Cancer treatment using combination of EGFR-binding antibodies

Abstract

The present disclosure relates to the field of therapeutic antibodies for the treatment of subjects with cancer. More specifically, the disclosure relates to the treatment of cancer using an antibody, or a functional portion, derivative, and / or analog of an antibody, comprising a variable domain that binds to the extracellular portion of EGFR, in combination therapy further comprising administration of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38. The cancer may be head and neck cancer, gastric cancer, esophageal cancer, gastroesophageal junction cancer, non-small cell lung cancer, or colorectal cancer. The disclosure also relates to pharmaceutical formulations, kit of parts and dosage regimens.

Description

Technical Field

[0001] The present disclosure relates to methods and means for treating cancer. In particular, the present disclosure relates to a method for treating cancer in a subject using a combination of an antibody that binds to EGFR and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38. The present disclosure further relates to combinations for use in such methods and in the manufacture of medicaments for treating cancer. Background Art

[0002] Colorectal cancer (CRC) is a leading cause of death worldwide, partly due to its increasing resistance to current treatments. While several new treatments have made progress in CRC, many fail in clinical trials, and metastatic CRC remains largely incurable. Current standard of care for advanced CRC includes chemotherapy regimens that block essential cancer cell functions and kill dividing cells.

[0003] Several anti-EGFR drugs have demonstrated some degree of efficacy as targeted therapies for metastatic CRC (mCRC). However, resistance to anti-EGFR therapies has been observed for various reasons, such as the emergence of resistance mutations or the presence of innate resistance to anti-EGFR therapies. Most patients whose cancers are sensitive to anti-EGFR therapy later develop resistant cancers.

[0004] Current standard of care treatment for CRC includes oxaliplatin, fluorouracil, and irinotecan. In the case of liver metastasis, standard of care treatment is combined with EGFR or VEGF blocking monoclonal antibodies.

[0005] Due to increasing resistance to current treatments, there remains a need to develop further therapeutics that target cancers such as CRC to improve cancer treatment or expand treatment options. Summary of the Invention

[0006] The present disclosure provides an antibody, or a functional portion, derivative, and / or analog thereof, comprising a variable domain that binds to the extracellular portion of EGFR, for use in treating cancer in a subject, wherein the treatment further comprises administering a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38. In certain aspects, the present disclosure provides an antibody, or a functional portion, derivative, and / or analog thereof, comprising a variable domain that binds to the extracellular portion of EGFR, for use in treating cancer in a subject, wherein the treatment further comprises administering fluorouracil, TAS-102, oxaliplatin, venetoclax, or SN-38.

[0007] The present disclosure shows that combination therapy comprising administering a multispecific antibody targeting EGFR and LGR5 in combination with a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 is more effective when compared to the effects of administering the multispecific antibody or the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 separately.

[0008] In certain aspects, the present disclosure further provides a combination of an antibody or a functional portion, derivative and / or analog thereof and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 for use in treating cancer.

[0009] In certain aspects, the present disclosure provides an antibody or a functional part, derivative and / or analogue thereof, comprising a variable domain that binds to the extracellular portion of EGFR, and for use in treating cancer in a subject, wherein the treatment further comprises administration of fluorouracil, oxaliplatin and folinic acid.

[0010] In certain aspects, the present disclosure provides an antibody or a functional part, derivative and / or analog thereof comprising a variable domain that binds to the extracellular portion of EGFR and for use in treating cancer in a subject, wherein the treatment further comprises administering fluorouracil, irinotecan and folinic acid.

[0011] Further provided are methods of treating cancer in a subject, comprising administering to the subject an effective amount of an antibody comprising a variable domain that binds to the extracellular portion of EGFR, or a functional portion, derivative, and / or analog thereof, and fluorouracil, TAS-102, oxaliplatin, venetoclax, or SN-38. In certain aspects, the fluorouracil, TAS-102, oxaliplatin, venetoclax, or SN-38 is administered to the subject in an effective amount.

[0012] Further provided is a combination therapy, wherein a combination therapy is administered to a subject having cancer, the combination comprising administering to the subject having cancer an effective amount of an antibody comprising a variable domain that binds to the extracellular portion of EGFR, or a functional part, derivative and / or analog thereof, and fluorouracil, TAS-102, oxaliplatin, venetoclax or SN-38.

[0013] In certain aspects, the antibody or its functional part, derivative and / or analog comprising a variable domain that binds to the extracellular portion of EGFR is administered together with fluorouracil, folinic acid and irinotecan (FOLFIRI) or folinic acid, fluorouracil and oxaliplatin (FOLFOX). In certain aspects, the subject has received previous treatment for the cancer. In certain aspects, the subject has received chemotherapy as a second-line treatment. In certain aspects, the subject has received chemotherapy as a third-line or subsequent treatment. In certain aspects, the subject suffers from colorectal cancer, particularly metastatic colorectal cancer (mCRC). In certain aspects, the treatment of the present disclosure further comprises administering 1500 mg of petosemtamab to the subject every two weeks (Q2W). In certain aspects, the subject suffers from mCRC and is RAS and / or RAF wild type (including Kristen rat sarcoma (KRAS) and / or B- rapidly accelerated fibrosarcoma (BRAF)). In certain aspects, the mCRC is genotypically wild-type for RAS and / or RAF, including KRAS and / or BRAF.

[0014] In some aspects, the cancer is adenocarcinoma or squamous cell carcinoma. In some aspects, the cancer is head and neck cancer, stomach cancer, esophageal cancer, gastroesophageal junction cancer, non-small cell lung cancer, or colorectal cancer. In some aspects, the cancer is colorectal cancer.

[0015] In certain aspects, the subject of the present disclosure is a mammalian subject, such as a human subject.

[0016] In certain aspects, the subject has progressed following a prior anti-cancer therapy, wherein the anti-cancer therapy is chemotherapy, targeted therapy, immunotherapy, or radiation therapy.

[0017] In certain aspects, the antibody or its functional part, derivative and / or analog is enhanced by ADCC.In addition, in certain aspects, the antibody or its functional part, derivative and / or analog is defucosylated.

[0018] In certain aspects, the antibodies of the present disclosure, or their functional portions, derivatives and / or analogs, are multispecific antibodies. In certain aspects, the antibodies, or their functional portions, derivatives and / or analogs, are bispecific antibodies. In certain aspects, the antibodies, or their functional portions, derivatives and / or analogs, are bispecific antibodies that bind at least to EGFR. In certain aspects, the antibodies, or their functional portions, derivatives and / or analogs, comprise a second variable domain that does not bind to EGFR. In certain aspects, the antibodies, or their functional portions, derivatives and / or analogs, monovalently bind to EGFR. In certain aspects, the antibodies, or their functional portions, derivatives and / or analogs, comprise a second variable domain that binds to LGR5.

[0019] In some aspects, the subject to be treated has not received previous anticancer therapy. In some aspects, the subject to be treated has not received previous chemotherapy, targeted therapy, immunotherapy or radiotherapy. In some aspects, the subject to be treated has not received previous therapy using fluorouracil, oxaliplatin, FOLFOX, FOLFIRI, TAS-102, trastuzumab, pembrolizumab, nivolumab, cetuximab or a combination thereof.

[0020] In certain aspects, the antibody or a functional portion, derivative, and / or analog thereof is administered to the subject simultaneously, separately, or sequentially with a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

[0021] In certain aspects, the present disclosure provides a pharmaceutical composition comprising an antibody or a functional portion, derivative and / or analog thereof and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

[0022] In certain aspects, the present disclosure further provides a kit of parts comprising an antibody or a functional portion, derivative, and / or analog thereof and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38, and instructions for use of the antibody or a functional portion, derivative, and / or analog thereof and instructions for use of the fluoropyrimidine, platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

[0023] In certain aspects, the present disclosure provides use of an antibody, or a functional part, derivative, and / or analog thereof, and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38, in the manufacture of a medicament for treating cancer in a subject, wherein the treatment comprises simultaneous, separate, or sequential administration of the antibody, or a functional part, derivative, and / or analog thereof, and the fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

[0024] In certain aspects, the present disclosure relates to the use of an antibody or a functional portion, derivative, and / or analog of an antibody comprising a variable domain that binds to the extracellular portion of EGFR and, optionally, a variable domain that binds to the extracellular portion of LGR5, and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 in the manufacture of one or more pharmaceutical agents for treating cancer in a subject. In certain aspects, the antibody or a functional portion, derivative, and / or analog of an antibody and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 are used to manufacture separate pharmaceutical agents, such as two separate pharmaceutical agents, one for the antibody or its functional portion, derivative, and / or analog, and one for the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38. The pharmaceutical agent comprising the antibody, functional portion, derivative, and / or analog thereof comprising a variable domain that binds to the extracellular portion of EGFR can be contained in a container that is separate from (meaning not physically connected to) a container containing the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 as a pharmaceutical agent.

[0025] In certain aspects, the antibody or functional part, derivative and / or analogue of the antibody comprising a variable domain that binds to the extracellular part of EGFR and optionally comprises a variable domain that binds to the extracellular part of LGR5 is or comprises pesentuzumab.

[0026] In certain aspects, the present disclosure provides use of an antibody or functional portion, derivative, and / or analog of the present disclosure comprising a variable domain that binds to the extracellular portion of EGFR in the manufacture of a medicament for increasing the efficacy of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 for treating cancer.

[0027] In certain aspects, the present disclosure provides the use of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 in the manufacture of a medicament for increasing the efficacy of an antibody or functional portion, derivative, and / or analog of the present disclosure comprising a variable domain that binds to an extracellular portion of EGFR for treating cancer.

[0028] In addition, the present disclosure provides an antibody or a functional portion, derivative, and / or analog of an antibody comprising a variable domain that binds to the extracellular portion of EGFR and a combination of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 as described herein for use in treating cancer in a subject in need thereof.

[0029] In addition, the present disclosure provides a combination comprising: a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38; instructions for use of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 for treating cancer in a subject; and instructions for use of an antibody comprising a variable domain that binds to the extracellular portion of EGFR, or a functional portion, derivative, and / or analog of the antibody, for treating the cancer in a subject.

[0030] In addition, the present disclosure provides a combination comprising: an antibody or a functional portion, derivative, and / or analog of an antibody comprising a variable domain that binds to the extracellular portion of EGFR, instructions for use of the antibody or the functional portion, derivative, and / or analog of the antibody in treating cancer in a subject, and instructions for use of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 in treating the cancer in a subject.

[0031] In certain aspects, the present disclosure provides a pharmaceutical composition comprising an antibody or a functional portion, derivative and / or analog of the present disclosure comprising a variable domain that binds to the extracellular portion of EGFR and instructions for use thereof with a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 for treating cancer.

[0032] In certain aspects, the present disclosure provides a pharmaceutical composition for treating cancer comprising an antibody comprising a variable domain that binds to the extracellular portion of EGFR or a functional portion, derivative and / or analog of the antibody of the present disclosure, and a pharmaceutical composition comprising a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 for treating the cancer.

[0033] In certain aspects, the present disclosure provides a pharmaceutical composition for treating cancer, comprising an antibody, or a functional portion, derivative, and / or analog thereof, comprising a variable domain of the present disclosure that can bind to the extracellular portion of EGFR, wherein the pharmaceutical composition is administered in combination with a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 of the present disclosure.

[0034] In certain aspects, the present disclosure relates to a pharmaceutical composition for treating cancer, comprising an antibody, or a functional portion, derivative, and / or analog thereof, comprising a variable domain of the present disclosure that binds to the extracellular portion of EGFR, wherein the subject to be treated is administered a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 prior to, concurrently with, or after administration of the antibody.

[0035] In certain aspects, the present disclosure relates to a pharmaceutical composition for treating cancer in a subject, comprising a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38, wherein an antibody or a functional portion, derivative, and / or analog thereof of the present disclosure comprising a variable domain that can bind to the extracellular portion of EGFR is administered to the subject to be treated before, simultaneously with, or after administration of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38.

[0036] Therefore, the present disclosure relates to a combination of agents for treating cancer in a subject, comprising administering to the subject a plurality of different agents to treat the cancer, wherein the treatment comprises administering the agents simultaneously, sequentially, or separately. In certain aspects, the agent comprises an antibody, or a functional portion, derivative, and / or analog thereof, comprising a variable domain of the present disclosure that binds to the extracellular portion of EGFR, and the other different agents comprise a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

[0037] In certain aspects, the antibodies or functional portions, derivatives, and / or analogs comprising the variable domains of the present disclosure that can bind to the extracellular portion of EGFR can be administered simultaneously, sequentially, or separately with the fluoropyrimidines, platinum-based chemotherapeutics, BCL-2 inhibitors, or SN-38 of the present disclosure. Thus, the combination of the antibodies or functional portions, derivatives, and / or analogs comprising the variable domains of the present disclosure that can bind to the extracellular portion of EGFR and fluoropyrimidines, platinum-based chemotherapeutics, BCL-2 inhibitors, or SN-38 encompasses simultaneous, sequential, or separate administration.

[0038] Thus, in certain aspects, the present disclosure provides an antibody or a functional part, derivative and / or analog thereof, comprising a variable domain of the present disclosure that can bind to the extracellular portion of EGFR, and for use in a method of treating cancer, wherein the treatment further comprises administering a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38, wherein optionally the antibody or the functional part, derivative and / or analog thereof is administered simultaneously, sequentially, or separately with the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38.

[0039] Thus, in certain aspects, the present disclosure provides a method of treating a subject having cancer, comprising administering to the subject an effective amount of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 and an antibody or a functional portion, derivative, and / or analog of the present disclosure comprising a variable domain that binds to the extracellular portion of EGFR, wherein optionally, the antibody or functional portion, derivative, and / or analog of the antibody is administered simultaneously, sequentially, or separately with the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38.

[0040] Thus, in certain aspects, the present disclosure provides the use of an antibody or functional portion, derivative, and / or analog comprising a variable domain of the present disclosure that binds to the extracellular portion of EGFR and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 in the manufacture of a medicament for treating cancer, wherein the antibody or functional portion, derivative, and / or analog is optionally administered simultaneously, sequentially, or separately with the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38. In certain aspects, the antibody or functional portion, derivative, and / or analog is administered before, simultaneously, or after administration of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38.

[0041] In certain aspects, an antibody or functional portion, derivative, and / or analog comprising a variable domain of the present disclosure that binds to the extracellular portion of EGFR is used in the manufacture of a medicament for treating cancer, and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 is used in the manufacture of a medicament for treating the cancer, wherein the antibody or functional portion, derivative, and / or analog is optionally administered simultaneously, sequentially, or separately with the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38. Alternatively, the antibody or functional portion, derivative, and / or analog is administered before, simultaneously, or after the administration of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38.

[0042] In certain aspects, an antibody or functional portion, derivative and / or analog comprising a variable domain that binds to the extracellular portion of EGFR of the present disclosure is used in the manufacture of a medicament for treating cancer, whereby the treatment is combined with FOLFIRI, wherein optionally the antibody or functional portion, derivative and / or analog is administered simultaneously, sequentially or separately with the FOLFIRI. Alternatively, the antibody or functional portion, derivative and / or analog is administered before, simultaneously with or after the administration of FOLFIRI.

[0043] In certain aspects, an antibody or functional portion, derivative and / or analog comprising a variable domain that binds to the extracellular portion of EGFR disclosed herein is used in the manufacture of a medicament for treating cancer, whereby the treatment is combined with FOLFOX, wherein optionally the antibody or functional portion, derivative and / or analog is administered simultaneously, sequentially or separately with the FOLFOX. Alternatively, the antibody or functional portion, derivative and / or analog is administered before, simultaneously with or after the administration of FOLFOX. BRIEF DESCRIPTION OF THE DRAWINGS

[0044] Figure 1 : Human LGR5 sequence; SEQ ID NO: 1.

[0045] Figure 2 : Human EGFR sequence; SEQ ID NO: 2.

[0046] Figure 3: (a) The amino acid sequences of the heavy chain variable regions (SEQ ID Nos: 3-15) and the common light chain variable regions (such as the human kappa light chain IGKV1-39 / jk1 variable region (SEQ ID NO: 107)) together form the variable domain that binds to LGR5 and EGFR. The heavy chain CDRs (according to the KABAT numbering system) and framework regions are shown in Figure 3. Figure 3b shown.

[0047] Figure 4 a) Common light chain amino acid sequence. b) Common light chain variable region (IgVk1-39 / jk1). c) Light chain constant region. d) V region IgVk1-39; e) Common light chain CDR1, CDR2, and CDR3 numbering according to IMGT.

[0048] Figure 5 IgG heavy chain used to generate bispecific molecules. a) CH1 domain DNA. b) Hinge region. c) CH2 domain. d) CH3 domain containing variants L351K and T366K (KK). e) CH3 domain containing variants L351D and L368E (DE). Residue positions are according to EU numbering. DETAILED DESCRIPTION

[0049] In certain aspects, the present disclosure provides an antibody or a functional portion, derivative and / or analog thereof, which comprises a variable domain that binds to the extracellular portion of EGFR and is used to treat cancer in a subject, wherein the treatment further comprises administering a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

[0050] In certain aspects, the present disclosure provides a method of treating cancer in a subject, comprising administering to the subject an effective amount of an antibody, or a functional portion, derivative, and / or analog of an antibody, comprising a variable domain that binds to an extracellular portion of EGFR, and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

[0051] In certain aspects, the present disclosure provides an antibody or a functional part, derivative and / or analog thereof, which comprises a variable domain that binds to the extracellular portion of EGFR and a variable domain that binds to the extracellular portion of LGR5, and which is used to treat cancer in a subject, wherein the treatment further comprises administering a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

[0052] In certain aspects, the present disclosure provides a method of treating cancer in a subject, the method comprising administering to the subject an effective amount of an antibody or a functional part, derivative and / or analog thereof, and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38, wherein the antibody or functional part, derivative and / or analog thereof comprises a variable domain that binds to the extracellular portion of EGFR and a variable domain that binds to the extracellular portion of LGR5.

[0053] Cancer type

[0054] In certain aspects, the cancer of the present disclosure is adenocarcinoma or squamous cell carcinoma.

[0055] In certain aspects, the cancer is selected from gastric cancer, esophageal cancer, gastroesophageal junction cancer, head and neck cancer, non-small cell lung cancer or colorectal cancer, in particular squamous cell carcinoma of the head and neck (SCCHN).

[0056] In certain aspects, the cancer is gastric cancer, esophageal cancer, or gastroesophageal junction cancer. Gastric cancer (also known as stomach cancer) is a cancer that develops from the lining of the stomach, particularly the mucus-producing glandular cells found therein. This type of cancer is also known as adenocarcinoma, or in this case, gastric adenocarcinoma develops from the lining of the stomach. In particular, the cancer is therefore gastric adenocarcinoma or cancer that develops from the lining of the stomach, which are used interchangeably herein. Esophageal cancer is a cancer that develops from the esophagus. The two main subtypes are ESCC (esophageal squamous cell carcinoma) and EAC (esophageal adenocarcinoma). Gastroesophageal junction cancer (also known as gastroesophageal junction adenocarcinoma) originates from the gastroesophageal junction.

[0057] Cancers collectively known as head and neck cancers typically originate in squamous cells that line the moist mucous membranes of the inside of the head and neck, such as the inside of the mouth, nose, and throat. These squamous cell carcinomas are often referred to as head and neck squamous cell carcinomas and are treated in certain aspects of the present disclosure. Although rare, head and neck cancers can also occur in the salivary glands. In particular, head and neck cancers can occur in the oral cavity. This includes the lips, the front two-thirds of the tongue, the gums, the lining of the cheeks and lips, the floor of the mouth under the tongue, the hard palate, and a small area of gums behind the wisdom teeth.

[0058] Thus, in certain aspects, the head and neck cancer is a squamous cell carcinoma, and includes nasopharyngeal cancer, laryngeal cancer, hypopharyngeal cancer, nasal cavity cancer, paranasal sinus cancer, oral cancer, oropharyngeal cancer, or salivary gland cancer. More particularly, the present disclosure relates to the treatment of cancers including squamous cell head and neck cancer, such as cancers located in the oropharynx, hypopharynx, pharynx, oral cavity, or tongue. Furthermore, the head and neck cancer is particularly squamous cell carcinoma of unknown primary (also referred to in the art as cancer of unknown primary or CUP).

[0059] In certain aspects, the cancer is non-small cell lung cancer.

[0060] In some aspects, the cancer is colorectal cancer. In some aspects, the cancer is metastatic colorectal cancer (mCRC). In some aspects, the subject being treated has been diagnosed with colon or rectal adenocarcinoma. In some aspects, the cancer has previously been treated, such as with a first-line standard of care suitable for CRC or mCRC. In some aspects, the subject or the cancer has been treated and received second-line treatment with an antibody or its functional portion, derivative and / or analog, and in combination with a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38, wherein the antibody or its functional portion, derivative and / or analog comprises a variable domain that binds to the extracellular portion of EGFR and optionally a variable domain that binds to the extracellular portion of LGR5.

[0061] In some aspects, the subject or cancer has been treated and treated with an antibody or its functional portion, derivative and / or analog and in combination with a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38 as a third-line or subsequent treatment, wherein the antibody or its functional portion, derivative and / or analog comprises a variable domain that binds to the extracellular portion of EGFR and optionally a variable domain that binds to the extracellular portion of LGR5. In some aspects, the previous treatment includes a standard of care treatment suitable for CRC or mCRC. In some aspects, the standard of care includes or is chemotherapy suitable for CRC or mCRC. In some aspects, the previous treatment includes or is fluoropyrimidine-oxaliplatin-based chemotherapy with or without bevacizumab for CRC or mCRC, or fluoropyrimidine-irinotecan-based chemotherapy with or without bevacizumab.

[0062] In some aspects, the subject to be treated suffers from CRC and is RAS wild-type. In some aspects, the subject to be treated suffers from CRC and is RAF wild-type. RAS includes Kristen rat sarcoma (KRAS), and RAF includes B-rapidly accelerated fibrosarcoma (BRAF). In some aspects, the genome of the CRC is RAS and / or RAF wild-type, including KRAS and / or BRAF. In some aspects, the genome of the sample obtained from the CRC is RAS and / or RAF wild-type, including KRAS and / or BRAF.

[0063] In some aspects, the subject has previously been diagnosed with unresectable or metastatic colon or rectal adenocarcinoma by histological or cytological methods. In some aspects, the subject is RAS and / or RAF wild type, as determined by next generation sequencing (NGS) of tumor tissue (primary or metastatic). In some aspects, the subject has not received a prior therapy using an anti-EGFR therapy. In other words, the subject has not received (naive) prior anti-EGFR therapy. In some aspects, the subject has radiographically confirmed progression of the disease, which occurred within six months during or after the prior (first-line) chemotherapy.

[0064] In some aspects, the cancer is a cancer that expresses EGFR. In some aspects, the cancer is a cancer that expresses LGR5. In some aspects, the cancer is a cancer that expresses EGFR and LGR5.

[0065] In some aspects, the cancer is gastric cancer that expresses EGFR. In some aspects, the cancer is esophageal cancer that expresses EGFR. In some aspects, the cancer is gastroesophageal junction cancer that expresses EGFR. In some aspects, the cancer is head and neck cancer that expresses EGFR. In some aspects, the cancer is non-small cell lung cancer that expresses EGFR. In some aspects, the cancer is colorectal cancer that expresses EGFR. In some aspects, the cancer is squamous cell carcinoma of the head and neck (SCCHN) that expresses EGFR.

[0066] As used herein, a cancer expresses EGFR if it contains cells that express EGFR. Cells that express EGFR contain detectable levels of RNA encoding EGFR. In some aspects, EGFR expression is measured by ISH. As used herein, a cancer expresses LGR5 if it contains cells that express LGR5. Cells that express LGR5 contain detectable levels of RNA encoding LGR5.

[0067] In some aspects, EGFR protein expression is detected by IHC. In some aspects, EGFR expression is detected using a commercially available EGFR detection kit, such as the EGFR pharmDx for Dako automated staining (Agilent). TM The assay was performed by IHC using the manufacturer's recommendations or a commercially available IHC EGFR detection kit based on EGFR clone 113 (which binds to the EGFR extracellular domain) (Leica, https: / / shop.leicabiosystems.com / us / ihc-ish / ihc-primary-antibodies / pid-epidermal-growth-factor-receptor). Alternatively, a Novocastra kit based on clone EGFR.113 was used. TM EGFR expression was determined using a liquid mouse monoclonal antibody against EGFR (product code: NCL-L-EGFR, EGFR-IHC primary antibody, from leicabiosystems.com).

[0068] In brief, the commercially available EGFR pharmDx TM The IHC kit system contains the reagents necessary to perform routine IHC staining procedures on fixed, paraffin-embedded samples. Following incubation with a primary, non-Her2, Her3, and Her4 cross-reactive monoclonal antibody (clone 2-18C9) against the human EGFR protein, the kit utilizes a ready-to-use visualization reagent based on dextran technology. This reagent consists of a secondary goat anti-mouse antibody molecule and a horseradish peroxidase molecule linked to a common dextran polymer backbone. The enzymatic conversion of the chromogen subsequently occurs, resulting in the formation of a visible reaction product at the antigenic site. Results are typically assessed using light microscopy. Control slides containing two formalin-fixed, paraffin-embedded human cell lines, scored as 2+ and 0 for staining intensity, serve as quality control for kit reagent performance.

[0069] Staining intensity was determined as follows: 3+ (strong staining): visible at low magnification, x5 objective, and confirmed at higher levels as needed; 2+ (moderate staining): visible at moderate magnification, x10 or x20 objective; 1+ (weak staining): reliably confirmed only at high magnification, x40 objective; 0 (no staining): no staining visible at high magnification.

[0070] In some aspects, EGFR expression is determined using immunohistochemistry (IHC), and the cancer is IHC positive for EGFR. In some aspects, the cancer is characterized by an EGFR IHC score of 2+ or 3+. In some aspects, the cancer is characterized by an H-score for EGFR greater than 50, greater than 80, or greater than 200, but not greater than 300.

[0071] In certain aspects, EGFR expression is determined using immunohistochemistry (IHC), after which an H-score for EGFR is assigned using a scale of 0-300. In certain aspects, the cancers of the present disclosure are characterized by an H-score for EGFR greater than 200 on a scale of 0-300. In certain aspects, the H-score for EGFR is therefore between 200 and 300. In certain aspects, the cancers of the present disclosure are cancers characterized by an H-score for EGFR greater than 50 on a scale of 0-300. In certain aspects, the H-score for EGFR is therefore between 50 and 300. In certain aspects, the cancers of the present disclosure are head and neck cancers characterized by an H-score for EGFR greater than 80 on a scale of 0-300. In certain aspects, the H-score for EGFR is therefore between 80 and 300.

[0072] Here, determining the H score to specify the EGFR expression status involves a first step of determining the membrane staining intensity (results are scored as 0, 1+, 2+, or 3+), which is measured for each cell within the predefined field of view described herein. Subsequently, the percentage of cells at each staining intensity level is calculated, and finally the H score is assigned using the following formula: [1×(% of cells with 1+ staining)+2×(% of cells with 2+ staining)+3×(% of cells with 3+ staining)], resulting in an H score for EGFR ranging from 0 to 300. Therefore, the H score gives greater relative weight to higher staining intensity or amount of staining in a given tumor sample.

[0073] Optionally, treatment with an antibody or a functional portion, derivative, and / or analog thereof includes (or in some aspects is preceded by) a step of diagnosing the subject's EGFR status. In some aspects, subjects with an IHC score of 3+ or cancer characterized by an EGFR H score greater than 200 in the range of 0-300 are selected for treatment. In some aspects, a step of diagnosing the subject's EGFR H score as greater than 50 in the range of 0-300 is performed before treating the subject. In some aspects, a step of diagnosing the subject's EGFR H score as greater than 80 in the range of 0-300 is performed before treating the subject. In some aspects, a step of diagnosing the subject's EGFR H score as greater than 200 in the range of 0-300 is performed before treating the subject. After diagnosis, subjects whose scores fall within the selected range (i.e., 50-300, 80-300, or 200-300) are selected for treatment according to the present disclosure.

[0074] In certain aspects, the cancer expresses LGR5. As used herein, a cancer expresses LGR5 if the cancer comprises cells that express LGR5. Cells that express LGR5 comprise detectable levels of RNA encoding LGR5.

[0075] Expression can also be detected by incubating cells with antibodies that bind to LGR5. However, some cells do not express enough protein to be tested for such antibody testing. In this case, detection using mRNA or other forms of nucleic acid sequences is preferred. In some aspects, LGR5 is detected by mRNA expression. In some aspects, LGR5 detection is performed by RNA sequencing. In some aspects, LGR5 detection is performed by tissue microarray (TMA) staining. In some aspects, LGR5 expression is measured using in situ hybridization (ISH). Therefore, preferably, the cancer is ISH positive for LGR5. ISH positive preferably means that the expression is characterized by an H score of 1 or higher.

[0076] Detection and scoring techniques based on TMA, ISH, and IHC are well known to those skilled in the art and are generally commercially available as standard kits. For example, for LGR5, mRNA levels are quantified using ISH and expressed as an H score, which can be expressed using commercially available kits such as Advanced Cell Diagnostics (Hayward, CA, USA). The ISHH score for LGR5 quantification ranges from 0 to 400. Alternatively, LGR5 expression can be determined by RNA sequencing (RNAseq).

[0077] In certain aspects, the cancer expresses LGR5 at a level sufficient for an antibody to bind to the LGR5 protein, such as an antibody comprising a variable domain comprising the amino acid sequence of the VH chain of MF5816 as described in Figure 3 , or a VH chain of other variable domains that bind to LGR5 listed herein, that binds LGR5. In certain aspects, the cancer expresses EGFR at a level sufficient for an antibody to bind to the EGFR protein, such as an antibody comprising a variable domain comprising the amino acid sequence of the VH chain of MF3755 as described in Figure 3 , or a VH chain of other variable domains that bind to EGFR listed herein, that binds EGFR.

[0078] Mutations and models

[0079] Cancers such as gastric cancer, esophageal cancer, gastroesophageal junction cancer, head and neck cancer, non-small cell lung cancer, or colorectal cancer may be associated with the presence of mutations. Such mutations include mutations in known oncogenes such as PIK3CA, RAS, KRAS, HRAS, MAP2K1, RAF, BRAF, and NOTCH1. Oncogenic mutations are often described as activating mutations or mutations that result in new functions. Another type of cancer mutation involves tumor suppressor genes such as TP53, MLH1, CDKN2A, and PTEN. Mutations in tumor suppressor genes are often inactivating mutations.

[0080] In certain aspects, the cancer and / or the subject suffering from the cancer has a mutation in one or more of the WNT, RTK / RAS, TP53 and / or TGF-β signaling pathway genes.

[0081] In some aspects, the cancer is colorectal cancer and is wild-type for RAS and / or RAF, including Kristen rat sarcoma (KRAS) and / or beta-rapidly accelerated fibrosarcoma (BRAF). In some aspects, the colorectal cancer is genotypically wild-type for RAS and / or RAF, including KRAS and / or BRAF. In some aspects, the cancer is mCRC and is wild-type for RAS and / or RAF, including Kristen rat sarcoma (KRAS) and / or beta-rapidly accelerated fibrosarcoma (BRAF). In some aspects, the mCRC is genotypically wild-type for RAS and / or RAF, including KRAS and / or BRAF.

[0082] In certain aspects, for subjects suffering from mCRC, a screening step is included to detect (and optionally exclude suffering from) somatic mutations in RAS and RAF family genes (i.e., KRAS, NRAS, HRAS, BRAF, ARAF, RAF1). In certain aspects, a screening step is included to detect mutations or other oncogenic drivers associated with resistance to antibody-based EGFR inhibition, such as EGFR extracellular domain or ERBB2 / HER2 amplification.

[0083] In some aspects, the cancer has an oncogenic mutation in one or more genes associated with colorectal cancer. In some aspects, the cancer has an oncogenic mutation in a gene selected from APC, FBXW7, KRAS, PIK3CA, SOX9, TP53, SMAD2, SMAD3, SMAD4, NRAS, or TCF7L2, and the corresponding encoded proteins. In some aspects, the cancer has a mutation in a gene selected from APC, FBXW7, KRAS, PIK3CA, SOX9, TP53, BRAF, SMAD2, SMAD3, SMAD4, NRAS, or TCF7L2, and the corresponding encoded proteins.

[0084] In certain aspects, the cancer has a mutation in the gene encoding APC. In certain aspects, the mutation is a missense mutation, a nonsense mutation, a frameshift mutation, an insertion, a deletion, or a mutation that results in a copy number change. In certain aspects, the mutation is a monoallelic loss-of-function mutation, a biallelic loss-of-function mutation, or a heterozygous loss-of-function mutation. In certain aspects, the mutation is a biallelic loss-of-function mutation. In certain aspects, APC contains a monoallelic loss-of-function missense mutation, R653K, in its protein structure, which results in an R>K amino acid change. In certain aspects, the mutation is a heterozygous loss-of-function mutation, which results in a copy number change. In certain aspects, APC contains a biallelic loss-of-function mutation at position 4704, resulting in a frameshift at position P1442fs*31 in its protein. In certain aspects, APC contains a biallelic loss-of-function mutation at positions 5040-5041, resulting in a frameshift at position T1556fs*3 in its protein. In certain aspects, APC comprises a mutation at position 4565-4566, resulting in a frameshift at position F1396fs*1 in its protein.

[0085] Any mutations mentioned herein follow standard nomenclature as understood by the skilled artisan. As mentioned herein in the context of a mutation, it will be clear to the skilled artisan that an asterisk (*) indicates the presence of a stop codon, thereby terminating translation. Additionally, frameshift mutations are described herein in the form X1fs*X2, where X1 represents the first affected amino acid in the protein and *X2 represents the position of the translation stop codon in the new reading frame.

[0086] In certain aspects, the cancer has a mutation in the gene encoding FBXW7. In certain aspects, the mutation is a missense mutation or a copy number change mutation. In certain aspects, the mutation is a monoallelic loss-of-function or heterozygous loss-of-function mutation.

[0087] In some aspects, the cancer has a mutation in the gene encoding KRAS. In some aspects, the mutation is a missense mutation. In some aspects, the mutation is a gain-of-function mutation. In some aspects, KRAS comprises a G12V mutation in its protein structure, resulting in a G>V amino acid change. In some aspects, KRAS comprises a Q61H mutation in its protein structure, resulting in a Q>H amino acid change. In some aspects, KRAS comprises a G12S mutation in its protein structure, resulting in a G>S amino acid change.

[0088] In some aspects, the cancer has a mutation in the gene encoding PIK3CA. In some aspects, the mutation is a missense mutation. In some aspects, the mutation is a gain-of-function mutation. In some aspects, PIK3CA includes a gain-of-function missense mutation E545K in its protein structure, resulting in an E>K amino acid change.

[0089] In some aspects, the cancer has a mutation in the gene encoding OX9. In some aspects, the mutation is a nonsense mutation or a frameshift mutation. In some aspects, the mutation is a monoallelic loss-of-function mutation. In some aspects, SOX9 contains a monoallelic loss-of-function nonsense mutation Q412* in its protein structure, resulting in a Q>* amino acid change. In some aspects, SOX9 contains a monoallelic loss-of-function mutation at positions 1898-1899, resulting in a frameshift at positions *510fs*68 in its protein.

[0090] In some aspects, the cancer has a mutation in the gene encoding TP53. In some aspects, the mutation is a missense mutation or a nonsense mutation. In some aspects, the mutation is a monoallelic loss-of-function mutation or a biallelic loss-of-function mutation. In some aspects, TP53 includes a missense mutation N239D in its protein structure, resulting in an N>D amino acid change. In some aspects, TP53 includes a biallelic loss-of-function nonsense mutation Y126* in its protein structure, resulting in a Y>* amino acid change. In some aspects, TP53 includes a monoallelic loss-of-function missense mutation R282W in its protein structure, resulting in an R>W amino acid change. In some aspects, TP53 includes a biallelic loss-of-function missense mutation M237I in its protein structure, resulting in an M>I amino acid change.

[0091] In some aspects, the cancer has a mutation in the gene encoding MAD4. In some aspects, the mutation is a missense mutation or a loss-of-heterozygosity mutation. In some aspects, SMAD4 comprises a missense mutation G510V in its protein structure, resulting in a G>V amino acid change.

[0092] In some aspects, the cancer has a mutation in a gene selected from SMAD2, SMAD3, SMAD4, NRAS or TCF7L2. In some aspects, the mutation is a loss of heterozygous mutation. In some aspects, the mutation causes a copy number change.

[0093] In some aspects, the present disclosure relates to a method for treating cancer in a subject, wherein the subject has a mutation in one or more genes selected from APC, SOX9, KRAS, NRAS, SMAD2 or SMAD4. In some aspects, cancer has a mutation in APC, SOX9, KRAS, NRAS, SMAD2 and SMAD4 genes. In some aspects, the mutation encoding the APC gene is a loss-of-function missense mutation R653K in its protein structure, resulting in an R>K amino acid change. In some aspects, the mutation encoding the APC gene is a mutation at positions 4565-4566, resulting in a frame shift at position F1396fs*1 in its protein. In some aspects, the mutation encoding the SOX9 gene is a loss-of-function nonsense mutation Q412* in its protein structure, resulting in a Q>* amino acid change. In some aspects, the mutation encoding the KRAS gene is a mutation G12V in its protein structure, resulting in a G>V amino acid change. In certain aspects, the mutations in the NRAS, SMAD2, and SMAD4 genes are loss-of-heterozygosity mutations, resulting in copy number changes.

[0094] In certain aspects, the present disclosure relates to a method of treating cancer in a subject having a mutation in APC.

[0095] In certain aspects, the present disclosure relates to a method for treating cancer in a subject, wherein the subject has a mutation in one or more genes selected from TP53, APC, NRAS, SMAD2, SMAD4, or FBXW7. In certain aspects, the cancer has a mutation in the TP53, APC, NRAS, SMAD2, SMAD4, and FBXW7 genes. In certain aspects, the mutation in the genes encoding APC, NRAS, SMAD2, SMAD4, and FBXW7 is a heterozygous loss-of-function mutation, resulting in a copy number change. In certain aspects, the mutation in the TP53 gene encoding is a missense mutation N239D in its protein structure, resulting in an N>D amino acid change. In certain aspects, the mutation in the TP53 gene encoding is a biallelic loss-of-function nonsense mutation Y126* in its protein structure, resulting in a Y>* amino acid change.

[0096] In certain aspects, the present disclosure relates to a method for treating cancer in a subject, wherein the subject has a mutation in one or more genes selected from TP53, APC, KRAS or PIK3CA. In certain aspects, the cancer has a mutation in the TP53, APC, KRAS and PIK3CA genes. In certain aspects, the mutation encoding the APC gene is a loss-of-function mutation of the double allelic gene at position 4704, resulting in a frame shift of the P1442fs*31 position in its protein. In certain aspects, the mutation encoding the KRAS gene is a mutation G12S in its protein structure, resulting in a G>S amino acid change. In certain aspects, the mutation encoding the PIK3CA gene is a gain-of-function missense mutation E545K in its protein structure, resulting in an E>K amino acid change. In certain aspects, the mutation encoding the TP53 gene is a loss-of-function missense mutation R282W in its protein structure, resulting in an R>W amino acid change.

[0097] In some aspects, the disclosure relates to a method for treating a subject's cancer, wherein the subject has a mutation of one or more genes selected from TP53, APC, SOX9, KRAS, PIK3CA, TCF7L2, NRAS, SMAD2, SMAD3 or SMAD4. In some aspects, cancer has a mutation in TP53, APC, SOX9, KRAS, PIK3CA, TCF7L2, NRAS, SMAD2, SMAD3 and SMAD4 genes. In some aspects, the mutation encoding APC, TCF7L2, NRAS, SMAD2, SMAD4 and TP53 genes is a heterozygous loss-of-function mutation, resulting in a copy number change. In some aspects, the mutation encoding APC gene is a double allelic loss-of-function mutation at position 5040-5041, resulting in a frame shift at position T1556fs*3 in its protein. In some aspects, the mutation encoding the SOX9 gene is a monoallelic loss-of-function mutation at position 1898-1899, resulting in a frame shift at position *510fs*68 in its protein. In some aspects, the mutation encoding the KRAS gene is a mutation Q61H in its protein structure, resulting in a Q>H amino acid change. In some aspects, the mutation encoding the PIK3Ca gene is a gain-of-function missense mutation E545K in its protein structure, resulting in an E>K amino acid change. In some aspects, the mutation encoding the SMAD4 gene is a missense mutation G510V in its protein structure, resulting in a G>V amino acid change. In some aspects, the mutation encoding the TP53 gene is a biallelic loss-of-function missense mutation M237I in its protein structure, resulting in an M>I amino acid change.

[0098] Standard of care molecules disclosed herein and their administration

[0099] Fluoropyrimidine

[0100] In certain aspects, the antibody or functional portion, derivative and / or analog thereof is used in a therapy further comprising administering a fluoropyrimidine. In certain aspects, the method of treating cancer in a subject further comprises administering to the subject an effective amount of a fluoropyrimidine.

[0101] Fluoropyrimidines are antimetabolites that inhibit thymidylate synthase and reduce the production of the pyrimidine thymidine. Without being bound by any theory, it is believed that fluoropyrimidines interfere with DNA synthesis, and to a lesser extent RNA synthesis, leading to cell death in rapidly growing cells.

[0102] Examples of fluoropyrimidines suitable for use in humans are capecitabine, carmofur (HCFU), doxifluridine, 5-FU, and tegafur. In certain aspects, as used herein, "fluoropyrimidines" include, but are not limited to, capecitabine, carmofur (HCFU), doxifluridine, 5-FU, and tegafur.

[0103] In certain aspects, the fluoropyrimidine is fluorouracil.

[0104] In certain aspects, the present disclosure provides pesentuzumab for use in treating cancer in a subject, wherein the treatment further comprises administering fluorouracil. In certain aspects, the present disclosure further provides a method of treating cancer in a subject, comprising administering to the subject an effective amount of pesentuzumab and fluorouracil.

[0105] Fluorouracil is a fluoropyrimidine with the chemical formula 5-fluoro-2,4(1H,3H)-pyrimidinedione. Fluorouracil is converted into three major active metabolites: 5-fluoro-2'-deoxyuridine-5'-monophosphate (FdUMP), 5-fluorouridine-5'-triphosphate (FUTP), and 5-fluoro-2'-deoxyuridine-5'-triphosphate (FdUTP). These metabolites have several effects, including inhibition of thymidylate synthase by FdUMP, incorporation of FUTP into RNA, and incorporation of FdUTP into DNA.

[0106] Fluorouracil is known by several synonyms, the most common of which are 5-fluorouracil, 5-fluoropyrimidine-2,4-dione, 5-FU, and 5-fluorouracil. Fluorouracil is sold under the trade name Adrucil, among others. Its IUPAC name is 5-fluoropyrimidine-2,4(1H,3H)-dione. It is used intravenously to treat colorectal cancer, esophageal cancer, gastric cancer, pancreatic cancer, breast cancer, and cervical cancer. As a cream, it is used to treat actinic keratosis, basal cell carcinoma, and skin warts.

[0107] Fluorouracil and other human fluoropyrimidines have been used clinically for a considerable time, and appropriate treatment regimens and dosage information are available to those of ordinary skill in the art. Fluorouracil can be administered by intravenous injection (such as bolus, infusion, or continuous infusion) for up to several days. Fluorouracil can be administered as a monotherapy, in combination with leucovorin alone, or in combination with leucovorin and oxaliplatin or irinotecan. Fluorouracil can also be administered as a component of a cyclophosphamide-based multidrug regimen or as a component of a platinum-containing multidrug chemotherapy regimen.

[0108] In certain aspects, the dosing regimen of fluorouracil comprises 200-3000 mg / m 2 Fluorouracil was administered intravenously within a range of 1:1.

[0109] In certain aspects, the dosing regimen of fluorouracil comprises 400 mg / m2 on day 1. 2 Intravenous bolus, followed by 2400 mg / m2 over 46 hours every 2 weeks 2 Up to 3000 mg / m 2 Fluorouracil was administered as an intravenous continuous infusion in combination with folinic acid alone or in combination with folinic acid and oxaliplatin or irinotecan.

[0110] In certain aspects, the dosing regimen of fluorouracil comprises 500 mg / m2 on days 1, 8, 15, 22, 29, and 36 of an 8-week cycle. 2 Fluorouracil in combination with folinic acid was administered as an intravenous bolus.

[0111] In certain aspects, the dosing regimen of fluorouracil comprises 500 mg / m2 on days 1 and 8 of every 28 days for 6 cycles. 2 Up to 600 mg / m 2 Fluorouracil was administered intravenously as a component of a cyclophosphamide-based multidrug regimen.

[0112] In certain aspects, the dosing regimen of fluorouracil comprises 200 mg / m2 over 24 hours. 2 Up to 1000 mg / m 2Fluorouracil is administered as an intravenous continuous infusion as a component of a platinum-containing multi-drug chemotherapy regimen.

[0113] In certain aspects, the dosing regimen of fluorouracil comprises 400 mg / m2 on day 1. 2 Intravenous bolus, followed by 2400 mg / m2 over 46 hours every 2 weeks 2 Fluorouracil is administered by continuous intravenous infusion in combination with folinic acid or as a component of a multidrug chemotherapy regimen that includes folinic acid.

[0114] TAS-102

[0115] In certain aspects, the antibody or functional part, derivative and / or analog thereof is used in a therapy further comprising administering a fluoropyrimidine. In certain aspects, the method of treating cancer in a subject further comprises administering an effective amount of a fluoropyrimidine.

[0116] In certain aspects, the antibody or functional portion, derivative and / or analog thereof is used in a therapy that further comprises administering a thymidine phosphorylase inhibitor. In certain aspects, the method of treating cancer in a subject further comprises administering an effective amount of thymidine phosphorylase.

[0117] In certain aspects, the antibody or its functional portion, derivative and / or analog is used in a treatment that further comprises administering a fluoropyrimidine and a thymidine phosphorylase inhibitor. In certain aspects, the method of treating cancer in a subject further comprises administering to the subject an effective amount of a fluoropyrimidine and an effective amount of thymidine phosphorylase.

[0118] In certain aspects, the fluoropyrimidine is trifluridine. In certain aspects, the thymidine phosphorylase inhibitor is tipiracil hydrochloride.

[0119] In certain aspects, the present disclosure provides pesentuzumab for use in treating cancer in a subject, wherein the treatment further comprises administering trifluridine and tipiracil hydrochloride. In certain aspects, the present disclosure further provides a method of treating cancer in a subject, comprising administering to the subject an effective amount of pesentuzumab, trifluridine, and tipiracil hydrochloride.

[0120] In certain aspects, the combination of trifluorothymidine and tipiracil in a molar ratio of 1:0.5 is TAS-102.

[0121] Trifluridine is a thymidine nucleoside analog with anti-malignant activity, and its chemical formula is 2'-deoxy-5-(trifluoromethyl)uridine. Tipiracil hydrochloride is a thymidine phosphorylase inhibitor, and its chemical formula is 5-chloro-6-[(2-iminopyrrolidin-1-yl)methyl]pyrimidine-2,4-(1H,3H)-dione monohydrochloride or 2,4(1H,3H)-pyrimidinedione, 5-chloro-6-[(2-imino-1-pyrrolidinyl)methyl]-, hydrochloride (1:1). TAS-102 contains trifluridine and tipiracil in a molar ratio of 1:0.5 (weight ratio, 1:00471). Without being bound by theory, it is believed that the inclusion of tipiracil increases trifluridine exposure by inhibiting its metabolism by thymidine phosphorylase. After uptake into cancer cells, trifluridine is incorporated into DNA, interferes with DNA synthesis, and inhibits cell proliferation.

[0122] TAS-102 is known by various synonyms, the most common of which are Lonsurf, TAS102, TAS-102 (trifluridine / tipiracil HCl), the hydrochloride salt of 4-hydroxy-1-((2R,4S,5R)-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-(trifluoromethyl)pyrimidin-2(1H)-one and 5-chloro-6-((2-iminopyrrolidin-1-yl)methyl)pyrimidine-2,4-diol (1:1), a mixture of Viroptic and 5-CIMU, a trifluridine-tipiracil hydrochloride mixture, tipiracil / trifluridine, and EX-A1755. TAS-102 is sold under the trade name Lonsurf.

[0123] TAS-102 has been in clinical use for a considerable period of time, and appropriate treatment regimens and dosage information are readily available to those of ordinary skill in the art. TAS-102 is indicated as a monotherapy for the treatment of adult patients with metastatic colorectal cancer (CRC) who have previously received or are not considered candidates for existing therapies, including fluoropyrimidine-, oxaliplatin-, and irinotecan-based chemotherapy, anti-VEGF agents, and anti-EGFR agents. TAS-102 is also indicated as a monotherapy for the treatment of adult patients with metastatic gastric cancer (including gastroesophageal junction adenocarcinoma) who have previously received at least two prior systemic treatment regimens for advanced disease.

[0124] In certain aspects, the dosing regimen of TAS-102 comprises 35-80 mg / m2 twice daily. 2 Trifluridine and tipiracil were administered at a molar ratio of 1:0.5.

[0125] In certain aspects, the dosing regimen of TAS-102 comprises 35 mg / m2 twice daily on days 1 to 5 and days 8 to 12 of each 28-day cycle.2 Trifluridine and tipiracil are administered orally at a molar ratio of 1:0.5, starting at a dose of 1 mg / dose to a maximum dose of 80 mg / dose, for as long as benefit is observed or until unacceptable toxicity occurs.

[0126] Oxaliplatin

[0127] In certain aspects, the antibody or functional portion, derivative and / or analog thereof is used in a therapy further comprising administering a platinum-based chemotherapeutic agent. In certain aspects, the method of treating cancer in a subject further comprises administering to the subject an effective amount of a platinum-based chemotherapeutic agent.

[0128] Platinum-based chemotherapeutics are widely used anti-tumor drugs for the treatment of cancer. They are coordination complexes of platinum. Without being bound by any theory, it is believed that platinum-based chemotherapeutics cause DNA crosslinking, which inhibits DNA repair and / or DNA synthesis.

[0129] Examples of platinum-based chemotherapeutic agents suitable for use in humans are cisplatin, oxaliplatin, carboplatin, nedaplatin, triplatin tetranitrate, phenanthriplatin, picoplatin, and satraplatin. In certain aspects, as used herein, "platinum-based chemotherapeutic agents" include, but are not limited to, cisplatin, oxaliplatin, carboplatin, nedaplatin, triplatin tetranitrate, phenanthriplatin, picoplatin, and satraplatin. While cisplatin, oxaliplatin, nedaplatin, and carboplatin are approved and widely used to treat several different cancer indications, triplatin tetranitrate, phenanthriplatin, picoplatin, and satraplatin are currently in clinical trials.

[0130] In certain aspects, the platinum-based chemotherapeutic agent is cisplatin, oxaliplatin, or carboplatin. In certain aspects, the platinum-based chemotherapeutic agent is oxaliplatin.

[0131] In certain aspects, the present disclosure provides pesentuzumab for use in treating cancer in a subject, wherein the treatment further comprises administering oxaliplatin. In certain aspects, the present disclosure further provides a method of treating cancer in a subject, comprising administering to the subject an effective amount of pesentuzumab and oxaliplatin.

[0132] Oxaliplatin is a platinum-based drug with the molecular formula C8H 14N2O4Pt and its chemical name is cis-[(1R,2R)-1,2-diaminocyclohexane-N,N'][(2-)-O,O']platinum. Oxaliplatin is an organoplatinum complex in which the platinum atom is complexed with 1,2-diaminocyclohexane (DACH) and complexed with an oxalate ligand as a leaving group. Oxaliplatin undergoes nonenzymatic conversion to active derivatives in physiological solution via displacement of the labile oxalate ligand. These derivatives form inter- and intra-strand DNA crosslinks, which inhibit DNA replication and transcription.

[0133] Oxaliplatin is known by various synonyms, the most common of which are diaminocyclohexane oxaliplatin, L-OHP, oxalatoplatin, oxalatoplatinum, oxaliplatin, Dacplat, Eloxatin, or Elplat. Oxaliplatin's trade name is Eloxatin. Oxaliplatin, in combination with fluorouracil and leucovorin, is indicated for the treatment of stage III (Duke's C) colon cancer and metastatic colorectal cancer after complete resection of the primary tumor.

[0134] Oxaliplatin and other platinum-based chemotherapeutic agents have been used clinically for a considerable period of time, and appropriate treatment regimens and dosage information are readily available to those skilled in the art. In certain aspects, the dosing regimen of oxaliplatin comprises about 65-160 mg / m 2 Oxaliplatin was administered within the range of

[0135] In certain aspects, the dosing regimen of oxaliplatin comprises 85 mg / m2 every two weeks. 2 Administer oxaliplatin.

[0136] In certain aspects, oxaliplatin is administered every two weeks in combination with fluorouracil and folinic acid.

[0137] In certain aspects, 85 mg / m 2 With 200mg / m 2 Oxaliplatin was administered as an intravenous infusion over 120 minutes in combination with folinic acid, followed by 400 mg / m2 of oxaliplatin over 2-4 minutes. 2 Fluorouracil was administered intravenously as a bolus, followed by 600 mg / m 2 Fluorouracil was administered as a 22-hour continuous infusion.

[0138] Venetoclax

[0139] In certain aspects, the antibody or functional portion, derivative and / or analog thereof is used in a therapy further comprising administering a BCL-2 inhibitor. In certain aspects, the method of treating cancer in a subject further comprises administering to the subject an effective amount of a BCL-2 inhibitor.

[0140] B-cell lymphoma 2 (BCL-2) is part of a family of apoptosis regulators. BCL-2 is a pro-survival protein that, when overexpressed, protects cells from apoptosis. BCL-2 is overexpressed in many cancers and plays a key role in negatively regulating apoptosis. Its expression is associated with drug resistance and increased tumor cell survival.

[0141] Mimetics comprise a class of BCL-2 inhibitors that have shown promising results in several hematological malignancies, either as single agents or in combination with other anticancer drugs. Without being bound by any theory, it is believed that BH3 mimetics inhibit the activity of BCL-2 and restore the apoptotic process in tumor cells. Examples of BCL-2 inhibitors suitable for use in humans are ABT-737, navitoclax (ABT-263), and venetoclax.

[0142] In certain aspects, as used herein, "BCL-2 inhibitors" include, but are not limited to, ABT-737, navitoclax (ABT-263), or venetoclax.

[0143] In certain aspects, the BLC-2 inhibitor is venetoclax.

[0144] In certain aspects, the present disclosure provides pesentuzumab for use in treating cancer in a subject, wherein the treatment further comprises administering venetoclax. In certain aspects, the present disclosure further provides a method of treating cancer in a subject, comprising administering to the subject an effective amount of pesentuzumab and venetoclax.

[0145] Venetoclax is an anti-apoptotic protein and a potent, selective inhibitor of BCL-2. Venetoclax binds directly to the BH3-binding groove of BCL-2, displacing pro-apoptotic proteins containing the BH3 motif, such as BIM, to initiate mitochondrial outer membrane permeabilization (MOMP), caspase activation, and programmed cell death. In nonclinical studies, venetoclax has demonstrated cytotoxic activity in tumor cells that overexpress BCL-2.

[0146] Venetoclax is known by various synonyms, the most common of which are Venclexta, Venclyxto, ABT-199, GDC-0199, ABT199, ABT 199, and RG7601. Venclexta is indicated for the treatment of patients with chronic lymphocytic leukemia (CLL) or small lymphocytic lymphoma (SLL). Venclexta is also indicated in combination with azacitidine, decitabine, or low-dose cytarabine for the treatment of adults with newly diagnosed acute myeloid leukemia (AML) who are ineligible for intensive chemotherapy.

[0147] Venetoclax has been used clinically for a considerable period of time, and appropriate treatment regimens and dosage information are readily available to persons of ordinary skill in the art.

[0148] In certain aspects, venetoclax is administered at a dose of 20-1200 mg per day.

[0149] In certain aspects, venetoclax is administered in a progressive or staged increasing dosage regimen. In certain aspects, the initial dose of venetoclax is gradually increased in a progressive increasing dosage manner until the full dose is reached. In certain aspects, the progressive increasing dosage is administered at regular intervals.

[0150] In certain aspects, venetoclax is administered at an initial dose of 20 mg in week 1, followed by a dose of 50 mg in week 2, followed by a dose of 100 mg in week 3, followed by a dose of 200 mg in week 4, and followed by a dose of 400 mg in week 5.

[0151] In certain aspects, venetoclax is administered at a dose of 400 mg daily following the escalating dosing regimen described herein. In certain aspects, venetoclax is administered at a dose of 400 mg once daily until disease progression or unacceptable toxicity is observed.

[0152] SN-38

[0153] In some aspects, the antibody or a functional portion, derivative, and / or analog thereof is used in a therapy that further comprises administering SN-38. In some aspects, the method of treating cancer in a subject further comprises administering an effective amount of SN-38 to the subject.

[0154] SN-38 is 7-ethyl-10-hydroxycamptothecin and is a compound that inhibits DNA topoisomerase 1 activity. It is a biologically active and water-insoluble metabolite of irinotecan (CPT-11). Without being bound by any theory, it is believed that irinotecan is converted to SN-38 by carboxylesterases in the liver and tumors. SN-38 has been shown to have 1,000-fold higher cytotoxic activity than irinotecan against various cancer cells in vitro. However, the metabolic conversion rate is very low, with only <10% of the original volume of irinotecan being metabolized to SN-38; the efficiency of irinotecan conversion to SN-38 also depends on individual genetic variation in carboxylesterase activity. Direct administration of SN-38 itself for clinical cancer treatment would obviate the need for conversion. SN-38 is also known to be comparable to irinotecan in efficacy and toxicity (Nakajima TE et al., Int J Cancer. 2008 May 1; 122(9): 2148-53). The skilled artisan will understand that references herein to “administering SN-38 to a subject” do not encompass administering to a subject irinotecan, which can subsequently be converted to SN-38. The skilled artisan will understand that all references herein to “administering SN-38 to a subject” also encompass administering to a subject a pharmaceutical composition comprising SN-38.

[0155] SN-38 is known by various synonyms, the most common of which are 7-ethyl-10-hydroxycamptothecin, SN-38, SN38, SN-38 lactone, NK012, NK-012, 7-ethyl-10-hydroxy-20(S)-camptothecin, and 10-hydroxy-7-ethylcamptothecin. Its IUPAC name is (4S)-4,11-diethyl-4,9-dihydroxy-1,4-dihydro-3H,14H-pyrano[3',4':6,7]indolizino[1,2-b]quinoline-3,14-dione. LE-SN-38 is a novel liposomal formulation containing liposomes with a uniform size distribution (<200 nm). The drug encapsulation efficiency of this formulation is >95%.

[0156] In certain aspects, the present disclosure provides pesentuzumab for use in treating cancer in a subject, wherein the treatment further comprises administering SN-38. In certain aspects, the present disclosure further provides a method of treating cancer in a subject, comprising administering to the subject an effective amount of pesentuzumab and SN-38.

[0157] In certain aspects, SN-38 is administered encapsulated or conjugated to a suitable carrier, including a soluble polymer, liposomes, micelles, antibodies, peptides, polymer-drug conjugates, nanoparticles, or polymer implants.

[0158] FOLFOX

[0159] In certain aspects, the antibody or its functional portion, derivative and / or analog is used in a treatment that further comprises administering oxaliplatin, folinic acid (leucovorin) and fluorouracil. In certain aspects, the method of treating cancer in a subject further comprises administering to the subject an effective amount of oxaliplatin, folinic acid and fluorouracil.

[0160] In certain aspects, the present disclosure provides pesentuzumab for use in treating cancer in a subject, wherein the treatment further comprises administering FOLFOX. In certain aspects, the present disclosure further provides a method of treating cancer in a subject, comprising administering to the subject an effective amount of pesentuzumab and FOLFOX.

[0161] In certain aspects, the combination of oxaliplatin, folinic acid, and fluorouracil is the FOLFOX regimen. FOLFOX is known by various synonyms, the most common of which are the FOLFOX regimen, the FOLFOX treatment regimen, FOLFOX-4, FOLFOX-6, FOLFOX-7, and the fluorouracil regimen with folinic acid and oxaliplatin. Folinic acid is also known by synonyms such as folinic acid, FA, or calcium folate.

[0162] In certain aspects, the present disclosure provides pesentuzumab for use in treating cancer in a subject, wherein the treatment further comprises administering oxaliplatin, folinic acid, and fluorouracil. In certain aspects, the present disclosure further provides a method of treating cancer in a subject, comprising administering to the subject an effective amount of pesentuzumab, oxaliplatin, folinic acid, and fluorouracil.

[0163] In certain aspects, an antibody or a functional portion, derivative, and / or analog thereof is used in a treatment that further comprises administering FOLFOX (fluorouracil, folinic acid, and oxaliplatin). In certain aspects, the subject or cancer being treated is RAS and / or RAF wild-type (including Kristen rat sarcoma (KRAS) and / or B-rapidly accelerated fibrosarcoma (BRAF)). In certain aspects, the genomics of the mCRC is RAS and / or RAF wild-type, including KRAS and / or BRAF. In certain aspects, the subject to be administered FOLFOX has received only a single prior anticancer treatment, such as chemotherapy for a metastatic setting, such as a fluoropyrimidine-irinotecan-based first-line chemotherapy with or without bevacizumab.

[0164] In certain aspects, the dosing regimen of FOLFOX comprises intravenous administration of oxaliplatin and folinic acid, followed by intravenous administration of fluorouracil.

[0165] In certain aspects, the FOLFOX dosing regimen includes oxaliplatin at 50-200 mg / m2 The dose of folinic acid is 200-600mg / m 2 administrated intravenously at a dose of 1200-3600 mg / m 2 The dose was administered intravenously.

[0166] In certain aspects, the FOLFOX dosing regimen includes oxaliplatin at 85 mg / m 2 and folinic acid at 400 mg / m 2 intravenously, followed by fluorouracil at 2400 mg / m 2 Intravenous administration.

[0167] In certain aspects, the FOLFOX dosing regimen comprises: on day 1, oxaliplatin at 85 mg / m 2 and folinic acid at 200 mg / m 2 Both were administered as a 2-hour infusion, followed by fluorouracil at 400 mg / m 2 A bolus injection was administered, followed by fluorouracil at 600 mg / m 2 Administer as a 22-hour infusion; on day 2, folinic acid 200 mg / m 2 Administer as a 2-hour infusion, followed by fluorouracil at 400 mg / m 2 A bolus injection was administered, followed by fluorouracil at 600 mg / m 2 Infusion is administered over 22 hours. In certain aspects, FOLFOX is administered every 2 weeks for 12 cycles.

[0168] In certain aspects, the FOLFOX dosing regimen comprises oxaliplatin at 85 mg / m2 on day 1 of a two-week cycle. 2 Administration (e.g., as an intravenous infusion over 2 hours), followed by or concurrently with folinic acid at 200 mg / m 2 Administration (e.g., as an intravenous infusion over 2 hours), followed by 5-FU at 400 mg / m 2 Administration (e.g., intravenous bolus administration), followed by 5-FU at 600 mg / m 2 In some aspects, the FOLFOX dosing regimen further comprises, on day 2 of a two-week cycle, folinic acid at 200 mg / m 2 Administration (e.g., as an intravenous infusion over 2 hours), followed by 5-FU at 400 mg / m 2 Administration (e.g., intravenous bolus administration), followed by 5-FU at 600 mg / m 2 Administration (e.g., as a continuous infusion over 22 hours).

[0169] In certain aspects, the FOLFOX dosing regimen includes oxaliplatin at 85 mg / m2 on day 1. 2 Fluorouracil was administered at 400 mg / m 2 Bolus injection and folinic acid at 400 mg / m 2 Administer as a 2-hour infusion followed by fluorouracil at 2400 mg / m 2 Administer as a 46-hour continuous infusion.

[0170] If a subject experiences an oxaliplatin-related adverse reaction (also known as an adverse event or AE) after starting any of the above-mentioned related methods, the method may involve adjusting the dosage regimen based on the approved oxaliplatin prescribing information. Dose adjustments for adverse reactions in advanced colorectal cancer are shown in Table 1.

[0171]

[0172]

[0173] Table 1: Suggested dose adjustments for oxaliplatin-related adverse reactions, according to the FDA-approved prescribing information for Eloxatin (oxaliplatin).

[0174] Adverse events should be managed according to their severity. For the following events, provide dose modifications and supportive measures as specified in the oxaliplatin prescribing information: allergic reactions, peripheral sensory neuropathy, acute neuropathy, delayed neuropathy, severe myelosuppression, posterior reversible encephalopathy syndrome (PRES), pulmonary toxicity, hepatotoxicity, QT prolongation and ventricular arrhythmia, rhabdomyolysis, and bleeding.

[0175] In certain aspects, the recommended dose adjustments for oxaliplatin drug-related adverse events follow the FDA-approved prescribing information for Eloxatin (Oxaliplatin for intravenous injection, revised 6 / 2023 or the current version at the time of administration) in Table 1.

[0176] In certain aspects, if a subject experiences a toxicity as mentioned in the table, the dose adjustment at the start of a subsequent cycle of therapy is adjusted relative to the starting dose used in the previous cycle following Tables 2 and 3.

[0177] In certain aspects, if the subject experiences acute and delayed neuropathy, the dose of oxaliplatin is reduced or permanently discontinued for persistent neurosensory reactions according to the schedule in Table 1, depending on the severity of the adverse reaction.

[0178] In certain aspects, if a subject experiences persistent Grade 2 neuropathy, oxaliplatin at 65 mg / m 2 In certain aspects, if the subject experiences persistent Grade 3 neuropathy, oxaliplatin is discontinued. In certain aspects, if the subject experiences Grade 4 neuropathy, oxaliplatin is discontinued.

[0179] In certain aspects, if the subject experiences Grade 4 neutropenia or febrile neutropenia, dosing is delayed until the neutrophil count is ≥ 1.5 × 10 9 / L and platelet count ≥75×10 9 In certain aspects, the oxaliplatin dose is (reduced to) 65 mg / m 2 In certain aspects, if the subject has Grade 4 thrombocytopenia, dosing is delayed until the neutrophil count is ≥ 1.5 × 10 9 / L and platelet count ≥75×10 9 In certain aspects, the oxaliplatin dose is (reduced to) 65 mg / m 2 .

[0180] In certain aspects, if a subject experiences a grade 3-4 gastrointestinal adverse reaction, the oxaliplatin dose is reduced after administration of oxaliplatin as part of the FOLFOX regimen and then resumed at 65 mg / m 2 The dose of fluorouracil was then reduced to 300 mg / m 2 Intravenous bolus and 500 mg / m 2 A 22-hour continuous infusion.

[0181] FOLFIRI

[0182] In certain aspects, the antibody or its functional portion, derivative and / or analog is used in a treatment that further comprises administering irinotecan, folinic acid and fluorouracil. In certain aspects, the method of treating cancer in a subject further comprises administering to the subject an effective amount of irinotecan, folinic acid and fluorouracil.

[0183] In certain aspects, the present disclosure provides pesentuzumab for use in treating cancer in a subject, wherein the treatment further comprises administering FOLFIRI. In certain aspects, the present disclosure further provides a method of treating cancer in a subject, comprising administering to the subject an effective amount of pesentuzumab and FOLFIRI.

[0184] In certain aspects, the combination of irinotecan, folinic acid, and fluorouracil is the FOLFIRI regimen. FOLFIRI is known by various synonyms, the most common being the FOLFIRI regimen, the FOLFIRI treatment regimen, 5-fluorouracil / folinic acid / irinotecan, and a mixture of irinotecan, folinic acid, and fluorouracil.

[0185] In certain aspects, an antibody or a functional portion, derivative and / or analog thereof is used in treatment, the treatment further comprising administering fluorouracil, folinic acid and irinotecan (FOLFIRI). In certain aspects, the subject or cancer being treated is RAS and / or RAF wild type (including Kristen rat sarcoma (KRAS) and / or B- rapidly accelerated fibrosarcoma (BRAF)). In certain aspects, the genome of the mCRC is RAS and / or RAF wild type, including KRAS and / or BRAF. In certain aspects, the subject being administered FOLFIRI has only received a single prior anticancer treatment, such as chemotherapy for a metastatic setting. In certain aspects, the prior treatment includes fluoropyrimidine-oxaliplatin-based chemotherapy with or without bevacizumab.

[0186] In certain aspects, the present disclosure provides pesentuzumab for use in treating cancer in a subject, wherein the treatment further comprises administering irinotecan, folinic acid, and fluorouracil. In certain aspects, the present disclosure further provides a method of treating cancer in a subject, comprising administering to the subject an effective amount of pesentuzumab, irinotecan, folinic acid, and fluorouracil.

[0187] In certain aspects, the dosing regimen of FOLFIRI comprises intravenous administration of irinotecan and leucovorin, followed by administration of fluorouracil.

[0188] In certain aspects, the dosing regimen of FOLFIRI comprises irinotecan at 180 mg / m 2 and folinic acid at 200-400 mg / m 2 Intravenous administration followed by fluorouracil at 400-2400 mg / m 2 Intravenous administration.

[0189] In certain aspects, the dosing regimen of FOLFIRI comprises irinotecan at 180 mg / m 2 and folinic acid at 200 mg / m 2 or 400 mg / m 2 intravenously, followed by fluorouracil at 400 mg / m 2 A bolus injection was administered, followed by fluorouracil at 2400 mg / m 2 It is administered as a 46-hour continuous infusion. In certain aspects, FOLFIRI administration is repeated every 14 days.

[0190] In certain aspects, the FOLFIRI regimen comprises irinotecan at 180 mg / m2 on day 1 of a 2-week cycle. 2 Administer as an intravenous infusion over 90 minutes, followed by or concurrently with folinic acid (leucovorin) at 200 mg / m 2 Administer as an intravenous infusion over 2 hours, followed by fluorouracil at 400 mg / m 2 Administer an intravenous bolus followed by fluorouracil at 600 mg / m 2 Administer as a 22-hour continuous infusion. On day 2 of a 2-week cycle: folinic acid 200 mg / m 2 Intravenous infusion over 2 hours, followed by fluorouracil at 400 mg / m 2 Intravenous bolus followed by fluorouracil at 600 mg / m 2 Give as a 22-hour continuous infusion.

[0191] If a subject has a drug-related adverse event (AE) after starting any of the above-mentioned related methods, the method may involve adjusting the dosage regimen based on the approved prescribing information for irinotecan (also known as irinotecan hydrochloride or Camptosar). Dose adjustments for adverse reactions in advanced colorectal cancer are shown in Table 2.

[0192]

[0193]

[0194] Table 2: Suggested dose modifications for irinotecan drug-related AEs.

[0195] Source: Approved prescribing information for irinotecan.

[0196] 1 National Cancer Institute – Common Toxicity Criteria (Version 1.0)

[0197] 2 Relative to the starting dose used in the previous cycle

[0198] 3 Preprocessing

[0199] 4 Does not include hair loss, anorexia, or weakness

[0200]

[0201] Table 3: Starting dose and dose adjustment levels for FOLFIRI.

[0202] Source: FDA-approved prescribing information for Camptosar (irinotecan hydrochloride). Camptosar = irinotecan hydrochloride; 5-FU = 5-fluorouracil; FOLFIRI = 5-FU, LV, and irinotecan; LV = folinic acid.

[0203] In certain aspects, the grading of adverse events (or also referred to herein as toxicity or adverse reactions) and their definitions follow Table 2, National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE) v.4.03 / v5.0, NCI-CTC v1.0, or the current version at the time of administration.

[0204] In certain aspects, the recommended dose modifications for irinotecan drug-related adverse events (AEs) are as described in the National Cancer Institute–Common Toxicity Criteria (Version 1.0).

[0205] In certain aspects, the recommended dose adjustments for irinotecan drug-related adverse events follow the approved Camptosar prescribing information (revised 1 / 2022 for intravenous injection or the current version at the time of administration).

[0206] In certain aspects, if a subject experiences a toxicity as mentioned in the table, the dose adjustment at the start of a subsequent cycle of therapy is adjusted relative to the starting dose used in the previous cycle following Tables 2 and 3.

[0207] In certain aspects, if the subject does not experience any toxicity, the administered dose level is maintained during the therapy cycle. In certain aspects, if the subject does not experience any toxicity, the dose level is maintained at the starting dose for subsequent therapy cycles.

[0208] In certain aspects, if a subject experiences Grade 1 neutropenia (1500 to 1999 / mm 3 ), the dosage level administered is maintained during the therapy cycle. In certain aspects, the dosage level is maintained at the beginning of subsequent therapy cycles.

[0209] In certain aspects, if a subject experiences Grade 2 neutropenia (1000 to 1499 / mm 3 ), the dosage level administered during the therapy cycle is reduced by 1 dosage level. In certain aspects, the dosage level is maintained at the start of subsequent therapy cycles.

[0210] In certain aspects, if a subject experiences Grade 3 neutropenia (500 to 999 / mm 3), the administered dose level is omitted during the therapy cycle until recovery to ≤ Grade 2, followed by reduction by 1 dose level. In certain aspects, the dose level is reduced by 1 dose level at the start of subsequent therapy cycles.

[0211] In certain aspects, if a subject experiences Grade 4 neutropenia (<500 mm 3 ), the administered dose is omitted during the therapy cycle until recovery to ≤ Grade 2, followed by a reduction of 2 dose levels. In certain aspects, the dose level is reduced by 2 dose levels at the start of subsequent therapy cycles.

[0212] In certain aspects, if the subject has neutropenic fever, the dose is omitted until recovery to ≤ Grade 2, followed by a 2-dose reduction.

[0213] In certain aspects, if a subject experiences other hematologic toxicities, dose adjustments during a treatment cycle and at the start of subsequent treatment cycles are also based on NCI toxicity criteria and are the same as those recommended above for neutropenia. In certain aspects, if a subject has leukopenia or thrombocytopenia, dose adjustments during a treatment cycle and at the start of subsequent treatment cycles are also based on NCI toxicity criteria and are the same as those recommended above for neutropenia.

[0214] In certain aspects, if a subject experiences Grade 1 diarrhea (2-3 bowel movements / day > pre-treatment), the dose administered is delayed during the therapy cycle until recovery to baseline, after which the subject is administered the same dose. In certain aspects, the dose administered is maintained at the start of subsequent therapy cycles.

[0215] In certain aspects, if a subject experiences Grade 2 diarrhea (4-6 bowel movements / day > pre-treatment), the administered dose is omitted during the therapy cycle until recovery to baseline, followed by a reduction of 1 dose level. In certain aspects, the administered dose is maintained at the start of subsequent therapy cycles.

[0216] In certain aspects, if a subject experiences Grade 3 diarrhea (7-9 bowel movements / day > pre-treatment), the administered dose is omitted during the therapy cycle until recovery to baseline, followed by a reduction of 1 dose level. In certain aspects, the dose administered at the start of a subsequent therapy cycle is reduced by 1 dose level.

[0217] In certain aspects, if a subject experiences Grade 4 diarrhea (>10 bowel movements / day>pretreatment), the administered dose is omitted during the therapy cycle until recovery to baseline, followed by a reduction of 2 dose levels. In certain aspects, the dose administered at the start of a subsequent therapy cycle is reduced by 2 dose levels.

[0218] In certain aspects, the subject may experience other non-hematologic toxicities. In certain aspects, the other non-hematologic toxicities do not include alopecia, anorexia, and asthenia.

[0219] In certain aspects, if the subject experiences other non-hematologic toxicity (Grade 1), the administered dose is maintained during the therapy cycle. In certain aspects, the administered dose is maintained at the start of subsequent therapy cycles.

[0220] In certain aspects, if a subject experiences other non-hematologic toxicity (Grade 2), the administered dose is omitted during the therapy cycle until recovery to Grade 1, followed by a reduction of 1 dose level. In certain aspects, the administered dose is maintained at the start of subsequent therapy cycles.

[0221] In certain aspects, if a subject experiences other non-hematologic toxicity (Grade 3), the administered dose is omitted during the therapy cycle until recovery to Grade 2, followed by a reduction of 1 dose level. In certain aspects, the administered dose is reduced by 1 dose level at the start of subsequent therapy cycles.

[0222] In certain aspects, if a subject experiences other non-hematologic toxicity (Grade 4), the administered dose is omitted during the therapy cycle until recovery to Grade 2, followed by a reduction of 2 dose levels. In certain aspects, the dose administered at the start of subsequent therapy cycles is reduced by 2 dose levels.

[0223] In certain aspects, if the subject experiences mucositis or stomatitis, only the dose of 5-FU is reduced during the therapy cycle, without reducing the dose of irinotecan. In certain aspects, only the dose of 5-FU is reduced at the beginning of subsequent therapy cycles, without reducing the dose of irinotecan.

[0224] Multispecific / bispecific antibodies

[0225] In some aspects, the antibodies or functional portions, derivatives and / or analogs thereof as disclosed herein are multispecific antibodies. In some aspects, the antibodies are bispecific antibodies. In some aspects, the multi- or bispecific antibodies or functional portions, derivatives and / or analogs thereof comprise a first variable domain that binds to the extracellular portion of the epidermal growth factor (EGF) receptor and a second variable domain that does not bind to EGFR in some aspects. In some aspects, the antibody or its functional portion, derivative and / or analog monovalently binds to EGFR. In addition, in some aspects, the multispecific or bispecific antibodies or its functional portion, derivative and / or analog comprise a second variable domain that binds to LGR5. In some aspects, the antibody or its functional portion, derivative and / or analog comprising a variable domain that binds to the extracellular portion of EGFR and optionally comprising a variable domain that binds to the extracellular portion of LGR5 is or comprises pesentumab.

[0226] EGFR

[0227] The epidermal growth factor (EGF) receptor (EGFR, ErbB1 or HER1) is a member of a family of four receptor tyrosine kinases (RTKs) called Her- or cErbB-1, -2, -3 and -4. EGFR has multiple names, the most common of which is EGFR. EGFR has an extracellular domain (ECD) consisting of four subdomains, two of which are involved in ligand binding and the other two subdomains are involved in homodimerization and heterodimerization. EGFR integrates extracellular signals from a variety of ligands, producing different intracellular responses. The main signaling pathway for EGFR activation consists of the Ras-mitogen-activated protein kinase (MAPK) mitogenic signaling cascade. Activation of this pathway is initiated by recruiting Grb2 to tyrosine-phosphorylate EGFR. This leads to the activation of Ras through the Grb2-bound Ras-guanine nucleotide exchange factor Son of Sevenless (SOS). Furthermore, the PI3-kinase-Akt signaling pathway is also activated by EGFR, but this activation is much stronger when ErbB-3 (HER3) is co-expressed. EGFR is associated with several human epithelial malignancies, particularly breast, bladder, non-small cell lung, lung, colon, ovarian, head and neck, and brain cancers. Activating mutations in the gene, as well as overexpression of the receptor and its ligands, have been found to generate an autocrine activation loop. Therefore, this RTK has been widely used as a target for cancer therapy. Small molecule inhibitors targeting RTKs and monoclonal antibodies (mAbs) directed against the extracellular ligand-binding domain have been developed and have demonstrated some clinical success to date, but primarily in selected patient groups. The database accession number for the human EGFR protein and its encoding gene is GenBank NM_005228.3. This accession number is primarily intended to provide methods for further identifying the EGFR protein as a target, but the actual sequence of the EGFR protein to which the antibody binds may vary, for example, due to mutations in the encoding gene, such as those that occur in certain cancers and similar conditions.

[0228] When EGFR is referred to herein, unless otherwise indicated, the reference is to human EGFR. The variable domain antigen binding sites that bind to EGFR bind to EGFR and its various variants, such as those expressed on certain EGFR-positive tumors.

[0229] In certain aspects, the EGFR is human EGFR. The EGFR bound by the antibodies, or functional portions, derivatives, and / or analogs thereof, of the present disclosure includes wild-type EGFR and EGFR with oncogenic driver mutations. In certain aspects, the oncogenic driver mutation is an activating EGFR mutation. In certain aspects, such mutations do not conformationally alter the epitope bound by the antibodies of the present disclosure. In certain aspects, the EGFR mutations disclosed herein include mutations such as exon 18 mutations including G719A, G719C, 2E709_T710D, E709A, G719S; exon 19 deletion mutations including deletion of LREA or VAIKEL; exon 19 point mutations G735S, P753L, L747S, D761Y; exon 20 in-frame 1-7 amino acid insertion mutations, exon 20 point mutations including V765A, T783A, V774A, S784P, V769M, T790M; exon 21 mutations including L858R, T854A, A871E, L861A, L861C, L861S, V843I or P848L. The antibodies disclosed herein bind to epitopes that are not immediately adjacent to the mutations. In particular, EGFR is mutated to S492R, which results in loss of binding of cetuximab to EGFR.The antibodies of the present disclosure bind to an epitope that is different from the epitope recognized by cetuximab.

[0230] Without being bound by any theory, it is believed that Figure 2 The amino acid residues shown are I462; G465; K489; I491; N493; and C499 that participate in binding of the antibodies of the present disclosure to the epitope. In certain aspects, participation in binding is determined by observing reduced binding of the variable domain to EGFR having one or more amino acid residue substitutions selected from the group consisting of: I462A; G465A; K489A; I491A; N493A; and C499A.

[0231] In one aspect, the variable domain that binds to an epitope on the extracellular portion of human EGFR is a variable domain that binds to a Figure 2 In some aspects, the binding of the variable domain to EGFR is reduced by substitution of one or more of the following amino acid residues: I462A; G465A; K489A; I491A; N493A; and C499A in EGFR. In some aspects, the binding of the antibody to human EGFR interferes with the binding of EGF to the receptor. In some aspects, the epitope on EGFR is a conformational epitope. In one aspect, the epitope is located at Figure 2 In some aspects, the epitope is located within amino acid residues 420-480, or 430-480 of the sequence shown. Figure 2 Within 438-469 of the sequence shown.

[0232] Without being bound by theory, it is believed that the epitope contact residues, ie, where the variable domain contacts human EGFR, are I462; K489; I491; and N493. Amino acid residues G465 and C499 are indirectly involved in the binding of the antibody to EGFR.

[0233] In certain aspects, the variable domain binds to LGR5. In certain aspects, the LGR5 is human LGR5. The multispecific or bispecific antibodies described herein, or functional parts, derivatives, and / or analogs thereof, comprise a variable domain that binds to the extracellular portion of the human epidermal growth factor (EGF) receptor and, in certain aspects, a variable domain that binds to LGR5.

[0234] In an exemplary method, CHO cells express LGR5, or alanine substitution mutants such as those comprising one or more of the substitutions M46A, F67A, R90A, or F91A, on the cell membrane. A test antibody is contacted with the CHO cells and binding of the antibody to the cells is compared. If the test antibody binds to LGR5 and binds less strongly to LGR5 with the substitutions M46A, F67A, R90A, or F91A, then the test antibody is bound to the epitope. Binding is preferably compared to a panel of mutants each comprising a single alanine residue substitution. Such binding studies are well known in the art. Typically, the panel comprises single alanine substitution mutants covering substantially all amino acid residues. For LGR5, when using cells, this panel need only cover the extracellular portion of the protein and the portion that is guaranteed to associate with the cell membrane.

[0235] Expression of a particular mutant may be impaired, but this can be detected using one or more EGFR antibodies that bind to different regions. If expression of these control antibodies is also reduced, the level or folding of the protein on the membrane of this particular mutant is impaired. Binding characteristics of the test antibody to this panel of mutants identify whether the test antibody exhibits reduced binding to mutants with the substitutions I462A; G465A; K489A; I491A; N493A; and C499A.

[0236] In certain aspects, the present disclosure provides an antibody or its functional portion, derivative and / or analog, which comprises a first variable domain that binds to the extracellular portion of EGFR, wherein the antibody is a monovalent antibody that does not contain a second variable domain, or wherein the antibody comprises an EGFR-binding variable domain as the only variable domain. In certain aspects, the present disclosure provides an antibody or its functional portion, derivative and / or analog, which comprises a first variable domain that binds to the extracellular portion of EGFR and comprises a further second variable domain that does not bind to EGFR. In certain aspects, the antibody or its functional portion, derivative and / or analog monovalently binds to EGFR. In certain aspects, the antibody or its functional portion, derivative and / or analog. In certain aspects, the antibody comprises a second variable domain that binds to LGR5.

[0237] In certain aspects, the present disclosure provides an antibody or a functional part, derivative and / or analogue thereof, comprising a first variable domain that binds to the extracellular portion of EGFR and a second variable domain that binds to the extracellular portion of LGR5.

[0238] LGR5

[0239] The term "LGR" refers to a protein of the leucine-rich repeat G protein-coupled receptor family. Several members of the family are known to be involved in the WNT signaling pathway, particularly LGR4, LGR5, and LGR6.

[0240] LGR5 is a G protein-coupled receptor with leucine-rich repeats 5. Alternative names for this gene or protein are G protein-coupled receptor with leucine-rich repeats 5; G protein-coupled receptor with leucine-rich repeats 5; G protein-coupled receptor HG38; G protein-coupled receptor 49; G protein-coupled receptor 67; GPR67; GPR49; orphan G protein-coupled receptor HG38; G protein-coupled receptor 49; GPR49; HG38; and FEX. The LGR5-binding proteins or antibodies disclosed herein bind to human LGR5. Due to the sequence and tertiary structure similarities between human and other mammalian orthologs, the LGR5-binding proteins or antibodies disclosed herein may also bind to such orthologs, but this is not required. The database accession numbers for the human LGR5 protein and its encoding gene are (NC_000012.12; NT_029419.13; NC_018923.2; NP_001264155.1; NP_001264156.1; NP_003658.1). The accession numbers are primarily intended to provide a method for further identifying LGR5 as a target. The actual sequence of the LGR5 protein may vary, for example, due to mutations in the encoding gene, such as those that occur in some cancers or similar diseases. The LGR5 antigen binding site binds to LGR5 and various variants thereof, such as those expressed by some LGR5-positive tumor cells.

[0241] The antibodies or functional parts, derivatives and / or analogs thereof as described herein comprise a variable domain that binds to the extracellular portion of LGR5. In certain aspects, a second variable domain binds to LGR5. In certain aspects, LGR5 is human LGR5. The multispecific or bispecific antibodies or functional parts, derivatives and / or analogs thereof as described herein comprise a variable domain that binds to the extracellular portion of the human epidermal growth factor (EGF) receptor, and in certain aspects, comprise a variable domain that binds to human LGR5.

[0242] In certain aspects, an antibody or functional portion, derivative and / or analog thereof as described herein comprises a variable domain that binds to the extracellular portion of the epidermal growth factor (EGF) receptor and interferes with the binding of EGF to the receptor, and a variable domain that binds to LGR5, wherein the interaction of the antibody with LGR5 on cells expressing LGR5 does not block the binding of R spondin (RSPO) to LGR5. Methods for determining whether an antibody blocks or does not block the binding of R spondin to LGR5 are described in WO2017069528, which is incorporated herein by reference.

[0243] In certain aspects, the variable domain that binds the extracellular portion of LGR5 binds to a region located Figure 1An epitope within amino acid residues 21-118 of the sequence of wherein amino acid residues D43; G44, M46, F67, R90 and F91 are involved in binding of the antibody to the epitope.

[0244] In certain aspects, the LGR5 variable domain is a variable domain wherein one or more of the amino acid residue substitutions D43A; G44A, M46A, F67A, R90A, and F91A in LGR5 reduce binding of the variable domain to LGR5.

[0245] In some aspects, the epitope on LGR5 is a conformational epitope. In some aspects, the epitope is located at Figure 1 In certain aspects, the binding of the antibody to LGR5 is reduced by one or more of the following amino acid residue substitutions: D43A; G44A, M46A, F67A, R90A, and F91A.

[0246] Without being bound by theory, it is believed that Figure 1The M46, F67, R90, and F91 of LGR5 shown are contact residues of the variable domains noted above, i.e., the antigen binding site of the variable domains that binds to the LGR5 epitope. Substitution of these amino acid residues for D43A and G44A reduces binding to the antibody, presumably due to the fact that they are also contact residues. However, substitution of these amino acid residues may also induce a (slight) change in the configuration of a portion of LGR5 that has one or more other contact residues (i.e., at positions 46, 67, 90, or 91) and this configurational change reduces antibody binding. The epitope is characterized by the amino acid substitutions mentioned. Whether an antibody binds to the same epitope can be determined in a variety of ways. In an exemplary method, CHO cells express LGR5, or alanine substitution mutants, such as mutants comprising one or more of the substitutions M46A, F67A, R90A, or F91A, on the cell membrane. The test antibody is contacted with the CHO cells and the binding of the antibody to the cells is compared. If a test antibody binds to LGR5 and binds less strongly to LGR5 with an M46A, F67A, R90A, or F91A substitution, then the test antibody binds to the epitope. Binding to a panel of mutants, each containing a single alanine residue substitution, is preferably compared. Such binding studies are well known in the art. Typically, the panel includes single alanine substitution mutants covering substantially all amino acid residues. For LGR5, this panel need only cover the extracellular portion of the protein and, when used with cells, the portion that is bound to the cell membrane. Expression of a particular mutant may be impaired, but this can be readily detected using one or more LGR5 antibodies that bind to different regions. If expression of these control antibodies is also reduced, then the level or folding of the protein on the membrane of that particular mutant is impaired. The binding profile of the test antibody to this panel can identify whether the test antibody exhibits reduced binding to a mutant with an M46A, F67A, R90A, or F91A substitution, and therefore whether the test antibody is an antibody of the present disclosure. Reduced binding to mutants with M46A, F67A, R90A, or F91A substitutions also identified a region located at Figure 1 The epitope is within amino acid residues 21-118 of the sequence of MF5816. In certain aspects, the panel includes a D43A substitution mutant; and both G44A substitution mutants. Antibodies having a VH sequence of the VH of MF5816 exhibit reduced binding to these substitution mutants.

[0247] Where accession numbers or alternative names for proteins / genes are given herein, they are primarily given to provide a further means of identifying the referenced protein as a target, and the actual sequence of the target protein to which the disclosed antibodies bind may vary, for example due to mutations and / or alternative splicing in the encoding gene, such as those that occur in some cancers or their analogous diseases. The target protein is bound by the antibody as long as the epitope is present in the protein and the epitope is accessible to the antibody.

[0248] Antibodies that bind to EGFR and LGR5

[0249] The present disclosure further provides an antibody having a variable domain that binds to the extracellular portion of EGFR and a variable domain that binds to the extracellular portion of LGR5, wherein the LGR5 variable domain binds to an epitope on LGR5 located at Figure 1 within amino acid residues 21-118 of the sequence.

[0250] Disclosed herein are suitable variable domains that bind to the extracellular portion of EGFR and suitable variable domains that bind to the extracellular portion of LGR5. In certain aspects, the first variable domain comprises at least a CDR3 sequence, or at least a CDR1, CDR2, and CDR3 sequence, of an EGFR-specific heavy chain variable region selected from the group consisting of MF3370; MF3755; MF4280, or MF4289 as shown in FIG3 . In certain aspects, the second variable domain comprises at least a CDR3 sequence, or at least a CDR1, CDR2, and CDR3 sequence, of an LGR5-specific heavy chain variable region selected from the group consisting of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in FIG3 .

[0251] In certain aspects, the variable domain that binds to human EGFR is a variable domain having a heavy chain variable region, wherein the heavy chain variable region comprises at least the CDR3 sequence of VH of MF3755 as shown in Figure 3, or a CDR3 sequence that has at most three, or at most two, or no more than one amino acid difference from the CDR3 sequence of VH of MF3755 as shown in Figure 3.

[0252] In certain aspects, the variable domain that binds to human EGFR is a variable domain having a heavy chain variable region, wherein the heavy chain variable region comprises at least the CDR1, CDR2 and CDR3 sequences of the VH of MF3755 as shown in Figure 3; or the CDR1, CDR2 and CDR3 sequences of the VH of MF3755 as shown in Figure 3 having at most three, or at most two, or at most one amino acid substitution.

[0253] In certain aspects, the variable domain that binds to human EGFR is a variable domain having a heavy chain variable region comprising the sequence of the VH chain of MF3755 as shown in Figure 3; or the amino acid sequence of the VH chain of MF3755 as shown in Figure 3 having up to 15 (or in certain aspects, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or in certain aspects, 1, 2, 3, 4 or 5) amino acid insertions, deletions, substitutions or a combination thereof relative to the VH chain of MF3755.

[0254] In certain aspects, the present disclosure provides an antibody comprising a variable domain that binds to the extracellular portion of EGFR and a variable domain that binds to the extracellular portion of LGR5, wherein the heavy chain variable region of the variable domain comprises a CDR3 sequence of at least an EGFR-specific heavy chain variable region selected from the group consisting of MF3370; MF3755; MF4280, or MF4289 as shown in Figure 3, or wherein the heavy chain variable region of the variable domain comprises a heavy chain CDR3 sequence that differs from a CDR3 sequence of a VH selected from the group consisting of MF3370; MF3755; MF4280, or MF4289 as shown in Figure 3 by at most three, or at most two, or no more than one amino acid. In certain aspects, the variable domain comprises a heavy chain variable region comprising a CDR3 sequence of at least MF3370; MF3755; MF4280, or MF4289 as shown in Figure 3.

[0255] In certain aspects, the variable domain comprises a heavy chain variable region comprising at least the CDR1, CDR2, and CDR3 sequences of an EGFR-specific heavy chain variable region selected from the group consisting of MF3370, MF3755, MF4280, or MF4289 as shown in Figure 3 , or a heavy chain variable region comprising at least the CDR1, CDR2, and CDR3 sequences that differ by at most three, at most two, or at most one amino acid sequence from the CDR1, CDR2, and CDR3 sequences of an EGFR-specific heavy chain variable region selected from the group consisting of MF3370, MF3755, MF4280, or MF4289 as shown in Figure 3 . In certain aspects, the variable domain comprises a heavy chain variable region comprising at least the CDR1, CDR2, and CDR3 sequences of MF3370, MF3755, MF4280, or MF4289 as shown in Figure 3 . In certain aspects, the heavy chain variable region is MF3755. In certain aspects, the heavy chain variable region is MF4280.

[0256] In certain aspects, the present disclosure provides an antibody comprising a variable domain that binds to the extracellular portion of EGFR and a variable domain that binds to the extracellular portion of LGR5, wherein the variable domain that binds to the extracellular portion of EGFR is a heavy chain variable region comprising the following: the amino acid sequence of the VH chain of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3, or the amino acid sequence of the VH chain of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3 having up to 15, or 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid insertions, deletions, substitutions or a combination thereof relative to the VH chain of MF3370; MF3755; MF4280 or MF4289.

[0257] In certain aspects, an antibody comprises a variable domain that binds to the extracellular portion of EGFR and a variable domain that binds to the extracellular portion of LGR5, wherein the variable domain that binds to EGFR has a CDR3, CDR1, CDR2 and CDR3 and / or a VH sequence as shown above, and has a variable domain that binds to LGR5 that comprises at least a CDR3 sequence of an LGR5-specific heavy chain variable region selected from the group consisting of: MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3, or a heavy chain CDR3 sequence that differs from the CDR3 sequence of a VH selected from the group consisting of: MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3. In certain aspects, the variable domain comprises a heavy chain variable region comprising at least the CDR3 sequence of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in FIG. 3 .

[0258] In certain aspects, the LGR5 binding variable domain comprises a heavy chain variable domain comprising at least the CDR1, CDR2 and CDR3 sequence of an LGR5-specific heavy chain variable region selected from the group consisting of: MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3, or a heavy chain CDR1, CDR2 and CDR3 sequence that differs from the CDR1, CDR2 and CDR3 sequence of an LGR5-specific heavy chain variable region selected from the group consisting of: MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3. In certain aspects, the variable domain comprises a heavy chain variable region comprising at least the CDR1, CDR2, and CDR3 sequences of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 shown in Figure 3 . In certain aspects, the heavy chain variable region is MF5790; MF5803; MF5814; MF5816; MF5817; or MF5818. In certain aspects, the heavy chain variable region is MF5790; MF5814; MF5816; or MF5818. In certain aspects, the heavy chain variable region is MF5814, MF5818, or MF5816. In certain aspects, the heavy chain variable region is MF5816. In certain aspects, the heavy chain variable region is MF5818.

[0259] In certain aspects, the present disclosure provides an antibody comprising a variable domain that binds to the extracellular portion of EGFR and a variable domain that binds to the extracellular portion of LGR5, wherein the variable domain that binds to the extracellular portion of LGR5 is a heavy chain variable region comprising: the amino acid sequence of the VH chain of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in FIG3 , or having a VH chain amino acid sequence relative to MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in FIG3 . 803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 with up to 15 or 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid insertions, deletions, substitutions or combinations thereof of the VH chain of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3.

[0260] It has been shown that antibodies comprising one or more variable domains comprising the heavy chain variable region MF3755 or one or more CDRs thereof have better effects when used to inhibit the growth of EGFR ligand-responsive cancers or cells. In the case of bispecific or multispecific antibodies, an antibody arm comprising a variable domain comprising the heavy chain variable region MF3755 or one or more CDRs thereof is well combined with an arm comprising a variable domain comprising the heavy chain variable region MF5818 or one or more CDRs thereof.

[0261] It has been shown that antibodies comprising one or more variable domains comprising the heavy chain variable region MF3755 or one or more CDRs thereof have better effects when used to inhibit the growth of EGFR ligand-responsive cancers or cells. In the case of bispecific or multispecific antibodies, an antibody arm comprising a variable domain comprising the heavy chain variable region MF3755 or one or more CDRs thereof is well combined with an arm comprising a variable domain comprising the heavy chain variable region MF5816 or one or more CDRs thereof.

[0262] The VH chain of the variable domain that binds to EGFR or LGR5 can have one or more amino acid substitutions relative to the sequence as shown in Figure 3. In certain aspects, the VH chain has the amino acid sequence of EGFR or LGR5 VH as shown in Figure 3, which has up to 15, or 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, and in certain aspects, 1, 2, 3, 4 or 5 amino acid insertions, deletions, substitutions or combinations thereof relative to the VH chain sequence shown in Figure 3.

[0263] The CDR sequences may have one or more amino acid residue substitutions relative to the CDR sequences in the figures. Such one or more substitutions may be made, for example, for optimization purposes, such as to improve the binding strength or stability of the antibody. Optimization may be performed, for example, by a mutagenesis program, wherein, for example, the stability and / or binding affinity of the resulting antibody is tested, and improved EGFR-specific CDR sequences or LGR5-specific CDR sequences are selected. A skilled person is fully capable of generating antibody variants comprising at least one altered CDR sequence according to the present disclosure. For example, conservative amino acid substitutions may be applied. Examples of conservative amino acid substitutions include substitution of one hydrophobic residue, such as isoleucine, valine, leucine, or methionine, for another hydrophobic residue, and substitution of one polar residue for another, such as substitution of arginine for lysine, substitution of glutamic acid for aspartic acid, or substitution of glutamine for asparagine.

[0264] In some aspects, up to 15 (or in some aspects, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10, or in some aspects, 1, 2, 3, 4, or 5) amino acid substitutions in the VH or VL indicated herein are conservative amino acid substitutions. In some aspects, the amino acid insertions, deletions, substitutions, or combinations thereof in the VH or VL indicated herein are not present in the CDR3 region. In some aspects, the amino acid insertions, deletions, and substitutions mentioned are also not present in the CDR1 and CDR2 regions. In some aspects, the amino acid insertions, deletions, and substitutions mentioned are also not present in the FR4 region.

[0265] In certain aspects, the insertion, deletion, substitution, or combination thereof is not present in the CDR3 region of the VH chain, in certain aspects, is not present in the CDR1, CDR2, or CDR3 regions of the VH chain, and in certain aspects, is not present in the FR4 region.

[0266] In certain aspects, the insertions, deletions, substitutions, or combinations thereof are not present in the CDR1, CDR2, and CDR3 regions of the VH chain.

[0267] In certain aspects, the present disclosure provides an antibody comprising a variable domain that binds to an extracellular portion of EGFR and, in certain aspects, a variable domain that binds to an extracellular portion of LGR5, comprising:

[0268] - the amino acid sequence of the VH chain MF3755 as shown in Figure 3; or

[0269] - the amino acid sequence of the VH chain MF3755 as shown in Figure 3, which has up to 15 (or in some aspects, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or in some aspects, 1, 2, 3, 4 or 5) amino acid insertions, deletions, substitutions or combinations thereof relative to said VH; and

[0270] The VH chain of the variable domain that binds to LGR5 comprises:

[0271] - the amino acid sequence of the VH chain MF5790 as shown in Figure 3; or

[0272] - the amino acid sequence of the VH chain MF5790 as shown in Figure 3, which has up to 15 (or in some aspects, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or in some aspects, 1, 2, 3, 4 or 5) amino acid insertions, deletions, substitutions or a combination thereof relative to said VH.

[0273] In certain aspects, the present disclosure provides an antibody comprising a variable domain that binds to an extracellular portion of EGFR and, in certain aspects, a variable domain that binds to an extracellular portion of LGR5, comprising:

[0274] - the amino acid sequence of the VH chain MF3755 as shown in Figure 3; or

[0275] - the amino acid sequence of the VH chain MF3755 as shown in Figure 3, which has up to 15 (or in some aspects, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or in some aspects, 1, 2, 3, 4 or 5) amino acid insertions, deletions, substitutions or combinations thereof relative to said VH; and

[0276] The VH chain of the variable domain that binds to LGR5 comprises:

[0277] - the amino acid sequence of the VH chain MF5803 as shown in Figure 3; or

[0278] - The amino acid sequence of the VH chain MF5803 as shown in Figure 3, which has up to 15 (or in some aspects, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or in some aspects, 1, 2, 3, 4 or 5) amino acid insertions, deletions, substitutions or combinations thereof relative to said VH.

[0279] In certain aspects, the present disclosure provides an antibody comprising a variable domain that binds to an extracellular portion of EGFR and, in certain aspects, a variable domain that binds to an extracellular portion of LGR5, comprising:

[0280] - the amino acid sequence of the VH chain MF3755 as shown in Figure 3; or

[0281] - the amino acid sequence of the VH chain MF3755 as shown in Figure 3, which has up to 15 (or in some aspects, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or in some aspects, 1, 2, 3, 4 or 5) amino acid insertions, deletions, substitutions or combinations thereof relative to said VH; and

[0282] The VH chain of the variable domain that binds to LGR5 comprises:

[0283] - the amino acid sequence of the VH chain MF5814 as shown in Figure 3; or

[0284] - The amino acid sequence of the VH chain MF5814 as shown in Figure 3, which has up to 15 (or in some aspects, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or in some aspects, 1, 2, 3, 4 or 5) amino acid insertions, deletions, substitutions or combinations thereof relative to said VH.

[0285] In certain aspects, the treatments of the present disclosure utilize an antibody comprising a variable domain that binds to the extracellular portion of EGFR and, in certain aspects, to the extracellular portion of LGR5, comprising:

[0286] - the amino acid sequence of the VH chain MF3755 as shown in Figure 3; or

[0287] - the amino acid sequence of the VH chain MF3755 as shown in Figure 3, which has up to 15 (or in some aspects, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or in some aspects, 1, 2, 3, 4 or 5) amino acid insertions, deletions, substitutions or combinations thereof relative to said VH; and

[0288] The VH chain of the variable domain that binds to LGR5 comprises:

[0289] - the amino acid sequence of the VH chain MF5816 as shown in Figure 3; or

[0290] - the amino acid sequence of the VH chain MF5816 as shown in Figure 3, which has up to 15 (or in some aspects, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or in some aspects, 1, 2, 3, 4 or 5) amino acid insertions, deletions, substitutions or a combination thereof relative to said VH.

[0291] In certain aspects, the treatments of the present disclosure utilize an antibody comprising a variable domain that binds to the extracellular portion of EGFR and, in certain aspects, to the extracellular portion of LGR5, comprising:

[0292] - the amino acid sequence of the VH chain MF3755 as shown in Figure 3; or

[0293] - the amino acid sequence of the VH chain MF3755 as shown in Figure 3, which has up to 15 (or in some aspects, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or in some aspects, 1, 2, 3, 4 or 5) amino acid insertions, deletions, substitutions or combinations thereof relative to said VH; and

[0294] The VH chain of the variable domain that binds to LGR5 comprises:

[0295] - the amino acid sequence of the VH chain MF5817 as shown in Figure 3; or

[0296] - The amino acid sequence of the VH chain MF5817 as shown in Figure 3, which has up to 15 (or in some aspects, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or in some aspects, 1, 2, 3, 4 or 5) amino acid insertions, deletions, substitutions or combinations thereof relative to said VH.

[0297] In certain aspects, the treatments of the present disclosure utilize an antibody comprising a variable domain that binds to the extracellular portion of EGFR and, in certain aspects, to the extracellular portion of LGR5, comprising:

[0298] - the amino acid sequence of the VH chain MF3755 as shown in Figure 3; or

[0299] - the amino acid sequence of the VH chain MF3755 as shown in Figure 3, which has up to 15 (or in some aspects, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or in some aspects, 1, 2, 3, 4 or 5) amino acid insertions, deletions, substitutions or combinations thereof relative to said VH; and

[0300] The VH chain of the variable domain that binds to LGR5 comprises:

[0301] - the amino acid sequence of the VH chain MF5818 as shown in Figure 3; or

[0302] - The amino acid sequence of the VH chain MF5818 as shown in Figure 3, which has up to 15 (or in some aspects, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, or in some aspects, 1, 2, 3, 4 or 5) amino acid insertions, deletions, substitutions or a combination thereof relative to said VH.

[0303] In some aspects, the antibody is pesentuzumab (see WHO Drug Information, Recommended INN: List 83, Vol. 34, No. 1, 2020, pp. 75-77). In some aspects, its functional portion, derivative and / or analog is a functional portion, derivative and / or analog of pesentuzumab. In some aspects, the EGFR / LGR5 antibody of the present disclosure is or includes pesentuzumab.

[0304] In certain aspects, the variable domain that binds to the extracellular portion of EGFR comprises a heavy chain variable region comprising a CDR1, CDR2 and CDR3 sequence of a variable region selected from the group consisting of: MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3, and wherein the variable domain that binds to the extracellular portion of LGR5 comprises a heavy chain variable region comprising a CDR1, CDR2 and CDR3 sequence of a variable region selected from the group consisting of: MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3.

[0305] In certain aspects, the variable domain that binds to the extracellular portion of EGFR comprises a heavy chain variable region comprising the CDR1, CDR2 and CDR3 sequences of the variable region of MF3755 as shown in Figure 3; and wherein the variable domain that binds to the extracellular portion of LGR5 comprises a heavy chain variable region comprising the CDR1, CDR2 and CDR3 sequences of the variable region of MF5816 as shown in Figure 3.

[0306] In certain aspects, the VH chain of the variable domain that binds to EGFR comprises the amino acid sequence of VH chain MF3370, MF3755, MF4280 or MF4289 as shown in Figure 3; or the amino acid sequence of VH chain MF3370, MF3755, MF4280 or MF4289 as shown in Figure 3, which has a maximum of 15, preferably no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, preferably no more than 5, 4, 3, 2 or 1 amino acid modifications relative to the VH, including insertions, deletions, substitutions or combinations thereof; and wherein the VH chain of the variable domain that binds to LGR5 comprises the VH chain MF579 as shown in Figure 3 0, MF5803, MF5805, MF5808, MF5809, MF5814, MF5816, MF5817, or MF5818; or the amino acid sequence of the VH chain MF5790, MF5803, MF5805, MF5808, MF5809, MF5814, MF5816, MF5817, or MF5818 as shown in Figure 3, which has a maximum of 15, preferably no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, preferably no more than 5, 4, 3, 2 or 1 amino acid modifications relative to the VH, including insertions, deletions, substitutions or a combination thereof.

[0307] In certain aspects, the VH chain of the variable domain that binds to EGFR comprises the amino acid sequence of VH chain MF3755 as shown in Figure 3; or the amino acid sequence of VH chain MF3755 as shown in Figure 3, which has a maximum of 15, preferably no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, preferably no more than 5, 4, 3, 2 or 1 amino acid modifications relative to the VH, including insertions, deletions, substitutions or combinations thereof; and wherein the VH chain of the variable domain that binds to LGR5 comprises the amino acid of VH chain MF5816 as shown in Figure 3; or the amino acid sequence of VH chain MF5816 as shown in Figure 3, which has a maximum of 15, preferably no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, preferably no more than 5, 4, 3, 2 or 1 amino acid modifications relative to the VH, including insertions, deletions, substitutions or combinations thereof.

[0308] In certain aspects, the VH chain of the variable domain that binds to EGFR comprises the amino acid sequence of VH chain MF3370, MF3755, MF4280 or MF4289 as shown in Figure 3; and wherein the VH chain of the variable domain that binds to LGR5 comprises the amino acid sequence of VH chain MF5790, MF5803, MF5805, MF5808, MF5809, MF5814, MF5816, MF5817, or MF5818 as shown in Figure 3.

[0309] In certain aspects, the VH chain of the variable domain that binds to EGFR comprises the amino acid sequence of VH chain MF3755 as shown in Figure 3; and wherein the VH chain of the variable domain that binds to LGR5 comprises the amino acid sequence of VH chain MF5816 as shown in Figure 3.

[0310] In certain aspects, the variable domain that binds EGFR and the variable domain that binds LGR5 both comprise Figure 4 b shows the CDR1, CDR2 and CDR3 regions of the light chain variable region.

[0311] In certain aspects, the variable domain that binds EGFR and the variable domain that binds LGR5 both comprise Figure 4 b, wherein the light chain variable region comprises 0 to 10 amino acid insertions, deletions, substitutions, additions or a combination thereof, wherein there are no amino acid insertions, deletions or substitutions in the CDR1, CDR2 and CDR3 light chain variable regions.

[0312] Additional variants of the disclosed amino acid sequences that retain EGFR or LGR5 binding can be obtained, for example, from phage display libraries containing rearranged human IGKV1-39 / IGKJ1 VL regions (De Kruif et al. Biotechnol Bioeng. 2010(106)741-50), and collections of VH regions incorporating amino acid substitutions into the amino acid sequences of the EGFR or LGR5 VH regions disclosed herein (as previously described (e.g., WO2017 / 069628). Phage encoding Fab regions that bind to EGFR or LGR5 can be selected and analyzed by flow cytometry and sequenced to identify variants with amino acid substitutions, insertions, deletions, or additions that retain antigen binding.

[0313] In certain aspects, the light chain variable regions of the VH / VL EGFR and LGR5 variable domains of the EGFR / LGR5 antibodies of the present disclosure may be the same or different. In certain aspects, the VL region of the VH / VL EGFR variable domain of the EGFR / LGR5 antibodies is similar to the VL region of the VH / VL LGR5 variable domain. In certain aspects, the VL regions in the first and second VH / VL variable domains are the same.

[0314] In certain aspects, the light chain variable region of one or both VH / VL variable domains of the EGFR / LGR5 antibody comprises a common light chain variable region. In certain aspects, the common light chain variable region of the one or both VH / VL variable domains comprises a germline IgVκ1-39 variable region V segment. In certain aspects, the light chain variable region of the one or both VH / VL variable domains comprises a kappa light chain V segment, IgVκ1-39*01. IgVκ1-39 is an abbreviation for the immunoglobulin variable kappa 1-39 gene. This gene is also known as immunoglobulin kappa variable 1-39; IGKV139; IGKV1-39. The external IDs for this gene are HGNC: 5740; Entrez Gene: 28930; Ensembl: ENSG00000242371. Figure 4 The amino acid sequences of suitable V regions are provided in . The V region can be combined with five J regions. In some aspects, the J regions are jk1 and jk5, and the connected sequences are represented as IGKV1-39 / jk1 and IGKV1-39 / jk5; alternative names are IgVκ1-39*01 / IGJκ1*01 or IgVκ1-39*01 / IGJκ5*01 (named according to the IMGT database global network imgt.org). In some aspects, the light chain variable region of the one or two VH / VL variable domains comprises a kappa light chain IgVκ1-39*01 / IGJκ1*01 or IgVκ1-39*01 / IGJκ1*05 (such as Figure 4 describe).

[0315] In some aspects, Figure 4 d) The light chain variable region comprises LCDR1, LCDR2 and LCDR sequences. Such sequences can be determined or annotated by a skilled artisan using, for example, an annotation system such as IMGT, Chothia, Kabat or other appropriate annotation systems.

[0316] The CDRs and framework regions of antibody heavy and light chains have been described and defined in the art using a number of different systems, including, for example, Kabat (see Sequences of Proteins of Immunological Interest (National Institute of Health, Bethesda, Md., 1987 and 1991); Kabat et al., J. Biol. Chem. 252:6609-6616 (1977)), IMGT (discussed in Giudicelli et al., Nucleic Acids Res. 25:206-211 (1997)), Chothia (Chothia and Lesk J. Mol. Biol. 196:901-917, 1987; Chothia et al., Nature 2003; 1996). 342:877-883, 1989; Al-Lazikani et al., J. Mol. Biol. 273:927-948, 1997) and Honnegher and Pluckthun nomenclature (Honnegher and Pluckthun, J. Mol. Biol. 309:657-670, 2001), MacCallum (MacCallum et al., J. Mol. Biol. 262:732-745 (1996); Abhinandan and Martin, Mol. Immunol., 45:3832-3839 (2008)), and Lefranc (Lefranc MP et al., Dev. Comp. Immunol., 27:55-77 (2003)). In general, it does not matter which numbering system is used, as antibodies exhibit their properties regardless of the numbering system used. Given the amino acid sequence of a variable region, a skilled artisan can readily identify its CDRs based on the different numbering systems indicated above. Thus, the present disclosure encompasses defining CDRs according to each numbering system available to the skilled artisan. In particular, the present disclosure encompasses defining CDRs according to the numbering systems of Kabat, IMGT, and Chothia. In certain aspects, heavy chain CDRs are defined using the Kabat definition and light chain CDRs using the IMGT definition. Amino acids in the constant region are indicated according to the EU numbering system.

[0317] In certain aspects, the light chain variable region is or includes one or two VH / VL variable domains, which are present in the EGFR binding antibodies of the present disclosure and include LCDR1, LCDR2, and LCDR sequences. Such sequences can be determined or annotated by a skilled artisan using, for example, an annotation system such as IMGT, Chothia, Kabat, or other appropriate annotation systems.

[0318] In certain aspects, the light chain variable region of one or both VH / VL variable domains of the bispecific antibodies of the present disclosure comprises a LCDR1 comprising the amino acid sequence QSISSY (e.g., Figure 4 Described), LCDR2 containing the amino acid sequence AAS (such as Figure 4 ), and LCDR3 containing the amino acid sequence QQSYSTPPT (as described in Figure 4 In certain aspects, the light chain variable region of one or both VH / VL variable domains of the EGFR / LGR5 binding antibodies of the present disclosure comprises a LCDR1 comprising the amino acid sequence QSISSY (e.g., Figure 4 Described), LCDR2 containing the amino acid sequence AAS (such as Figure 4 ), and LCDR3 containing the amino acid sequence QQSYSTPPT (as described in Figure 4 described).

[0319] In certain aspects, the light chain variable region of one or both VH / VL variable domains of the EGFR / LGR5 bispecific antibodies of the present disclosure comprises a LCDR1 comprising the amino acid sequence QSISSY (e.g., Figure 4 Described), LCDR2 containing the amino acid sequence AAS (such as Figure 4 ), and LCDR3 containing the amino acid sequence QQSYSTPPT (as described in Figure 4 In certain aspects, the light chain variable region of one or both VH / VL variable domains of the EGFR / LGR5 antibody comprises a LCDR1 comprising the amino acid sequence QSISSY (e.g., Figure 4 Described), LCDR2 containing the amino acid sequence AAS (such as Figure 4 ), and LCDR3 containing the amino acid sequence QQSYSTPPT (as described in Figure 4 described).

[0320] In certain aspects, one or both VH / VL variable domains of the EGFR / LGR5 antibody comprise a light chain variable region comprising Figure 4 The amino acid sequences shown have amino acid sequences that are at least 90%, in some aspects at least 95%, in some aspects at least 97%, in some aspects at least 98%, in some aspects at least 99%, or in some aspects 100% identical.

[0321] For example, relative to Figure 4In certain aspects, the variable light chain region of one or two VH / VL variable domains of the EGFR / LGR5 antibody comprises 0 to 9, 0 to 8, 0 to 7, 0 to 6, 0 to 5, 0 to 4, 0 to 3, 0 to 2, or in certain aspects 0 to 1, or in certain aspects 0 amino acid insertions, deletions, substitutions, additions, or a combination thereof relative to the indicated amino acid sequence.

[0322] In addition, the light chain variable region of one or both VH / VL variable domains of the EGFR / LGR5 antibody may comprise Figure 4 In some aspects, the two VH / VL variable domains of the EGFR / LGR5 antibody comprise the same VL region. In some aspects, the VL of the two VH / VL variable domains of the EGFR / LGR5 bispecific antibody comprises the amino acid sequence shown in FIG. Figure 4 The amino acid sequence shown.

[0323] In certain aspects, the VL of the two VH / VL of the EGFR / LGR5 bispecific antibody comprises Figure 4 The amino acid sequence shown.

[0324] In certain aspects, the antibody or functional portion, derivative and / or analog comprising a variable domain that binds to the extracellular portion of EGFR is a bispecific antibody having two variable domains, one of which binds to EGFR and the other binds to LGR5, as described herein. The antibody or functional portion, derivative and / or analog of an antibody can be provided in a variety of forms. Many different forms of bispecific antibodies are known in the art and have been reviewed in Kontermann (Drug Discov Today, 2015 Jul; 20(7):838-47; MAbs, 2012 Mar-Apr; 4(2):182-97) and Spiess et al. (Alternative molecular formats and therapeutic applications for bispecific antibodies. Mol. Immunol. (2015) http: / / dx.doi.org / 10.1016 / j.molimm.2015.01.003), each of which is incorporated herein by reference. For example, rather than a typical antibody with two VH / VL combinations, a bispecific antibody format has at least a variable domain comprising a heavy chain variable region and a light chain variable region. This variable domain can be linked to a single-chain Fv fragment, monomer, VH, and Fab fragment that provides a second binding activity.

[0325] In some aspects, the antibody or the functional part, derivative and / or analog of the antibody is a bispecific antibody of human IgG subclass (e.g., IgG1, IgG2, IgG3, IgG4). In some aspects, the antibody belongs to human IgG1 subclass. Full-length IgG antibodies are preferred because of their good half-life and low immunogenicity. Therefore, the bispecific antibody is a full-length IgG molecule in some aspects. In some aspects, the bispecific antibody is a full-length IgG1 molecule. Therefore, in some aspects, the bispecific antibody comprises a crystallizable fragment (Fc). In some aspects, the Fc of the EGFR / LGR5 bispecific antibody is composed of human constant regions. The constant region or Fc of the EGFR / LGR5 bispecific antibody can contain one or more, or no more than 10, or no more than 5 amino acid differences with the constant region of naturally occurring human antibodies. For example, each Fab arm of the bispecific antibody can also include an Fc region, which includes modifications that promote bispecific antibody formation, promote stability and / or other features described herein.

[0326] EGFR signaling inhibition

[0327] In certain aspects, antibodies as herein described or their functional parts, derivatives and / or analogs interfere with the binding of a ligand to EGFR. As used herein, the term "interference binding" refers to the binding of an antibody or its functional part, derivative and / or analog to EGFR that competes with the binding of a ligand to the EGF receptor. When a ligand has been bound to the EGF receptor, the antibody or its functional part, derivative and / or analog can reduce ligand binding, displace the ligand, or it can, for example, at least partially prevent the ligand from binding to the EGF receptor via steric hindrance.

[0328] In some aspects, EGFR antibodies disclosed herein inhibit EGFR ligand-induced signal transmission respectively, which is measured with the growth of ligand-induced BxPC3 cells (ATCC CRL-1687) or BxPC3-luc2 cells (Perkin Elmer 125058) or ligand-induced A431 (ATCC CRL-1555) cell death. EGFR can bind to a variety of ligands and stimulate the growth of above-mentioned BxPC3 cells or BxPC3-luc2 cells. When the EGFR ligand exists, the growth of BxPC3 or BxPC3-luc2 cells is stimulated. The BxPC3 cell growth induced by EGFR ligand can be measured by the growth of cells when there is no ligand and the presence of the ligand. In some aspects, the preferred EGFR ligand for measuring the BxPC3 or BxPC3-luc2 cell growth induced by EGFR ligand is EGF. In some aspects, the growth of ligand induction is measured using a saturated amount of ligand. In some aspects, EGF is used with the amount of 100ng / ml culture medium. In some aspects, the EGF is used in an amount of 100 ng / ml culture medium. In some aspects, the EGF is EGF from R&D Systems, catalog numbers: 396-HB and 236-EG (see also WO 2017 / 069628; incorporated herein by reference).

[0329] In some aspects, EGFR antibodies disclosed herein inhibit the growth of EGFR ligand-induced BxPC3 cells (ATCCCRL-1687) or BxPC3-luc2 cells (Perkin Elmer 125058). EGFR can bind to a variety of ligands and stimulate the growth of above-mentioned BxPC3 cells or BxPC3-luc2 cells. When the ligand is present, the growth of BxPC3 or BxPC3-luc2 cells is stimulated. The growth of BxPC3 cells induced by EGFR ligands can be measured by comparing the growth of cells in the absence and presence of the ligand. In some aspects, the preferred EGFR ligand for measuring the growth of BxPC3 or BxPC3-luc2 cells induced by EGFR ligands is EGF. In some aspects, the growth of ligand induction is measured using a saturated amount of ligand. In some aspects, EGF is used in an amount of 100ng / ml culture medium. In certain aspects, the EGF is EGF from R&D Systems, catalog numbers: 396-HB and 236-EG (see also WO 2017 / 069628; which is incorporated herein by reference).

[0330] For the avoidance of doubt, cell growth as used herein refers to a change in cell number. Inhibition of growth refers to a decrease in the number of cells that would otherwise be obtained. Increased growth refers to an increase in the number of cells that would otherwise be obtained. Cell growth generally refers to cell proliferation.

[0331] In some aspects, whether the antibodies described herein inhibit signaling or growth in a multispecific format is determined using a monospecific monovalent or monospecific bivalent form of the antibody as described herein above. In some aspects, such antibodies have a binding site for a signaling receptor to be determined. Monospecific monovalent antibodies can have variable domains with unrelated binding specificities (e.g., tetanus toxoid specificity). In some aspects, the antibody is a bivalent monospecific antibody in which the antigen-binding variable domains consist of variable domains that bind to members of the EGF receptor family.

[0332] Nucleic acids and cells

[0333] Bispecific antibodies are typically produced by cells expressing nucleic acids encoding the antibodies. Thus, in certain aspects, the bispecific EGFR / LGR5 antibodies disclosed herein are produced by providing cells containing one or more nucleic acids encoding the heavy and light chain variable regions and constant regions of the bispecific EGFR / LGR5 antibodies. In certain aspects, the cells are animal cells, such as mammalian cells or primate cells, and in certain aspects, human cells. Suitable cells are any cells capable of containing and producing EGFR / LGR5 bispecific antibodies.

[0334] Suitable cells for producing antibodies are known in the art and include hybridoma cells, Chinese hamster ovary (CHO) cells, NSO cells, or PER-C6 cells. Various institutions and companies have developed cell lines for large-scale production of antibodies, for example for clinical use. Non-limiting examples of such cell lines are CHO cells, NSO cells, or PER.C6 cells. In some aspects, the cells are human cells. In some aspects, the cells are transformed with an adenovirus E1 region or a functional equivalent thereof. An example of such a cell line is a PER.C6 cell line or its equivalent. In some aspects, the cells are CHO cells or variants thereof. In some aspects, the variants utilize a glutamine synthetase (GS) vector system to express the antibody. In some aspects, the cells are CHO cells.

[0335] In some aspects, the cell expresses different light chains and heavy chains constituting EGFR / LGR5 bispecific antibodies. In some aspects, the cell expresses two different heavy chains and at least one light chain. In some aspects, the cell expresses "common light chain" as described herein to reduce the number of different antibody species (combinations of different heavy and light chains). For example, methods known in the art for producing bispecific IgG (WO2013 / 157954; incorporated herein by reference) are used, combined with rearranged human IGKV1-39 / IGKJ1 (huVκ139) light chains, each VH region is cloned into an expression vector, and it has been previously shown that it can be paired with more than one heavy chain, thereby producing antibodies with different specificities, which promote the production of bispecific molecules (DeKruif et al. J.Mol.Biol.2009(387)548-58; WO2009 / 157771).

[0336] Antibody-producing cells expressing a common light chain and two heavy chains in equal amounts typically produce 50% of bispecific antibodies and 25% of each monospecific antibody (i.e., having the same heavy-light chain combination). Several methods have been published that can promote the production of bispecific antibodies rather than corresponding monospecific antibodies. This is typically achieved by modifying the constant region of the heavy chain so that they are conducive to heterodimerization (i.e., heavy chain dimerization in combination with other heavy / light chains) rather than homodimerization. In some aspects, the bispecific antibodies of the present disclosure comprise two different immunoglobulin heavy chains with compatible heterodimerization domains. Various compatible heterodimerization domains have been described in the art. In some aspects, the compatible heterodimerization domains are compatible immunoglobulin heavy chain CH3 heterodimerization domains. The art describes a variety of ways in which such heavy chain heterodimerization can be achieved.

[0337] A kind of preferred method for producing EGFR / LGR5 bispecific antibody is disclosed in US 9,248,181 and US 9,358,286.Particularly, the preferred mutation that only produces bispecific full-length IgG molecule is aminoacid replacement L351K and T366K (EU numbering) (" KK variant " heavy chain) in the first CH3 domain, and aminoacid replacement L351D and L368E (" DE variant " heavy chain) in the second CH3 domain, and vice versa.As previously mentioned, DE variant and KK variant are preferably paired to form heterodimer (so-called " DEKK " bispecific molecule).Due to the strong repulsive force between the charged residues in the CH3-CH3 interface between identical heavy chain, the homodimerization (DEDE homodimer) of DE variant heavy chain or the homodimerization of KK variant heavy chain (KKKK homodimer) hardly occur.

[0338] Thus, in certain aspects, a heavy chain / light chain combination comprising a variable domain that binds to EGFR comprises a DE variant of the heavy chain.In certain aspects, a heavy chain / light chain combination comprising a variable domain that binds to LGR5 comprises a KK variant of the heavy chain.

[0339] Any suitable assay can be used to test the binding of candidate EGFR / LGR5 IgG bispecific antibodies. For example, the binding to EGFR or LGR5 expressed on CHO cell membranes can be assessed by flow cytometry (according to the FACS procedure previously described in WO2017 / 069628). In some aspects, the binding of candidate EGFR / LGR5 bispecific antibodies to LGR5 on CHO cells is confirmed by flow cytometry performed by standard procedures known in the art. The binding to CHO cells is compared with CHO cells that are not transfected with EGFR and / or LGR5 expression cassettes. The binding of candidate bispecific IgG1 to EGFR is determined using CHO cells transfected with EGFR expression constructs; LGR5 monospecific antibodies and EGFR monospecific antibodies, as well as unrelated IgG1 isotype control mAbs are included in the assay as controls (e.g., antibodies that bind LGR5 and another antigen, such as tetanus toxin (TT)).

[0340] The affinity of LGR5 and EGFR Fab of the candidate EGFR / LGR5 bispecific antibody to its target can be measured using BIAcore T100 by surface plasmon resonance (SPR) technology. Briefly, anti-human IgG mouse monoclonal antibody (Becton and Dickinson, catalog number 555784) was coupled to the surface of a CM5 sensor chip using free amine chemistry (NHS / EDC). The bispecific antibody was then captured on the sensor surface. Subsequently, recombinant purified antigen human EGFR (Sino Biological Inc, catalog number 11896-H07H) and human LGR5 protein were flowed over the sensor surface at a certain concentration range to measure the association rate and dissociation rate. After each cycle, the sensor surface was regenerated by HCl pulse, and the bispecific antibody was captured again. Based on the obtained sensor graph, the association rate and dissociation rate and affinity value of binding to human LGR5 and EGFR were determined using BIAevaluation software, as previously described for CD3 in US2016 / 0368988.

[0341] ADCC

[0342] In certain aspects of the human IgG subclass, the antibodies disclosed herein are generally bispecific full-length antibodies. In certain aspects, the antibodies belong to the human IgG1 subclass. Such antibodies have good ADCC properties, which, if desired, can be enhanced by techniques known in the art to have a favorable half-life when administered to humans in vivo, or by CH3 engineering techniques to provide modified heavy chains that preferentially form heterodimers rather than homodimers when co-expressed in clonal cells.

[0343] When an antibody inherently has low ADCC activity, its ADCC activity can be enhanced by modifying its constant region. Another approach to enhancing antibody ADCC activity is to enzymatically interfere with the glycosylation pathway, resulting in a reduction in fucose. Several in vitro methods are used to measure the efficacy of antibodies or effector cells in inducing ADCC. These include chromium-51 (Cr51) release assays, europium-51 (Eu) release assays, and sulfur-35 (S35) release assays. Typically, a labeled target cell line expressing a surface-exposed antigen is incubated with an antibody specific for that antigen. After washing, effector cells expressing the Fc receptor CD16 are co-incubated with the antibody-labeled target cells. Target cell lysis is then measured by scintillation counting or spectrophotometry, measuring the released intracellular label.

[0344] The bispecific antibodies disclosed herein may be enhanced by ADCC. In some aspects, such bispecific antibodies are afucosylated. In some aspects, when compared to the same antibody produced in normal CHO cells, the bispecific antibody comprises a reduced amount of fucosylation of N-linked carbohydrate structures in the Fc region. Low fucose levels are associated with increased binding of CD16 (FcγRIIIa) on NK effector cells, resulting in increased ADCC activity. In some aspects, in addition to its direct anti-tumor activity, the bispecific antibodies of the present disclosure can eliminate tumor cells under opsonization and subsequent natural killer (NK) cell-mediated ADCC activity and complement dependent cytotoxicity (CDC) activity.

[0345] An antibody comprising a variable domain that binds to the extracellular portion of EGFR and a variable domain that binds to the extracellular portion of LGR5 may further comprise one or more additional variable domains that bind to one or more other targets. In certain aspects, the other targets are proteins, such as membrane proteins that comprise an extracellular portion. As used herein, a membrane protein is a cell membrane protein, such as a protein in the outer membrane of a cell, which separates the cell from the outside world. The membrane protein has an extracellular portion. If a membrane protein comprises a transmembrane region located in the cell membrane of the cell, the membrane protein is at least located on the cell.

[0346] Antibodies with more than two variable domains are known in the art. For example, additional variable domains may be attached. In certain aspects, antibodies with three or more variable domains are multivalent multimeric antibodies, as described in PCT / NL2019 / 050199, which is incorporated herein by reference.

[0347] In certain aspects, the antibody is a bispecific antibody comprising two variable domains, wherein one variable domain binds to the extracellular portion of EGFR and the other variable domain binds to the extracellular portion of LGR5. In certain aspects, the variable domains are the variable domains described herein.

[0348] The functional portion of the antibodies described herein comprises at least a variable domain that binds to the extracellular portion of EGFR and a variable domain that binds to the extracellular portion of LGR5 described herein. Thus, it comprises the antigen-binding portion of the antibodies described herein and typically comprises the variable domains of the antibodies. The variable domains of the functional portion may be a single-chain Fv fragment or a so-called single-domain antibody fragment. In certain aspects, the antibody portion or derivative has at least two variable domains of an antibody or its equivalent. Non-limiting examples of such variable domains or their equivalents are F(ab)-fragments and single-chain Fv fragments. The functional portion of a bispecific antibody comprises the antigen-binding portion of a bispecific antibody, or a derivative and / or analog of the binding portion. As described above, the binding portion of the antibody is contained in the variable domain.

[0349] Preparations and pharmaceutical compositions

[0350] Also provided is a pharmaceutical composition comprising an antibody or functional portion, derivative, and / or analog thereof of the present disclosure and a fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 as disclosed herein, and a pharmaceutically acceptable carrier. In certain aspects, the present disclosure provides a pharmaceutical composition comprising an EGFR / LGR5 bispecific antibody and a fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38. In certain aspects, the present disclosure provides a combination or kit-of-parts of a pharmaceutical composition comprising an EGFR / LGR5 bispecific antibody and a separate pharmaceutical composition comprising a fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38.

[0351] The term "pharmaceutically acceptable" as used herein refers to a substance approved by a government regulatory agency or listed in the U.S. Pharmacopoeia or another generally recognized pharmacopoeia for use in animals, particularly humans, and includes any and all solvents, salts, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like that are physiologically compatible. The term "carrier" refers to a diluent, adjuvant, excipient, or vehicle that is administered with the compound. Such pharmaceutical carriers can be sterile liquids, such as water and oils, including oils of petroleum, animal, plant, or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil, glycerol polyethylene glycol ricinoleate, and the like. Water or physiological saline solutions, as well as aqueous glucose solutions and glycerol solutions, can be used as carriers, particularly for injectable solutions. Liquid compositions for parenteral administration can be formulated for injection or continuous infusion. Routes of administration via injection or infusion include intravesical, intratumoral, intravenous, intraperitoneal, intramuscular, intrathecal, and subcutaneous administration. Depending on the route of administration (eg, intravenous, subcutaneous, intraarticular, and the like), the active compound may be coated with a material to protect the compound from the action of acids and other natural conditions that may inactivate the compound.

[0352] The pharmaceutical composition comprising the antibody comprising a variable domain that binds to the extracellular portion of EGFR, a functional portion, derivative, and / or analog thereof can be contained in a container that is separate from, meaning not physically connected to, the container comprising the pharmaceutical composition comprising the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38.

[0353] Pharmaceutical compositions suitable for administration to human patients are typically formulated for parenteral administration, for example in a liquid carrier, or are suitable for reconstitution into liquid solutions or suspensions for intravenous administration. Such compositions can be formulated in dosage unit form for ease of administration and uniformity of dosage. Also included are solid formulations that are intended to be converted into liquid formulations for oral or parenteral administration shortly before use. Such liquid forms include solutions, suspensions, and emulsions.

[0354] The compositions and methods provided herein are particularly suitable for treating cancer in a patient, particularly head and neck cancer, gastric cancer, esophageal cancer, gastroesophageal junction cancer, non-small cell lung cancer or colorectal cancer. Thus, the compositions and methods can be used to treat various malignancies.

[0355] In certain aspects, the subject or patient is a mammal, particularly a human.

[0356] Previous treatment

[0357] In some aspects, the subject has not been previously treated with an anticancer therapy. In other words, the subject's cancer has not been previously treated with an anticancer therapy. In some aspects, the subject has not received (naive) anticancer therapy. In some aspects, the treatment with the disclosed therapeutic agent is a first-line treatment. In some aspects, the subject has not been previously treated with neoadjuvant therapy. In some aspects, the cancer is head and neck cancer, gastric cancer, esophageal cancer, gastroesophageal junction cancer, non-small cell lung cancer, or colorectal cancer.

[0358] In certain aspects, the subject has previously been treated with an anti-cancer therapy. In other words, the subject's cancer has previously been treated with an anti-cancer therapy. In certain aspects, the subject has previously been treated with an anti-cancer therapy. In other words, the subject's cancer has previously been treated with an anti-cancer therapy.

[0359] In certain aspects, the subject has previously been treated with two or more anti-cancer therapies. In other words, the subject's cancer has previously been treated with two or more anti-cancer therapies. In certain aspects, the subject has previously been treated with two or more anti-cancer therapies. In other words, the subject's cancer has previously been treated with two or more anti-cancer therapies.

[0360] In certain aspects, the cancer is head and neck cancer, gastric cancer, esophageal cancer, gastroesophageal junction cancer, non-small cell lung cancer, or colorectal cancer. In certain aspects, the subject has previously been treated with neoadjuvant therapy.

[0361] In certain aspects, the anti-cancer therapy is chemotherapy, radiation therapy, targeted therapy, and / or immunotherapy.

[0362] The subject may have previously been treated with one or more standard approved therapies or standard of care. Although surgery or radiation therapy may be preferred for most patients with early-stage or localized disease and may be considered for locally advanced disease, they may not be appropriate for all patients, for example, due to the anatomical location of the cancer. In certain aspects, the standard approved therapy or standard of care herein includes treatment by administering a chemotherapeutic agent, such as a platinum-based chemotherapeutic agent (e.g., cisplatin, carboplatin, oxaliplatin), an anti-tumor compound (e.g., methotrexate), a fluoropyrimidine (e.g., fluorouracil, 5-FU, capecitabine, a taxane (e.g., docetaxel or paclitaxel), a nucleoside analog (e.g., gemcitabine), a BCL-2 inhibitor, or SN-38, or any combination thereof. In certain aspects, the chemotherapy is fluorouracil, TAS-102, oxaliplatin, venetoclax, SN-38, FOLFOX, or FOLFIRI.

[0363] Targeted therapies disclosed herein include treatment with agents that block the action of certain enzymes, proteins, or other molecules involved in the growth or spread of cancer cells. Targeted therapies include, but are not limited to, angiogenesis inhibitors (e.g., bevacizumab) or monoclonal antibodies (e.g., trastuzumab, cetuximab, afatinib). In certain aspects, the targeted therapy is cetuximab.

[0364] In some aspects, immunotherapy includes treatment with immune checkpoint inhibitors. In some aspects, immune checkpoint inhibitors of the present disclosure target immune checkpoint proteins selected from PD-L1, PD-1, CTLA-4, B7-1 or B7-2. In some aspects, immune checkpoint inhibitors include durvalumab, pembrolizumab, ipilimumab, nivolumab, atezolizumab, retifanlimab, cemiplimab or other approved or developed anti-PD1, anti-PD-L1 antibodies. In some aspects, immune checkpoint inhibitors include durvalumab or pembrolizumab.

[0365] In certain aspects, the subject to be treated has progressed following treatment with fluorouracil, oxaliplatin, FOLFOX, FOLFIRI, targeted therapy, immunotherapy, or a combination thereof. In certain aspects, the treatment is for colorectal cancer.

[0366] In certain aspects, the subject or cancer to be treated has previously received treatment with a first-line standard of care suitable for CRC or mCRC. In certain aspects, the subject or cancer to be treated has previously received treatment and is administered a second-line treatment with an antibody comprising a variable domain that binds to the extracellular portion of EGFR and, optionally, a variable domain that binds to the extracellular portion of LGR5, or a functional portion, derivative, and / or analog thereof, in combination with a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

[0367] In some aspects, the subject or cancer to be treated has previously been treated and a third-line treatment or subsequent treatment is administered using an antibody or its functional portion, derivative, and / or analog comprising a variable domain that binds to the extracellular portion of EGFR and optionally a variable domain that binds to the extracellular portion of LGR5, and in combination with a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38. In some aspects, the previous treatment includes a standard of care treatment suitable for CRC or mCRC. In some aspects, the standard of care includes or is chemotherapy suitable for CRC or mCRC. In some aspects, the previous treatment includes or is fluoropyrimidine-oxaliplatin-based chemotherapy with or without bevacizumab or fluoropyrimidine-irinotecan-based chemotherapy with or without bevacizumab for CRC or mCRC.

[0368] In certain aspects, the subject or cancer has previously been treated with a fluoropyrimidine-oxaliplatin-based chemotherapy with or without bevacizumab, and an antibody comprising a variable domain that binds to the extracellular portion of EGFR and, optionally, a variable domain that binds to the extracellular portion of LGR5, or a functional portion, derivative, and / or analog thereof, is administered, and FOLFIRI is administered. In certain aspects, the subject or cancer has previously been treated with only the oxaliplatin-based chemotherapy. Thus, the use of the antibody comprising a variable domain that binds to the extracellular portion of EGFR and, optionally, a variable domain that binds to the extracellular portion of LGR5, or a functional portion, derivative, and / or analog thereof, and the administration of FOLFIRI is provided as a second-line therapy.

[0369] In certain aspects, the subject or cancer has previously been treated with fluoropyrimidine-irinotecan-based chemotherapy with or without bevacizumab and is administered an antibody comprising a variable domain that binds to the extracellular portion of EGFR and, optionally, a variable domain that binds to the extracellular portion of LGR5, or a functional part, derivative and / or analog thereof, and is administered FOLFOX.

[0370] In certain aspects, the subject or cancer has previously been treated with only the irinotecan-based chemotherapy. Thus, the use of the antibody comprising a variable domain that binds to the extracellular portion of EGFR and optionally a variable domain that binds to the extracellular portion of LGR5, or a functional part, derivative and / or analog thereof, and the administration of FOLFIRI is provided as a second-line therapy.

[0371] In certain aspects, the subject to be treated has progressed following treatment with fluorouracil, oxaliplatin, FOLFOX, FOLFIRI, targeted therapy, immunotherapy, or a combination thereof. In certain aspects, the treatment is for gastric cancer.

[0372] In certain aspects, the subject to be treated has progressed following treatment with a fluoropyrimidine, oxaliplatin, FOLFOX, FOLFIRI, targeted therapy, immunotherapy, or a combination thereof. In certain aspects, the treatment is for esophageal cancer, gastroesophageal junction cancer.

[0373] In certain aspects, the subject to be treated has progressed following treatment with radiation therapy, a fluoropyrimidine, a platinum-based therapy, a targeted therapy, an immunotherapy, or a combination thereof. In certain aspects, the treatment is for head and neck cancer.

[0374] In certain aspects, the subject to be treated has progressed following treatment with platinum-based therapy, targeted therapy, immunotherapy, or a combination thereof. In certain aspects, the treatment is for non-small cell lung cancer.

[0375] Application

[0376] Merus in its Multispecific antibodies targeting EGFR and LGR5 (G protein-coupled receptors rich in leucine repeats) have been developed in the antibody program. The efficacy of such multispecific antibodies has been evaluated in vitro and in vivo using patient-derived CRC organoids and mouse PDX models, respectively (see, for example, WO2017 / 069628; which is incorporated herein by reference). Multispecific antibodies targeting EGFR and LGR5 have been shown to inhibit tumor growth. The efficacy of such inhibitory antibodies has been shown to be correlated with the level of LGR5 RNA expression in cells from cancer. In certain aspects, the multispecific antibodies targeting EGFR and LGR5 are as described in WO2017 / 069628.

[0377] The present disclosure further provides a method for treating such cancer in a subject, the method comprising administering an antibody or its functional portion, derivative and / or analog and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38 to a subject in need. In certain aspects, the method comprises providing a 1500 mg dose of the antibody or its functional portion, derivative and / or analog to the subject. In certain aspects, the method comprises providing a fixed dose of 1500 mg of the antibody or its functional portion, derivative and / or analog to the subject. In certain aspects, the therapeutic agent or combination of therapeutic agents may be administered to the subject weekly, biweekly or monthly. In certain aspects, the therapeutic agent or combination of therapeutic agents is administered once every two weeks. In certain aspects, the therapeutic agent is pesentuzumab and is administered at a dose of 1500 mg once every two weeks.

[0378] In some aspects, the antibody or its functional part, derivative and / or analog is used at a dosage between 5 and 2000mg. In some aspects, the antibody or its functional part, derivative and / or analog is used at a dosage of 5, 20, 50, 90, 150, 225, 335, 500, 750, 1100, 1500mg or 2000mg. In some aspects, the antibody or its functional part, derivative and / or analog is used at a dosage of 750, 1100, 1500mg or 2000mg. In some aspects, the antibody or its functional part, derivative and / or analog is used at a dosage of 750mg. In some aspects, the antibody or its functional part, derivative and / or analog is used at a dosage of 1100mg. In some aspects, the antibody or its functional part, derivative and / or analog is used at a dosage of 1500mg. In some aspects, the antibody or its functional portion, derivative and / or analog is administered at a dosage of 2000 mg. As will be appreciated by those skilled in the art, dosage can be administered over time. In some aspects, the dosage is administered intravenously, for example, by infusion over 1-6 hours, or by infusion over 2-4 hours. In some aspects, the dosage is administered once every two weeks. In some aspects, the dosage is a flat dosage suitable for adults and / or subjects weighing at least 35 kilograms.

[0379] In certain aspects, pesentuzumab is administered at a dose of between 5 and 2000 mg. In certain aspects, pesentuzumab is administered at a dose of 5, 20, 50, 90, 150, 225, 335, 500, 750, 1100, 1500 mg, or 2000 mg. In certain aspects, pesentuzumab is administered at a dose of 750 mg. In certain aspects, pesentuzumab is administered at a dose of 1100 mg. In certain aspects, pesentuzumab is administered at a dose of 1500 mg. In certain aspects, pesentuzumab is administered at a dose of 2000 mg. As will be appreciated by those skilled in the art, dosages may be administered over time. In certain aspects, dosages are administered intravenously, for example, by infusion over 1-6 hours or by infusion over 2-4 hours. In certain aspects, the dosage is administered once every two weeks. In certain aspects, the dosage is a fixed dose suitable for adults and / or subjects weighing at least 35 kg.

[0380] In certain aspects, pesentuzumab is administered at a dose of 750 mg weekly. In certain aspects, pesentuzumab is administered at a dose of 750 mg every two weeks. In certain aspects, pesentuzumab is administered at a dose of 750 mg every three weeks.

[0381] In certain aspects, pesentuzumab is administered at a dose of 1100 mg weekly. In certain aspects, pesentuzumab is administered at a dose of 1100 mg every two weeks. In certain aspects, pesentuzumab is administered at a dose of 1100 mg every three weeks.

[0382] In certain aspects, pesentuzumab is administered weekly at a dose of 1500 mg. In certain aspects, pesentuzumab is administered every two weeks at a dose of 1500 mg. In certain aspects, pesentuzumab is administered every three weeks at a dose of 1500 mg.

[0383] In certain aspects, the antibody or functional portion, derivative and / or analog thereof comprises pesentumab and is administered in an amount that achieves at least 90%, at least 95%, or at least 99% engagement of the human receptor targets of both EGFR and LGR5 in a statistically significant number of subjects per relevant body weight. The 90% amount can be achieved using a fixed dose of about 1000 mg Q2W. The 95% amount can be achieved using a fixed dose of about 1100 to about 1200 mg Q2W.

[0384] In certain aspects, a premedication scheme is used. Such schemes may be applicable to reduce the possibility or severity of the reaction associated with the infusion. In general, (for example, oral, intravenous) steroids, such as dexamethasone and / or antihistamines, such as dexchlorpheniramine, diphenhydramine or chlorpheniramine are administered before treatment with the therapeutic agents mentioned herein. In certain aspects, the treatment of the present disclosure includes the premedication of paracetamol / acetaminophen, antihistamines or corticosteroids. In certain aspects, according to local standard clinical practice, the premedication is administered in the case of infusion-related reactions, allergic reactions and / or anaphylaxis. In certain aspects, 1500mg of the Pesentuzumab dosage is administered with antihistamines, pain relievers, antipyretics and / or anti-inflammatory drugs as premedication. In certain aspects, the 750 mg dose of pesentuzumab is premedicated with an antihistamine, a pain reliever, a fever reducer, and / or an anti-inflammatory drug.

[0385] As used herein, combination therapy, administration, or co-administration includes simultaneous therapy or administration of an antibody or its functional portion, derivative, and / or analog of the present disclosure and a fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38, which are administered separately or sequentially in the same or different dosage forms. Thus, in certain aspects, the antibody or its functional portion, derivative, and / or analog can be used in a method of treating cancer in a subject, wherein the antibody or its functional portion, derivative, and / or analog and the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 are administered simultaneously, separately, or sequentially. In other aspects, the antibody or its functional portion, derivative, and / or analog can be used in the treatment of cancer in a subject, wherein the antibody or its functional portion, derivative, and / or analog and the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 are administered simultaneously, separately, or sequentially.

[0386] In some aspects, the antibody or its functional portion, derivative, and / or analog can be used to manufacture a medicament for treating cancer in a subject, wherein the antibody or its functional portion, derivative, and / or analog and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 are administered simultaneously, separately, or sequentially. In some aspects, the antibody or its functional portion, derivative, and / or analog can be used to manufacture a medicament for treating cancer in a subject, wherein the antibody or its functional portion, derivative, and / or analog and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 are administered simultaneously, separately, or sequentially.

[0387] In some aspects, the antibody or its functional part, derivative and / or analog and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 can be used in the manufacture of one or more medicaments for treating cancer in a subject, wherein the antibody or its functional part, derivative and / or analog and the fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 are administered simultaneously, separately, or sequentially.

[0388] A product comprising the antibody or a functional part, derivative and / or analog thereof and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38 can be a combined preparation for simultaneous, separate or sequential use in treating cancer in a subject.

[0389] The antibody or its functional portion, derivative, and / or analog and the pyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 can also be administered according to any applicable schedule. In certain aspects, the antibody or its functional portion, derivative, and / or analog and the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 can be administered simultaneously in a single formulation. In certain aspects, the antibody or its functional portion, derivative, and / or analog and the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 can be formulated for separate administration, wherein they can be administered simultaneously or sequentially.

[0390] For example, in certain aspects, the antibody or its functional portion, derivative and / or analog can be administered first, followed by administration of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38, or vice versa. The dosage regimen in the above-mentioned treatment methods and uses is adjusted to provide the optimal desired response (e.g., a therapeutic response).

[0391] The antibodies, or functional portions, derivatives, and / or analogs thereof disclosed herein, and fluoropyrimidines, platinum-based chemotherapeutic agents, BCL-2 inhibitors, or SN-38 can be administered according to an applicable dose and by an applicable route (e.g., intravenous, intraperitoneal, intramuscular, intrathecal, or subcutaneous). For example, a single bolus injection can be administered, several divided doses can be administered over time, or the dose can be proportionally reduced or increased as indicated by the exigencies of the therapeutic situation.

[0392] In certain aspects, the present disclosure provides a method of treating cancer in a subject, wherein the method comprises administering to the subject an effective amount of the antibody or a functional portion, derivative and / or analog thereof, and an effective amount of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

[0393] In certain aspects, the antibody or its functional portion, derivative, and / or analog is administered prior to the administration of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38, e.g., the antibody or its functional portion, derivative, and / or analog is administered to the patient first, followed by the administration of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38. In one aspect, the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 is administered prior to the administration of the antibody or its functional portion, derivative, and / or analog, e.g., the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 is administered to the patient first, followed by the administration of the antibody or its functional portion, derivative, and / or analog (e.g., one or more minutes, hours, or days later). Such simultaneous or sequential administration results in both the antibody or its functional portion, derivative, and / or analog and the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 being present in the treated patient at the same time. The simultaneous presence of the antibody or its functional part, derivative and / or analog and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38 will support both cancer treatment induced by the antibody or its functional part, derivative and / or analog and inhibition of EGFR / LGR5 signaling mediated by the antibody or its functional part, derivative and / or analog.

[0394] In certain aspects, a single dose of the antibody, or its functional portion, derivative, and / or analog, and a single dose of a fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38, as disclosed herein, is administered to a subject. In certain aspects, the antibody, or its functional portion, derivative, and / or analog, and the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 are administered repeatedly over the course of treatment. For example, in certain aspects, multiple (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, or more) doses of a fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 and multiple (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, or more) doses of the antibody, or its functional portion, derivative, and / or analog, are administered to a subject in need of treatment.

[0395] In certain aspects, the administration of the antibody, or its functional portion, derivative, and / or analog, and the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 can be weekly, biweekly, or monthly, in which case they can be administered on the same day (e.g., simultaneously), or sequentially (e.g., one or more minutes, hours, or days before or after each other). When administered separately, the antibody, or its functional portion, derivative, and / or analog, and the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 can (but not necessarily) be administered according to the same administration (i.e., dosing) schedule. For example, a treatment cycle can include one or more administrations of the antibody, or its functional portion, derivative, and / or analog, and the therapeutically effective dose of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 can be administered more or less frequently than the antibody, or its functional portion, derivative, and / or analog. In certain aspects, each dose of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 and the antibody, or a functional portion, derivative, and / or analog thereof, can be administered on the same day, or the fluoropyrimidine, platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 can be administered one or more days before or after administration of the antibody, or a functional portion, derivative, and / or analog thereof.

[0396] In certain aspects, the dosage of the antibody, or its functional portion, derivative, and / or analog, and / or fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 varies over time. For example, the antibody, or its functional portion, derivative, and / or analog, and / or fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 can be administered initially at a high dose and can be reduced over time. In another aspect, the antibody, or its functional portion, derivative, and / or analog, and / or fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 can be administered initially at a low dose and can be increased over time.

[0397] In certain aspects, the amount of the antibody or its functional portion, derivative and / or analog and / or fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor or SN-38 in each dose is fixed. In another aspect, the amount of the antibody or its functional portion, derivative and / or analog and / or fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor or SN-38 in each dose is variable. For example, each maintenance (or subsequent) dose may be higher than or equal to the loading dose administered for the first time. In another aspect, each maintenance dose may be lower than or equal to the loading dose. The clinician may use a dose that is optimal based on the condition of the patient to be treated. The dose may depend on a variety of factors, including the stage of the disease. The specific dose to be administered based on the presence of one or more of these factors is within the skill of the artisan. In general, treatment is initiated with a smaller dose that is less than the optimal dose of the compound. Thereafter, the dose is increased in small amounts until the optimal effect under the circumstances is achieved. For convenience, if necessary, the total daily dose may be divided into multiple parts and administered over the course of a day. Intermittent therapy (e.g., one week within three weeks or three weeks within four weeks) may also be used.

[0398] In some aspects, the antibody or its functional portion, derivative and / or analog is administered at a dosage of 0.1, 0.3, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 mg / kg body weight. Alternatively, the antibody or its functional portion, derivative and / or analog is administered at a dosage of 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 mg / kg body weight. In some aspects, a fixed dose of 1500 mg is used to provide the antibody or its functional portion, derivative and / or analog to the subject. Fixed doses provide several advantages over surface or body weight administration because they reduce preparation time and reduce potential dosage calculation errors. In some aspects, the antibody or its functional portion, derivative and / or analog is provided at a dosage of at least 500 mg. In some aspects, the dosage is between 1100 and 2000 mg. In some aspects, the dosage is between 1100 and 1800 mg. As will be appreciated by those skilled in the art, dosage can be administered over time. For example, the dose can be administered intravenously, for example, by infusion over 1-6 hours or over 2-4 hours. In certain aspects, the antibody or its functional portion, derivative and / or analog is administered once every 2 weeks. In certain aspects, the antibody is pesentumomab and is administered once every 2 weeks at a fixed dose of 1500 mg. In particular, the fixed doses disclosed herein are suitable for adults and / or subjects weighing at least 35 kg. In certain aspects, the subject suffers from head and neck cancer, gastric cancer, esophageal cancer, gastroesophageal junction cancer, non-small cell lung cancer, or colorectal cancer.

[0399] In certain aspects, a premedication regimen may be used. Such regimens may be useful in reducing the likelihood or severity of infusion-related reactions. Generally, steroids such as dexamethasone and / or antihistamines such as dexclofamine, diphenhydramine, or chlorpheniramine are administered (e.g., orally, intravenously) prior to antibody treatment.

[0400] The treatment methods described herein are generally continued as long as the clinician supervising the patient's care deems the treatment method effective, i.e., the patient is responding to the treatment. Non-limiting parameters indicating that the treatment method is effective may include one or more of the following: reduction in tumor cells; inhibition of tumor cell proliferation; elimination of tumor cells; progression-free survival; and appropriate response of appropriate tumor markers (if applicable).

[0401] Regarding the frequency of administration of the antibody, or its functional portion, derivative, and / or analog, and / or fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38, one of ordinary skill in the art will be able to determine the appropriate frequency. For example, a clinician may decide to administer the antibody, or its functional portion, derivative, and / or analog, and / or fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 relatively infrequently (e.g., once every two weeks) and gradually shorten the time period between doses as tolerated by the patient. Exemplary lengths of time associated with a course of treatment according to the claimed methods include: about one week; about two weeks; about three weeks; about four weeks; about five weeks; about six weeks; about seven weeks; about eight weeks; about nine weeks; about ten weeks; about eleven weeks; about twelve weeks; about thirteen weeks; about fourteen weeks; about fifteen weeks; about sixteen weeks; about seventeen weeks; about eighteen weeks; about nineteen weeks; about twenty weeks; about twenty-one weeks; about twenty-two weeks; about twenty-three weeks; about twenty-four weeks; about seven months; about eight months; about nineteen months; about ten months; about eleven months; about twelve months; about thirteen months; about fourteen months; about fifteen months; about sixteen months; about seventeen months; about eighteen months; about nineteen months; about twenty months; about twenty-one months; about twenty-two months; about twenty-three months; about twenty-four months; about thirty months; about three years; about four years; about five years; and permanently (e.g., ongoing maintenance therapy). The foregoing durations may be associated with one or more rounds / cycles of treatment.

[0402] Any suitable means can be used to assess the efficacy of the therapeutic methods provided herein. In certain aspects, a reduction in the number of cancer cells is used as an objective response criterion to analyze the clinical efficacy of treatment. Patients (e.g., humans) treated according to the methods disclosed herein experience improvement in at least one symptom of cancer. In certain aspects, one or more of the following may occur: the number of cancer cells may be reduced; cancer recurrence may be prevented or delayed; one or more symptoms associated with cancer may be alleviated to a certain extent. In addition, in vitro tests are performed to determine T cell-mediated target cell lysis. In certain aspects, tumor assessment is based on CT scans and / or MRI scans, see, for example, RECIST 1.1 guidelines (Response Evaluation Criteria in Solid Tumours) (Eisenhauer et al., 2009 Eur J Cancer 45: 228–247). Such assessments are generally performed every 4 to 8 weeks after treatment.

[0403] In some aspects, the tumor cells are no longer detectable after treatment as described herein. In some aspects, the subject is in partial or complete remission. In some aspects, the subject's overall survival, median survival rate and / or progression-free survival increase.

[0404] The combinations of the present disclosure (i.e., combinations of the antibodies or functional portions, derivatives, and / or analogs thereof with fluoropyrimidines, platinum-based chemotherapeutics, BCL-2 inhibitors, or SN-38) can also be used with other well-known therapies (e.g., chemotherapy or radiation therapy) that are selected for their particular usefulness against the cancer being treated.

[0405] Methods for the safe and effective administration of chemotherapeutic agents are known to those skilled in the art. In addition, their administration is described in standard literature. For example, the administration of many chemotherapeutic agents is described in the Physicians' Desk Reference (PDR), e.g., the 1996 edition (Medical Economics Company, Montvale, NJ 07645-1742, USA); the disclosure of which is incorporated herein by reference.

[0406] It will be apparent to those skilled in the art that the administration of chemotherapeutic agents and / or radiation therapy may vary depending on the disease being treated and the known effects of the chemotherapeutic agents and / or radiation therapy on the disease. In addition, according to the knowledge of the skilled clinician, the treatment regimen (e.g., dosage and time of administration) may vary in view of the observed effects of the administered therapeutic agent on the patient and in view of the observed response of the disease to the administered therapeutic agent.

[0407] In certain aspects, at the start of treatment, at least one, more than one or all of the following inclusion factors IF1-IF16 are applicable to the subject to be treated. In certain aspects, the subject comprises or meets all of IF1-IF16:

[0408] IF 1. Aged at least 18 years old.

[0409] IF 2. Patients with histologically or cytologically confirmed solid tumors with evidence of metastatic or locally advanced disease that is intolerant to standard therapy with curative intent, or with locally advanced unresectable or metastatic disease.

[0410] IF 3. Previously diagnosed with unresectable or metastatic adenocarcinoma of the colon or rectum, confirmed by histology or cytology. Subject is RAS and / or RAF wild-type, as determined by NGS on tumor tissue (primary or metastatic).

[0411] IF4. Subjects have not received prior anti-EGFR therapy (naive).

[0412] IF5. Disease progression must have occurred during or within 6 months of the previous first-line chemotherapy, as confirmed by appropriate radiological testing.

[0413] IF6.1. Subjects treated with the combination of pesentuzumab and FOLFIRI should have received only a single prior chemotherapy regimen for the metastatic setting, which was usually first-line fluoropyrimidine-oxaliplatin-based chemotherapy ± bevacizumab.

[0414] IF6.2. Subjects treated with the combination of pesentuzumab and FOLFOX should have received only a single prior chemotherapy regimen for the metastatic setting, which was usually first-line fluoropyrimidine-irinotecan-based chemotherapy with or without bevacizumab.

[0415] IF 7. Have a new baseline tumor sample (e.g., formalin-fixed, paraffin-embedded block [FFPE]) from a metastatic or primary site. If the subject has such a tumor sample from a sufficient collection of material (at least 20 slides with >20% tumor content) and has not received further anticancer treatment since the sample was collected, a new tumor biopsy is not required at baseline. Archival FFPE slides will not be accepted.

[0416] IF 8. Agree with biopsy.

[0417] IF 9. Patients have radiographically measurable disease as defined by RECIST version 1.1.

[0418] IF 10. Eastern Cooperative Oncology Group (ECOG) expression status is 0 or 1.

[0419] IF 11. Life expectancy ≥ 12 weeks, usually based on investigator assessment.

[0420] IF 12. Left ventricular ejection fraction (LVEF) of at least 50% as shown by echocardiography (ECHO) or multiple gated acquisition (MUGA).

[0421] IF 13. Adequate organ function:

[0422] IF 13.1 absolute neutrophil count (ANC) of at least 1.5 x 109 / L.

[0423] IF 13.2 Hemoglobin level is at least 9 g / dL.

[0424] IF 13.3 Platelet level is at least 100 × 109 / L.

[0425] Total serum calcium after correction for IF 13.4 was within the normal range.

[0426] IF 13.5 Serum magnesium, sodium, corrected total calcium, phosphate, and potassium are within normal ranges (or corrected with supplementation or appropriate treatment).

[0427] IF 13.6 alanine aminotransferase (ALT), aspartate aminotransferase (AST) equal to or less than 2.5 times the upper limit of normal (ULN) and total bilirubin equal to or less than 1.5 times ULN, provided that the subject has Gilbert's syndrome, total bilirubin less than or equal to 3.0 times ULN, or direct bilirubin less than or equal to 1.5 times ULN; provided that in the case of liver involvement, ALT / AST equal to or less than 5 times ULN and total bilirubin equal to or less than 2 times ULN.

[0428] IF 13.7 for patients older than 65 years with a serum creatinine level less than or equal to 1.5 times the ULN, or a creatinine clearance of at least 60 mL / min, calculated according to the Cockroft and Gault formula or the Modification of Diet in Renal Disease (MDRD) formula.

[0429] IF 13.8 Serum albumin level is at least 3 g / dL.

[0430] IF International Normalized Ratio (INR) 13.9 or prothrombin time (PT) level less than or equal to 1.5 x ULN, unless the patient is receiving anticoagulant therapy and is within the therapeutic range for the intended anticoagulant.

[0431] • IF 13.10 Activated partial thromboplastin time (APTT) or PTT is less than or equal to 1.5×ULN, unless the patient is receiving anticoagulant therapy and is within the therapeutic range of the intended anticoagulant.

[0432] IF 14. If not tested for human immunodeficiency virus (HIV) within the past 6 months, the subject is willing to undergo HIV testing. Known HIV-positive patients are eligible as long as the cluster of differentiation 4 (CD4+) count is greater than 300 / μL, the viral load is undetectable, and the subject is currently receiving highly active antiretroviral therapy (HAART).

[0433] • IF 15. Hepatitis B (HBsAg) positive but receiving antiviral treatment with lamivudine, tenofovir, entecavir or other antiviral agents, starting at least 7 days from the start of treatment according to the present disclosure.

[0434] IF 16. History of hepatitis B (anti-HBc positive, HBsAg and hepatitis B virus [HBV]-DNA negative).

[0435] IF 17. Hepatitis C virus (HCV) ribonucleic acid (RNA) test is positive, provided that the HCV infection resolves spontaneously (i.e., HCV antibodies are positive but HCV-RNA is undetectable), or subjects who have achieved sustained remission after antiviral treatment show no detectable HCV RNA when using an IFN-free regimen for at least 6 months or an IFN-based regimen for at least 12 months after stopping antiviral treatment.

[0436] In certain aspects, all values for organ function measures based on IF13 have an upper limit observed in healthy subjects.

[0437] In some aspects, the experimenter included in the treatment meets any one or more factors selected from IF1-IF16. In some aspects, the experimenter included in the treatment meets factors IF2, IF3, IF4, IF5, IF6, IF9, IF10, IF12, IF13, IF14, IF15, IF16 and IF17. In some aspects, the experimenter included in the treatment meets factors IF2, IF3, IF4, IF5 and IF6. In some aspects, the experimenter included in the treatment meets factors IF3 and IF6.

[0438] In certain aspects, at the start of treatment, at least one, more than one, or all of the following exclusion factors EF1-EF18 apply to subjects enrolled in treatment:

[0439] EF 1. Have central nervous system metastases that are untreated and symptomatic, or require radiation therapy or surgery, or require ongoing steroid therapy to control symptoms within 14 days of starting the disclosed treatment.

[0440] EF 2. With known leptomeningeal involvement.

[0441] EF 3. Participated in another clinical trial or received any investigational drug within 4 weeks before starting this treatment.

[0442] EF 4. Systemic anticancer therapy within 4 weeks or 5 half-lives (whichever is longer) after the first dose of the present disclosure. For cytotoxic agents with major delayed toxicity (e.g., mitomycin C, nitrosoureas), or anticancer immunotherapy, a six-week washout period is required before starting treatment with the present disclosure.

[0443] EF 5. Requires immunosuppressive drugs (eg, methotrexate, cyclophosphamide).

[0444] EF 6. Subjects who have received major surgery or radiation therapy within 3 weeks of starting treatment with the present disclosure are excluded if they have received prior radiation therapy that damaged at least 25% or more of the bone marrow at any time.

[0445] EF 7. Persistent grade >1 clinically significant toxicity related to prior antineoplastic therapy (except alopecia); provided that stable sensory neuropathy of National Cancer Institute - Common Terminology Criteria for Adverse Events (NCI-CTCAE) v4.03 or v5.0 grade equal to 2 (or lower) is allowed or as current status at the time of dosing.

[0446] EF 8. History of any excipients required for this study, including pesentuzumab, human proteins, or any non-IMP treatment.

[0447] EF 9. Uncontrolled hypertension (systolic blood pressure > 150 mmHg and / or diastolic blood pressure > 100 mmHg) or unstable angina despite appropriate treatment.

[0448] EF 10. A history of congestive heart failure of New York Heart Association (NYHA) class II-IV, or severe arrhythmia requiring treatment (excluding atrial fibrillation and paroxysmal supraventricular tachycardia).

[0449] EF 11. Suffering from a myocardial infarction within six months of initiation of treatment of the present disclosure.

[0450] EF 12. History of prior malignancy, excluding excised cervical intraepithelial neoplasia, or non-melanoma skin cancer, or a cancer that was curatively treated with low risk of recurrence and no evidence of disease for at least three years prior to initiation of the disclosed treatment.

[0451] EF 13. Patients with any cause of dyspnea at rest or other conditions requiring continuous oxygen therapy.

[0452] EF 14. History of interstitial lung disease (ILD) (e.g., pneumonia or pulmonary fibrosis) or evidence of ILD on baseline chest computed tomography (CT) scan.

[0453] EF 15. Presence of a serious disease or medical condition at the time of initiation of treatment of the present disclosure, including but not limited to: uncontrolled active infection, clinically significant pulmonary, metabolic, or psychiatric disease.

[0454] EF 16. Subjects with active hepatitis B surface antigen infection (HBsAg positive) who are not receiving antiviral treatment.

[0455] EF 17. Hepatitis C virus (HCV) test was positive.

[0456] EF 18. Subjects with Child-Pugh B or C cirrhosis status; subjects with fibrolamellar HCC, subjects with sarcomatoid HCC, or subjects with mixed cholangiocarcinoma and HCC.

[0457] EF 18. Pregnant or breastfeeding subjects; subjects of childbearing potential must use highly effective contraception prior to initiating treatment of the present disclosure, during treatment, and for six months after the last administration of petosemtamab.

[0458] In certain aspects, the subjects included in the treatment meet any one or more factors selected from EF1-EF18. In certain aspects, the subjects included in the treatment meet all EF1-EF18 factors. In certain aspects, the subjects included in the treatment meet IF13, IF16 factors.

[0459] The ECOG expression status score grades in this area are defined as follows, meaning: 0 Fully active, able to perform all pre-disease manifestations without restriction. 1 Vigorous physical activity is limited, but ambulatory and able to perform light or sedentary work, such as light housework and office work. 2 Able to ambulatory and able to take care of themselves, but unable to perform any work activities. At most or approximately more than 50% of waking time. 3 Only limited self-care can be performed, and more than 50% of waking time is confined to bed or chair. 4 Total disability. Unable to perform any self-care. Completely confined to bed or chair. 5 Death.

[0460] The Child-Pugh score, also known as the Child-Pugh classification, the Child-Turcotte-Pugh (CTP) calculator, or the Child criteria, is used herein according to standard clinical practice. The Child-Pugh score is determined by scoring five clinical indicators of liver disease and the likelihood of eventual liver failure. Each indicator is scored as 1, 2, or 3, with 3 being the most severe. The five clinical indicators are total bilirubin, serum albumin level, prothrombin time (or prolonged or INR, as blood clotting time), ascites, and hepatic encephalopathy. Grade A represents 5 to 6 points, the least severe liver disease, with a one to five year survival rate of 95%. Grade B: 7 to 9 points, moderate to severe liver disease, with a one to five year survival rate of 75%. Grade C: 10 to 15 points, the most severe liver disease, with a one to five year survival rate of 50%. Based on clinical measurements, the following scores are given. Encephalopathy: None = 1 point, Grades 1 and 2 = 2 points, and Grades 3 and 4 = 3 points. Ascites: None = 1 point, Mild = 2 points, Moderate = 3 points. Bilirubin: Less than 2 mg / ml = 1 point, 2 to 3 mg / ml = 2 points, More than 3 mg / ml = 3 points. Albumin: More than 3.5 mg / ml = 1 point, 2.8 to 3.5 mg / ml = 2 points, Less than 2.8 mg / ml = 3 points. Prothrombin time (PT, prolonged seconds): Less than 4 seconds = 1 point, 4 to 6 seconds = 2 points, More than 6 seconds = 3 points. Alternatively, the International Normalized Ratio (INR) can be used as an alternative to the PT: INR less than 1.7 = 1 point, INR 1.7 to 2.2 = 2 points, INR greater than 2.2 = 3 points.

[0461] For this article, renal clearance was calculated for subjects aged <65 years according to the Cockcroft and Gault formula: men = 1.25 x body weight (kg) x (140 - age) / serum creatinine (μmol / L). Women = 1.04 x body weight (kg) x (140 - age) / serum creatinine (μmol / L).

[0462] For this article, Modification of Diet in Renal Disease (MDRD) was calculated according to the following formula: Patient age > 65 years. For male subjects aged 65 years and older, the calculation is as follows: Male = 186 x (Serum creatinine (μmol / L) x 0.0113) - 1.154 x Age - 0.203. For subjects with darker skin, the result was multiplied by 1.21. For female subjects, the result was multiplied by 0.742.

[0463] Component kits and combinations of therapeutic agents

[0464] In certain aspects, the present disclosure provides a kit or product comprising a pharmaceutical composition comprising a therapeutically effective amount of an EGFR / LGR5 bispecific antibody and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 suitable for use in the aforementioned methods, and a pharmaceutically acceptable carrier. In certain aspects, the kit or product may optionally further comprise instructions for use, such as an administration plan, to allow a practitioner (e.g., a physician, nurse, or patient) to administer the composition contained therein to a patient suffering from cancer.

[0465] Thus, in certain aspects, the present disclosure provides a kit of parts comprising:

[0466] - antibodies or functional parts, derivatives and / or analogues thereof,

[0467] - A fluoropyrimidine, platinum-based chemotherapy agent, BCL-2 inhibitor, or SN-38; and

[0468] - Instructions for use of the EGFR / LGR5 bispecific antibody and instructions for use of a fluoropyrimidine, platinum-based chemotherapy agent, BCL-2 inhibitor, or SN-38.

[0469] In some aspects, the instructions for use of pesentuzumab include instructions for dosing at 1500 mg. In some aspects, the instructions for use are directed to the use of the antibody, or a functional portion, derivative, and / or analog thereof, a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38, in the treatment of adenocarcinoma or squamous cell carcinoma. In some aspects, the instructions for use are directed to the treatment of head and neck cancer, gastric cancer, esophageal cancer, gastroesophageal junction cancer, non-small cell lung cancer, or colorectal cancer. In some aspects, the instructions for use are directed to the treatment of colorectal cancer.

[0470] In certain aspects, the kit or product includes multiple packages of single-dose pharmaceutical compositions, each single-dose pharmaceutical composition containing an effective amount of an antibody or a functional portion thereof, derivative and / or analog and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38, which is administered single-time according to the above-described methods. The kit or product may also include an instrument or device required for administering the pharmaceutical composition. For example, the kit or product may provide one or more pre-filled syringes containing a unit dose of an antibody or a functional portion thereof, derivative and / or analog and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 in the same container or in separate containers to be administered as separate and distinct compositions.

[0471] In certain aspects, the antibody or functional portion, derivative, and / or analog thereof and one or both of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 are provided in a solid form suitable for reconstitution and subsequent administration according to accompanying instructions.

[0472] In certain aspects, the present disclosure provides a combination of an antibody, or a functional portion, derivative, and / or analog thereof, and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 for use in treating cancer in a subject in need thereof.

[0473] In certain aspects, the present disclosure provides a combination of:

[0474] - an antibody of the present disclosure or a functional part, derivative and / or analogue thereof,

[0475] - instructions for use of the antibody or a functional part, derivative and / or analogue thereof in the treatment of cancer in a subject,

[0476] - Instructions for use of a fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 in the subject's cancer treatment.

[0477] In certain aspects, the present disclosure provides a combination of:

[0478] - Fluoropyrimidines, platinum-based chemotherapy agents, BCL-2 inhibitors, or SN-38,

[0479] - Instructions for use of a fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 in the subject's cancer treatment,

[0480] - and instructions for use of the antibody or functional part, derivative and / or analogue thereof in the treatment of cancer in a subject.

[0481] In certain aspects, the instructions for use include the amount of the antibody, or functional part, derivative, and / or analog thereof, to be used and / or the amount of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 to be used, and / or the dosing interval and / or the cancer to be treated.

[0482] In a still further aspect, the composition or combination or kit or product includes one or more additional active agents.

[0483] The compounds and compositions disclosed herein are useful as therapy and in treatment, thereby as medicaments, and in methods of preparing medicaments.

[0484] In certain aspects, the present disclosure provides a pharmaceutical composition and instructions for use thereof in the treatment of such cancers, wherein the pharmaceutical composition comprises an antibody or a functional portion, derivative, and / or analog thereof, wherein the antibody or a functional portion, derivative, and / or analog thereof comprises a variable domain of the present disclosure that can bind to the extracellular portion of EGFR.

[0485] In certain aspects, the present disclosure provides: a pharmaceutical composition for treating cancer comprising an antibody or a functional portion, derivative, and / or analog thereof, wherein the antibody or functional portion, derivative, and / or analog thereof comprises a variable domain of the present disclosure that can bind to the extracellular portion of EGFR; and a pharmaceutical composition for treating the cancer comprising a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 of the present disclosure.

[0486] In certain aspects, the present disclosure provides a pharmaceutical composition for treating cancer, comprising an antibody or a functional portion, derivative, and / or analog thereof, wherein the antibody or functional portion, derivative, and / or analog thereof comprises a variable domain of the present disclosure that can bind to the extracellular portion of EGFR, wherein the pharmaceutical composition is administered in combination with a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 of the present disclosure.

[0487] In certain aspects, the present disclosure relates to a pharmaceutical composition for treating cancer, comprising an antibody or a functional portion, derivative and / or analog thereof, wherein the antibody or functional portion, derivative and / or analog thereof comprises a variable domain of the present disclosure that can bind to the extracellular portion of EGFR, wherein the subject to be treated is administered a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 before, simultaneously with, or after administration of the bispecific antibody.

[0488] In certain aspects, the present disclosure relates to a pharmaceutical composition for treating cancer in a subject, comprising a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38, wherein the subject to be treated is administered an antibody, or a functional portion, derivative, and / or analog thereof, comprising a variable domain of the present disclosure that binds to the extracellular portion of EGFR, prior to, concurrently with, or after administration of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38.

[0489] Therefore, the present disclosure relates to a combination of agents for treating cancer in a subject, comprising administering to the subject a plurality of different agents to treat the cancer, wherein the treatment comprises administering the agents simultaneously, sequentially, or separately. In certain aspects, the agent comprises an antibody, or a functional portion, derivative, and / or analog thereof, comprising a variable domain of the present disclosure that binds to the extracellular portion of EGFR, and the other different agents comprise a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

[0490] In certain aspects, the antibodies or functional portions, derivatives, and / or analogs comprising the variable domains of the present disclosure that can bind to the extracellular portion of EGFR can be administered simultaneously, sequentially, or separately with the fluoropyrimidines, platinum-based chemotherapeutics, BCL-2 inhibitors, or SN-38 disclosed herein. The combination of the antibodies or functional portions, derivatives, and / or analogs comprising the variable domains of the present disclosure that can bind to the extracellular portion of EGFR and the fluoropyrimidines, platinum-based chemotherapeutics, BCL-2 inhibitors, or SN-38 thus encompasses simultaneous, sequential, or separate administration.

[0491] Thus, in certain aspects, the present disclosure provides an antibody or a functional portion, derivative and / or analog of an antibody comprising a variable domain of the present disclosure that binds to the extracellular portion of EGFR, for use in a method of treating cancer, wherein the treatment further comprises administering a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38, wherein optionally the antibody or its functional portion, derivative and / or analog is administered simultaneously, sequentially, or separately with the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38.

[0492] Thus, in certain aspects, the present disclosure provides a method of treating a subject having cancer, comprising administering to the subject an effective amount of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 and an antibody comprising a variable domain of the present disclosure that binds to the extracellular portion of EGFR, or a functional portion, derivative, and / or analog of the antibody, wherein optionally the antibody or its functional portion, derivative, and / or analog is administered simultaneously, sequentially, or separately with the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38.

[0493] Thus, in certain aspects, the present disclosure provides the use of an antibody or functional portion, derivative, and / or analog comprising a variable domain of the present disclosure that binds to the extracellular portion of EGFR and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 in the manufacture of a medicament for treating cancer, wherein the antibody or functional portion, derivative, and / or analog is optionally administered simultaneously, sequentially, or separately with the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38. In certain aspects, the antibody or functional portion, derivative, and / or analog is administered before, simultaneously, or after administration of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38.

[0494] In certain aspects, an antibody or functional portion, derivative, and / or analog comprising a variable domain of the present disclosure that binds to the extracellular portion of EGFR is used in the manufacture of a medicament for treating cancer, and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 is used in the manufacture of a medicament for treating the cancer, wherein the antibody or functional portion, derivative, and / or analog is optionally administered simultaneously, sequentially, or separately with the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38. Alternatively, the antibody or functional portion, derivative, and / or analog is administered before, simultaneously, or after the administration of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38.

[0495] definition

[0496] In order to make this specification easier to understand, certain terms are defined below. Additional definitions may be set forth throughout the detailed description where deemed necessary.

[0497] As used herein, the singular forms "a," "an," and "the" include plural references. The use of the terms "include," "have," "includes," and other forms such as "includes (plural verb)," "includes (singular verb)," "includes (past tense verb)," "has (singular verb)," "has (plural verb)," "has (past tense verb)," "includes (plural verb)," "includes (singular verb)," and "includes (past tense verb)" is not limiting.

[0498] As used herein, the term "antibody" refers to a protein molecule belonging to the class of immunoglobulins that contains one or more domains that bind to an epitope on an antigen, wherein such domains are from or derived from, or share sequence homology with, the variable regions of an antibody. Antibodies are generally composed of basic structural units, each having two heavy chains and two light chains. Antibodies according to the present disclosure are not limited to any particular form or method of producing the same.

[0499] "Bispecific antibodies" are antibodies as described herein, wherein one domain of the antibody binds to a first antigen and a second domain of the antibody binds to a second antigen, wherein the first and second antigens are different, or wherein one domain binds to a first epitope on an antigen and the second domain binds to a second epitope on an antigen. The term "bispecific antibody" also encompasses antibodies in which one heavy chain variable region / light chain variable region (VH / VL) combination binds to a first antigen or an epitope on an antigen and a second VH / VL combination binds to a second antigen or an epitope on an antigen. The term further encompasses antibodies in which the VH specifically recognizes the first antigen and the VL (which pairs with the VH in an immunoglobulin variable region) specifically recognizes the second antigen. The resulting VH / VL pairing will bind to either antigen 1 or antigen 2. Such so-called "two-in-one antibodies" are described, for example, in WO 2008 / 027236, WO 2010 / 108127, and Schaefer et al. (Cancer Cell 20, 472-486, October 2011). The bispecific antibodies according to the present disclosure are not limited to any particular bispecific format or method of producing the same.

[0500] As used herein, the term "common light chain" means the two light chains (or their VL portions) in a bispecific antibody. The two light chains (or their VL portions) may be identical or have some amino acid sequence differences, while the binding specificity of the full-length antibody is unaffected. The terms "common light chain," "common VL," "single light chain," "single VL," whether or not the term "rearrangement" is added, are used interchangeably herein. "Common" also refers to the functional equivalents of light chains with different amino acid sequences. There are many variants of the light chain, in which there are mutations (deletions, substitutions, insertions, and / or additions) that do not affect the formation of the functional binding region. In certain aspects, the light chain of the present disclosure may also be a light chain as specified herein, having 0 to 10 amino acid insertions, deletions, substitutions, additions, or combinations thereof. In certain aspects, the light chain of the present disclosure may also be a light chain as specified herein, having 0 to 5 amino acid insertions, deletions, substitutions, additions, or combinations thereof. For example, non-identical but still functionally equivalent light chains can be made or found within the definition of a common light chain as used herein, e.g., by introducing and testing conservative amino acid changes, changes in amino acids in regions that do not or only partially contribute to binding specificity when paired with a heavy chain, etc.

[0501] As used herein, "comprising" and its conjunctions are used in a non-limiting sense, meaning that the items following the word are included, but items not specifically mentioned are not excluded. In addition, the verb "consisting of" can be replaced with "consisting essentially of", meaning that the compounds or auxiliary compounds as defined herein may contain additional components in addition to the specifically specified components, which additional components do not change the unique characteristics of the present disclosure.

[0502] "Antibody derivatives" are proteins that deviate from the amino acid sequence of a native antibody by up to 20 amino acids, excluding the CDR regions. Antibody derivatives disclosed herein are antibodies that deviate from the amino acid sequence by up to 20 amino acids. Functional parts, derivatives and / or analogs maintain the binding specificity of the (bispecific) antibody. An "antibody analog" is a protein that may not be identical in structure, form or origin, but maintains the binding specificity of the antibody to which it is an analog.

[0503] The "percent identity (%)" of nucleic acid or amino acid sequences referred to herein is defined as the percentage of residues in a candidate sequence that are identical to the residues in a selected sequence after alignment for optimal comparison purposes. The percent sequence identity of nucleic acid sequences was compared using Vector NTI Advance The results were determined using the AlignX application of 11.5.2 software, using the original settings and a modified ClustalW algorithm (Thompson, JD, Higgins, DG, and Gibson T.J., (1994) Nucl. Acid Res. 22(22): 4673-4680), the swgapdnamt scoring matrix, a gap opening penalty of 15 and a gap extension penalty of 6.66. Amino acid sequences were aligned using the AlignX application of 11.5.2 software using native settings and a modified ClustalW algorithm (Thompson, JD, Higgins, DG, and Gibson TJ, (1994) Nucl. Acid Res. 22(22):4673-4680), a blosum62mt2 scoring matrix, a gap opening penalty of 10, and a gap extension penalty of 0.1.

[0504] Since antibodies generally recognize an epitope of an antigen, and this epitope may also be present in other compounds, an antibody of the present disclosure that "specifically recognizes" an antigen (e.g., EGFR or LGR5) may also recognize other compounds if such other compounds contain the same type of epitope. Therefore, the term "specifically recognizes" with respect to the interaction between an antigen and an antibody does not exclude the binding of an antibody to other compounds containing the same type of epitope.

[0505] The term "epitope" or "antigenic determinant" refers to a site on an antigen to which an immunoglobulin or antibody specifically binds. An epitope can be formed by contiguous amino acids or non-contiguous amino acids juxtaposed by tertiary folding of the protein (so-called linear and conformational epitopes). Epitopes formed by contiguous, linear amino acids are generally retained upon exposure to denaturing solvents, whereas epitopes formed by tertiary folded conformations are generally lost upon treatment with denaturing solvents. An epitope can typically include 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 amino acids in a unique spatial conformation.

[0506] As used herein, the terms "subject" and "patient" are used interchangeably and refer to mammals, such as humans, mice, rats, hamsters, guinea pigs, rabbits, cats, dogs, monkeys, cows, horses, pigs, etc. (e.g., a patient suffering from cancer, such as a human patient).

[0507] Unless otherwise specified, the terms cancer and tumor are used herein to refer generally to cancer.

[0508] As used herein, the terms "treat (verb)", "treat (gerund)" and "treatment (noun)" mean any type of intervention or procedure on a subject, or the administration of an active agent or combination of active agents to a subject, for the purpose of reversing, alleviating, ameliorating, inhibiting or slowing or preventing the progression, development, severity or recurrence of symptoms, complications, conditions or biochemical markers associated with a disease.

[0509] As used herein, "effective treatment" or "positive therapeutic response" means a treatment that produces a beneficial effect, e.g., an improvement in at least one symptom of a disease or disorder (e.g., cancer). A beneficial effect can take the form of an improvement over baseline, including improvements that were measured or observed before starting therapy according to the present method. For example, a beneficial effect can take the form of slowing, stabilizing, halting, or reversing the progression of a cancer in a subject at any clinical stage, as evidenced by a reduction or elimination of clinical or diagnostic symptoms of the disease or markers of the cancer. An effective treatment can, for example, reduce tumor size, reduce the presence of circulating tumor cells, reduce or prevent metastasis of a tumor, slow or stop tumor growth, and / or prevent or delay tumor recurrence or recurrence.

[0510] The term "effective amount" or "therapeutically effective amount" means the amount of a pharmaceutical agent or pharmaceutical composition that provides the desired biological, therapeutic and / or preventive result. The result can be one or more of a reduction, improvement, alleviation, reduction, delay and / or alleviation of the signs, symptoms or causes of a disease, or any other desired change in a biological system. In terms of tumor development, an effective amount is an amount sufficient to delay tumor development. In terms of tumor recurrence, an effective amount is an amount sufficient to prevent or delay tumor recurrence. The effective amount can be administered once or multiple times. An effective amount of a pharmaceutical agent or pharmaceutical composition can: (i) reduce the number of cancer cells; (ii) reduce tumor size; (iii) inhibit, delay, slow down and prevent cancer cells from infiltrating into surrounding organs to a certain extent; (iv) inhibit tumor metastasis; (v) inhibit tumor growth; (vi) prevent or delay the occurrence and / or recurrence of tumors; and / or (vii) alleviate one or more symptoms associated with cancer to a certain extent. In one aspect, an "effective amount" is an amount of an antibody disclosed herein and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 as a composition to affect a reduction in cancer (e.g., a reduction in the number of cancer cells), slow the progression of cancer, or prevent the regeneration or recurrence of cancer. As previously described herein, the antibodies or functional portions, derivatives, and / or analogs thereof that bind to EGFR or to both EGFR and LGR5 and the fluoropyrimidine, platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 (as a composition) disclosed herein are also referred to herein as "therapeutic agents." In certain aspects, an effective amount of an antibody or functional portion, derivative, and / or analog thereof that binds to EGFR or to both EGFR and LGR5 disclosed herein is a fixed dose of 1500 mg administered biweekly to a subject suffering from a cancer disclosed herein.

[0511] The term "fixed dose" herein means a dosing regimen in which a fixed amount of a therapeutic substance is administered to a subject over multiple administrations, regardless of the subject's weight. A fixed dose is typically abbreviated as qnw, where n is an integer representing an interval, and w is the number of weeks. For example, a q2w fixed dose administration regimen of 1500 mg of an antibody means that a fixed amount of 1500 mg of the antibody is administered every 2 weeks. In certain aspects, the therapeutic substance is an antibody that binds to EGFR or EGFR and LGR5, which is administered at a q2w dosing regimen of 1500 mg. In certain aspects, the subject has been administered at least 3 q2w fixed doses of 1500 mg. In certain aspects, the administration is at least 4 doses or more and may be continued until the patient shows sufficient clinical or radiological progress.

[0512] The fixed dose can be premedicated, meaning that a drug is administered to the subject before administration of the disclosed antibody. In certain aspects, a fixed dose of 1500 mg of the antibody is premedicated with an antihistamine, a pain reliever, a fever reducer, and / or an anti-inflammatory drug.

[0513] An effective amount of the combination therapy is administered according to the methods described herein in an "effective regimen," meaning a combination of an antibody, or functional portion, derivative, and / or analog thereof, and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38, wherein the order of administration, the amounts administered, and the frequency of the doses are sufficient to achieve treatment.

[0514] As used herein, the terms "synergy," "therapeutic synergy," and "synergistic effect" refer to the phenomenon that the therapeutic outcome of a patient treated with a combination of therapeutic agents (e.g., an EGFR / LGR5 binding antibody, such as pesentumab, in combination with a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38) is superior to the outcome achieved when each individual component of the combination is used alone (see, e.g., TH Corbett et al., 1982, Cancer Treatment Reports, 66, 1187). In this context, a therapeutically superior outcome includes one or more of the following: (a) an increase in the therapeutic response that is greater than one or both of the individual effects of each agent in the combination at the same dose; (b) a reduction in the dose of one or more agents in the combination without diminishing the therapeutic efficacy; (c) a reduction in the incidence of adverse events, while achieving a therapeutic benefit equal to or greater than that achieved with monotherapy of each agent at the same dose in the combination; (d) a reduction in dose-limiting toxicity, while achieving a therapeutic benefit greater than that achieved with monotherapy of each agent; (e) a delay or minimization of the development of drug resistance. In an organoid model, a combination used at its maximum tolerated dose (wherein the dose of each component will generally not exceed its individual maximum tolerated dose) exhibits therapeutic synergy when the reduction in organoid growth achieved by administration of the combination is greater than the reduction in organoid growth achieved by the optimal component when administered alone.

[0515] All documents and references mentioned herein, including Genbank entries, patents and published patent applications, and websites, are expressly incorporated by reference to the same extent as if fully or partially written into this document.

[0516] For purposes of clarity and concise description, features herein will be described as part of the same or separate parts of the present invention; however, it will be understood that the scope of the present disclosure may include preferred aspects having a combination of all or some of the features.

[0517] The present disclosure will now be described with reference to the following examples, which are illustrative only and are not intended to limit the present disclosure. Although the present disclosure has been described in detail with reference to specific aspects of the present disclosure, it will be apparent to those skilled in the art that various changes and modifications may be made thereto without departing from the spirit and scope of the present disclosure.

[0518] List of Articles

[0519] 1. An antibody or a functional part, derivative and / or analogue thereof comprising a variable domain that binds to the extracellular portion of EGFR for use in treating cancer in a subject, wherein the treatment further comprises administration of a fluoropyrimidine.

[0520] 2. A method for treating cancer in a subject, the method comprising administering to the subject an effective amount of an antibody or a functional portion, derivative and / or analog thereof and a fluoropyrimidine, wherein the antibody or the functional portion, derivative and / or analog thereof comprises a variable domain that binds to the extracellular portion of EGFR.

[0521] 3. The antibody or functional part, derivative and / or analogue thereof, or method as described in clause 1 or 2, wherein the fluoropyrimidine is fluorouracil.

[0522] 4. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of the preceding clauses, wherein fluorouracil is administered at a dose of 200-3000 mg / m 2 Administer intravenously within the range.

[0523] 5. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein fluorouracil is administered at 400 mg / m on day 1. 2 Administer intravenously as a bolus, followed by 2400 mg / m² every two weeks 2 Up to 3000 mg / m 2 Administer intravenously as a continuous infusion over 46 hours.

[0524] 6. The antibody or functional part, derivative and / or analogue thereof, or the method according to any of the preceding clauses, wherein fluorouracil is administered at 500 mg / m2 on days 1, 8, 15, 22, 29 and 36 of an 8-week cycle. 2 Administer as an intravenous bolus.

[0525] 7. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the cancer has a mutation in one or more genes selected from APC, KRAS, NRAS, SMAD2, SMAD4, SOX9, PIK3CA or TP53.

[0526] 8. An antibody or a functional part, derivative and / or analogue thereof comprising a variable domain that binds to the extracellular portion of EGFR, for use in treating cancer in a subject, wherein the treatment further comprises administration of a fluoropyrimidine and a thymidine phosphorylase inhibitor.

[0527] 9. A method for treating cancer in a subject, the method comprising administering to the subject an effective amount of an antibody or a functional portion, derivative and / or analog thereof, a fluoropyrimidine and a thymidine phosphorylase inhibitor, wherein the antibody or the functional portion, derivative and / or analog thereof comprises a variable domain that binds to the extracellular portion of EGFR.

[0528] 10. The antibody or functional part, derivative and / or analogue thereof, or method according to clause 8 or 9, wherein the fluoropyrimidine is trifluorothymidine and the thymidine phosphorylase inhibitor is tipiracil.

[0529] 11. The antibody or functional part, derivative and / or analogue thereof, or the method according to any one of clauses 8 to 10, wherein trifluorothymidine and tipiracil are administered at a molar ratio of 1:0.5 twice daily at a dose of 35-80 mg / m 2 Application.

[0530] 12. The antibody or functional part, derivative and / or analogue thereof, or method of any one of clauses 8 to 11, wherein the cancer has a mutation in one or more genes selected from APC, FBXW7, KRAS, NRAS, SMAD2, SMAD3, SMAD4, SOX9, PIK3CA or TP53.

[0531] 13. An antibody or a functional part, derivative and / or analogue thereof comprising a variable domain that binds to the extracellular portion of EGFR, for use in treating cancer in a subject, wherein the treatment further comprises administering a platinum-based chemotherapeutic agent.

[0532] 14. A method of treating cancer in a subject, the method comprising administering to the subject an effective amount of an antibody, or a functional portion, derivative and / or analog thereof, comprising a variable domain that binds to the extracellular portion of EGFR and a platinum-based chemotherapeutic agent.

[0533] 15. The antibody or functional part, derivative and / or analogue thereof, or method according to clause 13 or 14, wherein the platinum-based chemotherapeutic agent is oxaliplatin.

[0534] 16. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of clauses 13 to 15, wherein oxaliplatin is administered at 65-130 mg / m 2 Apply within the range.

[0535] 17. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of clauses 13 to 16, wherein oxaliplatin is administered at 85 mg / m2 every two weeks. 2 Application.

[0536] 18. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of clauses 13 to 17, wherein the cancer has a mutation in one or more genes selected from APC, FBXW7, KRAS, NRAS, SMAD2, SMAD4, SOX9, PIK3CA or TP53.

[0537] 19. An antibody or a functional part, derivative and / or analogue thereof comprising a variable domain that binds to the extracellular portion of EGFR, for use in treating cancer in a subject, wherein the treatment further comprises administering a BCL-2 inhibitor.

[0538] 20. A method of treating cancer in a subject, the method comprising administering to the subject an effective amount of an antibody or a functional part, derivative and / or analog thereof comprising a variable domain that binds to the extracellular portion of EGFR and a BCL-2 inhibitor.

[0539] 21. The antibody or functional part, derivative and / or analogue thereof, or method according to clause 19 or 20, wherein the BCL-2 inhibitor is venetoclax.

[0540] 22. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of clauses 19 to 21, wherein venetoclax is administered at a dose of 20-1200 mg / m2 per day. 2 Application.

[0541] 23. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of clauses 19 to 22, wherein the cancer has a mutation in one or more genes selected from APC, KRAS, NRAS, SMAD2, SMAD4, SOX9, PIK3CA or TP53.

[0542] 24. An antibody or a functional part, derivative and / or analog thereof, comprising a variable domain that binds to the extracellular portion of EGFR, for use in treating cancer in a subject, wherein the treatment further comprises administering SN-38.

[0543] 25. A method of treating cancer in a subject, the method comprising administering to the subject an effective amount of an antibody, or a functional portion, derivative, and / or analog thereof, comprising a variable domain that binds to the extracellular portion of EGFR, and SN-38.

[0544] 26. The antibody or functional part, derivative and / or analogue thereof, or method according to clause 24 or 25, wherein the cancer has a mutation in one or more genes selected from APC, FBXW7, KRAS, NRAS, PIK3CA, SMAD2, SMAD3, SMAD4, SOX9, TCF7L2 and TP53.

[0545] 27. An antibody or a functional part, derivative and / or analogue thereof comprising a variable domain that binds to the extracellular portion of EGFR, for use in treating cancer in a subject, wherein the treatment further comprises administration of oxaliplatin, folinic acid and fluorouracil.

[0546] 28. A method of treating cancer in a subject, the method comprising administering to the subject an effective amount of an antibody or a functional portion, derivative and / or analog thereof comprising a variable domain that binds to the extracellular portion of EGFR and oxaliplatin, folinic acid and fluorouracil.

[0547] 29. The antibody or functional part, derivative and / or analogue thereof, or method according to clause 27 or 28, wherein oxaliplatin is administered at a dose of 50-200 mg / m 2 And folinic acid at 200-600mg / m 2 Administer intravenously together, followed by fluorouracil at 1200-3600 mg / m 2 Intravenous administration.

[0548] 30. An antibody or a functional part, derivative and / or analogue thereof comprising a variable domain that binds to the extracellular portion of EGFR, for use in treating cancer in a subject, wherein the treatment further comprises administration of irinotecan, folinic acid and fluorouracil.

[0549] 31. A method of treating cancer in a subject, the method comprising administering to the subject an effective amount of an antibody or a functional portion, derivative and / or analog thereof comprising a variable domain that binds to the extracellular portion of EGFR, and irinotecan, folinic acid and fluorouracil.

[0550] 32. The antibody or functional part, derivative and / or analogue thereof, or method according to clause 30 or 31, wherein irinotecan is administered at 180 mg / m 2 And folinic acid at 200-400mg / m 2 Administer intravenously together, followed by fluorouracil at 400-2400 mg / m 2 Intravenous administration.

[0551] 33. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of clauses 27 to 32, wherein the cancer has a mutation in one or more genes selected from APC, KRAS, NRAS, SMAD2, SMAD4 or SOX9.

[0552] 34. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the cancer is adenocarcinoma or squamous cell carcinoma.

[0553] 35. The antibody or functional part, derivative and / or analogue thereof, or method of any of the preceding clauses, wherein the cancer is head and neck cancer, gastric cancer, esophageal cancer, gastroesophageal junction cancer, non-small cell lung cancer or colorectal cancer.

[0554] 36. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the cancer is colorectal cancer.

[0555] 37. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the antibody is enhanced by ADCC.

[0556] 38. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the antibody is afucosylated.

[0557] 39. The antibody or functional part, derivative and / or analogue thereof, or method as described in any of the preceding clauses, wherein the antibody is a multispecific antibody.

[0558] 40. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the antibody is a bispecific antibody.

[0559] 41. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the variable domain that binds to the extracellular portion of EGFR is a heavy chain variable region comprising:

[0560] - a CDR3 sequence of at least the VH of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3 , or a CDR3 sequence that differs from the VH CDR3 sequence of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3 by at most three, two or one amino acid; or

[0561] - at least the CDR1, CDR2 and CDR3 sequence of the VH of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3 ; or the CDR1, CDR2 and CDR3 sequence of the VH of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3 with up to three, two or one amino acid substitutions; or

[0562] - the amino acid sequence of the VH chain of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3; or

[0563] - the amino acid sequence of the VH chain of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3, which has up to 15, or 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid insertions, deletions, substitutions or a combination thereof relative to the VH chain of MF3370; MF3755; MF4280 or MF4289.

[0564] 42. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the antibody comprises a second variable domain that does not bind to EGFR.

[0565] 43. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the antibody comprises a second variable domain that binds LGR5.

[0566] 44. An antibody or functional part, derivative and / or analogue thereof, or method as described in any of the preceding clauses, wherein the variable domain that binds to LGR5 binds to a region located at Figure 1 Epitope within amino acid residues 21-118 of the human LGR5 sequence shown.

[0567] 45. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the variable domain that binds to LGR5 is a heavy chain variable region comprising:

[0568] - a CDR3 sequence of at least the VH of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3 , or a CDR3 sequence that differs from a CDR3 sequence of the VH of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3 by at most three, two, or one amino acid; or

[0569] - at least the CDR1, CDR2 and CDR3 sequence of the VH of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3 ; or the CDR1, CDR2 and CDR3 sequence of the VH of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3 with up to three, two or one amino acid substitutions; or

[0570] - the sequence of the VH chain of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3; or

[0571] - the amino acid sequence of the VH chain of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3, which has up to 15, or 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid insertions, deletions, substitutions or a combination thereof relative to the VH chain of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818.

[0572] 46. An antibody or functional part, derivative and / or analogue thereof, or use or method as described in any of the preceding clauses, wherein the variable domain that binds to the extracellular part of EGFR comprises a heavy chain variable region comprising the CDR1, CDR2 and CDR3 sequences of the variable region selected from the group consisting of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3, and wherein the variable domain that binds to the extracellular part of LGR5 comprises a heavy chain variable region comprising the CDR1, CDR2 and CDR3 sequences of the variable region selected from the group consisting of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3.

[0573] 47. An antibody or functional part, derivative and / or analogue thereof, or use or method as described in any of the preceding clauses, wherein the variable domain that binds to the extracellular portion of EGFR comprises a heavy chain variable region comprising the CDR1, CDR2 and CDR3 sequences of the variable region of MF3755 as shown in Figure 3, and wherein the variable domain that binds to the extracellular portion of LGR5 comprises a heavy chain variable region comprising the CDR1, CDR2 and CDR3 sequences of the variable region of MF5816 as shown in Figure 3.

[0574] 48. An antibody or functional part, derivative and / or analogue thereof, or use or method according to any of the preceding clauses, wherein the VH chain of the variable domain that binds to EGFR comprises the amino acid sequence of the VH chain of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3 ; or the amino acid sequence of the VH chain of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3 , which has at most 15, preferably no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, and preferably no more than 5, 4, 3, 2 or 1 amino acid modifications relative to said VH, including insertions, deletions, substitutions or combinations thereof; and wherein the VH chain of the variable domain that binds to LGR5 comprises MF5790; M as shown in Figure 3 . or the amino acid sequence of the VH chain of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818; or the amino acid sequence of the VH chain of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3, which has up to 15, preferably no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, and preferably no more than 5, 4, 3, 2 or 1 amino acid modifications relative to the VH, including insertions, deletions, substitutions or a combination thereof, preferably wherein the amino acid insertions, deletions and substitutions are not present in the CDR1, CDR2 and CDR3 light chain variable regions.

[0575] 49. The antibody or functional part, derivative and / or analogue thereof, or use or method according to any of the preceding clauses, wherein the VH chain of the variable domain that binds to EGFR comprises the amino acid sequence of the VH chain of MF3755 as shown in Figure 3; or the amino acid sequence of the VH chain of MF3755 as shown in Figure 3, which has at most 15, preferably no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, and preferably no more than 5, 4, 3, 2 or 1 amino acid modifications relative to said VH, including insertions, deletions, substitutions or combinations thereof. ; and wherein the VH chain of the variable domain that binds to LGR5 comprises the amino acid sequence of the VH chain of MF5816 as shown in Figure 3; or the amino acid sequence of the VH chain of MF5816 as shown in Figure 3, which has at most 15, preferably no more than 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, and preferably no more than 5, 4, 3, 2 or 1 amino acid modifications relative to said VH, including insertions, deletions, substitutions or a combination thereof, preferably wherein the amino acid insertions, deletions and substitutions are not present in the CDR1, CDR2 and CDR3 light chain variable regions.

[0576] 50. An antibody or functional part, derivative and / or analogue thereof, or use or method as described in any of the preceding clauses, wherein the VH chain of the variable domain that binds to EGFR comprises the amino acid sequence of the VH chain of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3; and wherein the VH chain of the variable domain that binds to LGR5 comprises the amino acid sequence of the VH chain of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3.

[0577] 51. The antibody or functional part, derivative and / or analogue thereof, or use or method according to any of the preceding clauses, wherein the VH chain of the variable domain that binds to EGFR comprises the amino acid sequence of the VH chain of MF3755 as shown in Figure 3; and wherein the VH chain of the variable domain that binds to LGR5 comprises the amino acid sequence of the VH chain of MF5816 as shown in Figure 3.

[0578] 52. The antibody or functional part, derivative and / or analogue thereof, or use or method according to any of the preceding clauses, wherein both the variable domains that bind to EGFR and the variable domains that bind to LGR5 comprise Figure 4 b shows the CDR1, CDR2 and CDR3 regions of the light chain variable region.

[0579] 53. An antibody or functional part, derivative and / or analogue thereof, or use or method as described in any of the preceding clauses, wherein both the variable domains that bind to EGFR and the variable domains that bind to LGR5 comprise Figure 4 b) The light chain variable region comprises 0 to 10 amino acid insertions, deletions, substitutions, additions or a combination thereof, wherein the amino acid insertions, deletions and substitutions are not present in the CDR1, CDR2 and CDR3 light chain variable regions.

[0580] 54. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the antibody is pesentumomab.

[0581] 55. An antibody or functional part, derivative and / or analog thereof, or method as described in any of the preceding clauses, wherein the antibody is a monovalent antibody without a second variable domain, or wherein the antibody comprises the EGFR binding variable domain as the only variable domain.

[0582] 56. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the subject is a mammal, particularly a human.

[0583] 57. The antibody or functional part, derivative and / or analog thereof, or method as described in any of the preceding clauses, wherein the treatment comprises providing the subject with 1500 mg of the antibody or functional part, derivative and / or analog thereof.

[0584] 58. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the subject to be treated has not received previous anti-cancer therapy.

[0585] 59. The antibody or functional part, derivative and / or analogue thereof, or the method according to any one of clauses 1 to 57, wherein the subject to be treated has progressed following a previous anti-cancer therapy.

[0586] 60. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the previous anti-cancer therapy is chemotherapy, targeted anti-cancer therapy, immunotherapy or radiotherapy.

[0587] 61. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the anti-cancer therapy is fluorouracil, a platinum-based chemotherapy agent, oxaliplatin, FOLFOX, FOLFIRI, TAS-102, trastuzumab, pembrolizumab, nivolumab, cetuximab or a combination thereof.

[0588] 62. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the subject has progressed following fluoropyrimidine-oxaliplatin-based chemotherapy with or without bevacizumab.

[0589] 63. The antibody or functional part, derivative and / or analogue thereof, or the method according to clause 62, wherein the subject is administered FOLFIRI.

[0590] 64. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the subject has progressed following fluoropyrimidine-irinotecan-based chemotherapy with or without bevacizumab.

[0591] 65. The antibody or functional part, derivative and / or analogue thereof, or method according to clause 64, wherein the subject is administered FOLFOX.

[0592] 66. The antibody or functional part, derivative and / or analogue thereof, or method according to any of the preceding clauses, wherein the cancer is metastatic colorectal cancer (mCRC), such as colon or rectal adenocarcinoma.

[0593] 67. The antibody or functional part, derivative and / or analog thereof, or the method according to any of the preceding clauses, wherein the antibody or functional part, derivative and / or analog thereof and the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor or SN-38 are administered to the subject simultaneously, separately or sequentially.

[0594] 68. The method of treating cancer in a subject as described in any of the preceding clauses, wherein the method comprises administering to the subject an effective amount of an antibody or a functional portion, derivative and / or analog thereof and an effective amount of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

[0595] 69. A pharmaceutical composition comprising an antibody as defined in any one of the preceding clauses, or a functional part, derivative and / or analog thereof; and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

[0596] 70. The pharmaceutical composition of clause 69, wherein the antibody or functional part, derivative and / or analog thereof is provided in a single formulation comprising a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38.

[0597] 71. The pharmaceutical composition of clause 69, wherein the antibody or functional part, derivative and / or analog thereof and the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor or SN-38 are provided in separate formulations.

[0598] 72. A kit of parts comprising:

[0599] - an antibody as defined in any of the preceding clauses, or a functional part, derivative and / or analogue thereof;

[0600] - A fluoropyrimidine, platinum-based chemotherapy agent, BCL-2 inhibitor, or SN-38; and

[0601] - Instructions for use of the antibody, or functional portion, derivative, and / or analog thereof, and instructions for use of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38.

[0602] 73. The kit of parts according to clause 72, wherein the instructions for use of the antibody or its functional part, derivative and / or analogue comprise instructions to administer it at 1500 mg.

[0603] 74. The kit of parts of clause 72 or 73, wherein the kit comprises the antibody or functional portion, derivative, and / or analog thereof and instructions for use of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 in the treatment of adenocarcinoma or squamous cell carcinoma.

[0604] 75. The kit of parts of any one of clauses 72 to 74, wherein the kit comprises the antibody or functional part, derivative, and / or analog thereof and instructions for use of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 in the treatment of head and neck cancer, gastric cancer, esophageal cancer, gastroesophageal junction cancer, non-small cell lung cancer, or colorectal cancer.

[0605] 76. The kit of parts of any one of clauses 72 to 75, wherein the kit comprises the antibody or functional part, derivative and / or analog thereof and instructions for use of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 in the treatment of colorectal cancer.

[0606] 77. An antibody as defined in any one of the preceding clauses, or a functional part, derivative and / or analog thereof, in combination with a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38, for use in treating cancer in a subject in need thereof.

[0607] 78. The following combination:

[0608] - an antibody as defined in any of the preceding clauses, or a functional part, derivative and / or analogue thereof,

[0609] - instructions for use of the antibody in the treatment of cancer in a subject, and

[0610] - Instructions for use of a fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 as defined in any of the preceding clauses for the treatment of cancer in a subject.

[0611] 79. The combination of clause 78, wherein the instructions for use comprise the amount of the antibody, or functional part, derivative, and / or analog thereof, to be used and / or the amount of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 to be used, and / or the dosing interval and / or the cancer to be treated.

[0612] 80. The following combination:

[0613] - a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 as defined in any of the preceding clauses,

[0614] - Instructions for use of a fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 in the subject's cancer treatment,

[0615] - Instructions for use of an antibody as defined in any of the preceding clauses, or a functional part, derivative and / or analogue thereof, for the treatment of cancer in a subject.

[0616] 81. The combination of clause 80, wherein the instructions for use comprise the amount of the antibody, or functional part, derivative, and / or analog thereof, to be used and / or the amount of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor, or SN-38 to be used, and / or the dosing interval and / or the cancer to be treated.

[0617] 82. Use of an antibody as defined in any of the preceding clauses, or a functional part, derivative, and / or analog thereof, and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 as defined in any of the preceding clauses, in the manufacture of one or more medicaments for treating cancer in a subject.

[0618] 83. The use of clause 82, wherein the treatment comprises administering to the subject an effective amount of an antibody or a functional portion, derivative and / or analog thereof and an effective amount of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

[0619] 84. The use of clause 82 or 83, wherein the treatment comprises administering to the subject the antibody or its functional part, derivative and / or analog and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38 simultaneously, separately or sequentially.

[0620] Examples

[0621] As used herein, "MFXXXX," wherein X is independently a number 0-9, refers to a Fab comprising a variable domain wherein VH has an amino acid sequence identified by the four digits shown in Figure 3. Unless otherwise indicated, the light chain variable region of the variable domain typically has Figure 4 b. The light chain in the example has Figure 4 The sequence shown in a. "MFXXXX VH" refers to the amino acid sequence of the VH identified by a 4-digit number. The MF also contains a light chain constant region and a heavy chain constant region that typically interacts with the light chain constant region. The heavy chain VH / variable regions are different and typically the CH3 regions are also different, with one heavy chain having a KK mutation in the CH3 domain and the other heavy chain having a complementary DE mutation in the CH3 domain (see reference PCT / NL2013 / 050294 (published as WO2013 / 157954) and Figure 5 d and Figure 5 e). The bispecific antibody in the example has Figure 5 The Fc tail with KK / DE CH3 heterodimerization domain, CH2 domain and CH1 domain is shown. Figure 4 The common light chain shown in a and the VH designated by MF numbering. For example, the bispecific antibody represented by MF3755 x MF5816 has the above general sequence and a VH variable domain having the sequence of MF3755 and a VH variable domain having the sequence of MF5816.

[0622] The amino acid sequence of each heavy chain variable region (VH) is shown in Figure 3. The bispecific antibody EGFR / LGR5, MF3755xMF5816, comprising the heavy chain variable regions MF3755 and MF5816 and a common light chain and including the ADCC-enhancing modification from afucosylation, as well as other LGR5 and EGFR combinations as shown in Figure 3, has been shown to be effective in WO2017 / 069628.

[0623] Example 1. Generation of bispecific antibodies

[0624] Bispecific antibodies are produced by transient co-transfection of two plasmids encoding IgG with different VH domains, using proprietary CH3 engineering technology to ensure effective heterodimerization and the formation of bispecific antibodies. The common light chain is also co-transfected on the same plasmid or another plasmid in the same cell. In our applications (e.g., WO2013 / 157954 and WO2013 / 157953; incorporated herein by reference), we have disclosed methods and means for making bispecific antibodies from single cells, thereby providing a means for forming bispecific antibodies rather than monospecific antibodies. These methods may also be advantageously used in the present disclosure. In particular, preferred mutations that substantially only generate bispecific full-length IgG molecules are amino acid substitutions at positions 351 and 366, e.g., L351K and T366K (numbering according to the EU numbering system) in the first CH3 domain (referred to as the "KK-variant" heavy chain), and amino acid substitutions at positions 351 and 368, e.g., L351D and L368E, in the second CH3 domain (referred to as the "DE-variant" heavy chain), or vice versa (see Figure 5 d and Figure 5 e). Previously, in the aforementioned applications, it was demonstrated that negatively charged DE-variant heavy chains and positively charged KK-variant heavy chains preferentially pair to form heterodimers (referred to as "DEKK" bispecific molecules). Homodimerization of DE-variant heavy chains (DE-DE homodimers) or KK-variant heavy chains (KK-KK homodimers) is difficult to occur due to the strong repulsion between charged residues in the CH3-CH3 interface between the same heavy chains.

[0625] The VH gene of the variable domain that binds to LGR5 was cloned into a vector encoding a positively charged CH3 domain. The VH gene of the variable domain that binds to EGFR, such as those disclosed in WO 2015 / 130172 (incorporated herein by reference), was cloned into a vector encoding a negatively charged CH3 domain. 293F Freestyle cells adapted for suspension growth were cultured in a T125 flask on a shaker platform until the density reached 3.0 x 10e 6 cells / ml. Cells were cultured at 0.3-0.5 x 10e 6 Cells were seeded at a density of 10 viable cells / ml per well of a 24-deep-well plate. These cells were transiently transfected with a mixture of two plasmids encoding different antibodies cloned into a proprietary vector system. Seven days after transfection, cell supernatants were collected and filtered through a 0.22 μM filter (Sartorius). Sterile supernatants were stored at 4°C until antibody purification.

[0626] IgG purification and quantification

[0627] Under sterile conditions, Protein-A affinity chromatography was used to purify the protein in a filter disc. First, the pH of the culture medium was adjusted to pH 8.0, followed by incubation of the supernatant containing IgG with Protein A Sepharose CL-4B microbeads (50% v / v) (Pierce) on a shaking platform at 25°C, 600rpm for 2 hours. Next, the microbeads were harvested by filtration. The microbeads were washed twice with PBS pH 7.4. The bound IgG was then eluted with 0.1M citrate buffer at pH 3.0, and the eluate was immediately neutralized with Tris pH 8.0. Buffer exchange was performed by centrifugation using a multi-sieve Ultracel 10 multi-sieve disc (Millipore). Samples were finally harvested in PBS at pH 7.4. IgG concentration was measured using Octet. Protein samples were stored at 4°C.

[0628] To determine the amount of purified IgG, antibody concentration was determined by Octet analysis using a protein A biosensor (Forte-Bio, according to the supplier's recommendations) using total human IgG (Sigma Aldrich, catalog number 14506) as a standard.

[0629] The following bispecific antibodies are suitable for use in this example and in the methods of the present disclosure: MF3370xMF5790, MF3370xMF5803, MF3370xMF5805, MF3370xMF5808, MF3370xMF5809, MF3370xMF5814, MF3370xMF5816, MF3370xMF5817, MF3370xMF5818, MF3755xMF5790, MF3755xMF5803, MF3755xMF5805, MF3755xMF5808, MF3755xMF5809, MF3755xMF5814, MF3755xMF5816, MF3755xMF5817, F3755xMF5818, MF4280xMF5790, MF4280xMF5803, MF4280xMF5805, MF4280xMF5808, MF4280xMF5809, MF4280xMF5814, MF4280xMF5816, MF4280xMF5817, MF4280x MF5818, MF4289xMF5790, MF4289xMF5803, MF4289xMF5805, MF4289xMF5808, MF4289xMF5809, MF4289xMF5814, MF4289xMF5816, MF4289xMF5817 and MF4289xMF5818. Each bispecific antibody comprises two VHs designated by MF numbering capable of binding to EGFR and LGR5, respectively, and further comprises VHs having the sequences of SEQ ID NO: 117 ( Figure 5 d) and SEQ ID NO: 118 ( Figure 5 e) the Fc tail of the KK / DE CH3 heterodimerization domain, as shown in SEQ ID NO: 116 ( Figure 5 c) CH2 domain as shown in SEQ ID NO: 115 ( Figure 5 b) and the hinge domain shown in SEQ ID NO: 114 ( Figure 5 a) CH1 domain, and SEQ ID NO: 107 ( Figure 4 a) Common light chain shown.

[0630] Example 2: Materials and Methods

[0631] Organoid Models:

[0632] Tumor organoid models were obtained from HUB organoids (see huborganoids.nl) and described in van de Wetering, M. et al., Prospective derivation of a living organoid biobank of colorectal cancer patients, Cell 161, 933–945 (2015). Table 4 summarizes the mutations of components of the four major CRC driving pathways (WNT, RTK / RAS, TP53, and transforming growth factor β) in each model. Further details of the clinicopathological characteristics can be found in Herpers B. et al., Nat Cancer. 2022 Apr; 3(4): 418-436.

[0633] Table 4: Organoid Models

[0634]

[0635]

[0636] Culture medium:Organoids were expanded and exposed to droplets of Matrigel matrix (Corning, catalog number 354230) in organoid culture medium (OCM) and seeded into 384-well plates (Greiner, catalog number 781091) using a liquid handler. Briefly, OCM contained RSPO3 (BioTechne, 3500-RS / CF), Noggin (Peprotech, 120-10C), B27 (Thermo Fisher Scientific, Catalog No. 17504001), Nicotinamide (Sigma Aldrich, Catalog No. N0636), N-acetylcysteine (Sigma Aldrich, Catalog No. A9165), SB-202190 (Sigma Aldrich, Catalog No. Aldrich, catalog number S7067), gastrin (Merck, catalog number G9145), and A83-01 (Tocris, catalog number 2939). OCM was supplemented with WNT3A-conditioned medium (OL-016_v2) for the healthy colon organoid model. During passaging, 10 μM Y27632 (RhoKi) (Bioconnect, catalog number M1817) was used.

[0637] Colon organoids were seeded in hydrogels in 384-well plates as clusters of 300 or 600 (model P18T) 10-20 cell stage cells. After 30 minutes of gelation, culture medium was added. For colon tumors, organoid culture medium was used. For normal colon organoids, organoid culture medium supplemented with WNT3A was used. Growth factors were added to the culture medium at specified concentrations on the day of inoculation. Bispecific antibodies were added to the culture medium containing growth factors on the day of inoculation. Compound exposure was performed on the day of inoculation or 24 hours after inoculation. Organoids were exposed to growth factors, antibodies and compounds for 5-8 days (depending on the growth rate of the organoid model - normal organoids exhaust the culture medium faster than colon tumors). At the end of the experiment, the organoids were fixed to stain the actin cytoskeleton and nuclei to provide a reference for the development of new compounds. In preparation for image-based analysis, 25–40 sections per z-stack with a 50 μm step size were captured per well using a 4x objective (1080 x 1080 pixels). Data from each well were pooled, and the mean and standard deviation for each morphological feature (>500 features) were recorded and processed.

[0638] Growth factors: 10 ng / ml of EGF (Peprotech, catalog number AF-100-15) was used during expansion of tumor-like models, while 50 ng / ml of EGF was used in healthy organoid models.

[0639] Test compound:

[0640] Pesentuzumab: 22.9 mg / ml (Lot No. FB01701; stored at 4°C); PG2708p218: 2.5 mg / ml (stored at 4°C); 5-FU (F-2449): 10 mM (DMSO) (Lot No. G1264; stored at -20°C); Oxaliplatin (15604985): 12.59 mM (PBS) (Lot No. 211E0081; stored at 4°C); SN-38 (S4908): 10 mM (DMSO) ( Batch number: 86639-52-3-02; stored at -20°C); TAS-102 (S8539): 10 mM (DMSO) (Batch number: S853901; stored at -20°C); Venetoclax (S8048): 10 mM (DMSO) (Batch number: S804806; stored at -20°C); Staurosporine (HY-15141): 10 mM (DMSO) (Batch number: CS2716; stored at -20°C).

[0641] Image analysis

[0642] After incubation, organoids were fixed and stained to visualize the nucleus and actin cytoskeleton, respectively. After washing with PBS, 25–40 sections per well were imaged at 4x magnification using a Molecular Devices ImageXpress Micro XLS system with a 50 μm z-step size. Image processing was performed to quantify the shape, size, location, number, and intensity of individual nuclei, organoids, lumen, and epithelium within the organoids.

[0643] QC was performed on the segmented images and data by guiding the data from the positive and negative control groups to calculate the random Z' factor and one-way analysis of variance (ANOVA) as a measure of data consistency. The summary data were normalized (z score and percentage of control (POC), with DMSO+PBS+PG2708 as the negative control reference) and analyzed in TIBCO The data were formatted in GraphPad Prism v8.4 for graphical visualization and curve fitting was performed.

[0644] Example 3: Dose ranging testing of individual compounds

[0645] In preparation for the combination study in Experiment 4, a dose range of pesentuzumab, 5-FU, oxaliplatin, SN-38, TAS-102, and venetoclax was tested in the presence of 5 ng / ml EGF to estimate the IC20. The models tested were P18T, C55T, C82T, and C82N. PBS, DMSO, and the PG2708 negative control antibody were used as negative controls. 1 μM staurosporine and 0 ng / ml EGF were used as positive controls. Compound exposure was performed on the day of inoculation for organoid models C82N, C82T, C55T, and P18T for 8 days. Table 5 indicates the different concentrations tested for the compounds.

[0646] Table 5: Concentrations of compounds tested

[0647]

[0648] The control performed as expected. As compound doses increased, all compounds reduced proliferation, increased cell death, and reduced tube formation in organoids. Based on the collected results, IC20 values were determined for all compounds in the preparation of Example 4. IC20 values are summarized in Table 6.

[0649] Table 6: Summary of IC20 and suggested dose ranges for individual compounds

[0650]

[0651] Example 4: Combination study of pesentumomab and individual compounds

[0652] In models P18T, C20T, C55T, C82T, C92T, and C92N, we tested whether pesentuzumab treatment sensitized organoids to treatment with 5-FU, oxaliplatin, SN-38, TAS-102, or venetoclax in a 6x6 combination matrix in the presence of 5 ng / ml EGF.

[0653] The following combinations were added on the day of inoculation: pesentumomab (0–0.031–0.1–0.316–1–3.162 μg / ml); containing 5-FU, TAS-102, or venetoclax (0–0.1–0.316–1–3.162–10 μM); or SN38 (0–0.1–0.316–1–3.162–10 nM); or oxaliplatin (0–0.316–1–3.162–10–31.62 μM).

[0654] The antibody-drug combination was cultured for 7 days before fixation to visualize 3D actin and nuclear organization in organoids. PBS, DMSO, and PG2708 negative control antibody were used as negative controls. 1 μM staurosporine and 0 ng / ml EGF were used as positive controls.

[0655] For each model and combination, three features were evaluated: nucleus count, total progeny count (sum childcount) (total number of viable nuclei), and total organoid size. Total organoid size is known to decrease due to growth inhibition and induction of cell death.

[0656] The control performed as expected and was consistent across the experiments. All compound combinations reduced organoid growth and increased cell death. Total organoid size was further used to assess synergy between different combinations.

[0657] Data Analysis:

[0658] The data were analyzed using the online tool SynergyFinder.org (Zheng, S. et al., bioRxiv2021.06.01.446564 (2021); Zheng S, et al., SynergyFinder Plus: Toward Better Interpretation and Annotation of Drug Combination Screening Datasets. Genomics Proteomics Bioinformatics. 2022 Jan 25: S1672-0229 (22) 00008-0). The SynergyFinder tool uses four methods, HSA, LOEWE, BLISS, and ZIP, to indicate the degree of interaction. After SynergyFinder analysis, the CI of each data point was calculated using the Chou-Talalay method (see Chou TCDrug combination using the Chou-Talalay method. Cancer research 70, 440-446 (2010)). The CI for each data point was calculated using the maximum value of the measured single agent effect or the maximum value predicted by the model based on the single agent effect and divided by the actual measured effect using all four methods (ZIP, BLISS, LOEWE and HSA). Table 7 summarizes the resulting synergy scores and CI scores for different models.

[0659] Table 7: Synergy scores and combination index (CI) scores of the combination therapy of pesentuzumab with 5-FU, oxaliplatin, SN-38, TAS-102 or venetoclax in different models

[0660] Model Drug 1 Drug 2 ZIP HAS Loewe Bliss CI score C82T Pesentumomab 5-FU 4.90 3.35 2.97 4.84 0.572 C55T Pesentumomab 5-FU 4.83 8.68 8.78 2.85 0.672 C55T Pesentumomab Oxaliplatin 14.44 16.22 15.55 14.09 0.410 C82T Pesentumomab Oxaliplatin 7.46 6.38 5.63 7.77 0.481 C92N Pesentumomab Oxaliplatin 0.34 0.84 0.83 0.38 0.675 C20T Pesentumomab Oxaliplatin 1.48 1.41 1.13 0.48 0.725 P18T Pesentumomab Oxaliplatin 2.69 4.81 2.42 2.41 0.809 C55T Pesentumomab SN38 14.55 14.03 12.65 15.09 0.365 C92N Pesentumomab SN38 1.45 1.94 1.22 1.80 0.461 C82T Pesentumomab SN38 5.19 4.31 3.82 5.01 0.506 C20T Pesentumomab SN38 0.79 1.05 0.90 1.07 0.530 C92T Pesentumomab SN38 4.48 7.02 6.63 4.17 0.594 C82T Pesentumomab TAS-102 6.09 6.00 5.92 6.83 0.303 C55T Pesentumomab TAS-102 6.73 9.38 8.43 6.68 0.515 C82T Pesentumomab Venetoclax 6.80 7.13 6.79 6.85 0.198 C55T Pesentumomab Venetoclax 1.79 6.08 5.13 1.49 0.766 C20T Pesentumomab Venetoclax -0.01 0.93 0.43 0.01 0.854

[0661] In conclusion, combination treatment with pesentuzumab and each of the five tested standard-of-care drugs resulted in synergistic growth inhibition in the indicated models.

[0662] Example 5: Combination study of pesentumomab and Folfox / Folfiri

[0663] We also tested whether the combination of pesentuzumab with FOLFOX and FOLFIRI could improve tumor regression. The individual components of the FOLFOX and FOLFIRI combination are described elsewhere herein. 2Intravenous injection for 2 hours. This corresponds to a plasma Cmax of 1.61-2.2 μg / ml or 4-5 μM. Next, folinic acid was combined with irinotecan at 180 mg / m 2 Intravenous injection over 90-120 minutes. 150 mg / m 2 Irinotecan resulted in a plasma Cmax of approximately 2 μg / ml irinotecan, which provided an average of 36 ng / ml of its metabolite SN-38 (92 nM). 2 A bolus of 500 mg / ml was provided, resulting in a mean Cmax of 55 μg / ml, followed by 2400 mg / m 2 Provided as an intravenous infusion over 46 hours. 2 Intravenous infusion over 3 days resulted in a mean plasma concentration of 4.6 uM (peak 7.3 uM). Folinic acid was used at a concentration where it is soluble (50 μg / ml). This resulted in the following combination:

[0664] -5 μM 5-FU + 5 μM oxaliplatin + 50 μg / ml folinic acid to simulate FOLFOX (5 μM) or

[0665] - 5 μM 5-FU + 100 nM SN-38 + 50 μg / ml folinic acid to mimic FOLFIRI (5 μM).

[0666] The combination was tested in the C55T model in the presence of 5 ng / ml EGF.

[0667] The synergy of the combination of FOLFIRI and FOLFOX was calculated using total organoid size and the methods described in Example 4. The synergy scores and combination index (CI) are summarized in Table 8. The synergy scores and CI indicate that both FOLFIRI and FOLFOX have a synergistic effect in combination with pesentumab in the indicated models.

[0668] Table 8: Synergy scores and combined sensitivity scores for the combination treatment of pesentuzumab with FOLFOX or FOLFIRI for model C55T calculated in SynergyFinder.

[0669] Model Drug 1 Drug 2 ZIP HAS Loewe Bliss CI score 1 C55T Pesentumomab FOLFIRI 3.69 7.68 5.62 4.05 0.49 2 C55T Pesentumomab FOLFOX 3.37 8.94 7.81 3.65 0.62

[0670] Example 6: Clinical research plan

[0671] Study Design

[0672] This is a first-in-human, Phase 1 / 2, open-label, multicenter study combining an initial dose-escalation portion with a dose-expansion portion in mCRC. The initial dose-escalation portion is complete, and the preliminary RP2D is set at 1500 mg Q2W.

[0673] The combination of pesentuzumab and selected chemotherapeutic agents is being evaluated in the dose expansion portion of the study, which is intended to treat mCRC.Pesentuzumab will be administered intravenously as a fixed dose over a 2- to 6-hour infusion period in 4-week cycles (28 days) Q2W.

[0674] Patients will be assigned to dose-expansion treatment arms, which include a single-agent pesentuzumab arm and one arm using pesentuzumab in combination with a chemotherapy agent. Specifically, pesentuzumab is being tested in combination with FOLFOX and, in another study arm, in combination with FOLFIRI.

[0675] Safety, PK, immunogenicity, and antitumor activity will be characterized in all cohorts of patients, and retrospective biomarker analyses including EGFR and LGR5 status will be performed.

[0676] Research group

[0677] Inclusion criteria

[0678] 1. Sign the Informed Consent Form (ICF) before starting any study procedures.

[0679] 2. Age ≥18 at the time of ICF.

[0680] 3. Histologically or cytologically confirmed solid tumors with evidence of metastatic or locally advanced disease that is not amenable to standard treatment with curative intent:

[0681] · Expanding the Queue : Patients with locally advanced unresectable or metastatic disease for the following indications:

[0682] Patients with mCRC who are receiving second-line therapy and have a prior histological or cytological diagnosis of unresectable or metastatic colon or rectal adenocarcinoma. Patients must be RAS / RAF wild-type, as determined by NGS on tumor tissue (primary or metastatic). NGS data reports are provided to the sponsor for confirmation prior to study enrollment. Patients must be naive to prior anti-EGFR therapy. Radiographically confirmed disease progression during or within 6 months of prior 1L chemotherapy is required.

[0683] o Group to be treated with pesentuzumab and FOLFIRI: Patients have received only one prior chemotherapy regimen for the metastatic setting, which included 1L fluoropyrimidine-oxaliplatin-based chemotherapy with or without bevacizumab.

[0684] o Group to be treated with pesentuzumab and FOLFOX: Patients should have received only one prior chemotherapy regimen for the metastatic setting, which included 1L fluoropyrimidine-irinotecan-based chemotherapy with or without bevacizumab.

[0685] 4. A new tumor sample at baseline (e.g., formalin-fixed, paraffin-embedded block [FFPE]) from a metastatic or primary site. If the subject has a tumor sample available as an FFPE block with sufficient material (at least 20 slides with >20% tumor content) and has not received further anticancer treatment since the sample was collected, a new tumor biopsy is not required at baseline. Archival FFPE slides will not be accepted.

[0686] 5. Measurable disease as defined by RECIST version 1.1 radiologically.

[0687] 6.ECOGPS is 0 or 1.

[0688] 7. According to the investigator's assessment, life expectancy is ≥12 weeks.

[0689] 8. Echocardiogram (ECHO) or multiple gated acquisition (MUGA) scan showing left ventricular ejection fraction (LVEF) ≥ 50%.

[0690] 9. Adequate organ function:

[0691] Absolute neutrophil count (ANC) ≥1.5 × 10 9 / L

[0692] Hemoglobin ≥9 g / dL

[0693] ·Platelets ≥100×10 9 / L

[0694] Serum magnesium, sodium, corrected total calcium, phosphate, and potassium are within normal ranges (or corrected with supplements or otherwise managed appropriately). If these electrolyte ranges remain outside normal ranges despite corrective therapy, the sponsor should be consulted to confirm eligibility.

[0695] Alanine aminotransferase (ALT), aspartate aminotransferase (AST) ≤ 2.5 x upper limit of normal (ULN) and total bilirubin ≤ 1.5 x ULN (in the case of total bilirubin > 3.0 x ULN or direct bilirubin > 1.5 x ULN, unless due to known Gilbert's syndrome, which is excluded); in the case of liver involvement, ALT / AST ≤ 5 x ULN and total bilirubin ≤ 2 x ULN are allowed, unless due to known Gilbert's syndrome, when total bilirubin ≤ 3.0 x ULN or direct bilirubin ≤ 1.5 x ULN is allowed.

[0696] For patients >65 years of age, serum creatinine ≤1.5 x ULN, or creatinine clearance (CrCl) ≥60 mL / min, calculated using the Cockroft and Gault formula or the Modification of Diet in Renal Disease (MDRD) formula.

[0697] International normalized ratio (INR) or prothrombin time (PT) level ≤ 1.5 x ULN, unless the patient is receiving anticoagulant therapy and is within the therapeutic range of the intended anticoagulant.

[0698] Activated partial thromboplastin time (APTT) or partial thromboplastin time (PTT) ≤ 1.5 x ULN, unless the patient is receiving anticoagulant therapy and is within the therapeutic range of the intended anticoagulant.

[0699] Exclusion criteria

[0700] 1. Have central nervous system metastases that are untreated and symptomatic, or require radiation therapy, surgery, or continuous steroid therapy to control symptoms within 14 days of the start of the study.

[0701] 2. Known leptomeningeal involvement.

[0702] 3. Participated in another clinical trial or used any investigational drug for treatment within 4 weeks before the start of the study.

[0703] 4. Systemic anticancer therapy within 4 weeks or 5 half-lives (whichever is longer) after the first dose of study treatment. For cytotoxic agents with major delayed toxicity (e.g., mitomycin C, nitrosoureas), or anticancer immunotherapy, a six-week washout period is required.

[0704] 5. Requires immunosuppressive drug treatment (e.g., methotrexate, cyclophosphamide).

[0705] 6. Major surgery or radiation therapy within 3 weeks of the first dose of study treatment. Patients who have received prior radiation therapy that affected ≥25% of the bone marrow at any time are ineligible.

[0706] 7. Persistent clinically significant toxicity related to previous anti-tumor therapy > Grade 1 (except alopecia); stable sensory neuropathy grade ≤ Grade 2 of the National Cancer Institute-Common Terminology Criteria for Adverse Events (NCI-CTCAE) v4.03 / v5.0 or the current criteria at the time of administration is allowed.

[0707] 8. History of hypersensitivity reaction to petosemtamab, human protein, or any excipient of any non-IMP treatment required for this study.

[0708] 9. Uncontrolled hypertension (systolic BP>150 mmHg and / or diastolic BP>100 mmHg) or unstable angina after appropriate treatment; history of congestive heart failure of New York Heart Association (NYHA) class II-IV, or severe arrhythmia requiring treatment (except atrial fibrillation and paroxysmal supraventricular tachycardia); or history of myocardial infarction within six months of the start of the study.

[0709] 10. A history of prior malignancy, excluding excised cervical intraepithelial neoplasia or non-melanoma skin cancer, or a cancer treated with curative intent that is considered to have a low risk of recurrence with no evidence of disease within three years or more, or a secondary primary malignancy.

[0710] 11. Patients currently experiencing dyspnea at rest for any reason, or other conditions requiring continuous oxygen therapy, including patients with a history of interstitial lung disease (ILD) (e.g., pneumonia or pulmonary fibrosis), or evidence of ILD on baseline chest computed tomography (CT) scan.

[0711] 12. Current serious illness or medical condition, including but not limited to: uncontrolled active infection, clinically significant pulmonary, metabolic, or psychiatric disease.

[0712] 13. Patients suffering from known infectious diseases.

[0713] 14. Pregnant subjects or patients who are breastfeeding; patients of childbearing potential must use highly effective contraceptive methods before the start of the study, during the study, and within six months after the last dose of petosemtamab.

[0714] 15. Patients with mCRC that have RAS / RAF mutations identified by local ctDNA or tumor testing at screening or identified in disease history are not eligible for this study.

[0715] 16. Patients with active inflammatory bowel disease or other bowel diseases causing chronic diarrhea (defined as NCI-CTCAE grade ≥2) are not eligible for this study.

[0716] 17. Patients with peripheral sensory neuropathy and functional impairment (defined as NCI CTCAE grade ≥3) are not eligible for this study.

[0717] Pre-screening of patients

[0718] Pre-screening can be performed to exclude somatic mutations in RAS and RAF family genes (ie, KRAS, NRAS, HRAS, BRAF, ARAF, RAF1 wild type).

[0719] Prescreening of patients with mCRC to be treated in the second-line setting in combination with FOLFIRI or FOLFOX: A validated tumor NGS assay report confirming the absence of RAS / RAF or other relevant mutations in the patient's tumor can be used to determine eligibility. This report can be from any time during the natural history of the disease. Knowledge of the patient's RAS / RAF status is essential for inclusion in this group.

[0720] Pesentuzumab administration

[0721] Pesentuzumab is administered by intravenous infusion Q2W using a 1500 mg Q2W dosing regimen. For the Day 1 infusion of Cycle 1, administration is performed by intravenous infusion over approximately 6 hours. At the investigator's discretion and in the absence of an IRR, subsequent infusions after Day 1 of Cycle 1 may be reduced to 2 hours (±15 minutes). Alternatively, Pesentuzumab is administered at a fixed dose of 1100 mg Q2W, or in an amount that achieves at least 90%, at least 95%, or at least 99% human receptor target engagement for both EGFR and LGR5 in a statistically significant number of subjects at the relevant body weight. 90% of this target engagement can be achieved using a fixed dose of approximately 1000 mg Q2W. 95% of this amount can be achieved using a fixed dose of approximately 1100 to approximately 1200 mg Q2W.

[0722] One cycle is considered to be 4 weeks. For each patient, a 1-hour observation period will be implemented after the end of infusion (EOI).

[0723] Duration of treatment

[0724] Study treatment was administered until progressive disease (PD) confirmed by independent review (according to RECIST v1.1), unacceptable toxicity, withdrawal of patient consent, patient noncompliance, investigator decision (e.g., clinical worsening), or discontinuation of pesentuzumab for more than 6 consecutive weeks. All patients entered the survival follow-up period and survival status will continue to be tracked until the end of the study, and all patients have the opportunity to be followed for ≥18 months, even after starting new anticancer treatment.

[0725] Efficacy evaluation

[0726] Tumor assessments will be based on CT / magnetic resonance imaging (MRI) with appropriate anatomical contrast of the head, neck, chest, abdomen, and tumor location according to RECIST v1.1, Q8W, for up to 12 months after the start of treatment. Tumor assessments can be performed after 12 months (Q12W). Brain scans will be performed at baseline if clinically indicated and repeatedly at the same frequency as head, neck, chest, and abdomen scans if metastases are present. For patients with suspected bone metastases outside the CT scan area at baseline or suspected disease on study, bone scans will be performed as clinically indicated. Imaging for all patients will be performed by a central imaging center for BICR and reviewed by local investigators.

[0727] Concomitant medication

[0728] Pesentuzumab is metabolized by lysosomal enzymes in the liver and / or kidney to amino acids, which are then reabsorbed and / or reincorporated into endogenous proteins. Therefore, drug-drug interactions with drugs metabolized by cytochrome P450 (CYP450), including components of the FOLFIRI and FOLFOX regimens, are not expected.

[0729] An appropriate recommended premedication regimen should be followed prior to the first infusion. For subsequent infusions, investigators are advised to administer an appropriate premedication regimen at their discretion. In the event of an IRR, hypersensitivity reaction, or anaphylaxis, symptomatic treatment should be administered according to local standard clinical practice and dosing instructions.

[0730] The investigator may administer, at his or her discretion, all medications necessary for the patient's safety and well-being that are not expected to interfere with the evaluation of the study drug.

[0731] Concurrent radiation therapy for symptom control without evidence of progression during the study period was performed only with sponsor authorization.

[0732] Prohibited medications include concomitant long-term oral corticosteroids (>10 mg / day prednisone equivalents), tumor necrosis factor (TNF)-α inhibitors, anti-T cell antibodies, or other immunosuppressive drugs. Strong CYP3A4 inducers, strong CYP3A4 or UGT1A1 inhibitors will not be administered with irinotecan unless there is no therapeutic alternative. Any study drug or other anticancer therapy during the study. Optionally, start herbal therapy for cancer treatment. Herbal therapy is allowed to be started before entry into the study and continued during the study. No major surgery or radiation therapy may be performed without the consent of the sponsor except in emergencies. Prior radiation therapy to ≥25% of the bone marrow.

Claims

1. An antibody or a functional part, derivative and / or analog thereof, comprising a variable domain that binds to the extracellular portion of EGFR, for use in treating cancer in a subject, wherein the treatment further comprises administration of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

2. A method for treating cancer in a subject, comprising administering to the subject an effective amount of an antibody comprising a variable domain that binds to the extracellular portion of EGFR, or a functional portion, derivative, and / or analog thereof, and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38.

3. The antibody or functional part, derivative and / or analogue thereof, or method according to claim 1 or 2, wherein the cancer is adenocarcinoma or squamous cell carcinoma.

4. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of the preceding claims, wherein the cancer is head and neck cancer, gastric cancer, esophageal cancer, gastroesophageal junction cancer, non-small cell lung cancer or colorectal cancer.

5. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of the preceding claims, wherein the cancer is colorectal cancer.

6. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of the preceding claims, wherein the fluoropyrimidine is fluorouracil.

7. The antibody or functional part, derivative and / or analogue thereof, or method according to claim 6, wherein fluorouracil is administered at 200 mg / m 2 Up to 3000 mg / m 2 Administer intravenously within the range of 8. The antibody or functional part, derivative and / or analogue thereof, or method according to claim 6 or 7, wherein fluorouracil is administered at 400 mg / m on day 1. 2 Administer intravenously as a bolus, followed by 2400 mg / m² every two weeks 2 Up to 3000 mg / m 2 Administer intravenously as a continuous infusion over 46 hours.

9. The antibody or functional part, derivative and / or analogue thereof, or the method according to claim 6 or 7, wherein fluorouracil is administered at 500 mg / m2 on days 1, 8, 15, 22, 29 and 36 of an 8-week cycle. 2 Administer as an intravenous bolus.

10. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of claims 1 to 5, wherein the fluoropyrimidine is trifluridine and the treatment further comprises the administration of a thymidine phosphorylase inhibitor.

11. The antibody or functional part, derivative and / or analogue thereof, or method according to claim 10, wherein the thymidine phosphorylase inhibitor is tipiracil hydrochloride.

12. The antibody or functional part, derivative and / or analogue thereof, or method according to claim 10 or 11, wherein trifluridine and tipiracil hydrochloride are administered at a molar ratio of 1:0.5 twice daily at a dose of 35-80 mg / m 2 Application.

13. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of claims 1 to 5, wherein the platinum-based chemotherapeutic agent is oxaliplatin.

14. The antibody or functional part, derivative and / or analogue thereof, or method according to claim 13, wherein oxaliplatin is administered at 85 mg / m2 every two weeks. 2 Application.

15. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of claims 1 to 5, wherein the BCL-2 inhibitor is venetoclax.

16. The antibody or functional part, derivative and / or analogue thereof, or method according to claim 15, wherein venetoclax is administered at 20-1200 mg per day.

17. The antibody or functional part, derivative and / or analog thereof, or method according to any one of claims 1 to 5, wherein the treatment comprises administering SN-38.

18. The antibody or functional part, derivative and / or analogue thereof, or method according to claim 6, wherein the treatment further comprises the administration of oxaliplatin and folinic acid.

19. The antibody or functional part, derivative and / or analogue thereof, or method according to claim 18, wherein oxaliplatin is administered at a dose of 50-200 mg / m 2 With folinic acid at 200-600 mg / m 2 Administer intravenously together, followed by fluorouracil at 1200-3600 mg / m 2 Intravenous administration.

20. The antibody or functional part, derivative and / or analogue thereof, or method according to claim 6, wherein the treatment further comprises administration of irinotecan and folinic acid.

21. The antibody or functional part, derivative and / or analogue thereof, or method according to claim 20, wherein irinotecan is administered at 180 mg / m 2 With folinic acid at 200-400 mg / m 2 Administer intravenously together, followed by fluorouracil at 400-2400 mg / m 2 Intravenous administration.

22. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of the preceding claims, wherein the antibody is enhanced by ADCC.

23. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of the preceding claims, wherein the antibody is defucosylated.

24. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of the preceding claims, wherein the antibody is a multispecific antibody.

25. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of the preceding claims, wherein the antibody is a bispecific antibody.

26. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of the preceding claims, wherein the variable domain that binds to the extracellular part of EGFR is a heavy chain variable region comprising: - a CDR3 sequence of at least the VH of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3 , or a CDR3 sequence that differs from the VH CDR3 sequence of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3 by at most three, two or one amino acid; or - at least the CDR1, CDR2 and CDR3 sequence of the VH of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3 ; or the CDR1, CDR2 and CDR3 sequence of the VH of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3 with up to three, two or one amino acid substitutions; or - the amino acid sequence of the VH chain of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3; or - the amino acid sequence of the VH chain of MF3370; MF3755; MF4280 or MF4289 as shown in Figure 3, which has up to 15, or 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid insertions, deletions, substitutions or a combination thereof relative to the VH chain of MF3370; MF3755; MF4280 or MF4289.

27. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of the preceding claims, wherein the antibody comprises a second variable domain that does not bind to EGFR.

28. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of the preceding claims, wherein the antibody comprises a second variable domain that binds to LGR5.

29. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of the preceding claims, wherein the variable domain that binds to LGR5 binds to an epitope located within amino acid residues 21-118 of the human LGR5 sequence shown in Figure 1 .

30. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of the preceding claims, wherein the variable domain that binds to LGR5 is a heavy chain variable region comprising: - a CDR3 sequence of at least the VH of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3 , or a CDR3 sequence that differs from a CDR3 sequence of the VH of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3 by at most three, two, or one amino acid; or - at least the CDR1, CDR2 and CDR3 sequence of the VH of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3 ; or the CDR1, CDR2 and CDR3 sequence of the VH of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3 with up to three, two or one amino acid substitutions; or - the sequence of the VH chain of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3; or - the amino acid sequence of the VH chain of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818 as shown in Figure 3, which has up to 15, or 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 amino acid insertions, deletions, substitutions or a combination thereof relative to the VH chain of MF5790; MF5803; MF5805; MF5808; MF5809; MF5814; MF5816; MF5817; or MF5818.

31. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of the preceding claims, wherein the antibody is pesentumab.

32. An antibody or functional part thereof, derivative and / or analog, or method as described in any one of claims 1 to 26, wherein the antibody is a monovalent antibody that does not contain a second variable domain, or wherein the antibody comprises the EGFR binding variable domain as its only variable domain.

33. The antibody or functional part, derivative and / or analogue thereof, or the method according to any one of the preceding claims, wherein the subject is a mammal, in particular a human.

34. The antibody or functional part, derivative and / or analogue thereof, or the method according to any one of the preceding claims, wherein the treatment comprises providing 1500 mg of the antibody or functional part, derivative and / or analogue thereof to the subject.

35. The antibody or functional part, derivative and / or analogue thereof, or the method according to any one of the preceding claims, wherein the subject to be treated has not received previous anti-cancer therapy.

36. The antibody or functional part, derivative and / or analogue thereof, or method according to any one of claims 1 to 34, wherein the cancer or the subject to be treated has progressed following a previous anti-cancer therapy.

37. The antibody or functional part, derivative and / or analogue thereof, or method according to claim 36, wherein the previous anti-cancer therapy is chemotherapy, targeted therapy, immunotherapy or radiotherapy.

38. The antibody or functional part, derivative and / or analogue thereof, or method according to claim 36 or 37, wherein the previous anti-cancer therapy is fluorouracil, a platinum-based chemotherapy agent, oxaliplatin, FOLFOX, FOLFIRI, TAS-102, trastuzumab, pembrolizumab, nivolumab, cetuximab or a combination thereof.

39. The antibody or functional part, derivative and / or analog thereof, or the method according to any one of the preceding claims, wherein the antibody or functional part, derivative and / or analog thereof and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38 are administered to the subject simultaneously, separately or sequentially.

40. The method for treating cancer in a subject according to any one of the preceding claims, wherein the method comprises administering to the subject an effective amount of the antibody or a functional part, derivative and / or analog thereof, and an effective amount of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38.

41. A pharmaceutical composition comprising an antibody as defined in any one of the preceding claims, or a functional part, derivative and / or analogue thereof, and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38.

42. The pharmaceutical composition according to claim 41, wherein the antibody or functional part, derivative and / or analog thereof and the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor or SN-38 are provided in a single formulation.

43. The pharmaceutical composition according to claim 41, wherein the antibody or functional part, derivative and / or analog thereof and the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor or SN-38 are provided in separate formulations.

44. A kit of parts comprising: - an antibody as defined in any of the preceding claims, or a functional part, derivative and / or analogue thereof; - A fluoropyrimidine, platinum-based chemotherapy agent, BCL-2 inhibitor, or SN-38; and - Instructions for use of the antibody or functional part, derivative and / or analog thereof, and instructions for use of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor or SN-38.

45. The kit of parts according to claim 44, wherein the instructions for use of the antibody or a functional part, derivative and / or analogue thereof comprise instructions to administer it at 1500 mg.

46. The kit of parts according to claim 44 or 45, wherein the kit comprises the antibody or functional part, derivative and / or analog thereof and instructions for use of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38 in the treatment of adenocarcinoma or squamous cell carcinoma.

47. The kit of parts according to any one of claims 44 to 46, wherein the kit comprises the antibody or a functional part, derivative and / or analog thereof and instructions for use of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38 in the treatment of head and neck cancer, gastric cancer, esophageal cancer, gastroesophageal junction cancer, non-small cell lung cancer or colorectal cancer.

48. The kit of parts according to any one of claims 44 to 47, wherein the kit comprises the antibody or a functional part, derivative and / or analog thereof and instructions for use of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38 in the treatment of colorectal cancer.

49. A combination of an antibody or a functional part, derivative and / or analogue thereof as defined in any of the preceding claims and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38 as defined in any of the preceding claims for use in treating cancer in a subject in need thereof.

50. The following combination: - an antibody as defined in any of the preceding claims, or a functional part, derivative and / or analogue thereof; - instructions for use of the antibody in the treatment of cancer in a subject, and - Instructions for use of a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor, or SN-38 as defined in any one of the preceding claims for the treatment of cancer in a subject.

51. The combination according to claim 50, wherein the instructions for use include the amount of the antibody or functional part, derivative and / or analog thereof to be used and / or the amount of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor or SN-38 to be used, and / or the dosing interval and / or the cancer to be treated.

52. The following combination: - a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38 as defined in any one of the preceding claims, - Instructions for use of a fluoropyrimidine, platinum-based chemotherapy agent, BCL-2 inhibitor, or SN-38 in the subject's cancer treatment, and - Instructions for use of an antibody as defined in any of the preceding claims, or a functional part, derivative and / or analogue thereof, for the treatment of cancer in a subject.

53. The combination according to claim 52, wherein the instructions for use include the amount of the antibody or functional part, derivative and / or analog thereof to be used and / or the amount of the fluoropyrimidine, platinum-based chemotherapeutic agent, BCL-2 inhibitor or SN-38 to be used, and / or the dosing interval and / or the cancer to be treated.

54. Use of an antibody or a functional part, derivative and / or analog thereof as defined in any one of the preceding claims and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38 in the manufacture of one or more medicaments for treating cancer in a subject, wherein the treatment comprises the simultaneous, separate or sequential administration of the antibody or a functional part, derivative and / or analog thereof and a fluoropyrimidine, a platinum-based chemotherapeutic agent, a BCL-2 inhibitor or SN-38.

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