodiment, the subject requiring such treatment is resistant to one or more of the prior treatments. In one embodiment, the prior treatment is selected from: trastuzumab, pertuzumab, tucatinib, neratinib, pyrotinib, paclitaxel, capecitabine, palbociclib, letrozole, or combinations thereof; preferably, the prior treatment is selected from: trastuzumab, pertuzumab, pyrotinib, capecitabine, palbociclib, letrozole, or combinations thereof; more preferably, the prior treatment is selected from: trastuzumab, pertuzumab, pyrotinib, palbociclib, or combinations thereof. In one embodiment, the prior treatment includes treatment with trastuzumab, pertuzumab, pyrotinib, and capecitabine. In another embodiment, the prior treatment includes treatment with palbociclib and letrozole.

[0166] In one embodiment, the treatment methods and uses described herein prolong overall survival (OS). In one embodiment, the drug combinations described herein produce a synergistic effect, such as a synergistic effect on tumor suppression. In one embodiment, the treatment methods and uses described herein do not result in an increase in side effects compared to the corresponding monotherapy. In one embodiment, the treatment methods and uses described herein improve overall response rate (ORR), complete response (CR), or partial response (PR) compared to the corresponding monotherapy. In one embodiment, the treatment methods and uses described herein prolong progression-free survival (PFS). In one embodiment, the treatment methods and uses described herein improve tumor growth inhibition rate (TGI).

[0167] Regarding the treatment methods and uses described in this article, the following implementation methods for other cancer therapeutic agents are covered and illustrated. ● These other cancer therapeutic agents are selected from: kinase inhibitors and chemotherapeutic agents. The cell proliferation condition is, for example, a cancerous tumor.

[0168] ●The other cancer treatment agents are selected from: tyrosine kinase inhibitors, taxanes, and pyrimidine antimetabolites. The proliferative disease is, for example, gastric cancer, esophageal cancer, or breast cancer.

[0169] ●The other cancer treatment agents mentioned are selected from: HER family tyrosine kinase inhibitors. The cell proliferation disease mentioned is, for example, gastric cancer, esophageal cancer, or breast cancer.

[0170] ●The other cancer treatment agents mentioned are selected from: tucatinib, neratinib, pyrotinib, paclitaxel, and capecitabine. The proliferative disease mentioned is, for example, breast cancer.

[0171] ●The other cancer treatment agents mentioned are selected from: tucatinib and pyrotinib. The proliferative disease mentioned is, for example, gastric cancer, esophageal cancer, or breast cancer.

[0172] ● The other cancer treatment agent is pyrotinib. The proliferative disease is, for example, breast cancer. In one embodiment, the breast cancer is metastatic, recurrent, refractory, or advanced. In one embodiment, the subject requiring treatment for breast cancer is resistant to antibody therapy, particularly trastuzumab plus pertuzumab therapy. Preferably, this resistance is acquired after trastuzumab plus pertuzumab therapy. In one embodiment, the subject requiring treatment for breast cancer is resistant to kinase inhibitor therapy, preferably CDK kinase inhibitor (e.g., CDK4 / 6 inhibitor) therapy, particularly palbociclib therapy. In one embodiment, the subject requiring treatment for breast cancer is resistant to a combination of trastuzumab, pertuzumab, pyrotinib, and capecitabine. In one embodiment, the subject requiring treatment for breast cancer is resistant to a combination of palbociclib and letrozole.

[0173] ●The other cancer treatment agents mentioned are pyrotinib or tucatinib. The proliferative disease mentioned is, for example, gastric cancer or esophageal gastric cancer. In one embodiment, a subject requiring treatment for gastric cancer is resistant to a combination of trastuzumab, pertuzumab, pyrotinib, and capecitabine. In one embodiment, a subject requiring treatment for breast cancer is resistant to a combination of palbociclib and letrozole.

[0174] ● The other cancer treatment agent is tucatinib. The proliferative disease is, for example, breast cancer. In one embodiment, the breast cancer is metastatic, recurrent, refractory, or advanced. In one embodiment, the breast cancer is metastatic. In one embodiment, the subject requiring treatment for breast cancer is resistant to antibody therapy, particularly trastuzumab plus pertuzumab therapy. Preferably, this resistance is acquired after trastuzumab plus pertuzumab treatment. In one embodiment, the subject requiring treatment for breast cancer is resistant to kinase inhibitor therapy, preferably CDK kinase inhibitor (e.g., CDK4 / 6 inhibitor) therapy, particularly palbociclib therapy. In one embodiment, the subject requiring treatment for breast cancer is resistant to pyrotinib therapy. In one embodiment, the subject requiring treatment for breast cancer is resistant to a combination of trastuzumab, pertuzumab, pyrotinib, and capecitabine. In one embodiment, the subject requiring treatment for breast cancer is resistant to a combination of palbociclib and letrozole.

[0175] ●The other cancer treatment agent mentioned is tucatinib. The proliferative disease mentioned is, for example, gastric cancer or esophageal cancer.

[0176] ●The other cancer treatment agents are selected from taxanes. The proliferative disease is, for example, breast cancer. In one embodiment, the taxane is paclitaxel.

[0177] ●The other cancer treatment agents are selected from: pyrimidine antimetabolites. The proliferative disease is, for example, breast cancer. In one embodiment, the pyrimidine antimetabolite is capecitabine.

[0178] Beneficial effects

[0179] The drug combination described in this paper achieves at least one of the following technical effects:

[0180] (1) Good anti-cancer effects, especially in improving overall response rate (ORR), complete response (CR), partial response (PR), or tumor growth inhibition rate (TGI).

[0181] (2) Good physicochemical properties (e.g., solubility, physical and / or chemical stability).

[0182] (3) Good pharmacokinetic properties (e.g., good stability in plasma, appropriate half-life and duration of action).

[0183] (4) Good safety profile (low toxicity to non-target normal cells or tissues, and / or fewer side effects, wider therapeutic window), etc.

[0184] (5) It can be used to prevent, alleviate or treat subjects who are resistant to antibody therapy, especially trastuzumab plus pertuzumab therapy.

[0185] (6) It can be used to treat subjects who are resistant to kinase inhibitor therapy, preferably CDK kinase inhibitor therapy, especially palbociclib therapy.

[0186] Example

[0187] Preparation Examples

[0188] To more clearly illustrate the objectives and technical solutions, the following description further illustrates specific embodiments. It should be understood that these embodiments are not intended to limit the scope of this document. Specific experimental methods not mentioned in the following embodiments were performed according to conventional experimental methods.

[0189] abbreviation

[0190] CR (Complete Remission); PR (Partial Remission); SD (Stable Disease); PD (Progressive Disease); OS (Overall Survival); PFS (Progression-Free Survival); IHC (Immunohistochemistry); DAR (Drug-Antibody Ratio); TGI (Tumor Growth Inhibition Rate).

[0191] Instruments, materials and reagents

[0192] Unless otherwise stated, the instruments and reagents used in the examples are commercially available. The reagents can be used directly without further purification.

[0193] In HER2 testing via IHC staining, the IHC antibodies used are typically non-mutation-specific antibodies.

[0194] GQ01 is a mixture of conjugates of formula (i-1) and (i-2), wherein the structures of formulas (i-1) and (i-2) are as described above. The total drug-antibody ratio (DAR) of GQ01 was determined to be approximately 1.8 using HIC-HPLC with a butyl column.

[0195] Example 1: In vivo efficacy evaluation of the combination of GQ01 and TKI in a human breast cancer PDX model.

[0196] Experimental objective: To evaluate the in vivo antitumor efficacy of the combination of ADC drug GQ01 and TKI in mice carrying a human breast cancer HER2 IHC 3+PDX model.

[0197] I. Tumor tissue from the PDX model was extracted from tumor-bearing mice, transferred to culture medium, cut into small pieces, and subcutaneously injected into the right scapula of 6-8 week old SPF-grade female NOD SCID mice.

[0198] II. Measure the tumor diameter with calipers and calculate it according to the formula V = 0.5a × b. 2 Calculate the tumor volume (where a is the long axis of the tumor and b is the short axis). The average tumor volume is approximately 100 mm². 3 -300mm 3 Mice were randomly assigned to the following groups: solvent group, GQ01 5 mg / kg group, tucatinib 20 mg / kg group, neratinib 20 mg / kg group, pyrotinib 20 mg / kg group, GQ01 + tucatinib combination group, GQ01 + neratinib combination group, and GQ01 + pyrotinib combination group. Mice in the solvent group were administered GQ01 solvent and pyrotinib solvent at the same frequency and via the same route. Tumor volume was measured twice weekly in each group. The experiment ended on day 77, and the tumor growth inhibition rate (TGI) was calculated as follows: TGI(%) = [1 - (mean tumor volume of the treatment group on the end day - mean tumor volume of the treatment group on day 1) / (mean tumor volume of the solvent group on the end day - mean tumor volume of the solvent group on day 1)] × 100%.

[0199] III. Figure 1a Tumor volume changes in tumor-bearing NOD SCID female mice treated with the following: (1) solvent, (2) GQ01 5 mg / kg (Q3W×2), (3) tucatinib 20 mg / kg (BID×2W+BID×2W), (4) GQ01 5 mg / kg (Q3W×2) + tucatinib 20 mg / kg (BID×2W+BID×2W). Table 1a shows that on the end day (day 77), the mean tumor volume was 950 mmHg in the GQ01 5 mg / kg group, the tucatinib 20 mg / kg group, and the GQ01 5 mg / kg + tucatinib 20 mg / kg group. 3 879mm 3 307mm 3 The TGI values ​​were 46.22%, 51.08%, and 91.20%, respectively.

[0200] These data indicate that both GQ01 5 mg / kg and Tucatinib 20 mg / kg, when used as monotherapy, can inhibit tumor growth, while the combination of GQ01 and Tucatinib has better anti-tumor efficacy than monotherapy.

[0201] Table 1a. Tumor growth inhibition based on tumor volume in a human breast cancer PDX model using the combination of GQ01 and tucatinib.

[0202]

[0203] a. Mean ± SEM; measured on the last day;

[0204] b. TGI(%)=[1-(T 77 -T0) / (V 77 -V0)]×100%. T0 is the mean tumor volume in the treatment group on the first day of administration, T 77 V0 is the mean tumor volume in the treatment group on day 77 after administration; V0 is the mean tumor volume in the solvent group on day 1 after administration. 77 This represents the average tumor volume in the solvent group on day 77 after application.

[0205] IV. Figure 1b Tumor volume changes in tumor-bearing NOD SCID female mice treated with the following: (1) solvent, (2) GQ01 5 mg / kg (Q3W×2), (3) neratinib 20 mg / kg (QD×2W+QD×2W), and (4) GQ01 5 mg / kg (Q3W×2) + neratinib 20 mg / kg (QD×2W+QD×2W). Table 1b shows that the mean tumor volume at the end of day 77 was 950 mmHg in the GQ01 5 mg / kg group, the neratinib 20 mg / kg group, and the GQ01 5 mg / kg + neratinib 20 mg / kg group. 3 724mm 3 220mm 3 The TGI values ​​were 46.22%, 61.80%, and 97.91%, respectively.

[0206] These data indicate that both GQ01 5 mg / kg and neratinib 20 mg / kg, when used as monotherapy, can inhibit tumor growth. Compared with the monotherapy group, the combination of GQ01 and neratinib showed superior tumor suppression efficacy and could more effectively inhibit tumor growth.

[0207] Table 1b. Tumor growth inhibition based on tumor volume in a human breast cancer PDX model using the combination of GQ01 and neratinib.

[0208]

[0209] a. Mean ± SEM; measured on the last day;

[0210] b. TGI(%)=[1-(T77 -T0) / (V 77 -V0)]×100%. T0 is the mean tumor volume in the treatment group on day 1 of administration, and T 77 V0 is the mean tumor volume in the treatment group on day 77 after administration; V0 is the mean tumor volume in the solvent group on day 1 after administration. 77 This represents the average tumor volume in the solvent group on day 77 after application.

[0211] V. Figure 1c Tumor volume changes in tumor-bearing NOD SCID female mice treated with the following: (1) solvent, (2) GQ01 5 mg / kg (Q3W×2), (3) pyrotinib 20 mg / kg (QD×2W+QD×2W), and (4) GQ01 5 mg / kg (Q3W×2) + pyrotinib 20 mg / kg (QD×2W+QD×2W). Table 1c shows that the mean tumor volume at the end of day 77 was 950 mmHg in the GQ01 5 mg / kg group, the pyrotinib 20 mg / kg group, and the GQ01 5 mg / kg + pyrotinib 20 mg / kg group. 3 1108mm 3 202mm 3 The TGI values ​​were 46.22%, 34.93%, and 98.74%, respectively.

[0212] These data indicate that both GQ01 5 mg / kg and pyrotinib 20 mg / kg, when used as monotherapy, can inhibit tumor growth to some extent; the combination of GQ01 and pyrotinib significantly improves anti-tumor efficacy compared to GQ01 or pyrotinib monotherapy.

[0213] Table 1c. Tumor growth inhibition based on tumor volume in a human breast cancer PDX model using the combination of GQ01 and pyrotinib.

[0214]

[0215] a. Mean ± SEM; measured on the last day;

[0216] b. TGI(%)=[1-(T 77 -T0) / (V 77 -V0)]×100%. T0 is the mean tumor volume in the treatment group on day 1 of administration, and T 77 V0 is the mean tumor volume in the treatment group on day 77 after administration; V0 is the mean tumor volume in the solvent group on day 1 after administration. 77 This represents the average tumor volume in the solvent group on day 77 after application.

[0217] The results showed that in tumor-bearing mice (HER2-positive (IHC 3+) human breast cancer PDX model), GQ01 or TKI monotherapy could inhibit tumor cell proliferation with varying degrees of efficacy. Furthermore, the combination of GQ01 with tucatinib, neratinib, or pyrotinib significantly enhanced the antitumor efficacy.

[0218] Example 2: In vivo efficacy evaluation of the combination of GQ01 and TKI in a human breast cancer PDX model.

[0219] Objective: To evaluate the in vivo antitumor efficacy of the combination ADC drug GQ01 and a TKI in mice carrying a HER2 IHC 3+ PDX model of breast cancer. This model was derived from patients with stage III invasive breast cancer who developed lung and bone metastases post-surgery. After treatment with palbociclib and letrozole, the disease was classified as progressive (PD), and the model was identified as exhibiting initial resistance to palbociclib.

[0220] I. Tumor tissue from the PDX model was removed from tumor-bearing mice, transferred to culture medium, cut into small pieces, and subcutaneously injected into the right scapula of 6-8 week old SPF-grade female NCG mice.

[0221] II. Measure the tumor diameter with calipers 30 days after inoculation, and calculate the value according to the formula V = 0.5a × b. 2 Calculate the tumor volume (where a is the long axis of the tumor and b is the short axis). The average tumor volume is approximately 100 mm². 3 -300mm 3 Mice were randomly assigned to four groups: solvent group, GQ01 5 mg / kg group, tucatinib 50 mg / kg group, pyrotinib 20 mg / kg group, GQ01 + tucatinib combination group, and GQ01 + pyrotinib combination group. Each group consisted of 6 mice. The day of the first administration was defined as day 0. Mice in the solvent group were administered GQ01 solvent and tucatinib solvent at the same frequency and via the same route. Tumor volume was measured twice weekly. The experiment ended on day 49, and the tumor growth inhibition rate (TGI) was calculated as follows: TGI(%) = [1 - (mean tumor volume of the treatment group on the end day - mean tumor volume of the treatment group on day 1) / (mean tumor volume of the solvent group on the end day - mean tumor volume of the solvent group on day 1)] × 100%.

[0222] III. Figure 2aTumor volume changes in tumor-bearing female NCG mice treated with the following: (1) solvent, (2) GQ01 5 mg / kg (days 0 and 28), (3) tucatinib 50 mg / kg (QD, days 0–13, 28–41), and (4) GQ01 5 mg / kg (days 0 and 28) + tucatinib 50 mg / kg (QD, days 0–13, 28–41). Table 2a shows that the mean tumor volume at the end of day (day 49) was 648 mm in the GQ01 5 mg / kg group, the tucatinib 50 mg / kg group, and the GQ01 5 mg / kg + tucatinib 50 mg / kg group. 3 732mm 3 485mm 3 The TGI values ​​were 12.92%, -1.09%, and 40.16%, respectively.

[0223] The results showed that the combination of GQ01 and tucatinib significantly inhibited the growth of the tumor compared with monotherapy.

[0224] Table 2a. Tumor growth inhibition based on tumor volume in a human breast cancer PDX model using the combination of GQ01 and tucatinib.

[0225]

[0226] a. Mean ± SEM; measured on the last day;

[0227] b. TGI(%)=[1-(T 49 -T0) / (V 49 -V0)]×100%. T0 is the mean tumor volume in the treatment group on day 1 of administration, and T 49 V0 is the mean tumor volume in the treatment group on day 49 after administration; V0 is the mean tumor volume in the solvent group on day 1 after administration. 49 This represents the average tumor volume in the solvent group on day 49 after application.

[0228] IV. Figure 2b Tumor volume changes in tumor-bearing female NCG mice treated with the following: (1) solvent, (2) GQ01 5 mg / kg (days 0 and 28), (3) pyrotinib 20 mg / kg (QD, days 0–13, 28–41), and (4) GQ01 5 mg / kg (days 0 and 28) + pyrotinib 20 mg / kg (QD, days 0–13, 28–41). Table 2b shows that the mean tumor volume at the end of day (day 49) was 648 mm in the GQ01 5 mg / kg group, the pyrotinib 20 mg / kg group, and the GQ01 5 mg / kg + pyrotinib 20 mg / kg group. 3741mm 3 338mm 3 The TGI values ​​were 12.92%, -2.65%, and 64.71%, respectively.

[0229] The results showed that the combination of GQ01 and pyrotinib was more effective in inhibiting tumor growth than GQ01 5 mg / kg or pyrotinib 20 mg / kg monotherapy.

[0230] Table 2b. Tumor growth inhibition based on tumor volume in a human breast cancer PDX model using the combination of GQ01 and pyrotinib.

[0231]

[0232] a. Mean ± SEM; measured on the last day;

[0233] b. TGI(%)=[1-(T 49 -T0) / (V 49 -V0)]×100%. T0 is the mean tumor volume in the treatment group on day 1 of administration, and T 49 V0 is the mean tumor volume in the treatment group on day 49 after administration; V0 is the mean tumor volume in the solvent group on day 1 after administration. 49 This represents the average tumor volume in the solvent group on day 49 after application.

[0234] The results showed that in a metastatic and drug-resistant human breast cancer PDX model, TKIs as monotherapy failed to inhibit tumor growth, while the combination of GQ01 with tucatinib or pyrotinib significantly improved antitumor efficacy. The combination of the ADC drug GQ01 with TKIs can benefit these patients.

[0235] Example 3: In vivo efficacy evaluation of the combination of GQ01 and TKI in a human breast cancer PDX model.

[0236] Objective: To evaluate the in vivo antitumor efficacy of the combination of the ADC drug GQ01 and a TKI in mice carrying a human breast cancer HER2 IHC 3+ PDX model. This model was derived from patients with stage III invasive breast cancer who were assessed as having acquired resistance to trastuzumab and pertuzumab based on their treatment history, and initial resistance to pyrotinib.

[0237] I. Tumor tissue from the PDX model was removed from tumor-bearing mice, transferred to culture medium, cut into small pieces, and then subcutaneously injected into the right scapula of 6-8 week old SPF-grade female NCG mice.

[0238] II. Measure the tumor diameter with calipers 29 days after inoculation, and calculate the value according to the formula V = 0.5a × b. 2Calculate the tumor volume (where a is the long axis of the tumor and b is the short axis). The average tumor volume is approximately 100 mm². 3 -300mm 3 Mice were randomly assigned to four groups: solvent group, GQ01 3 mg / kg, 5 mg / kg, tucatinib 50 mg / kg, pyrotinib 20 mg / kg, GQ01 + tucatinib combination group, and GQ01 + pyrotinib combination group. Each group consisted of 6 mice. The day of the first administration was defined as day 0. Mice in the solvent group were administered GQ01 and tucatinib solvents at the same frequency and via the same route. Tumor volume was measured twice weekly in each group. The experiment ended on day 42, and the tumor growth inhibition rate (TGI) was calculated as follows: TGI(%) = [1 - (mean tumor volume of the treatment group on the end day - mean tumor volume of the treatment group on day 1) / (mean tumor volume of the solvent group on the end day - mean tumor volume of the solvent group on day 1)] × 100%.

[0239] III. Figure 3a Tumor volume changes in tumor-bearing mice treated with the following are shown: (1) solvent, (2) GQ01 3 mg / kg / 5 mg / kg (3 mg / kg on day 0 and 5 mg / kg on day 21), (3) tucatinib 50 mg / kg (QD, days 0-13, 21-34), and (4) GQ01 3 mg / kg / 5 mg / kg (3 mg / kg on day 0 and 5 mg / kg on day 21) + tucatinib 50 mg / kg (QD, days 0-13, 21-34). Table 3a shows that on the end day (day 42), the mean tumor volume was 920 mm in the GQ01 3 mg / kg / 5 mg / kg group, the tucatinib 50 mg / kg group, and the GQ01 3 mg / kg / 5 mg / kg + tucatinib 50 mg / kg group. 3 1558mm 3 857mm 3 The TGI values ​​were 40.97%, -6.46%, and 45.44%, respectively.

[0240] Based on changes in tumor volume and TGI, GQ01 monotherapy can inhibit tumor growth to some extent. The model was unresponsive to tucatinib treatment, and the combination of GQ01 and tucatinib showed better anti-tumor efficacy.

[0241] Table 3a. Tumor growth inhibition based on tumor volume in the combination of GQ01 and tucatinib in a human breast cancer PDX model.

[0242]

[0243] a. Mean ± SEM; measured on the last day;

[0244] b. TGI(%)=[1-(T 42 -T0) / (V 42 -V0)]×100%. T0 is the mean tumor volume in the treatment group on day 1 of administration, and T 42 V0 is the mean tumor volume in the treatment group on day 42 after administration; V0 is the mean tumor volume in the solvent group on day 1 after administration. 42 This represents the average tumor volume in the solvent group on day 42 after application.

[0245] IV. Figure 3b Tumor volume changes in tumor-bearing mice treated with the following are shown: (1) solvent, (2) GQ01 3 mg / kg / 5 mg / kg (3 mg / kg on day 0 and 5 mg / kg on day 21), (3) pyrotinib 20 mg / kg (QD, days 0-13, 21-34), and (4) GQ01 3 mg / kg / 5 mg / kg (3 mg / kg on day 0 and 5 mg / kg on day 21) + pyrotinib 20 mg / kg (QD, days 0-13, 21-34). Table 3b shows that on the end day (day 42), the mean tumor volume was 920 mm in the GQ01 3 mg / kg / 5 mg / kg group, the pyrotinib 20 mg / kg group, and the GQ01 3 mg / kg / 5 mg / kg + pyrotinib 20 mg / kg group. 3 1333mm 3 634mm 3 The TGI values ​​were 40.97%, 10.08%, and 62.09%, respectively.

[0246] Based on changes in tumor volume and TGI, GQ01 monotherapy demonstrated tumor-suppressing efficacy. In this pyrotinib-resistant breast cancer model, the combination of GQ01 and pyrotinib significantly inhibited tumor growth, demonstrating better anti-tumor efficacy than monotherapy.

[0247] Table 3b. Tumor growth inhibition based on tumor volume in a human breast cancer PDX model using the combination of GQ01 and pyrotinib.

[0248]

[0249] a. Mean ± SEM; measured on the last day;

[0250] b. TGI(%)=[1-(T 42 -T0) / (V 42 -V0)]×100%. T0 is the mean tumor volume in the treatment group on day 1 of administration, and T 42V0 is the mean tumor volume in the treatment group on day 42 after administration; V0 is the mean tumor volume in the solvent group on day 1 after administration. 42 This represents the average tumor volume in the solvent group on day 42 after application.

[0251] The results showed that in a TKI-resistant HER2-positive (IHC 3+) breast cancer model, GQ01 monotherapy tended to improve efficacy compared with solvent therapy, and the combination of GQ01 and TKI showed stronger tumor suppression.

[0252] Example 4: In vivo efficacy evaluation of the combination of GQ01 and TKI in a human gastric cancer PDX model.

[0253] Experimental Objective: To evaluate the in vivo antitumor efficacy of the combination of ADC drug GQ01 and TKI in mice carrying a HER2 IHC 3+ PDX model of gastric cancer.

[0254] I. Tumor tissue from the PDX model was extracted from tumor-bearing mice, transferred to RPMI-1640 medium, cut into small pieces, and then subcutaneously injected into the right scapula of 6-8 week old SPF-grade female Nu / Nu mice.

[0255] II. Measure the tumor diameter with calipers 14 days after inoculation, and calculate the value according to the formula V = 0.5a × b. 2 Calculate the tumor volume (where a is the long axis of the tumor and b is the short axis). The average tumor volume is approximately 100 mm². 3 -300mm 3 Mice were randomly assigned to four groups: solvent group, GQ01 10 mg / kg group, tucatinib 50 mg / kg group, pyrotinib 20 mg / kg group, GQ01 + tucatinib combination group, and GQ01 + pyrotinib combination group. Each group consisted of 6 mice. The day of the first administration was defined as day 0. Mice in the solvent group were administered both GQ01 solvent and pyrotinib solvent at the same frequency and via the same route of administration. Tumor volume was measured twice weekly in each group. The experiment ended on day 35, and the tumor growth inhibition rate (TGI) was calculated as follows: TGI(%) = [1 - (mean tumor volume of the treatment group on the end day - mean tumor volume of the treatment group on day 1) / (mean tumor volume of the solvent group on the end day - mean tumor volume of the solvent group on day 1)] × 100%.

[0256] III. Figure 4aTumor volume changes in tumor-bearing mice treated with the following are shown: (1) solvent, (2) GQ01 10 mg / kg (single dose), (3) tucatinib 50 mg / kg (QD, 2 weeks), and (4) GQ01 10 mg / kg (single dose) + tucatinib 50 mg / kg (QD, 2 weeks). Table 4a shows that on the end day (day 35), the mean tumor volume was 255 mm in the GQ01 10 mg / kg group, the tucatinib 50 mg / kg group, and the GQ01 10 mg / kg + tucatinib 50 mg / kg group. 3 400mm 3 107mm 3 The TGI values ​​were 78.83%, 48.94%, and 108.94%, respectively.

[0257] Tumor volume changes and TGI rates indicated that both GQ01 10 mg / kg and tucatinib 50 mg / kg, when used as monotherapy, could inhibit tumor growth to some extent. The combination of GQ01 and tucatinib was significantly superior to either GQ01 or tucatinib monotherapy and could induce 1 / 6 PR and even 1 / 6 CR.

[0258] Table 4a. Tumor growth inhibition based on tumor volume in a human gastric cancer PDX model using the combination of GQ01 and tucatinib.

[0259]

[0260] a. Mean ± SEM; measured on the last day;

[0261] b. TGI(%)=[1-(T 35 -T0) / (V 35 -V0)]×100%. T0 is the mean tumor volume in the treatment group on day 1 of administration, and T 35 V0 is the mean tumor volume in the treatment group on day 35 after administration; V0 is the mean tumor volume in the solvent group on day 1 after administration. 35 This represents the average tumor volume in the solvent group on day 35 after application.

[0262] Remission status was calculated based on the diameter or disappearance of the target lesion. Results are provided in the table below:

[0263]

[0264] IV. Figure 4bTumor volume changes in tumor-bearing mice treated with the following are shown: (1) solvent, (2) GQ01 10 mg / kg (single dose), (3) pyrotinib 20 mg / kg (QD, 2 weeks), and (4) GQ01 10 mg / kg (single dose) + pyrotinib 20 mg / kg (QD, 2 weeks). Table 4b shows that on the end day (day 35), the mean tumor volume was 255 mm in the GQ01 10 mg / kg group, the pyrotinib 20 mg / kg group, and the GQ01 10 mg / kg + pyrotinib 20 mg / kg group. 3 234mm 3 0mm 3 The TGI values ​​were 78.83%, 83.09%, and 131.17%, respectively.

[0265] The results showed that both GQ01 and pyrotinib monotherapy could inhibit tumor growth, and the combination of GQ01 and pyrotinib had stronger anti-tumor activity compared with monotherapy, and caused complete tumor regression in all mice.

[0266] Table 4b. Tumor growth inhibition based on tumor volume in a human gastric cancer PDX model using the combination of GQ01 and pyrotinib.

[0267]

[0268] a.Mean±SEM; measured on the end day;

[0269] b. TGI(%)=[1-(T 35 -T0) / (V 35 -V0)]×100%. T0 is the mean tumor volume in the treatment group on day 1 of administration, and T 35 V0 is the mean tumor volume in the treatment group on day 35 after administration; V0 is the mean tumor volume in the solvent group on day 1 after administration. 35 This represents the average tumor volume in the solvent group on day 35 after application.

[0270] Remission status was calculated based on the diameter or disappearance of the target lesion. Results are provided in the table below:

[0271]

[0272] The results showed that in a HER2-positive (IHC 3+) gastric cancer model, GQ01 and TKI as monotherapy could inhibit the proliferation of tumor cells, and the combination of GQ01 and TKI showed significantly enhanced antitumor activity, resulting in an increase in overall response rate (ORR), complete response (CR), partial response (PR), or tumor growth inhibition rate (TGI).

[0273] Example 5: In vivo efficacy evaluation of the combination of GQ01 and chemotherapy in a human breast cancer CDX model.

[0274] Experimental objective: To evaluate the in vivo antitumor efficacy of the combination of ADC drug GQ01 and chemotherapy in mice with a HER2-overexpressing CDX breast cancer model.

[0275] I. Breast cancer tumor cells were passaged in vivo using BT-474 cells (ATCC, batch number HTB-20). Cells in the exponential growth phase were collected and counted for tumor inoculation. 2 × 10⁶ cells were inoculated with 0.2 mL of Matrigel buffer (PBS:Matrigel = 1:1). 6 One cell was subcutaneously injected into the right scapula of 6-8 week old SPF-grade female BALB / c nude mice. Four days prior to cell inoculation, BALB / c nude mice were subcutaneously injected with estrogen tablets.

[0276] II. Ten days after inoculation, measure the tumor diameter with calipers and calculate it according to the formula V = 0.5a × b. 2 Calculate the tumor volume (where a is the long axis of the tumor and b is the short axis). The average tumor volume is approximately 100 mm². 3 -300mm 3 Mice were randomly assigned to four groups: a solvent group, a GQ01 5 mg / kg group, a paclitaxel 10 mg / kg group, a capecitabine 300 mg / kg group, a GQ01 + paclitaxel combination group, and a GQ01 + capecitabine combination group. Each group consisted of 6 mice. The day of the first administration was defined as day 0. Mice in the solvent group were administered GQ01 solvent and capecitabine solvent at the same frequency and via the same route of administration. Tumor volume was measured twice a week in each group. The experiment ended on day 25, and the tumor growth inhibition rate (TGI) was calculated as follows: TGI(%) = [1 - (mean tumor volume of the treatment group on the end day - mean tumor volume of the treatment group on day 1) / (mean tumor volume of the solvent group on the end day - mean tumor volume of the solvent group on day 1)] × 100%.

[0277] III. Figure 5a Tumor volume changes in tumor-bearing mice treated with the following are shown: (1) solvent, (2) GQ01 5 mg / kg (single dose), (3) paclitaxel 10 mg / kg (BIW, 4 weeks), and (4) GQ01 5 mg / kg (single dose) + paclitaxel 10 mg / kg (BIW, 4 weeks). Table 5a shows that the mean tumor volume was 452 mm² after 25 days of administration in the GQ01 5 mg / kg group, the paclitaxel 10 mg / kg group, and the GQ01 5 mg / kg + paclitaxel 10 mg / kg group. 3 520mm 3 and 99mm 3The TGI values ​​were 66.04%, 58.54%, and 105.27%, respectively.

[0278] These data indicate that all monotherapy groups can inhibit tumor growth, and the combination of GQ01 and paclitaxel can significantly reduce tumor volume, indicating that this combination has superior anti-tumor efficacy compared with paclitaxel or GQ01 monotherapy.

[0279] Table 5a. Tumor growth inhibition based on tumor volume in the combination of GQ01 and chemotherapy in a breast cancer CDX model.

[0280]

[0281] a. Mean ± SEM; measured on the last day;

[0282] b. TGI(%)=[1-(T 25 -T0) / (V 25 -V0)]×100%. T0 is the mean tumor volume in the treatment group on day 1 of administration, and T 25 V0 is the mean tumor volume in the treatment group on day 25 after administration; V0 is the mean tumor volume in the solvent group on day 1 after administration. 25 This represents the average tumor volume in the solvent group on day 25 after application.

[0283] IV. Figure 5b Tumor volume changes in tumor-bearing mice treated with the following are shown: (1) solvent, (2) GQ01 5 mg / kg (single dose), (3) capecitabine 300 mg / kg (QD, 2 weeks), and (4) GQ01 5 mg / kg (single dose) + capecitabine 300 mg / kg (QD, 2 weeks). Table 5b shows that on the end day (day 25), the mean tumor volume in the GQ01 5 mg / kg group, the capecitabine 300 mg / kg group, and the GQ01 5 mg / kg + capecitabine 300 mg / kg group was 452 mm. 3 858mm 3 89mm 3 The TGI values ​​were 66.04%, 21.01%, and 106.41%, respectively.

[0284] The results showed that GQ01 monotherapy could inhibit tumor growth, and the combination of GQ01 and capecitabine had better antitumor activity compared with capecitabine or GQ01 monotherapy.

[0285] Table 5b. Tumor growth inhibition based on tumor volume in the combination of GQ01 and chemotherapy in a breast cancer CDX model.

[0286]

[0287] a. Mean ± SEM; measured on the last day;

[0288] b. TGI(%)=[1-(T 25 -T0) / (V 25 -V0)]×100%. T0 is the mean tumor volume in the treatment group on day 1 of administration, and T 25 V0 is the mean tumor volume in the treatment group on day 25 after administration; V0 is the mean tumor volume in the solvent group on day 1 after administration. 25 This represents the average tumor volume in the solvent group on day 25 after application.

[0289] The results showed that in the CDX model of HER2-positive (IHC 3+) breast cancer, both GQ01 and chemotherapy as monotherapy could inhibit the proliferation of tumor cells, and the combination of GQ01 and chemotherapy showed significantly increased anti-tumor activity compared with monotherapy.

[0290] Example 6: In vivo efficacy evaluation of the combination of GQ01 and chemotherapy in a human breast cancer CDX model.

[0291] Experimental objective: To evaluate the in vivo antitumor efficacy of the combination of ADC drug GQ01 and chemotherapy in mice with a HER2-overexpressing CDX breast cancer model.

[0292] I. Collect and count HCC1954 cells in the exponential growth phase for tumor inoculation. Use 0.2 mL of Matrigel buffer (PBS:Matrigel = 1:1) to mix 1 × 10⁶ cells with the target cell. 6 One cell was subcutaneously injected into the right flank of 6-8 week old SPF-grade female BALB / c nude mice.

[0293] II. Nine days after inoculation, measure the tumor diameter with calipers and calculate it according to the formula V = 0.5a × b. 2 Calculate the tumor volume (where a is the long axis of the tumor and b is the short axis). The average tumor volume is approximately 100 mm². 3 -300mm 3Mice were randomly assigned to four groups: a solvent group, a GQ01 2 mg / kg group, a paclitaxel 10 mg / kg group, a capecitabine 300 mg / kg group, a GQ01 + paclitaxel combination group, a GQ01 + capecitabine combination group, and a trastuzumab 5 mg / kg + pertuzumab 5 mg / kg + paclitaxel 10 mg / kg group. Each group consisted of 6 mice. The day of the first administration was defined as day 0. Mice in the solvent group were administered GQ01 solvent and capecitabine solvent at the same frequency and via the same route. Tumor volume was measured twice weekly in each group. The experiment ended on day 32, and the tumor growth inhibition rate (TGI) was calculated as follows: TGI(%) = [1 - (mean tumor volume of the treatment group on the end day - mean tumor volume of the treatment group on day 1) / (mean tumor volume of the solvent group on the end day - mean tumor volume of the solvent group on day 1)] × 100%.

[0294] III. Figure 6a Tumor volume changes in tumor-bearing BALB / c nude mice treated with the following: (1) solvent, (2) GQ01 2 mg / kg (single dose), (3) paclitaxel 10 mg / kg (BIW, 4 doses), (4) GQ01 2 mg / kg (single dose) + paclitaxel 10 mg / kg (BIW, 4 doses), (5) trastuzumab 5 mg / kg (single dose) + pertuzumab 5 mg / kg (single dose) + paclitaxel 10 mg / kg (BIW, 4 doses). Table 6a shows that on the end day (day 32), the mean tumor volume was 774 mm in the GQ01 2 mg / kg group, the paclitaxel 10 mg / kg group, the GQ01 2 mg / kg + paclitaxel 10 mg / kg group, and the trastuzumab 5 mg / kg + pertuzumab 5 mg / kg + paclitaxel 10 mg / kg group. 3 1219mm 3 632mm 3 and 909mm 3 The TGI values ​​were 56.93%, 26.60%, 66.62%, and 47.73%, respectively.

[0295] The results showed that GQ01 2 mg / kg as monotherapy could inhibit tumor cell growth. The combination of GQ01 and paclitaxel significantly inhibited tumor growth and was more effective than the combination therapy of trastuzumab, pertuzumab and paclitaxel.

[0296] Table 6a. Tumor growth inhibition based on tumor volume in the combination of GQ01 and chemotherapy in a breast cancer CDX model.

[0297]

[0298] a. Mean ± SEM; measured on the last day;

[0299] b. TGI(%)=[1-(T 32 -T0) / (V 32 -V0)]×100%. T0 is the mean tumor volume in the treatment group on day 1 of administration, and T 32 V0 is the mean tumor volume in the treatment group on day 32 after administration; V0 is the mean tumor volume in the solvent group on day 1 after administration. 32 This represents the average tumor volume in the solvent group on day 32 after application.

[0300] IV. Figure 6b Tumor volume changes in tumor-bearing female BALB / c mice treated with the following: (1) solvent, (2) GQ01 2 mg / kg (single dose), (3) capecitabine 300 mg / kg (QD, 2 weeks), and (4) GQ01 2 mg / kg (single dose) + capecitabine 300 mg / kg (QD, 2 weeks). Table 6b shows that on the end day (day 32), the mean tumor volume was 774 mm in the GQ01 2 mg / kg group, the capecitabine 300 mg / kg group, and the GQ01 2 mg / kg + capecitabine 300 mg / kg group. 3 1183mm 3 238mm 3 The TGI values ​​were 56.93%, 29.05%, and 93.50%, respectively.

[0301] The results showed that GQ01 had anti-tumor efficacy as a monotherapy, and the combination of GQ01 and capecitabine significantly improved tumor suppression compared with capecitabine or GQ01 monotherapy.

[0302] Table 6b. Tumor growth inhibition based on tumor volume in the combination of GQ01 and chemotherapy in a CDX model of breast cancer.

[0303]

[0304] a. Mean ± SEM; measured on the last day;

[0305] b. TGI(%)=[1-(T 25 -T0) / (V 25 -V0)]×100%. T0 is the mean tumor volume in the treatment group on day 1 of administration, and T 25 V0 is the mean tumor volume in the treatment group on day 25 after administration; V0 is the mean tumor volume in the solvent group on day 1 after administration. 25 This represents the average tumor volume in the solvent group on day 25 after application.

[0306] The results showed that in the CDX model of HER2-positive (IHC 3+) breast cancer, both GQ01 and chemotherapy, when used as monotherapy, could inhibit the proliferation of tumor cells, and the combination of GQ01 and chemotherapy had a significantly increased anti-tumor efficacy compared with monotherapy.

[0307] Example 7: Safety and preliminary efficacy of the combination of GQ01 and pyrotinib in a clinical trial.

[0308] Experimental objective:

[0309] Design a clinical-stage study to investigate the safety and preliminary efficacy of the combination of GQ01 and pyrotinib in previously treated patients with HER2-positive metastatic breast cancer.

[0310] method:

[0311] During the dose escalation phase, the starting dose of GQ01 was 6.0 mg / kg, administered intravenously every 21 days (Q3W); and the starting dose of pyrotinib was 320 mg orally once daily (QD), with a cycle of 28 days. The combination dose exploration of GQ01 and pyrotinib followed a "3+3" principle, with subsequent dose escalation or reduction depending on the occurrence of specific dose-limiting toxicities (DLTs) in the starting dose group. The DLT observation period was 21 days. The doses of GQ01 were 6.0 mg / kg, 7.2 mg / kg, and 8.4 mg / kg. Even at the highest tested dose of 8.4 mg / kg, the maximum tolerated dose (MTD) was not reached.

[0312] result:

[0313] A total of 15 patients with HER2-positive metastatic breast cancer received GQ01 and pyrotinib. All patients (15 / 15) had failed prior trastuzumab (or its biosimilar) therapy and chemotherapy. 33.3% of patients (5 / 15) had previously been treated with pertuzumab. No dose-limited leukemia (DLT) was reported upon completion of dose escalation, and the maximum tolerated dose (MTD) was not reached. Grade 3-4 treatment-related adverse events (TRAEs) occurred in 73.3% of patients (13 / 15), including diarrhea (66.7%), anemia (13.3%), elevated alanine aminotransferase (6.7%), thrombocytopenia (6.7%), and urticaria (6.7%).

[0314] At the completion of dose escalation, nine patients were still undergoing study treatment, with the longest treatment duration being 348 days. The objective response rate (ORR) for all enrolled patients was 66.7% (10 / 15), and 71.4% (10 / 14) of patients had undergone tumor evaluation. Analysis showed that the ORRs at dose levels of 6.0 mg / kg, 7.2 mg / kg, and 8.4 mg / kg were 75% (3 / 4), 100% (5 / 5), and 33.3% (2 / 6), respectively.

[0315] Ten days after dose escalation was completed, nine patients were still undergoing study treatment, with the longest treatment duration being 358 days. Analysis showed that the objective response rate (ORR) for all enrolled patients was 73.3% (11 / 15). The ORRs at dose levels of 6.0 mg / kg, 7.2 mg / kg, and 8.4 mg / kg were 75% (3 / 4), 100% (5 / 5), and 50% (3 / 6), respectively.

[0316] in conclusion

[0317] The combination of GQ01 and pyrotinib showed acceptable toxicity and preliminary antitumor activity in previously treated patients with HER2-positive metastatic breast cancer.

[0318] sequence list

[0319] SEQ ID NO:1: Trastuzumab LCDR1

[0320] RASQDVNTAVA

[0321] SEQ ID NO:2: Trastuzumab LCDR2

[0322] SASFLYS

[0323] SEQ ID NO:3: Trastuzumab LCDR3

[0324] QQHYTTPPT

[0325] SEQ ID NO:4: Trastuzumab HCDR1

[0326] DTYIH

[0327] SEQ ID NO:5: Trastuzumab HCDR2

[0328] RIYPTNGYTRYADSVKG

[0329] SEQ ID NO:6: Trastuzumab HCDR3

[0330] WGGDGFYAMDY

[0331] SEQ ID No:7: Trastuzumab light chain variable region:

[0332] DIQMTQSPSSSLSASVGDRVTITCRASQDVNTAVAWYQQKPGKAPKLLIYSASFLYSGV

[0333] PSRFSGSRSGTDFTLTISSLQPEDFATYYCQQHYTTPPTFGQGTKVEIKRTV

[0334] SEQ ID NO:8: Trastuzumab heavy chain variable region:

[0335] EVQLVESGGGLVQPGGSLRLSCAASGFNIKDTYIHWVRQAPGKGLEWVARIYPTNGY

[0336] TRYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCSRWGGDGFYAMDYWGQ

[0337] GTLVTVSS

[0338] SEQ ID No:9: Trastuzumab light chain full length:

[0339] DIQMTQSPSSSLSASVGDRVTITCRASQDVNTAVAWYQQKPGKAPKLLIYSASFLYSGV

[0340] PSRFSGSRSGTDFTLTISSLQPEDFATYYCQQHYTTPPTFGQGTKVEIKRTVAAPSVFIFP

[0341] PSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLS

[0342] STLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0343] SEQ ID No:10: Trastuzumab heavy chain full length:

[0344] EVQLVESGGGLVQPGGSLRLSCAASGFNIKDTYIHWVRQAPGKGLEWVARIYPTNGY

[0345] TRYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCSRWGGDGGFYAMDYWGQ

[0346] GTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGV

[0347] HTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT

[0348] CPPCPAPELLGGPSVFLFPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVE

[0349] VHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAK

[0350] GQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVL

[0351] DSDGSFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK SEQ ID No:11:

[0352] LPET

Claims

1. A drug combination comprising the following (a) and (b): (a) One or more couplings having the structure of formula (i-1) and / or formula (i-2). Preferably, trastuzumab is connected to the remainder of the conjugate via the C-terminus of the two light chains of trastuzumab; (b) One or more other cancer treatment agents, said one or more other cancer treatment agents being selected from: tyrosine kinase inhibitors and chemotherapeutic agents.

2. The drug combination according to claim 1, wherein the tyrosine kinase inhibitor is selected from: anaplastic lymphoma kinase (ALK) inhibitors, c-Met inhibitors, HER family inhibitors, FMS-like tyrosine kinase 3 (FLT3) inhibitors, vascular endothelial growth factor (VEGF) / vascular endothelial growth factor receptor (VEGFR) inhibitors, fibroblast growth factor receptor (FGFR) inhibitors, platelet-derived growth factor receptor (PDGFR) inhibitors, transfection rearrangement (RET) kinase inhibitors, tropomyosin receptor kinase (TRK) inhibitors, Bcr-Abl1 inhibitors, Bruton's tyrosine kinase (BTK) inhibitors, and Janus kinase (JAK) inhibitors.

3. The drug combination according to any one of claims 1 or 2, wherein the tyrosine kinase inhibitor is selected from: tucatinib, neratinib, and pyrotinib, preferably tucatinib or pyrotinib.

4. The drug combination according to claim 1, wherein the chemotherapeutic agent is selected from taxanes.

5. The pharmaceutical combination according to claim 4, wherein the taxanes are selected from docetaxel and paclitaxel, preferably paclitaxel.

6. The drug combination according to claim 1, wherein the chemotherapeutic agent is selected from pyrimidine antimetabolites, and the pyrimidine antimetabolites are selected from: cytarabine, capecitabine, ancitabine, gemcitabine, enoxatabine, carmoflu, dideoxyuridine, deoxyfluorouridine and fluorouridine, preferably capecitabine.

7. The pharmaceutical combination according to any one of claims 1-6, wherein the one or more conjugates and the one or more other cancer therapeutic agents are administered simultaneously, separately, or sequentially.

8. A kit or pharmaceutical composition comprising the pharmaceutical composition of any one of claims 1-7.

9. A method for preventing, alleviating, or treating a proliferative disorder, the method comprising administering to a subject requiring such treatment an effective amount of any one of claims 1-7, or a kit or pharmaceutical composition as described in claim 8.

10. The method of claim 9, wherein the cell proliferation disorder is cancer.

11. The method of claim 10, wherein the cancer is HER2-positive cancer.

12. The method of claim 11, wherein the HER2-positive cancer is selected from: gastric cancer or esophageal cancer, breast cancer, vaginal cancer, salivary gland cancer, endometrial cancer, bladder cancer, non-small cell lung cancer (NSCLC), cervical cancer, ovarian cancer, colorectal cancer, and pancreatic cancer.

13. The method according to any one of claims 10-12, wherein the cancer is metastatic, recurrent, refractory, or advanced.

14. The method according to any one of claims 9 to 13, wherein the subject requiring such treatment has received prior treatment; The aforementioned prior treatment includes antibody therapy, chemotherapy, and / or kinase inhibitor therapy; preferably antibody therapy and / or chemotherapy.

15. The method of claim 14, wherein the prior treatment is selected from: trastuzumab, pertuzumab, tucatinib, neratinib, pyrotinib, paclitaxel, capecitabine, letrozole, and combinations thereof.

16. The method according to claim 14 or 15, wherein the proliferative disease is cancer; preferably, the cancer is selected from: gastric cancer or esophageal cancer, breast cancer, vaginal cancer, salivary gland cancer, endometrial cancer, bladder cancer, non-small cell lung cancer (NSCLC), cervical cancer, ovarian cancer, colorectal cancer, and pancreatic cancer.

17. The method of claim 16, wherein The cancer is breast cancer, and the subject requiring this treatment has received prior treatment; preferably, the prior treatment is selected from: trastuzumab, pertuzumab, pyrotinib, capecitabine, palbociclib, letrozole, and combinations thereof; and / or The cancer is gastric cancer, and the subject requiring such treatment has received prior treatment; preferably, the prior treatment is selected from: trastuzumab, pertuzumab, pyrotinib, capecitabine, palbociclib, and combinations thereof.

18. The method of claim 17, wherein the subject requiring such treatment is resistant to one or more of the prior treatments.

19. The method according to any one of claims 9 to 18, wherein one or more conjugates and one or more other cancer therapeutic agents are administered simultaneously, separately, or sequentially.

20. The method according to any one of claims 9 to 19, wherein the conjugate is administered at an effective amount of about 5 mg / kg to 10 mg / kg per treatment cycle, and the other cancer treatment agent is administered at an effective amount of about 100 mg to 600 mg once daily per treatment cycle.

21. The method of claim 20, wherein each treatment cycle of conjugate administration is independently about 1 week, about 2 weeks, about 3 weeks, about 4 weeks, about 5 weeks, about 6 weeks, about 7 weeks; and / or Each treatment cycle of the other cancer treatment agents mentioned therein is independently about 1 week, about 2 weeks, about 3 weeks, about 4 weeks, about 5 weeks, about 6 weeks, and about 7 weeks.

22. The method according to claims 20 to 21, wherein the conjugate is administered at an effective amount of about 6 mg / kg to 8.4 mg / kg per treatment cycle, and the other cancer treatment agent is administered at an effective amount of about 320 mg once daily for 28 days per treatment cycle.

23. The method according to claims 20 to 21, wherein the conjugate is a mixture of the conjugate of formula (i-1) and the conjugate of formula (i-2); and the other cancer treatment agent is pyrotinib.

24. The kit of claim 8, further comprising instructions for administering the compound to a patient in need; wherein the dosage of the conjugate is approximately 5 mg / kg to 10 mg / kg per treatment cycle, and the dosage of the other cancer treatment agent is approximately 100 mg to 600 mg once daily per treatment cycle.

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