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WO2020181080A1 - Biomarqueurs pour le traitement du cancer - Google Patents

Biomarqueurs pour le traitement du cancer Download PDF

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WO2020181080A1
WO2020181080A1 PCT/US2020/021181 US2020021181W WO2020181080A1 WO 2020181080 A1 WO2020181080 A1 WO 2020181080A1 US 2020021181 W US2020021181 W US 2020021181W WO 2020181080 A1 WO2020181080 A1 WO 2020181080A1
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mutation
her3
gene
seq
cancer
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Richard W. GEDRICH
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Celldex Therapeutics Inc
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Celldex Therapeutics Inc
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Definitions

  • HER3 -associated cancer expressing specific biomarkers with HER3 inhibitors and provided herein are also biomarkers and uses thereof in determining likelihood of effective cancer treatment with HER3 inhibitors. Also provided herein are methods of treating a HER3 -associated head and neck cancer with HER3 inhibitors, wherein the primary tumor site is oral cavity.
  • the human epidermal growth factor receptor 3 (HER3, also known as Erbb3) is a receptor protein tyrosine and belongs to the epidermal growth factor receptor (EGFR) EGFR/HER subfamily of receptor protein tyrosine kinases (RTK), consisting of EGFR (HERl/Erbbl), HER2/Erbb2, HER3/Erbb3 and HER4/Erbb4.
  • EGFR and HER2 are among the most well-established oncogenic RTKs driving the tumorigenesis of multiple types of solid tumors, including major categories such as breast, colorectal, and lung cancers.
  • the tyrosine kinase activities of EGFR and HER2 have been shown to be essential for their oncogenic activities.
  • the transmembrane receptor HER3 consists of an extracellular ligand-binding domain (ECD), a dimerization domain within the ECD, an transmembrane domain, and intracellular protein tyrosine kinase domain (TKD) and a C- terminal phosphorylation domain (see, e.g., Kim et al. (1998), Biochem. J. 334, 189-195;
  • ECD extracellular ligand-binding domain
  • TKD intracellular protein tyrosine kinase domain
  • C- terminal phosphorylation domain see, e.g., Kim et al. (1998), Biochem. J. 334, 189-195;
  • HER3 has been shown to lack detectable tyrosine kinase activity, likely due to a non-conservative replacement of certain key residues in the tyrosine kinase domain. Therefore, a consequence of this kinase-deficiency, HER3 needs to form hetero-dimers with other RTKs, especially EGFR and HER2, to undergo phosphorylation and be functionally active.
  • HER3 The central role for HER3 in oncogenesis is acting as a scaffolding protein to enable the maximum induction of the PI3K/AKT pathway.
  • HER3 promotes tumor growth in unstressed conditions, and has also been found to be highly involved in conferring therapeutic resistances to many targeted drugs, including EGFR tyrosine kinase inhibitors, HER2 monoclonal antibodies such as trastuzumab, as well as small molecule inhibitors of PI3K or AKT or MEK.
  • HER3 has been found to be overexpressed and/or overactivated in several types of cancers such as breast cancer, ovarian cancer, prostate cancer, liver cancer, kidney and urinary bladder cancers, pancreatic cancers, brain cancers, hematopoietic neoplasms, retinoblastomas, melanomas, colorectal cancers, gastric cancers, head and neck cancers, lung cancer, etc. (see, e.g. , Sithanandam & Anderson (2008) Cancer Gene Ther. 15, 413-448). In general, HER3 is frequently activated in EGFR, HER2, C-Met, and FGFRII- expressing cancers.
  • a method of treating a HER3 -associated cancer comprising administering to a patient diagnosed with said cancer a therapeutically effective amount of a HER3 inhibitor, wherein the cancer is characterized in that cells from the cancer comprise a mutation in the FAT1 gene.
  • a method of determining whether a patient diagnosed with a HER3 -associated cancer is indicated as likely to be responsive to treatment with a HER3 inhibitor comprising detecting a mutation in the FAT1 gene in a sample, wherein the sample comprises or is derived from the patient’s cells, e.g, cancerous cells, and wherein the presence of a mutation in the FAT1 gene in the sample indicates that the patient is likely to be responsive to treatment with a HER3 inhibitor.
  • a method for treating a patient with a HER3 inhibitor comprising the steps of: determining whether cells of the cancer comprise a mutation in the FAT1 gene; and if the cells of the cancer comprise a mutation in the FAT1 gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • a method of treating a HER3 -associated cancer in a patient diagnosed therewith comprising the steps of: determining whether cells of the cancer comprise a mutation in the FAT1 gene; and if the cells of the cancer comprise a mutation in the FAT1 gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the step of determining whether cells of the cancer comprise a mutation in the FAT1 gene is performed by: performing an assay on a sample that comprises or is derived from the patient’s cancerous cells to determine whether a mutation in the FAT1 gene is present in the sample.
  • the step of determining whether cells of the cancer comprise a mutation in the FAT1 gene is performed by: obtaining a sample, wherein the sample comprises or is derived from the patient’s cancerous cells; and performing an assay on the sample to determine whether a mutation in the FAT1 gene is present in the sample.
  • a method of detecting a mutation in the FAT1 gene in a patient diagnosed with a HER3 -associated cancer comprising: performing an assay on a sample that comprises or is derived from the patient’s cells, e.g. , cancerous cells, to determine whether a mutation in the FAT1 gene is present in the sample.
  • a method of detecting a mutation in the FAT1 gene in a patient diagnosed with a HER3 -associated cancer comprising: obtaining a sample, wherein the sample comprises or is derived from the patient’s cells, e.g. , cancerous cells; and performing an assay on the sample to determine whether a mutation in the FAT1 gene is present in the sample.
  • one or more additional biomarkers such as the mutation(s) in additional gene(s) are employed besides the mutation in the FAT1 gene.
  • a method of treating a HER3 -associated cancer comprising administering to a patient diagnosed with said cancer a therapeutically effective amount of a HER3 inhibitor, wherein the cancer is characterized in that cells from the cancer comprise at least one mutation in at least one NOTCH gene.
  • a method of determining whether a patient diagnosed with a HER3 -associated cancer is indicated as likely to be responsive to treatment with a HER3 inhibitor comprising detecting at least one mutation in at least one NOTCH gene in a sample, wherein the sample comprises or is derived from the patient’s cells, e.g ., cancerous cells, and wherein the presence of at least one mutation in the at least one NOTCH gene in the sample indicates that the patient is likely to be responsive to treatment with a HER3 inhibitor.
  • a method for treating a patient with a HER3 inhibitor wherein the patient is suffering from a HER3 -associated cancer, the method comprising the steps of: determining whether cells of the cancer comprise at least one mutation in at least one NOTCH gene; and if the cells of the cancer comprise at least one mutation in the at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • a method of treating a HER3 -associated cancer in a patient diagnosed therewith comprising the steps of: determining whether cells of the cancer comprise at least one mutation in at least one NOTCH gene; and if the cells of the cancer comprise at least one mutation in the at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the step of determining whether cells of the cancer comprise at least one mutation in at least one NOTCH gene is performed by: performing an assay on a sample that comprises or is derived from the patient’s cancerous cells to determine whether at least one mutation in the at least one NOTCH gene is present in the sample.
  • the step of determining whether cells of the cancer comprise at least one mutation in at least one NOTCH gene is performed by: obtaining a sample, wherein the sample comprises or is derived from the patient’s cancerous cells; and performing an assay on the sample to determine whether at least one mutation in the at least one NOTCH gene is present in the sample.
  • a method of detecting at least one mutation in at least one NOTCH gene in a patient diagnosed with a HER3 -associated cancer comprising: performing an assay on a sample that comprises or is derived from the patient’s cells, e.g ., cancerous cells, to determine whether at least one mutation in the at least one NOTCH gene is present in the sample.
  • a method of detecting at least one mutation in at least one NOTCH gene in a patient diagnosed with a HER3 -associated cancer comprising: obtaining a sample, wherein the sample comprises or is derived from the patient’s cells, e.g. , cancerous cells; and performing an assay on the sample to determine whether at least one mutation in the at least one NOTCH gene is present in the sample.
  • one or more additional biomarkers are employed besides the at least one mutation in the at least one NOTCH gene.
  • a method of treating a HER3 -associated cancer comprising administering to a patient diagnosed with said cancer a therapeutically effective amount of a HER3 inhibitor, wherein the cancer is characterized in that cells from the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene.
  • a method of treating a HER3 -associated cancer comprising administering to a patient diagnosed with said cancer a therapeutically effective amount of a HER3 inhibitor, wherein the cancer is characterized in that tissue from the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene.
  • a method of determining whether a patient diagnosed with a HER3 -associated cancer is indicated as likely to be responsive to treatment with a HER3 inhibitor comprising detecting a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene in a sample, wherein the sample comprises or is derived from the patient’s cells, e.g. , cancerous cells, and wherein the presence of a mutation in the FAT1 gene and at least one mutation in the at least one NOTCH gene in the sample indicates that the patient is likely to be responsive to treatment with a HER3 inhibitor.
  • a method for treating a patient with a HER3 inhibitor wherein the patient is suffering from a HER3 -associated cancer, the method comprising the steps of: determining whether cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and if the cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in the at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • a method of treating a HER3 -associated cancer in a patient diagnosed therewith comprising the steps of: determining whether cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and if the cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in the at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the step of determining whether cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene is performed by: performing an assay on a sample that comprises or is derived from the patient’s cancerous cells to determine whether a mutation in the FAT1 gene and at least one mutation in the at least one NOTCH gene are present in the sample.
  • the step of determining whether cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene is performed by: obtaining a sample, wherein the sample comprises or is derived from the patient’s cancerous cells; and performing an assay on the sample to determine whether a mutation in the FAT1 gene and at least one mutation in the at least one NOTCH gene are present in the sample.
  • a method for treating a patient with a HER3 inhibitor wherein the patient is suffering from a HER3 -associated cancer, the method comprising the steps of: determining whether tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and if the tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in the at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • a method of treating a HER3 -associated cancer in a patient diagnosed therewith comprising the steps of: determining whether tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and if the tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in the at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the step of determining whether tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene is performed by: performing an assay on a sample that comprises or is derived from the patient’s cancerous cells to determine whether a mutation in the FAT1 gene and at least one mutation in the at least one NOTCH gene are present in the sample.
  • the step of determining whether tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene is performed by: obtaining a sample, wherein the sample comprises or is derived from the patient’s cancerous cells; and performing an assay on the sample to determine whether a mutation in the FAT1 gene and at least one mutation in the at least one NOTCH gene are present in the sample.
  • a method of detecting a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene in a patient diagnosed with a HER3 -associated cancer comprising: performing an assay on a sample that comprises or is derived from the patient’s cells, e.g ., cancerous cells, to determine whether a mutation in the FAT1 gene and at least one mutation in the at least one NOTCH gene are present in the sample.
  • a method of detecting a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene in a patient diagnosed with a HER3 -associated cancer comprising: obtaining a sample, wherein the sample comprises or is derived from the patient’s cells, e.g. , cancerous cells; and performing an assay on the sample to determine whether a mutation in the FAT1 gene and at least one mutation in the at least one NOTCH gene are present in the sample.
  • one or more additional biomarkers are employed besides the mutation in the FAT1 gene and the at least one mutation in the at least one NOTCH gene.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in the at least one NOTCH gene is one or more loss-of-function mutations. In certain embodiments of the methods presented herein, the at least one mutation in the at least one NOTCH gene comprises one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation
  • the at least one mutation in the at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation
  • the at least one mutation in the at least one NOTCH gene comprises one or more loss-of- function mutations.
  • the at least one NOTCH gene is the NOTCH1, the NOTCH2, the NOTCH3, and/or the NOTCH4 gene.
  • the at least one NOTCH gene is the NOTCH1 gene.
  • the at least one NOTCH gene comprises the NOTCH1 gene.
  • the at least one NOTCH gene is the NOTCH2 gene.
  • the at least one NOTCH gene comprises the NOTCH2 gene.
  • the at least one NOTCH gene is the NOTCH3 gene.
  • the at least one NOTCH gene comprises the NOTCH3 gene. In a specific embodiment of the methods presented herein, the at least one NOTCH gene is the NOTCH4 gene. In another specific embodiment of the methods presented herein, the at least one NOTCH gene comprises the NOTCH4 gene.
  • the at least one mutation in at least one NOTCH gene is one mutation. In other embodiments of the methods presented herein, the at least one mutation in at least one NOTCH gene is more than one mutations. [0047] In some embodiments of the methods presented herein, the at least one NOTCH gene is one NOTCH gene. In particular embodiments, the one NOTCH gene is the NOTCH2 gene. In other embodiments of the methods presented herein, the at least one NOTCH gene is more than one NOTCH gene. In particular embodiments, one of the NOTCH genes is the NOTCH2 gene.
  • the at least one mutation in at least one NOTCH gene is one mutation in one NOTCH gene.
  • the one mutation in one NOTCH gene is one mutation in the NOTCH2 gene.
  • the at least one mutation in at least one NOTCH gene is more than one mutation in one NOTCH gene.
  • the one NOTCH gene is the NOTCH2 gene.
  • the at least one mutation in at least one NOTCH gene is more than one mutation in more than one NOTCH gene, for example, one mutation in one NOTCH gene and another mutation in another NOTCH gene.
  • one of the NOTCH genes is the NOTCH2 gene.
  • the at least one mutation in at least one NOTCH gene is one or more homozygous mutations. In other embodiments, the at least one mutation in at least one NOTCH gene is one or more heterozygous mutations. In other embodiments, the at least one mutation in at least one NOTCH gene is one or more homozygous mutations and one or more heterozygous mutations.
  • the at least one NOTCH gene comprises or is the NOTCH2 gene, and the at least one mutation in the at least one NOTCH gene comprises or is an R91 * truncation mutation in the NOTCH2 gene.
  • R91 * denotes a mutation that changes R91 of NP 077719.2 to a stop codon (for example, a CGA to TGA mutation at the DNA level).
  • the at least one NOTCH gene comprises or is the NOTCH2 gene, and the at least one mutation in the at least one NOTCH gene comprises or is a W16* truncation mutation in the NOTCH2 gene.
  • W16* denotes a mutation that changes W16 of NP 077719.2 to a stop codon (for example, a TGG to TGA mutation at the DNA level).
  • the at least one NOTCH gene comprises or is the NOTCH2 gene
  • the at least one mutation in the at least one NOTCH gene comprises or is a splice acceptor mutation in exon 6 in the NOTCH2 gene.
  • a method of treating a HER3 -associated cancer in a patient diagnosed therewith comprising: (1) administering to the patient an HER3 inhibitor, and (2) administering to the patient a FAT1 inhibitor.
  • the FAT1 inhibitor is an antibody, a small molecule, or an oligonucleotide.
  • the cancer is a malignant melice.
  • tissue or cells from the cancer also express a neuregulin or neuregulin fusion protein, and/or comprise a mutation in the BRAF and/or MEK gene.
  • the HER3 -associated cancer is head and neck cancer, breast cancer, ovarian cancer, prostate cancer, liver cancer, kidney cancer, bladder cancer, pancreatic cancer, brain cancer, hematopoietic neoplasms, retinoblastoma, melanoma, colorectal cancer, gastric cancers, or lung cancer.
  • the HER3 -associated cancer is head and neck cancer.
  • the HER3 -associated cancer is squamous cell carcinoma of the head and neck.
  • the primary tumor site is oral cavity.
  • a method of treating a HER3 -associated cancer comprising administering to a patient diagnosed with said cancer a therapeutically effective amount of a HER3 inhibitor, wherein the HER3 -associated cancer is head and neck cancer, and wherein the primary tumor site is oral cavity.
  • the HER3 -associated cancer is squamous cell carcinoma of the head and neck.
  • the HER3 inhibitor is an anti-HER3 antibody or antigen-binding fragment thereof.
  • the anti- HER3 antibody or antigen-binding fragment thereof specifically binds to the same HER3 epitope as an antibody or antigen-binding fragment thereof comprising the heavy chain variable region (VH) and light chain variable region (VL) of CL16 or 2C2.
  • the VH and VL of CL16 comprise SEQ ID NOs: 2 and 1, respectively, and the VH and VL of 2C2 comprise SEQ ID NOs: 2 and 3, respectively.
  • the anti-HER3 antibody or antigen-binding fragment thereof is affinity matured.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises an antibody VL and an antibody VH, wherein the VL comprises the amino acid sequence:
  • FW1 comprises SEQ ID NO: 40 or 44
  • FW2 comprises SEQ ID NO: 41
  • FW3 comprises SEQ ID NO: 42
  • FW4 comprises SEQ ID NO: 43
  • FW5 comprises SEQ ID NO: 36
  • FW6 comprises SEQ ID NO: 37
  • FW7 comprises SEQ ID NO: 38
  • FW8 comprises SEQ ID NO: 39.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises a VL and a VH comprising VL-CDR1, VL-CRD2, VL-CDR3, VH-CDRl, VH-CDR2, and VH-CDR3 amino acid sequences identical or identical except for four, three, two, or one amino acid substitutions in one or more CDRs to: SEQ ID NOs: 18, 21, 22, 31, 32, and 35, SEQ ID NOs: 18, 21, 26, 31, 32 and 35, SEQ ID NOs: 18, 21, 27, 31, 32 and 35, SEQ ID NOs: 20, 21, 22, 31, 32 and 35, SEQ ID NOs: 19, 21, 22, 31, 32 and 35, SEQ ID NOs: 18, 21, 25, 31, 32 and 35, SEQ ID NOs: 18, 21, 28, 31, 32 and 35, SEQ ID NOs: 18, 21, 29, 31, 32 and 35, SEQ ID NOs: 18, 21, 30, 31, 32 and 35, SEQ ID NOs: 18, 21, 23, 31, 32 and 35, SEQ ID NOs: 19, 21, 23, 31, 32 and 35, SEQ ID NOs
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises an antibody VL and an antibody VH, wherein the VL comprises an amino acid sequence at least about 90% to about 100% identical to a reference amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, and SEQ ID NO: 17.
  • a reference amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, and SEQ ID NO: 17.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises an antibody VL and an antibody VH, wherein the VH comprises an amino acid sequence at least about 90% to about 100% identical to a reference amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 12 and SEQ ID NO: 13.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises a VL comprising a sequence at least about 90% to about 100% identical to a reference amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, and SEQ ID NO: 17, and wherein the anti-HER3 antibody or antigen- binding fragment thereof comprises a VH comprising a sequence at least about 90% to about 100% identical to a reference amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 12 and SEQ ID NO: 13.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises a VL-CDRl, a VL-CDR2 and a VL-CDR3 comprising SEQ ID NO: 19, SEQ ID NO: 21 and SEQ ID NO: 23, respectively, and a VH-CDR1, a VH-CDR2, and a VH-CDR3 comprising SEQ ID NO: 31, SEQ ID NO: 32, and SEQ ID NO: 35, respectively.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises a VL comprising SEQ ID NO: 3 and a VH comprising SEQ ID NO: 2.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises a human heavy chain constant region or fragment thereof.
  • the heavy chain constant region or fragment thereof is an IgG constant region.
  • the IgG constant region is selected from an IgGl constant region, an IgG2 constant region, an IgG3 constant region and an IgG4 constant region.
  • the IgG constant region is an IgGl constant region.
  • the IgG constant domain comprises one or more amino acid substitutions relative to a wild-type IgG constant domain wherein the modified IgG has an increased half-life compared to the half-life of an IgG having the wild- type IgG constant domain.
  • the IgG constant domain comprises one or more amino acid substitutions of amino acid residues at positions 251-257, 285-290, 308-314, 385-389, and 428-436, wherein the numbering is according to the EU index as set forth in Kabat.
  • At least one IgG constant domain amino acid substitution is selected from the group consisting of: (a) substitution of the amino acid at position 252 with Tyrosine (Y), Phenylalanine (F), Tryptophan (W), or Threonine (T), (b) substitution of the amino acid at position 254 with Threonine (T), (c) substitution of the amino acid at position 256 with Serine (S), Arginine (R), Glutamine (Q), Glutamic acid (E), Aspartic acid (D), or Threonine (T), (d) substitution of the amino acid at position 257 with Leucine (L), (e) substitution of the amino acid at position 309 with Proline (P), (f) substitution of the amino acid at position 311 with Serine (S), (g) substitution of the amino acid at position 428 with
  • the human IgG constant domain comprises amino acid substitutions relative to a wild-type human IgG constant domain at positions 252, 254, and 256, wherein (a) the amino acid at position 252 is substituted with Tyrosine (Y), (b) the amino acid at position 254 is substituted with Threonine (T), and (c) the amino acid at position 256 is substituted with Glutamic acid (E), wherein the numbering is according to the EU index as set forth in Kabat.
  • the amino acid at position 434 is substituted with an amino acid selected from the group consisting of Tryptophan (W), Methionine (M), Tyrosine (Y), and Serine (S), and wherein the numbering is according to the EU index as set forth in Kabat.
  • the amino acid at position 428 is substituted with an amino acid selected from the group consisting of Threonine (T), Leucine (L), Phenylalanine (F), and Serine (S), and wherein the numbering is according to the EU index as set forth in Kabat.
  • the amino acid at position 257 is substituted with Leucine (L), and the amino acid at Kabat position 434 is substituted with Tyrosine (Y), and wherein the numbering is according to the EU index as set forth in Kabat.
  • the amino acid at Kabat position 428 is substituted with Leucine (L), and the amino acid at Kabat position 434 is substituted with Serine (S).
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises a light chain constant region selected from the group consisting of a human kappa constant region and a human lambda constant region. In some embodiments, the anti- HER3 antibody or antigen-binding fragment thereof comprises a human lambda constant region.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises an antibody VL of SEQ ID NO:3, an antibody VH of SEQ ID NO:2, and an IgGl constant region of SEQ ID NO:46.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises a human IgGl constant region and a human lambda constant region.
  • the anti-HER3 antibody or antigen-binding fragment thereof is a human antibody, a humanized antibody, a chimeric antibody, a monoclonal antibody, a polyclonal antibody, a recombinant antibody, a multispecific antibody, or an antigen-binding fragment thereof.
  • the anti-HER3 antibody or antigen binding fragment thereof is a human antibody or antigen-binding fragment thereof.
  • the antigen-binding fragment is a Fv, Fab, F(ab')2, Fab', dsFv, scFv, or sc(Fv)2.
  • the method comprises administering an EGFR inhibitor.
  • the EGFR inhibitor is an anti-EGFR antibody or an antigen-binding fragment thereof.
  • the anti-EGFR antibody is cetuximab.
  • the method comprises administering a BRAF inhibitor.
  • the method comprises administering a MEK inhibitor.
  • a kit comprising components for performing the method as described herein.
  • a HER3 inhibitor for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, comprising: determining whether cells of the cancer comprise a mutation in the FAT1 gene; and, if the cells of the cancer comprise a mutation in the FAT1 gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • a HER3 inhibitor for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the cancer is characterized in that cells from the cancer comprise a mutation in the FAT1 gene, said method comprising administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • one or more additional biomarkers are employed besides the mutation in the FAT1 gene.
  • a HER3 inhibitor for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, comprising: determining whether cells of the cancer comprise at least one mutation in at least one NOTCH gene; and, if the cells of the cancer comprise at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of- function mutations.
  • a HER3 inhibitor for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the cancer is characterized in that cells from the cancer comprise at least one mutation in at least one NOTCH gene, said method comprising administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • one or more additional biomarkers such as the mutation(s) in additional gene(s) are employed besides the at least one mutation in the at least one NOTCH gene.
  • a HER3 inhibitor for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, comprising: determining whether cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and, if the cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • a HER3 inhibitor for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, comprising: determining whether tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and, if the tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one
  • NOTCH gene then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of- function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • a HER3 inhibitor for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the cancer is characterized in that cells from the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, said method comprising administering a
  • a HER3 inhibitor for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the cancer is characterized in that tissue from the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, said method comprising administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • a HER3 inhibitor for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the HER3- associated cancer is head and neck cancer, and wherein the primary tumor site is oral cavity.
  • a HER3 inhibitor for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the anti-HER3 antibody is administered in combination with a FAT 1 inhibitor.
  • FIG. 1 depicts the treatment cycles of clinical trial CDX3379-04
  • FIG. 2 shows patients’ exposure to study drug in the CDX3379-04 trial.
  • FIG. 3 shows the results of a biomarker analysis of 18 tumor samples from CDX- 3379 studies. Shaded cells indicate presence of a mutation in TP53, PIK3CA, FAT1, NOTCH1,
  • FIG. 4 shows that FAT1 knockdown enhanced the anti-proliferative activity of CDX-3379 in head and neck squamous cell carcinoma (HNSCC) cells.
  • HER3 -associated cancer comprising administering a therapeutically effective amount of a HER3 inhibitor as well as methods of determining whether a patient is likely to be responsive to such a method of treatment.
  • methods of treating a HER3 -associated cancer expressing specific biomarkers with HER3 inhibitors and provided herein are also biomarkers and uses therefor in determining likelihood of effective HER3 -associated cancer treatment with HER3 inhibitors.
  • the HER3 -associated cancer is characterized in that tissue or cells from the cancer comprise a mutation in the FAT1 gene.
  • the HER3 -associated cancer is characterized in that tissue or cells from the cancer comprise a loss-of-function mutation in the FAT1 gene.
  • tissue or cells from the cancer comprise a loss-of-function mutation in the FAT1 gene.
  • the present invention also contemplates similar methods wherein the abnormal expression level (for example, lower expression level) of the FAT1 gene, abnormal activity (for example, lower activity) of the FAT1 protein, or abnormal level of signaling (for example, lower level of signaling) downstream of the FAT1 protein, as a biomarker for determining the likelihood of effective HER3 -associated cancer treatment with HER3 inhibitors.
  • Such lower expression level, lower activity, or lower level of signaling can be, for example, at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 2-fold, 5-fold, 10-fold, 50-fold, or 100-fold lower relative to the non-mutant or wild-type FAT1 gene/protein.
  • the HER3 -associated cancer is characterized in that tissue or cells from the cancer comprise at least one mutation in at least one NOTCH gene.
  • the HER3 -associated cancer is characterized in that tissue or cells from the cancer comprise one or more loss-of-function mutations in at least one NOTCH gene.
  • the present invention also contemplates similar methods wherein the abnormal expression level(s) (for example, lower expression level(s)) of a NOTCH gene(s), abnormal activity(ies) (for example, lower activity(ies)) of a NOTCH protein(s), or abnormal level(s) of signaling (for example, lower level(s) of signaling) downstream of a NOTCH protein(s), as a biomarker for determining the likelihood of effective HER3 -associated cancer treatment with HER3 inhibitors.
  • the abnormal expression level(s) for example, lower expression level(s) of a NOTCH gene(s
  • abnormal activity(ies) for example, lower activity(ies)
  • signaling for example, lower level(s) of signaling
  • Such lower expression level(s), lower activity(ies), or lower level(s) of signaling can be, for example, at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 2-fold, 5-fold, 10-fold, 50-fold, or 100-fold lower relative to a non-mutant or wild-type NOTCH gene/protein.
  • the HER3 -associated cancer is characterized in that tissue or cells from the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene.
  • the HER3- associated cancer is characterized in that tissue or cells from the cancer comprise a loss-of- function mutation in the FAT1 gene and one or more loss-of-function mutations in at least one NOTCH gene.
  • the present invention also contemplates similar methods wherein the abnormal expression levels (for example, lower expression levels) of the FAT1 gene and a NOTCH gene(s), abnormal activities (for example, lower activities) of the FAT1 protein and a NOTCH protein(s), or abnormal level of signaling (for example, lower level of signaling) downstream of the FAT1 protein and abnormal level of signaling (for example, lower level of signaling) downstream of a NOTCH protein(s), as a biomarker for determining the likelihood of effective HER3 -associated cancer treatment with HER3 inhibitors.
  • the abnormal expression levels for example, lower expression levels
  • abnormal activities for example, lower activities
  • abnormal level of signaling for example, lower level of signaling
  • abnormal level of signaling for example, lower level of signaling downstream of the FAT1 protein and abnormal level of signaling (for example, lower level of signaling) downstream of a NOTCH protein(s)
  • Such lower expression level(s), lower activity(ies), or lower level(s) of signaling can be, for example, at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 2-fold, 5-fold, 10-fold, 50-fold, or 100-fold lower relative to a non-mutant or wild-type FAT1 or NOTCH gene/protein (as the case may be).
  • one or more additional biomarkers are employed, besides the mutation in the FAT1 gene and/or the at least one mutation in the at least one NOTCH gene.
  • HER3 -associated cancer comprising administering a therapeutically effective amount of a HER3 inhibitor, wherein the HER3 -associated cancer is head and neck cancer and wherein the primary tumor site is oral cavity.
  • B, and/or C is intended to encompass each of the following aspects: A, B, and C; A, B, or C;
  • HER3 refers to the ErbB3 protein (also referred to as HER3, ErbB3 receptor in the literature), which is well known in the art; see, e.g ., U.S. Pat No. 5,480,968 Plowman et al. (1990) Proc. Natl. Acad. Sci. USA 87, 4905-4909; see also, Kani et al. (2005) Biochemistry 44, 15842-15857, and Cho & Leahy (2002) Science 297, 1330-1333.
  • a representative full-length, mature HER3 protein sequence corresponds to the sequence shown in FIG. 4 and SEQ ID NO: 4 of U.S. Pat. No. 5,480,968 minus the 19 amino acid leader sequence that is cleaved from the mature protein.
  • HER and“HER receptor” are used interchangeably herein, and refer to one or more, or all, members of the epidermal growth factor receptor (EGFR)
  • EGFR/HER subfamily of receptor protein tyrosine kinases consisting of EGFR (HERl/Erbbl), HER2/Erbb2, HER3/Erbb3 and HER4/Erbb4.
  • RTK receptor protein tyrosine kinases
  • EGFR EGFR protein
  • HERl HERl receptor
  • EGFR receptor EGFR receptor
  • a non-limiting example of an amino acid sequence of human EGFR is provided with GenBank Accession No. NP_005219.2.
  • a non-limiting example of a nucleotide sequence encoding human EGFR is provided with GenBank Accession No. NM_005228.3.
  • FAT1 is used herein to refer to human FAT atypical cadherin 1 (FAT1), which is a type 1 transmembrane protein belonging to the FAT gene family.
  • FAT1 human FAT atypical cadherin 1
  • Non limiting examples of a nucleotide sequence of the FAT1 gene are provided with GenBank Accession Nos. NG_046994.1 : 8083-144133 and NC_000004.12: cl86726696-186587783, as shown in Table 1 below.
  • Non-limiting examples of an mRNA sequence (depicted as DNA base) encoding the FAT1 protein are provided with GenBank Accession Nos.
  • NM_005245.4 XM_006714139.3, XM_005262834.3, and XM_005262835.2, as shown in Table 1 below.
  • GenBank mRNA sequences provided by GenBank are depicted as DNA bases (GCAT) rather than RNA bases (GCAU); a person of ordinary skill in the art would understand that the mRNA nucleotide sequences are identical to the versions depicted as DNA bases, except that U is used instead of T.
  • Non-limiting examples of an amino acid sequence of the FAT1 protein are provided with GenBank Accession Nos. NP_005236.2, XP_006714202.1, XP_005262891.1, and XP_005262892.1, as shown in Table 1 below.
  • Table 1 Non-limiting examples of genomic DNA sequence of the FAT1 gene, mRNA sequence (depicted as DNA bases) encoding the FAT1 protein (start codons are in bold, underlined and italicized, and stop codons are in bold and underlined), and amino acid sequence of the FAT1 protein.
  • a mutation in the FAT1 gene may be heterozygous or homozygous. In some embodiments of the methods presented herein, the mutation in the FAT1 gene is a homozygous mutation. In other embodiments, the mutation in the FAT1 gene is a heterozygous mutation.
  • NOTCH the human Type-1 transmembrane proteins
  • Non-limiting examples of nucleotide sequences of the human NOTCH1, NOTCH2, NOTCH3, and NOTCH4 genes are provided with GenBank Accession Nos. NG_007458.1 :4739-56354, NC_000009.12x136545786-136494433, NG_008163.1 :4960-163101,
  • NT_167248.2x3447269-3418042 NT_167249.2x3540275-3511048, respectively, as shown in Table 2 below.
  • mRNA sequences (depicted as DNA bases) encoding human NOTCH1, NOTCH2, NOTCH3 and NOTCH4 proteins are provided with GenBank Accession Nos. NM_017617.5, XM_011518717.2, NM_024408.4,
  • GCAT DNA bases
  • GCAU RNA bases
  • Non-limiting examples of amino acid sequences of NOTCH1, NOTCH2, NOTCH3 and NOTCH 4 proteins are provided with GenBank Accession Nos.
  • Table 2 Non-limiting examples of genomic DNA sequences of NOTCH1, NOTCH2, NOTCH3 and NOTCH4 genes, mRNA sequences (depicted as DNA bases) encoding NOTCH1, NOTCH2, NOTCH3 and NOTCH4 proteins (start codons are in bold, underlined and italicized, and stop codons are in bold and underlined), and amino acid sequences of NOTCH1, NOTCH2, NOTCH3 and NOTCH4 proteins.
  • the at least one NOTCH gene is one NOTCH gene (i.e., one of the NOTCH1, NOTCH2, NOTCH3 and NOTCH4 genes).
  • the one NOTCH gene is the NOTCH2 gene.
  • the at least one NOTCH gene is more than one NOTCH genes (i.e., more than one of the NOTCH1, NOTCH2, NOTCH3 and NOTCH4 genes).
  • one of the NOTCH genes is the NOTCH2 gene).
  • the at least one mutated NOTCH gene is a mutated NOTCH1 gene.
  • the at least one mutated NOTCH gene comprises a mutated NOTCH1 gene.
  • the at least one mutated NOTCH gene is a mutated NOTCH2 gene. In another preferred embodiment, the at least one mutated NOTCH gene comprises a mutated NOTCH2 gene.
  • the at least one mutated NOTCH gene is a mutated NOTCH3 gene. In another further embodiment, the at least one mutated NOTCH gene comprises a mutated NOTCH3 gene.
  • the at least one mutated NOTCH gene is a mutated NOTCH4 gene. In another further embodiment, the at least one mutated NOTCH gene comprises a mutated NOTCH4 gene.
  • the at least one mutated NOTCH gene is a mutated NOTCH1 gene and a mutated NOTCH2 gene. In another specific embodiment, the at least one mutated NOTCH gene is a mutated NOTCH1 gene and a mutated NOTCH3 gene. In another specific embodiment, the at least one mutated NOTCH gene is a mutated NOTCH1 gene and a mutated NOTCH4 gene. In another specific embodiment, the at least one mutated NOTCH gene is a mutated NOTCH2 gene and a mutated NOTCH3 gene. In another specific embodiment, the at least one mutated NOTCH gene is a mutated NOTCH2 gene and a mutated NOTCH4 gene.
  • the at least one mutated NOTCH gene is a mutated NOTCH3 gene and a mutated NOTCH4 gene. In another specific embodiment, the at least one mutated NOTCH gene is a mutated NOTCH1 gene, a mutated NOTCH2 gene and a mutated NOTCH3 gene. In another specific embodiment, the at least one mutated NOTCH gene is a mutated NOTCH1 gene, a mutated NOTCH2 gene and a mutated NOTCH4 gene. In another specific embodiment, the at least one mutated NOTCH gene is a mutated NOTCH1 gene, a mutated NOTCH3 gene and a mutated NOTCH4 gene. In another specific specific
  • the at least one mutated NOTCH gene is a mutated NOTCH2 gene, a mutated NOTCH3 gene and a mutated NOTCH4 gene.
  • the at least one mutation in at least one NOTCH gene is one mutation. In other embodiments of the methods presented herein, the at least one mutation in at least one NOTCH gene is more than one mutations.
  • the at least one mutation in at least one NOTCH gene is one mutation in one NOTCH gene.
  • the one NOTCH gene is the NOTCH2 gene.
  • the at least one mutation in at least one NOTCH gene is more than one mutation in one NOTCH gene.
  • the one NOTCH gene is the NOTCH2 gene.
  • the at least one mutation in at least one NOTCH gene is more than one mutation in more than one NOTCH gene, for example, one mutation in one NOTCH gene and another mutation in another NOTCH gene.
  • one of the NOTCH genes is the NOTCH2 gene.
  • a mutation in a NOTCH gene may be heterozygous or homozygous.
  • the at least one mutation in at least one NOTCH gene is one or more homozygous mutations.
  • the at least one mutation in at least one NOTCH gene is one or more heterozygous mutations.
  • the at least one mutation in at least one NOTCH gene is one or more homozygous mutations and one or more heterozygous mutations.
  • B-Raf “BRAF,” “B-raf,”“B-Rafl,”“BRAF1,” and“RAFBl” are used interchangeably herein, and refer to the B-Raf protein, for example, as described in Stephens et al., 1992, Mol. Cell. Biol. 12(9):3733-3742.
  • a non-limiting example of an amino acid sequence of human B-Raf is provided with GenBank Accession No. NP 004324.2.
  • a non-limiting example of a nucleotide sequence encoding human B-Raf is provided with GenBank Accession No. NM_004333.4.
  • MEK “mitogen-activated protein kinase (MAPK) kinase” and “MAPKK” are used interchangeably herein, and refer to the MEK protein, for example, as described in Zheng and Guan, 1994, EMBO J. 13(5): 1123-31.
  • a non-limiting example of an amino acid sequence of human MEK is provided with GenBank Accession No. AAI37460.1.
  • a non-limiting example of a nucleotide sequence encoding human MEK is provided with GenBank Accession No. BC137459.1.
  • neuregulin and “heregulin (HRG)” are used interchangeably herein, and refer to the family of neuregulin proteins, including isoforms type I to type III and subtypes (including, for example, NRGl, NRG2, NRG3, NRG4, and the subtypes or isoforms thereof, such as NRGl a, NRG l b, NRG2a, and NRG2P).
  • Other synonyms for neuregulin include acetylcholine receptor inducing activity (ARIA), breast cancer cell differentiation factor p45, glial growth factor, Neu differentiation factor, and sensory and motor neuron-derived factor.
  • a non-limiting example of an amino acid sequence of a human neuregulin, human NRGl is provided with GenBank Accession No. AAI50610.1.
  • a non-limiting example of a nucleotide sequence encoding a human neuregulin, human NRGl is provided with GenBank Accession No. NM 013958.3.
  • a non-limiting example of an amino acid sequence of a human neuregulin, human NRG2 is provided with GenBank Accession No. AAF28848.1.
  • a non limiting example of a nucleotide sequence encoding a human neuregulin, human NRG2 is provided with GenBank Accession No. NM_004883.2.
  • a non-limiting example of an amino acid sequence of a human neuregulin, human NRGl a is provided with GenBank Accession No. DAA00048.1.
  • a non-limiting example of an amino acid sequence encoding a human neuregulin, human NRG l b is provided with GenBank Accession No. AAA58639.1.
  • a non limiting example of an amino acid sequence of a human neuregulin, human NRG2a is provided with GenBank Accession No. AAF28848.1.
  • a non-limiting example of an amino acid sequence encoding a human neuregulin, human NRG2P is provided with GenBank Accession No. AAF28849.1.
  • Levels of NRG isoform expression in a cell or tumor sample may be assayed, for example, by using RNAscope ® technology (see Section VI) using an RNAscope ® probe developed for the specific isoform.
  • “Expressed” or“expression” may refer to: the transcription from a genetic sequence to give an RNA nucleic acid, the translation from an RNA molecule to give a protein, a polypeptide, or portion thereof, or the detectable presence or manifestation of a biomolecule or biomolecular complex, for example, a protein-protein complex.
  • the “expression” of a FAT1 or NOTCH polypeptide may refer to the detectable level of a FAT1 or NOTCH polypeptide as measured, for example, by measuring the detectable level of a FAT1 or NOTCH RNA or polypeptide in a sample.
  • A“high” level refers to a level (e.g ., of expression) that is greater than normal, for example, greater than a level in a“reference sample,” or a level that is greater than a particular standard.
  • the reference sample may be normal/healthy cells, or may be all cancer cells, or may be a particular subset of cancer cells.
  • A“high” level may also refer to a level that is higher than a predetermined amount or measure, such as a predetermined cutoff amount.
  • a group of samples may be divided around a particularly determined value (for example, a median, a mean, or a quartile), or a particularly determined threshold (such as, for example, a line on a graph), such that two subgroups exist, one that is considered to have a“high” level (e.g., of expression) and one that is considered to have a“low” level (or, that does not have a “high” level).
  • a particularly determined value for example, a median, a mean, or a quartile
  • a particularly determined threshold such as, for example, a line on a graph
  • A“reference population” refers to a representative group of reference samples, and can be 1, 5, 10, 25, 50, 75, 100, 200, 250, 300, 400, 500, 1000, 2000, 5000, 10,000, 20,000, 30,000 or more samples.
  • the reference population may be a group of tumor samples, across cancer types.
  • the reference population might also be tissue samples from a population of healthy individuals.
  • a mean, median, quartile, or a line on a graph as determined from a reference population or a reference sample may determine a value or threshold against which an expression level may be“high.”
  • a reference population may be the Affymetrix U133A, Affymetrix U133 2.0, and Affymetrix U133 Plus 2.0 platforms from ONCOMINE®.
  • inhibitors and “suppression” are used interchangeably herein and refer to any statistically significant decrease in biological activity, including full blocking of the activity.
  • inhibition can refer to a decrease of about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 100% in biological activity.
  • the term refers to the ability of an antibody or antigen-binding fragment thereof to statistically significantly decrease the phosphorylation of HER3 induced by an EGF- like ligand, relative to the phosphorylation in an untreated (control) cell.
  • the cell which expresses HER3 can be a naturally occurring cell or cell line (e.g, a cancer cell) or can be recombinantly produced by introducing a nucleic acid encoding HER3 into a host cell.
  • the anti-HER3 binding molecule e.g, an antibody or antigen binding fragment thereof inhibits ligand mediated phosphorylation of HER3 by at least 10%, or at least 20%, or at least 30%, or at least 40%, or at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 905, or about 100%, as determined, for example, by Western blotting followed by probing with an anti-phosphotyrosine antibody or by ELISA.
  • biomarker refers to a measurable indicator of some biological state or condition.
  • a biomarker has a high predictive value for a meaningful outcome measure and likelihood of therapeutic response.
  • biomarkers can be used in conjunction with other diagnostic tools or used alone.
  • the biomarker e.g, mutant FAT1 and/or mutant NOTCH
  • the biomarker is used to assess a pathological state. Measurements of the biomarker may be used alone or combined with other data obtained regarding a patient in order to determine the state of the patient.
  • a“mutation” or a“gene comprising a mutation” or the like has the meaning known to one of skill in the art.
  • a mutation refers to alterations in one or more nucleic acids in a genomic sequence, including one or more base substitutions, deletions, and/or insertions, duplications, amplifications, repeat expansions, rearrangements, e.g, inversions and/or translocations, and deletions of one or more bases that result in silent mutations, non-sense mutations, missense mutations, and/or frameshift mutations.
  • a mutation can, for example, result in premature stop codons, aberrant splicing, transcription and/or translation.
  • the mutation in the FAT1 gene and/or the at least one mutation in the at least one NOTCH gene is an inactivating mutation(s).
  • a gene comprising a mutation can have more than one mutation.
  • a mutation in the genetic sequence is relative to a non-mutant or wild-type genetic sequence.
  • a biomarker that can be used in accordance with a method described herein is a mutation in the FAT1 gene.
  • a biomarker that can be used in accordance with a method described herein is a mutation in a NOTCH gene, such as a
  • a biomarker that can be used in accordance with a method described herein is the combination of a mutation in the FAT1 gene and a mutation in a NOTCH gene, such as a NOTCHl, NOTCH2, NOTCH3 or NOTCH4 gene.
  • a mutation in a NOTCH gene is a truncation mutation.
  • a gene mutation may be present in the protein-coding region of the gene. In some embodiments, a gene mutation may be present in a promoter (regulatory) region of the gene. In some embodiments, a gene mutation may be present in a post-translational region of the gene. In certain embodiments, a gene mutation may be present in intron sequences of the gene. In certain embodiments, a gene mutation may be present in exon sequences of the gene. In certain embodiments, a gene mutation may be present within an intron-exon boundary sequence of the gene. In certain embodiments, a gene mutation may be a splice acceptor mutation. In some embodiments, the at least one mutation is a dominant or recessive mutation.
  • a mutation results in a change in the function of the gene.
  • such a functional mutation in a gene results in a change in the function of the gene relative to the function of a corresponding non-mutant or wild-type gene.
  • a functional mutation in a gene results in a change in the function of the protein encoded by the gene.
  • the change in the function of the protein is relative to the function of a protein encoded by the corresponding non-mutant or wild-type gene.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in the at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation
  • the at least one mutation in the at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a gain-of-function mutation.
  • the at least one mutation in the at least one NOTCH gene is one or more gain-of-function mutations.
  • the mutation in the FAT1 gene is a gain-of-function mutation, and the at least one mutation in the at least one NOTCH gene is one or more loss-of-function mutations. In some embodiments of the methods presented herein, the mutation in the FAT1 gene is a loss- of-function mutation, and the at least one mutation in the at least one NOTCH gene is one or more gain-of-function mutations. In some embodiments of the methods presented herein, the mutation in the FAT1 gene is a gain-of-function mutation, and the at least one mutation in the at least one NOTCH gene is one or more gain-of-function mutations.
  • “Loss-of-function” as used herein refers to a reduction or elimination of the normal activity of a gene or gene product.
  • the reduction or elimination of the activity of the gene or gene product can be due to a decrease in transcription and/or processing of the RNA, a decrease in translation, stability, transport, or function of the gene product, or any combination thereof.
  • a loss-of-function mutation can result in a gene product having less or no function (being partially or wholly inactivated). That is, a loss-of-function mutation can result in a partial loss of function or a complete loss of function.
  • a loss-of-function mutation results in a gene product exhibiting no activity, or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85% of normal activity.
  • a loss-of- function mutation in the FAT1 gene results in the FAT1 polypeptide exhibiting no activity, or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85% of normal activity.
  • a loss-of function mutation in a NOTCH gene results in a NOTCH polypeptide exhibiting no activity, or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85% of normal activity.
  • a loss-of-function mutation may lead to an abnormal level of expression a gene of interest, e.g., no expression, or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, or 80% of normal expression levels.
  • a loss-of-function mutation in the FAT1 gene leads to an abnormal level of expression of the FAT1 gene, e.g., no expression, or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, or 80% of normal FAT1 gene expression levels (FAT1 RNA and/or FAT1 polypeptide levels).
  • an abnormal level of expression of the FAT1 gene e.g., no expression, or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, or 80% of normal FAT1 gene expression levels (FAT1 RNA and/or FAT1 polypeptide levels).
  • a loss-of-function mutation in a NOTCH gene leads to an abnormal level of expression of a NOTCH gene, e.g., no expression, or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, or 80% of normal NOTCH gene expression levels (NOTCH RNA and/or NOTCH polypeptide levels).
  • the change in gene expression levels is due to abnormal upstream signaling of the gene.
  • the at least one NOTCH gene comprises or is the NOTCH2 gene, and the at least one mutation in the at least one NOTCH gene comprises or is an R91 * truncation mutation in the NOTCH2 gene.
  • R91 * denotes a mutation that changes R91 of NP 077719.2 to a stop codon (for example, a CGA to TGA mutation at the DNA level).
  • the at least one NOTCH gene comprises or is the NOTCH2 gene, and the at least one mutation in the at least one NOTCH gene comprises or is a W16* truncation mutation in the NOTCH2 gene.
  • the at least one NOTCH gene comprises or is the NOTCH2 gene
  • the at least one mutation in the at least one NOTCH gene comprises or is a splice acceptor mutation (for example, a splice acceptor mutation in exon 6) in the NOTCH2 gene.
  • “Gain-of-function” as used herein refers to any mutation in a gene in which the protein encoded by said gene acquires a function not normally associated with the protein causes or contributes to a disease or disorder.
  • the gain-of-function mutation changes the function of the resulting protein or causes interactions with other proteins.
  • a gain-of-function mutation changes the gene product such that its effect gets stronger (enhanced activation) or even is superseded by a different, abnormal function.
  • a gain-of-function mutation is a gain-of-function mutation in a NOTCH gene, for example, the NOTCH1, NOTCH2, NOTCH3 or NOTCH4 gene.
  • a gain-of-function mutation is a gain-of-function mutation in the FAT1 gene.
  • a gain-of-function mutation can result in a gene product having abnormal functions or increased activity (compared to the wild type protein).
  • a gain-of-function mutation results in a gene product exhibiting an abnormal function(s) or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85% higher than normal activity.
  • a gain-of-function mutation is a gain-of function mutation in a NOTCH gene that results in a NOTCH polypeptide exhibiting an abnormal function(s) or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or 85% higher than normal activity.
  • a gain-of-function mutation is a gain-of function mutation in the FAT1 gene that results in a FAT1 polypeptide exhibiting an abnormal function(s) or an enhanced activity that is about 120%, 130%, 150%, 200%, 250%, 300%, 400%, or more of normal activity.
  • a gain-of-function mutation may lead to an abnormal level of expression of a gene of interest, e.g., about 5%, 10%, 15%, 20%, 25%, 30%, 35%,
  • the gene of interest is a NOTCH gene
  • a gain-of-function mutation may lead to an abnormal level of expression of a NOTCH gene, e.g, about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, or 80% higher than normal NOTCH gene expression levels (NOTCH RNA and/or NOTCH polypeptide levels).
  • NOTCH RNA and/or NOTCH polypeptide levels normal NOTCH gene expression levels
  • the expression level of the NOTCH RNA and/or NOTCH polypeptide is normal (compared to wild type), but the resultant protein has abnormal or higher signaling abilities than the wild type protein.
  • the gene of interest is the FAT1 gene
  • a gain-of-function mutation may lead to an abnormal level of expression of the FAT1 gene, e.g, about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%,
  • FAT1 RNA and/or FAT1 polypeptide levels 70%, 75%, or 80% higher than normal FAT1 gene expression levels (FAT1 RNA and/or FAT1 polypeptide levels).
  • the expression level of the FAT1 RNA and/or FAT1 polypeptide is normal (compared to wild type), but the resultant protein has abnormal or higher signaling abilities than the wild type protein.
  • the change in gene expression levels is due to abnormal upstream signaling of the gene.
  • Changes in gene expression levels or activity can be detected by methods known in the art (e.g, ELISA, PCR, mass spectrometry based methods, immunohistochemistry, RIA, functional assays).
  • antibody or “immunoglobulin,” as used interchangeably herein, include whole antibodies and any antigen-binding fragment or single chains thereof.
  • a typical antibody comprises at least two heavy (H) chains and two light (L) chains interconnected by disulfide bonds.
  • Each heavy chain is comprised of a heavy chain variable region (abbreviated herein as VH) and a heavy chain constant region.
  • the heavy chain constant region is comprised of three domains, CHI, CH2, and CH3.
  • Each light chain is comprised of a light chain variable region (abbreviated herein as VL) and a light chain constant region.
  • the light chain constant region is comprised of one domain, CL.
  • the VH and VL regions can be further subdivided into regions of hypervariability, termed Complementarity Determining Regions (CDR), interspersed with regions that are more conserved, termed framework regions (FW).
  • CDR Complementarity Determining Regions
  • Each VH and VL is composed of three CDRs and four FWs, arranged from amino-terminus to carboxy -terminus in the following order: FW1, CDR1, FW2, CDR2, FW3, CDR3, FW4.
  • the variable regions of the heavy and light chains contain a binding domain that interacts with an antigen.
  • the constant regions of the antibodies can mediate the binding of the immunoglobulin to host tissues or factors, including various cells of the immune system (e.g ., effector cells) and the first component (Clq) of the classical complement system.
  • Exemplary antibodies of the present disclosure include the Clone 16 (CL16) anti-HER3 antibodies (original and germlined), affinity optimized clones including for example, the anti-HER3 2C2 antibody, and serum half-life-optimized anti-HER3 antibodies including for example the anti-HER3 2C2-YTE antibody.
  • CL16 Clone 16
  • affinity optimized clones including for example, the anti-HER3 2C2 antibody
  • serum half-life-optimized anti-HER3 antibodies including for example the anti-HER3 2C2-YTE antibody.
  • antibody means an immunoglobulin molecule that recognizes and specifically binds to a target, such as a protein, polypeptide, peptide, carbohydrate,
  • antibody encompasses intact polyclonal antibodies, intact monoclonal antibodies, antibody fragments (such as Fab, Fab', F(ab')2, and Fv fragments), single chain Fv (scFv) mutants, multispecific antibodies such as bispecific antibodies generated from at least two intact antibodies, chimeric antibodies, humanized antibodies, human antibodies, fusion proteins comprising an antigen determination portion of an antibody, and any other modified
  • An antibody can be of any the five major classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, or subclasses (isotypes) thereof (e.g., IgGl, IgG2, IgG3, IgG4, IgAl and IgA2), based on the identity of their heavy-chain constant domains referred to as alpha, delta, epsilon, gamma, and mu, respectively.
  • the different classes of immunoglobulins have different and well known subunit structures and three-dimensional configurations.
  • Antibodies can be naked or conjugated to other molecules such as toxins, radioisotopes, etc.
  • a “blocking" antibody or an “antagonist” antibody is one which inhibits or reduces biological activity of the antigen it binds, such as HER3.
  • blocking antibodies or antagonist antibodies substantially or completely inhibit the biological activity of the antigen. Desirably, the biological activity is reduced by 10%, 20%, 30%, 50%, 70%, 80%, 90%, 95%, or even 100%.
  • HER3 antibody or “an antibody that binds to HER3” or “anti-HER3” refers to an antibody that is capable of binding HER3 with sufficient affinity such that the antibody is useful as a therapeutic agent or diagnostic reagent in targeting HER3.
  • the extent of binding of an anti-HER3 antibody to an unrelated, non-HER3 protein is less than about 10% of the binding of the antibody to HER3 as measured, e.g, by a radioimmunoassay (RIA), BIACORETM (using recombinant HER3 as the analyte and antibody as the ligand, or vice versa), or other binding assays known in the art.
  • RIA radioimmunoassay
  • BIACORETM using recombinant HER3 as the analyte and antibody as the ligand, or vice versa
  • an antibody that binds to HER3 has a dissociation constant (KD) of ⁇ 1 mM, ⁇ 100 nM, ⁇ 10 nM, ⁇ 1 nM, ⁇ 0.1 nM, ⁇ 10 pM, ⁇ 1 pM, or ⁇ 0.1 pM.
  • KD dissociation constant
  • antigen binding fragment refers to a portion of an intact antibody and refers to the antigenic determining variable regions of an intact antibody. It is known in the art that the antigen binding function of an antibody can be performed by fragments of a full- length antibody. Examples of antibody fragments include, but are not limited to Fab, Fab', F(ab')2, and Fv fragments, linear antibodies, single chain antibodies, and multispecific antibodies formed from antibody fragments.
  • a “monoclonal antibody” refers to a homogeneous antibody population involved in the highly specific recognition and binding of a single antigenic determinant, or epitope.
  • polyclonal antibodies that typically include different antibodies directed against different antigenic determinants.
  • the term "monoclonal antibody” encompasses both intact and full-length monoclonal antibodies as well as antibody fragments (such as Fab, Fab', F(ab')2, Fv), single chain (scFv) mutants, fusion proteins comprising an antibody portion, and any other modified immunoglobulin molecule comprising an antigen recognition site.
  • monoclonal antibody refers to such antibodies made in any number of ways including, but not limited to, by hybridoma, phage selection, recombinant expression, and transgenic animals.
  • humanized antibody refers to an antibody derived from a non-human (e.g., murine) immunoglobulin, which has been engineered to contain minimal non-human (e.g. , murine) sequences.
  • humanized antibodies are human immunoglobulins in which residues from the complementary determining region (CDR) are replaced by residues from the CDR of a non-human species (e.g, mouse, rat, rabbit, or hamster) that have the desired specificity, affinity, and capability (Jones et al. , 1986, Nature , 321 :522-525;
  • Fv framework region (FW) residues of a human immunoglobulin are replaced with the corresponding residues in an antibody from a non-human species that has the desired specificity, affinity, and capability.
  • the humanized antibody can be further modified by the substitution of additional residues either in the Fv framework region and/or within the replaced non-human residues to refine and optimize antibody specificity, affinity, and/or capability.
  • the humanized antibody will comprise substantially all of at least one, and typically two or three, variable domains containing all or substantially all of the CDR regions that correspond to the non-human immunoglobulin whereas all or substantially all of the FR regions are those of a human immunoglobulin consensus sequence.
  • the humanized antibody can also comprise at least a portion of an immunoglobulin constant region or domain (Fc), typically that of a human immunoglobulin. Examples of methods used to generate humanized antibodies are described in U.S. Pat. Nos. 5,225,539 or 5,639,641.
  • variable region of an antibody refers to the variable region of the antibody light chain or the variable region of the antibody heavy chain, either alone or in combination.
  • the variable regions of the heavy and light chain each consist of four framework regions (FW) connected by three complementarity determining regions (CDRs) also known as hypervariable regions.
  • the CDRs in each chain are held together in close proximity by the FW regions and, with the CDRs from the other chain, contribute to the formation of the antigen-binding site of antibodies.
  • There are at least two techniques for determining CDRs (1) an approach based on cross-species sequence variability (; i.e ., Rabat et al.
  • the Kabat numbering system is generally used when referring to a residue in the variable domain (approximately residues 1-107 of the light chain and residues 1-113 of the heavy chain) (e.g .,, Kabat et al ., Sequences of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md. (1991)).
  • amino acid position numbering refers to the numbering system used for heavy chain variable domains or light chain variable domains of the compilation of antibodies in Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md. (1991). Using this numbering system, the actual linear amino acid sequence can contain fewer or additional amino acids corresponding to a shortening of, or insertion into, a FW or CDR of the variable domain.
  • a heavy chain variable domain can include a single amino acid insert (residue 52a according to Kabat) after residue 52 of H2 and inserted residues (e.g., residues 82a, 82b, and 82c, etc. according to Kabat) after heavy chain FW residue 82.
  • the end of the Chothia CDR-H1 loop when numbered using the Kabat numbering convention varies between H32 and H34 depending on the length of the loop (this is because the Kabat numbering scheme places the insertions at H35A and H35B; if neither 35 A nor 35B is present, the loop ends at 32; if only 35 A is present, the loop ends at 33; if both 35A and 35B are present, the loop ends at 34).
  • the AbM hypervariable regions represent a compromise between the Kabat CDRs and Chothia structural loops, and are used by Oxford Molecular's AbM antibody modeling software. See Table 3, above.
  • IMGT (ImMunoGeneTics) also provides a numbering system for the
  • immunoglobulin variable regions including the CDRs. See e.g ., Lefranc, M.P. et al ., Dev. Comp. Immunol. 27: 55-77(2003), which is herein incorporated by reference.
  • the IMGT numbering system was based on an alignment of more than 5,000 sequences, structural data, and characterization of hypervariable loops and allows for easy comparison of the variable and CDR regions for all species.
  • VH-CDR1 is at positions 26 to 35
  • VH-CDR2 is at positions 51 to 57
  • VH-CDR3 is at positions 93 to 102
  • VL- CDR1 is at positions 27 to 32
  • VL-CDR2 is at positions 50 to 52
  • VL-CDR3 is at positions 89 to 97.
  • VH CDRs sequences described correspond to the classical Kabat numbering locations, namely Kabat VH-CDR1 is at positions 31-35, VH-CDR2 is a positions 50-65, and VH-CDR3 is at positions 95-102.
  • VL-CDR2 and VL-CDR3 also correspond to classical Kabat numbering locations, namely positions 50-56 and 89-97, respectively.
  • VL-CDR1 or "light chain CDR1" correspond to sequences located at Kabat positions 23-34 in the VL (in contrast, the classical VL-CDR1 location according to the Kabat numbering schema corresponds to positions 24-34).
  • the Fc region includes the polypeptides comprising the constant region of an antibody excluding the first constant region immunoglobulin domain.
  • Fc refers to the last two constant region immunoglobulin domains of IgA, IgD, and IgG, and the last three constant region immunoglobulin domains of IgE and IgM, and the flexible hinge N- terminal to these domains.
  • IgA and IgM Fc may include the J chain.
  • Fc comprises immunoglobulin domains Cgamma2 and Cgamma3 (Cy2 and Cy3) and the hinge between Cgammal (Cyl) and Cgamma2 (Cy2)
  • the human IgG heavy chain Fc region is usually defined to comprise residues C226 or P230 to its carboxyl-terminus, wherein the numbering is according to the EU index as set forth in Kabat (Kabat et al ., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md. (1991)).
  • Fc may refer to this region in isolation, or this region in the context of an antibody, antibody fragment, or Fc fusion protein. Polymorphisms have been observed at a number of different Fc positions, including but not limited to positions 270, 272, 312, 315, 356, and 358 as numbered by the EU index, and thus slight differences between the presented sequence and sequences in the prior art may exist.
  • human antibody means an antibody produced by a human or an antibody having an amino acid sequence corresponding to an antibody produced by a human made using any technique known in the art. This definition of a human antibody includes intact or full-length antibodies, fragments thereof, and/or antibodies comprising at least one human heavy and/or light chain polypeptide such as, for example, an antibody comprising murine light chain and human heavy chain polypeptides.
  • chimeric antibodies refers to antibodies wherein the amino acid sequence of the immunoglobulin molecule is derived from two or more species.
  • the variable region of both light and heavy chains corresponds to the variable region of antibodies derived from one species of mammals (e.g ., mouse, rat, rabbit, etc) with the desired specificity, affinity, and capability while the constant regions are homologous to the sequences in antibodies derived from another (usually human) to avoid eliciting an immune response in that species.
  • YTE refers to a mutation in IgGl Fc that results in an increase in the binding to human FcRn and improves the serum half-life of the antibody having the mutation.
  • a YTE mutant comprises a combination of three mutations,
  • Binding affinity generally refers to the strength of the sum total of non- covalent interactions between a single binding site of a molecule (e.g ., an antibody) and its binding partner (e.g., an antigen). Unless indicated otherwise, as used herein, "binding affinity” refers to intrinsic binding affinity which reflects a 1 : 1 interaction between members of a binding pair (e.g, antibody and antigen).
  • the affinity of a molecule X for its partner Y can generally be represented by the dissociation constant (K D ). Affinity can be measured by common methods known in the art, including those described herein.
  • Low-affinity antibodies generally bind antigen slowly and tend to dissociate readily, whereas high-affinity antibodies generally bind antigen faster and tend to remain bound longer.
  • a variety of methods of measuring binding affinity are known in the art, any of which can be used for purposes described herein.
  • IC50 is the median inhibitory concentration of an antibody molecule. In functional assays, IC50 is the concentration that reduces a biological response by 50% of its maximum. In ligand binding studies, IC50 is the concentration that reduces receptor binding by 50% of maximal specific binding level. IC50 can be calculated by any number of means known in the art.
  • Improvement in potency can be determined by measuring, e.g, against the parent CL 16 (Clone 16) monoclonal antibody.
  • the fold improvement in potency for the antibodies or polypeptides described herein as compared to a Clone 16 antibody can be at least about 2-fold, at least about 4-fold, at least about 6-fold, at least about 8-fold, at least about 10-fold, at least about 20-fold, at least about 30-fold, at least about 40-fold, at least about 50-fold, at least about 60-fold, at least about 70-fold, at least about 80-fold, at least about 90-fold, at least about 100-fold, at least about 110- fold, at least about 120-fold, at least about 130-fold, at least about 140-fold, at least about 150- fold, at least about 160-fold, at least about 170-fold, or at least about 180-fold or more.
  • ADCC antibody-dependent cell-mediated cytotoxicity
  • FcRs Fc receptors
  • cytotoxic cells e.g, Natural Killer (NK) cells, neutrophils, and macrophages
  • NK Natural Killer
  • macrophages e.g, NK-derived cytotoxic cells
  • Specific high-affinity IgG antibodies directed to the surface of target cells "arm" the cytotoxic cells and are absolutely required for such killing. Lysis of the target cell is extracellular, requires direct cell-to-cell contact, and does not involve complement.
  • ADCC activity the cell-mediated cytotoxicity resulting from the activity of an Fc fusion protein is also referred to herein as ADCC activity.
  • a polypeptide, antibody, polynucleotide, vector, cell, or composition which is "isolated” is a polypeptide, antibody, polynucleotide, vector, cell, or composition which is in a form not found in nature.
  • Isolated polypeptides, antibodies, polynucleotides, vectors, cells or compositions include those which have been purified to a degree that they are no longer in a form in which they are found in nature.
  • an antibody, polynucleotide, vector, cell, or composition which is isolated is substantially pure.
  • subject refers to any animal (e.g ., a mammal), including, but not limited to humans, non-human primates, rodents, and the like, which is to be the recipient of a particular treatment.
  • subject and patient are used interchangeably herein in reference to a human subject.
  • composition refers to a preparation which is in such form as to permit the biological activity of the active ingredient to be effective, and which contains no additional components which are unacceptably toxic to a subject to which the composition would be administered.
  • Such composition can be sterile.
  • an “effective amount” of an antibody as disclosed herein is an amount sufficient to carry out a specifically stated purpose.
  • An “effective amount” can be determined empirically and in a routine manner, in relation to the stated purpose.
  • terapéuticaally effective amount refers to an amount of an antibody or other drug effective to "treat” a disease or disorder in a subject or mammal.
  • label when used herein refers to a detectable compound or composition which is conjugated directly or indirectly to the antibody so as to generate a "labeled" antibody.
  • the label can be detectable by itself (e.g., radioisotope labels or fluorescent labels) or, in the case of an enzymatic label, can catalyze chemical alteration of a substrate compound or composition which is detectable.
  • Terms such as “treating” or “treatment” or “to treat” or “alleviating” or “to alleviate” refer to therapeutic measures that cure, slow down, lessen symptoms of, and/or halt progression of a diagnosed pathologic condition or disorder and slow the development of a targeted pathologic condition or disorder.
  • those in need of treatment include those already with the disorder; those prone to have the disorder; and those in whom the disorder is to be prevented.
  • a subject is successfully "treated” for cancer according to the methods described herein if the patient shows, e.g ., total, partial, or transient remission of a certain type of cancer.
  • diagnosis refers to a clinical or other assessment of the condition of a subject based on observation, testing, or circumstances for identifying a subject having a disease, disorder, or condition based on the presence of at least one sign or symptom of the disease, disorder, or condition.
  • cancer refers to or describe the physiological condition in mammals that is typically characterized by unregulated cell growth.
  • cancers include but are not limited to, carcinoma including
  • adenocarcinomas lymphomas, blastomas, melanomas, sarcomas, and leukemias. More particular examples of such cancers include brain cancer, CNS cancer, squamous cell cancer, small-cell lung cancer, non-small cell lung cancer, gastrointestinal cancer, Hodgkin's and non- Hodgkin's lymphoma, pancreatic cancer, glioblastoma, glioma, cervical cancer, ovarian cancer, liver cancer such as hepatic carcinoma and hepatoma, bladder cancer, breast cancer (including hormonally mediated breast cancer, see, e.g. , Innes et al. (2006) Br. J.
  • Cancer 94: 1057-1065 colon cancer, colorectal cancer, endometrial carcinoma, myeloma (such as multiple myeloma), salivary gland carcinoma, kidney cancer such as renal cell carcinoma and Wilms' tumors, basal cell carcinoma, melanoma, prostate cancer, vulval cancer, thyroid cancer, testicular cancer, esophageal cancer, various types of head and neck cancer and cancers of mucinous origins, such as, mucinous ovarian cancer, cholangiocarcinoma (liver) and renal papillary carcinoma.
  • myeloma such as multiple myeloma
  • salivary gland carcinoma such as multiple myeloma
  • kidney cancer such as renal cell carcinoma and Wilms' tumors
  • basal cell carcinoma melanoma
  • prostate cancer vulval cancer
  • thyroid cancer testicular cancer
  • esophageal cancer various types of head and neck cancer and cancers of mucinous origins, such as, mucinous
  • carcinomas refers to cancers of epithelial cells, which are cells that cover the surface of the body, produce hormones, and make up glands. Examples of carcinomas are cancers of the skin, lung, colon, stomach, breast, prostate and thyroid gland.
  • HER3 -associated cancer is used to refer to any cancer that is completely or partially caused by, associated with, or is the result of, HER3 expression and/or activity or lack thereof.
  • a HER3 -associated cancer can be known to one of skill in the art or can be ascertained by one of skill in the art.
  • a HER3 -associated cancer is associated with HER3 expression and/or activity.
  • HER3 expression and/or activity may contribute, in combination with one or more other factors (e.g ., mutation or expression and/or activity of another gene), to
  • a HER3 -associated cancer is associated with one or more mutations of HER3.
  • the HER3 -associated cancer referred to in this disclosure is head and neck cancer.
  • the HER3 -associated cancer referred to in this disclosure is squamous cell carcinoma of the head and neck.
  • the primary tumor site is oral cavity.
  • HER3 protein activity level in a first biological sample is intended qualitatively or quantitatively measuring or estimating the activity of HER3 protein in a first biological sample either directly (e.g., by determining or estimating absolute activity level) or relatively (e.g, by comparing to the activity level in a second biological sample).
  • HER3 protein activity level in the first biological sample can be measured or estimated and compared to a standard HER3 protein activity, the standard being taken from a second biological sample obtained from an individual not having the disorder or being determined by averaging levels from a population of individuals not having the disorder or from an individual prior to treatment.
  • the "standard" HER3 protein activity level is known, it can be used repeatedly as a standard for comparison.
  • the activity level of HER3 in a biological sample is measured or estimated or compared by detecting phosphorylated HER3 in a biological sample.
  • the activity level of HER3 in a biological sample is measured or estimated or compared by detecting phosphorylated HER3 in a skin biopsy, wherein the skin is stimulated with HRG prior to or after biopsy.
  • positive therapeutic response refers to an improvement in the disease in association with the activity of these anti-HER3 binding molecules, e.g, antibodies or antigen-binding fragments, variants, or derivatives thereof, and/or an improvement in the symptoms associated with the disease.
  • an improvement in the disease can be characterized as a complete response.
  • complete response is intended an absence of clinically detectable disease with normalization of any previously test results.
  • an improvement in the disease can be categorized as being a partial response.
  • a "positive therapeutic response” encompasses a reduction or inhibition of the progression and/or duration of cancer, the reduction or amelioration of the severity of cancer, and/or the amelioration of one or more symptoms thereof resulting from the administration of an anti-HER3 binding molecule described herein.
  • such terms refer to one, two or three or more results following the administration of anti-HER3 binding molecules described herein: (1) a stabilization, reduction or elimination of the cancer cell population; (2) a stabilization or reduction in cancer growth; (3) an impairment in the formation of cancer; (4) eradication, removal, or control of primary, regional and/or metastatic cancer; (5) a reduction in mortality; (6) an increase in disease-free, relapse-free, progression- free, and/or overall survival, duration, or rate; (7) an increase in the response rate, the durability of response, or number of patients who respond or are in remission; (8) a decrease in hospitalization rate, (9) a decrease in hospitalization lengths, (10) the size of the cancer is maintained and does not increase or increases by less than 10%, preferably less than 5%, preferably less than 4%, preferably less than 2%, and (12) an increase in the number of patients in remission.
  • sample from a patient or "biological sample” is intended any biological sample obtained from an individual, cell line, tissue culture, or other source of cells potentially expressing HER3.
  • tissue biopsies and body fluids from mammals are well known in the art.
  • a combination therapy can provide "synergy” and prove “synergistic", i.e., the effect achieved when the active ingredients used together is greater than the sum of the effects that results from using the compounds separately.
  • a synergistic effect can be attained when the active ingredients are: (1) co-formulated and administered or delivered simultaneously in a combined, unit dosage formulation; (2) delivered by alternation or in parallel as separate formulations; or (3) by some other regimen.
  • a synergistic effect can be attained when the compounds are administered or delivered sequentially, e.g ., by different injections in separate syringes.
  • an effective dosage of each active ingredient is administered sequentially, i.e., serially
  • effective dosages of two or more active ingredients are administered together.
  • Polynucleotide or “nucleic acid,” as used interchangeably herein, refer to polymers of nucleotides of any length, and include DNA and RNA.
  • the nucleotides can be deoxyribonucleotides, ribonucleotides, modified nucleotides or bases, and/or their analogs, or any substrate that can be incorporated into a polymer by DNA or RNA polymerase.
  • a polynucleotide can comprise modified nucleotides, such as methylated nucleotides and their analogs. The preceding description applies to all polynucleotides referred to herein, including RNA and DNA.
  • vector means a construct, which is capable of delivering, and in some aspects, expressing, one or more gene(s) or sequence(s) of interest in a host cell.
  • vectors include, but are not limited to, viral vectors, naked DNA or RNA expression vectors, plasmid, cosmid or phage vectors, DNA or RNA expression vectors associated with cationic condensing agents, DNA or RNA expression vectors encapsulated in liposomes, and certain eukaryotic cells, such as producer cells.
  • polypeptide polypeptide
  • peptide protein
  • the terms “polypeptide,” “peptide,” and “protein” are used interchangeably herein to refer to polymers of amino acids of any length.
  • the polymer can be linear or branched, it can comprise modified amino acids, and it can be interrupted by non-amino acids.
  • the terms also encompass an amino acid polymer that has been modified naturally or by intervention; for example, disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, or any other manipulation or modification, such as conjugation with a labeling component.
  • polypeptides containing one or more analogs of an amino acid including, for example, unnatural amino acids, etc.
  • the polypeptides described herein are based upon antibodies, in certain aspects, the polypeptides can occur as single chains or associated chains.
  • nucleic acids or polypeptides refer to two or more sequences or subsequences that are the same or have a specified percentage of nucleotides or amino acid residues that are the same, when compared and aligned (introducing gaps, if necessary) for maximum correspondence, not considering any conservative amino acid substitutions as part of the sequence identity.
  • percent identity can be measured using sequence comparison software or algorithms or by visual inspection.
  • sequence alignment algorithm is the algorithm described in Karlin et al, 1990, Proc. Natl. Acad. Sci., 87:2264-2268, as modified in Karlin et al., 1993, Proc. Natl. Acad. Sci., 90:5873-5877, and incorporated into the NBLAST and XBLAST programs (Altschul et al., 1991, Nucleic Acids Res., 25:3389-3402).
  • Gapped BLAST can be used as described in Altschul et al, 1997, Nucleic Acids Res.
  • BLAST-2 Altschul et al., 1996, Methods in Enzymology, 266:460-480
  • ALIGN ALIGN-2
  • ALIGN-2 Genentech, South San Francisco, California
  • Megalign Megalign
  • the GAP program in the GCG software package which incorporates the algorithm of Needleman and Wunsch (J. Mol. Biol. (48):444-453 (1970)) can be used to determine the percent identity between two amino acid sequences (e.g, using either a BLOSUM 62 matrix or a PAM250 matrix, and a gap weight of 16, 14, 12, 10, 8, 6, or 4 and a length weight of 1, 2, 3, 4, 5).
  • the percent identity between nucleotide or amino acid sequences is determined using the algorithm of Myers and Miller (CABIOS, 4: 11-17 (1989)).
  • the percent identity can be determined using the ALIGN program (version 2.0) and using a PAM120 with residue table, a gap length penalty of 12 and a gap penalty of 4.
  • Appropriate parameters for maximal alignment by particular alignment software can be determined by one skilled in the art. In certain aspects, the default parameters of the alignment software are used.
  • the percentage identity "X" of a first amino acid sequence to a second sequence amino acid is calculated as 100 x (Y/Z), where Y is the number of amino acid residues scored as identical matches in the alignment of the first and second sequences (as aligned by visual inspection or a particular sequence alignment program) and Z is the total number of residues in the second sequence. If the length of a first sequence is longer than the second sequence, the percent identity of the first sequence to the second sequence will be higher than the percent identity of the second sequence to the first sequence.
  • a "conservative amino acid substitution” is one in which one amino acid residue is replaced with another amino acid residue having a similar side chain.
  • Families of amino acid residues having similar side chains have been defined in the art, including basic side chains (e.g ., lysine, arginine, histidine), acidic side chains (e.g, aspartic acid, glutamic acid), uncharged polar side chains (e.g, asparagine, glutamine, serine, threonine, tyrosine, cysteine), nonpolar side chains (e.g, glycine, alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), beta-branched side chains (e.g, threonine, valine, isoleucine) and aromatic side chains (e.g, tyrosine, phenylalanine, tryptophan, histidine).
  • basic side chains e.g ly
  • substitution of a phenylalanine for a tyrosine is a conservative substitution.
  • conservative substitutions in the sequences of the polypeptides and antibodies described herein do not abrogate the binding of the polypeptide or antibody containing the amino acid sequence, to the antigen(s), i.e., the HER3 to which the polypeptide or antibody binds.
  • Methods of identifying nucleotide and amino acid conservative substitutions which do not eliminate antigen binding are well-known in the art (see, e.g., Brummell el al, Biochem. 32: 1180-1 187 (1993); Kobayashi et al., Protein Eng. 12(10):879-884 (1999); and Burks et al., Proc. Natl. Acad. Sci. USA 94:.412-417 (1997)).
  • VL and heavy chain (VH) variable regions refers to a composite or genericized VL or VH sequence defined based on information as to which amino acid residues within the VL or VH chain are amenable to modification without detriment to antigen binding.
  • certain amino acid positions are occupied by one of multiple possible amino acid residues at that position. For example, if an arginine (R) or a serine (S) occur at a particular position, then that particular position within the consensus sequence can be either arginine or serine (R or S).
  • Consensus sequences for VH and VL chain can be defined, for example, by in vitro affinity maturation (e.g, randomizing every amino acid position in a certain CDR using degenerate coding primers), by scanning mutagenesis (e.g, alanine scanning mutagenesis) of amino acid residues within the antibody CDRs, or any other methods known in the art, followed by evaluation of the binding of the mutants to the antigen to determine whether the mutated amino acid position affects antigen binding.
  • mutations are introduced in the CDR regions.
  • mutations are introduced in framework regions.
  • mutations are introduced in CDR and framework regions.
  • a method of treating a HER3 -associated cancer comprising administering to a patient diagnosed with said cancer a therapeutically effective amount of a HER3 inhibitor, for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL16), wherein the cancer is characterized in that cells from the cancer comprise a mutation in the FAT1 gene.
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL16)
  • the cancer is characterized in that cells from the cancer comprise a mutation in the FAT1 gene.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • a method for treating a patient with a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16), wherein the patient is suffering from a HER3 -associated cancer, the method comprising the steps of: determining whether cells of the cancer comprise a mutation in the FAT1 gene; and if the cells of the cancer comprise a mutation in the FAT1 gene, then administering a therapeutically effective amount of the HER3 inhibitor to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • a method of treating a HER3 -associated cancer in a patient diagnosed therewith comprising the steps of: determining whether cells of the cancer comprise a mutation in the FAT1 gene; and if the cells of the cancer comprise a mutation in the FAT1 gene, then administering a therapeutically effective amount of the HER3 inhibitor, for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16), to the patient.
  • a therapeutically effective amount of the HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16)
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the step of determining whether cells of the cancer comprise a mutation in the FAT1 gene is performed by: obtaining a sample, wherein the sample comprises or is derived from the patient’s cancerous cells; and performing an assay on the sample to determine whether a mutation in the FAT1 gene is present in the sample.
  • the step of determining whether cells of the cancer comprise a mutation in the FAT1 gene is performed by performing an assay on a sample that comprises or is derived from the patient’s cancerous cells to determine whether a mutation in the FAT1 gene is present in the sample.
  • a method of treating a HER3 -associated cancer comprising administering to a patient diagnosed with said cancer a therapeutically effective amount of a HER3 inhibitor, for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16), wherein the cancer is characterized in that cells from the cancer comprise at least one mutation in at least one NOTCH gene.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the method further comprises administering to the patient an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab).
  • a method for treating a patient with a HER3 inhibitor for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL16), wherein the patient is suffering from a HER3 -associated cancer, the method comprising the steps of: determining whether cells of the cancer comprise at least one mutation in at least one NOTCH gene; and if the cells of the cancer comprise at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of the HER3 inhibitor to the patient.
  • the at least one mutation in at least one NOTCH gene is one or more loss- of-function mutations.
  • the method further comprises administering to the patient an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab).
  • a method of treating a HER3 -associated cancer in a patient diagnosed therewith comprising the steps of: determining whether cells of the cancer comprise at least one mutation in at least one NOTCH gene; and if the cells of the cancer comprise at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of the HER3 inhibitor, for example, an anti- HER3 antibody (e.g, 2C2-YTE or CL 16), to the patient.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the method further comprises administering to the patient an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab).
  • the step of determining whether cells of the cancer comprise at least one mutation in at least one NOTCH gene is performed by: obtaining a sample, wherein the sample comprises or is derived from the patient’s cancerous cells; and performing an assay on the sample to determine whether at least one mutation in at least one NOTCH gene is present in the sample.
  • the step of determining whether cells of the cancer comprise at least one mutation in at least one NOTCH gene is performed by performing an assay on a sample that comprises or is derived from the patient’s cancerous cells to determine whether at least one mutation in at least one NOTCH gene is present in the sample.
  • a method of treating a HER3 -associated cancer comprising administering to a patient diagnosed with said cancer a therapeutically effective amount of a HER3 inhibitor, for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL16), wherein the cancer is characterized in that cells from the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene.
  • a method of treating a HER3 -associated cancer comprising administering to a patient diagnosed with said cancer a therapeutically effective amount of a HER3 inhibitor, for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16), wherein the cancer is characterized in that tissue from the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene.
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16)
  • the cancer is characterized in that tissue from the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene.
  • the mutation in the FAT1 gene is a loss- of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the method further comprises administering to the patient an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab).
  • a method for treating a patient with a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16), wherein the patient is suffering from a HER3 -associated cancer, the method comprising the steps of: determining whether cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and if the cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of the HER3 inhibitor to the patient.
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16)
  • the patient is suffering from a HER3 -associated cancer
  • the method comprising the steps of: determining whether cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and if the cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in
  • a method for treating a patient with a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16), wherein the patient is suffering from a HER3 -associated cancer, the method comprising the steps of: determining whether tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and if the tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the method further comprises administering to the patient an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab).
  • a method of treating a HER3 -associated cancer in a patient diagnosed therewith comprising the steps of: determining whether cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and if the cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of the HER3 inhibitor, for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL16), to the patient.
  • a method of treating a HER3- associated cancer in a patient diagnosed therewith comprising the steps of:
  • tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and if the tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of the HER3 inhibitor, for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL 16), to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the method further comprises administering to the patient an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab).
  • the step of determining whether tissue or cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene is performed by: obtaining a sample, wherein the sample comprises or is derived from the patient’s cancerous cells; and performing an assay on the sample to determine whether a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene are present in the sample.
  • the step of determining whether tissue or cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene is performed by performing an assay on a sample that comprises or is derived from the patient’s cancerous cells to determine whether a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene are present in the sample.
  • a method of treating a HER3 -associated cancer in a patient diagnosed therewith comprising: (1) administering to the patient an HER3 inhibitor, for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16), and (2) administering to the patient a FAT1 inhibitor.
  • an HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16)
  • administering to the patient a FAT1 inhibitor.
  • the FAT1 inhibitor may be a pharmaceutical agent that reduces the expression level of FAT 1, a pharmaceutical agent that inhibits the activity of FAT 1, or a pharmaceutical agent that inhibits FAT1 signaling, relative to the FAT1 expression level, FAT1 activity level, or FAT1 signaling level (as the case may be) without treatment with the FAT1 inhibitor.
  • the FAT1 inhibitor is an antibody, a small molecule, or an
  • the FAT1 inhibitor is an antibody (for example, an anti-FATl antibody). In a particular embodiment, the FAT1 inhibitor is a monoclonal antibody (for example, an anti-FATl monoclonal antibody). In another specific embodiment, the FAT1 inhibitor is a small molecule. In another specific embodiment, the FAT1 inhibitor is an oligonucleotide. In one embodiment, the FAT1 inhibitor is an aptamer (for example, a FAT1 aptamer). In another embodiment, the FAT1 inhibitor is an shRNA (for example, a FAT1 shRNA). In another embodiment, the FAT1 inhibitor is an miRNA (for example, a FAT1 miRNA). In another embodiment, the FAT1 inhibitor is an siRNA (for example, a FAT1 siRNA). In another embodiment, the FAT1 inhibitor is an antisense DNA (for example, a FAT1 antisense DNA).
  • the FAT1 inhibitor is administered concurrently with the HER3 inhibitor. In other embodiments, the FAT1 inhibitor is administered sequentially with the HER3 inhibitor. In a specific embodiment, the FAT1 inhibitor is administered before the HER3 inhibitor (e.g, about 0.5 day, about 1 day, about 2 days, about 3 days, about 4 days, about 5 days, about 6 days, or about a week before). In another specific embodiment, the FAT1 inhibitor is administered after the HER3 inhibitor (e.g, about 0.5 day, about 1 day, about 2 days, about 3 days, about 4 days, about 5 days, about 6 days, or about a week after). One or both of the HER3 inhibitor and the FAT1 inhibitor can be administered once, or more than once, e.g. , in an alternating manner.
  • the HER3-associated cancer is head and neck cancer, breast cancer, ovarian cancer, prostate cancer, liver cancer, kidney cancer, bladder cancer, pancreatic cancer, brain cancer, hematopoietic neoplasms, retinoblastoma, melanoma, colorectal cancer, gastric cancers, or lung cancer.
  • the HER3- associated cancer is head and neck cancer.
  • the HER3- associated cancer is squamous cell carcinoma of the head and neck.
  • the primary tumor site is oral cavity.
  • a method of treating a HER3 -associated cancer comprising administering to a patient diagnosed with said cancer a therapeutically effective amount of a HER3 inhibitor, wherein the HER3 -associated cancer is head and neck cancer, and wherein the primary tumor site is oral cavity.
  • the HER3- associated cancer is squamous cell carcinoma of the head and neck.
  • the HER3 -associated cancer is characterized in that tissue or cells from the cancer also express a neuregulin or neuregulin fusion protein, and/or comprise a mutation in the BRAF and/or MEK gene.
  • the cancer is characterized in that tissue or cells from the cancer also express a neuregulin or neuregulin fusion protein (e.g. NRG1 and/or NRG2) at high levels.
  • a neuregulin or neuregulin fusion protein e.g. NRG1 and/or NRG2
  • the neuregulin is NRG1.
  • the neuregulin is NRG2 (e.g, NRG2A and/or NRG2B).
  • the cancer is characterized by a BRAF mutation (e.g, V600E or V600K).
  • the cancer is resistant to treatment with a BRAF inhibitor (e.g, vemurafenib or dabrafenib).
  • the cancer is resistant to treatment with a MEK inhibitor (e.g, selumetinib or trametinib).
  • the cancer is resistant to treatment with a BRAF inhibitor (e.g, vemurafenib or dabrafenib) and a MEK inhibitor (e.g, selumetinib or trametinib).
  • the cancer is melanoma.
  • the cancer is B-Raf mutated melanoma. In certain embodiments, the cancer is thyroid cancer. In specific embodiments, the cancer is B-Raf mutated thyroid cancer. In certain embodiments, the cancer is colorectal cancer. In specific embodiments, the cancer is B-Raf mutated colorectal cancer. In certain embodiments, the cancer is lung cancer. In specific embodiments, the cancer is B-Raf mutated lung cancer. In certain embodiments, the cancer is hairy cell leukemia. In specific embodiments, the cancer is B-Raf mutated hairy cell leukemia. In particular embodiments, the lung cancer is non- small cell lung carcinoma.
  • the cancer is a squamous cell carcinoma of the head and neck (SCCHN).
  • the cancer is ovarian cancer.
  • the cancer is pancreatic cancer.
  • the cancer is bladder cancer.
  • the cancer is testicular cancer.
  • the cancer is endometrial cancer.
  • the cancer is hepatocellular carcinoma.
  • the cancer is breast cancer.
  • the cancer is gastric cancer.
  • the cancer is prostate cancer.
  • the cancer is characterized by a HER2 or HER3 mutation.
  • the HER3 inhibitor is a monoclonal, human anti- HER3 antibody, which comprises an antibody VL of SEQ ID NO:3 and a human lambda light chain constant region, and an antibody VH of SEQ ID NO: 2 and a human IgGl constant region comprising amino acid substitutions relative to a wild-type human IgG constant domain at positions 252, 254, and 256, wherein the numbering is according to the EU index as set forth in Rabat, and wherein the amino acid at position 252 is substituted with Tyrosine (Y), the amino acid at position 254 is substituted with Threonine (T), and the amino acid at position 256 is substituted with Glutamic acid (E).
  • Y Tyrosine
  • T Threonine
  • E Glutamic acid
  • an inhibitor described herein e.g ., an anti-HER3 antibody or antigen-binding fragment thereof
  • which specifically binds to HER3 comprises a VH or VL of CL16 or 2C2 with one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15 or 20) amino acid substitutions (e.g, conservative amino acid substitutions), deletions, or additions.
  • the VH and VL of CL16 comprise SEQ ID NOs: 2 and 1, respectively
  • the VH and VL of 2C2 comprise SEQ ID NOs: 2 and 3, respectively.
  • an inhibitor described herein e.g, an anti-HER3 antibody or antigen-binding fragment thereof
  • which specifically binds to HER3 comprises a heavy chain constant region or fragment thereof which is an IgG constant region.
  • an inhibitor described herein e.g, an anti-HER3 antibody or antigen-binding fragment thereof
  • which specifically binds to HER3 comprises an IgG constant region with one or more (e.g ., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15 or 20) amino acid substitutions (e.g ., conservative amino acid substitutions), deletions, or additions.
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL16), for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, comprising: determining whether cells of the cancer comprise a mutation in the FAT1 gene; and if the cells of the cancer comprise a mutation in the FAT1 gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16), for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the cancer is characterized in that cells from the cancer comprise a mutation in the FAT1 gene, said method comprising administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16), for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, comprising: determining whether cells of the cancer comprise at least one mutation in at least one NOTCH gene; and if the cells of the cancer comprise at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the method of threating further comprises administering to the patient an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab).
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16), for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the cancer is characterized in that cells from the cancer comprise at least one mutation in at least one NOTCH gene, said method comprising administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of- function mutations.
  • the method of treating further comprises administering to the patient an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab).
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g ., 2C2-YTE or CL16), for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, comprising: determining whether cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and if the cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • an anti-HER3 antibody e.g ., 2C2-YTE or CL16
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL16), for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, comprising: determining whether tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and if the tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the method of treating further comprises administering to the patient an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab).
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16), for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the cancer is characterized in that cells from the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, said method comprising administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • an anti-HER3 antibody e.g, 2C2-YTE or CL16
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16), for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the cancer is characterized in that tissue from the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, said method comprising administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the method of treating further comprises administering to the patient an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab).
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g ., 2C2-YTE or CL16), for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the HER3 -associated cancer is head and neck cancer, and wherein the primary tumor site is oral cavity.
  • an anti-HER3 antibody e.g ., 2C2-YTE or CL16
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL16), for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the anti-HER3 antibody is administered in combination with a FAT 1 inhibitor.
  • an anti-HER3 antibody e.g., 2C2-YTE or CL16
  • the FAT1 inhibitor may be a pharmaceutical agent that reduces the expression level of FAT 1, a pharmaceutical agent that inhibits the activity of FAT 1, or a pharmaceutical agent that inhibits FAT1 signaling, relative to the FAT1 expression level, FAT1 activity level, or FAT1 signaling level (as the case may be) without treatment with the FAT1 inhibitor.
  • the FAT1 inhibitor is an antibody, a small molecule, or an
  • the FAT1 inhibitor is an antibody (for example, an anti-FATl antibody). In a particular embodiment, the FAT1 inhibitor is a monoclonal antibody (for example, an anti-FATl monoclonal antibody). In another specific embodiment, the FAT1 inhibitor is a small molecule. In another specific embodiment, the FAT1 inhibitor is an oligonucleotide. In one embodiment, the FAT1 inhibitor is an aptamer (for example, a FAT1 aptamer). In another embodiment, the FAT1 inhibitor is an shRNA (for example, a FAT1 shRNA). In another embodiment, the FAT1 inhibitor is an miRNA (for example, a FAT1 miRNA).
  • the FAT1 inhibitor is an siRNA (for example, a FAT1 siRNA).
  • the FAT1 inhibitor is an antisense DNA (for example, a FAT1 antisense DNA).
  • the FAT1 inhibitor is administered concurrently with the HER3 inhibitor.
  • the FAT1 inhibitor is administered sequentially with the HER3 inhibitor.
  • the FAT1 inhibitor is administered before the HER3 inhibitor (e.g, about 0.5 day, about 1 day, about 2 days, about 3 days, about 4 days, about 5 days, about 6 days, or about a week before).
  • the FAT1 inhibitor is administered after the HER3 inhibitor (e.g, about 0.5 day, about 1 day, about 2 days, about 3 days, about 4 days, about 5 days, about 6 days, or about a week after).
  • the HER3 inhibitor and the FAT1 inhibitor can be administered once, or more than once, e.g, in an alternating manner.
  • the HER3 inhibitor is an anti-HER3 antibody or antigen binding fragment thereof.
  • the anti-HER3 antibody or antigen-binding fragment thereof specifically binds to the same HER3 epitope as an antibody or antigen binding fragment thereof comprising the heavy chain variable region (VH) and light chain variable region (VL) of CL16 or 2C2.
  • the VH and VL of CL16 comprise SEQ ID NOs: 2 and 1, respectively, and the VH and VL of 2C2 comprise SEQ ID NOs: 2 and 3, respectively.
  • the anti-HER3 antibody or antigen-binding fragment thereof is affinity matured.
  • FAT1 and/or NOTCH mutations may be detected in accordance with any method described in this disclosure in DNA, mRNA or protein from samples by means well known in the art, for example, by means described in Section VI below.
  • a method of determining whether a patient diagnosed with a HER3 -associated cancer is indicated as likely to be responsive to treatment with a HER3 inhibitor for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL16), comprising detecting a mutation in the FAT1 gene in a sample, wherein the sample comprises or is derived from the patient’s cancerous cells, and wherein the presence of a mutation in the FAT1 gene in the sample indicates that the patient is likely to be responsive to treatment with the HER3 inhibitor.
  • the mutation in the FAT1 gene is a loss-of- function mutation.
  • the disclosure provides methods for determining whether a patient diagnosed with a HER3 -associated cancer is indicated as likely to be responsive to treatment with a combination of a HER3 inhibitor (for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL 16)) and an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab), comprising detecting a mutation in the FAT1 gene in a sample, wherein the sample comprises or is derived from the patient’s cancerous cells.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the disclosure provides methods for determining whether a patient diagnosed with a HER3 -associated cancer is indicated as likely to be responsive to treatment with a combination of a HER3 inhibitor (for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16)) and a B-Raf inhibitor, comprising detecting a mutation in the FAT1 gene in a sample, wherein the sample comprises or is derived from the patient’s cancerous cells.
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16)
  • a B-Raf inhibitor comprising detecting a mutation in the FAT1 gene in a sample, wherein the sample comprises or is derived from the patient’s cancerous cells.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the disclosure provides methods for determining whether a patient diagnosed with a HER3 -associated cancer (e.g, thyroid cancer or melanoma) is indicated as likely to be responsive to treatment with a combination of a HER3 inhibitor (for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16)) and a MEK inhibitor, comprising detecting a mutation in the FAT1 gene in a sample, wherein the sample comprises or is derived from the patient’s cancerous cells.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the cancer is resistant to treatment with a MEK inhibitor (e.g, selumetinib or trametinib).
  • a method of determining whether a patient diagnosed with a HER3 -associated cancer is indicated as likely to be responsive to treatment with a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16), comprising detecting at least one mutation in at least one NOTCH gene in a sample, wherein the sample comprises or is derived from the patient’s cancerous cells, and wherein the presence of at least one mutation in at least one NOTCH gene in the sample indicates that the patient is likely to be responsive to treatment with the HER3 inhibitor.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the disclosure provides methods for determining whether a patient diagnosed with a HER3 -associated cancer is indicated as likely to be responsive to treatment with a combination of a HER3 inhibitor (for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL 16)) and an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab), comprising detecting at least one mutation in at least one NOTCH gene in a sample, wherein the sample comprises or is derived from the patient’s cancerous cells.
  • the at least one mutation in at least one NOTCH gene is one or more loss- of-function mutations.
  • the disclosure provides methods for determining whether a patient diagnosed with a HER3 -associated cancer is indicated as likely to be responsive to treatment with a combination of a HER3 inhibitor (for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL 16)) and a B-Raf inhibitor, comprising detecting at least one mutation in at least one NOTCH gene in a sample, wherein the sample comprises or is derived from the patient’s cancerous cells.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the disclosure provides methods for determining whether a patient diagnosed with a HER3 -associated cancer (e.g, thyroid cancer or melanoma) is indicated as likely to be responsive to treatment with a combination of a HER3 inhibitor (for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL16)) and a MEK inhibitor, comprising detecting at least one mutation in at least one NOTCH gene in a sample, wherein the sample comprises or is derived from the patient’s cancerous cells.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the cancer is resistant to treatment with a MEK inhibitor (e.g, selumetinib or trametinib).
  • a method of determining whether a patient diagnosed with a HER3 -associated cancer is indicated as likely to be responsive to treatment with a HER3 inhibitor for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL16), comprising detecting a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene in a sample, wherein the sample comprises or is derived from the patient’s cancerous cells, and wherein the presence of a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene in the sample indicates that the patient is likely to be responsive to treatment with the HER3 inhibitor.
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL16)
  • the mutation in the FAT1 gene is a loss-of-function mutation. In certain embodiments, the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations. In certain embodiments, the mutation in the FAT1 gene is a loss-of-function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the disclosure provides methods for determining whether a patient diagnosed with a HER3 -associated cancer is indicated as likely to be responsive to treatment with a combination of a HER3 inhibitor (for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL 16)) and an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab), comprising detecting a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene in a sample, wherein the sample comprises or is derived from the patient’s cancerous cells.
  • the mutation in the FAT1 gene is a loss-of- function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the disclosure provides methods for determining whether a patient diagnosed with a HER3 -associated cancer is indicated as likely to be responsive to treatment with a combination of a HER3 inhibitor (for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16)) and a B-Raf inhibitor, comprising detecting a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene in a sample, wherein the sample comprises or is derived from the patient’s cancerous cells.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the disclosure provides methods for determining whether a patient diagnosed with a HER3 -associated cancer (e.g, thyroid cancer or melanoma) is indicated as likely to be responsive to treatment with a combination of a HER3 inhibitor (for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16)) and a MEK inhibitor, comprising detecting a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene in a sample, wherein the sample comprises or is derived from the patient’s cancerous cells.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of- function mutations.
  • the mutation in the FAT1 gene is a loss-of- function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the cancer is resistant to treatment with a MEK inhibitor (e.g ., selumetinib or trametinib).
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the loss-of-function mutation in the FAT1 gene is in the protein coding region of the FAT1 gene.
  • the loss-of-function mutation in the FAT1 gene is present in a promoter (regulatory) region of the FAT1 gene.
  • the loss-of-function mutation in the FAT1 gene is in a post-translational region of the FAT1 gene.
  • the loss-of-function mutation in the FAT1 gene is present in an intron sequence of the FAT1 gene.
  • the loss-of-function mutation in the FAT1 gene is present in an exon sequence of the FAT1 gene. In certain embodiments, the loss-of-function mutation in the FAT1 gene is present within an intron-exon boundary sequence of the FAT1 gene.
  • the loss-of-function mutation in the FAT1 gene is a dominant mutation. In some embodiments, the loss-of-function mutation in the FAT1 gene is a recessive mutation.
  • the loss-of function mutation in the FAT1 gene results in the FAT1 polypeptide exhibiting no activity, or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, or 80%, 85% of normal activity.
  • the loss-of-function mutation in the FAT1 gene results in an abnormal level of expression of the FAT1 gene, e.g., no expression, or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, or 80% of normal FAT1 gene expression levels (FAT1 RNA and/or FAT1 polypeptide levels).
  • an abnormal level of expression of the FAT1 gene e.g., no expression, or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, or 80% of normal FAT1 gene expression levels (FAT1 RNA and/or FAT1 polypeptide levels).
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the loss-of-function mutation in a NOTCH gene is in the protein coding region of a NOTCH gene.
  • the loss-of-function mutation in a NOTCH gene is present in a promoter (regulatory) region of a NOTCH gene.
  • the loss-of-function mutation in a NOTCH gene is in a post-translational region of a NOTCH gene.
  • the loss-of-function mutation in a NOTCH gene is present in an intron sequence of a NOTCH gene.
  • the loss-of-function mutation in a NOTCH gene is present in an exon sequence of a NOTCH gene. In certain embodiments, the loss-of-function mutation in a NOTCH gene is present within an intron-exon boundary sequence of a NOTCH gene. In certain embodiments, the loss-of-function mutation in a NOTCH gene is a splice acceptor mutation.
  • a loss of function mutation in a NOTCH gene may be a truncation mutation in the protein coding region of the gene.
  • Examples of particular truncations in NOTCH2 genes include without limitation: R91*, W16* and splice acceptor mutations ( e.g ., in Exon 6).
  • R91* denotes a mutation that changes R91 of NP 077719.2 to a stop codon (for example, a CGA to TGA mutation at the DNA level).
  • W16* denotes a mutation that changes W16 of NP 077719.2 to a stop codon (for example, a TGG to TGA mutation at the DNA level).
  • the loss-of-function mutation in a NOTCH gene is a dominant mutation. In some embodiments, the loss-of-function mutation in the NOTCH gene is a recessive mutation.
  • the loss-of function mutation in a NOTCH gene results in a NOTCH polypeptide exhibiting no activity, or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, or 80%, 85% of normal activity.
  • the loss-of-function mutation in a NOTCH gene results in an abnormal level of expression of a NOTCH gene, e.g., no expression, or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, or 80% of normal NOTCH gene expression levels (NOTCH RNA and/or NOTCH polypeptide levels).
  • an abnormal level of expression of a NOTCH gene e.g., no expression, or about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, or 80% of normal NOTCH gene expression levels (NOTCH RNA and/or NOTCH polypeptide levels).
  • one or more measuring steps are performed in vitro.
  • such methods for determining whether a patient diagnosed with a HER3 -associated cancer comprise a first step of obtaining the sample from a tumor from the patient.
  • the method comprises an additional step of administering to the patient a therapeutically effective amount of a HER3 inhibitor.
  • the HER3 inhibitor is an anti-HER3 antibody or antigen binding fragment thereof (e.g. 2C2-YTE).
  • the HER3 inhibitor is a 2C2-YTE anti-HER3 antibody.
  • the HER3 inhibitor is a 2C2-YTE anti-HER3.
  • the anti-HER3 antibody is a monoclonal, human anti- HER3 antibody, which comprises an antibody VL of SEQ ID NO:3 and a human lambda light chain constant region, and an antibody VH of SEQ ID NO: 2 and a human IgGl constant region comprising amino acid substitutions relative to a wild-type human IgG constant domain at positions 252, 254, and 256, wherein the numbering is according to the EU index as set forth in Rabat, and wherein the amino acid at position 252 is substituted with Tyrosine (Y), the amino acid at position 254 is substituted with Threonine (T), and the amino acid at position 256 is substituted with glutamic acid (E).
  • Y Tyrosine
  • T Threonine
  • E glutamic acid
  • the cancer is characterized in that tissue or cells from the cancer also express a neuregulin or neuregulin fusion protein, and/or comprise a mutation in the BRAF and/or MEK gene.
  • FAT1 and/or NOTCH mutations may be detected in accordance with any method described in this disclosure in DNA, mRNA or protein from samples by means well known in the art, for example, by means described in Section VI below.
  • the methods for treating a cancer are methods for treating a cancer associated with HER3 expression or HER3 -expressing cells, such as head and neck cancer, e.g ., squamous cell carcinoma of the head and neck (SCCHN), sarcoma, melanoma, lung cancer such as non-small cell lung cancer, breast cancer, ovarian cancer, cervical cancer, prostate cancer, retinoblastoma, melanoma, central nervous system cancers such as brain cancer, pancreatic cancer, thyroid cancer, gastric cancer, esophageal cancer, colon cancer, bladder cancer, liver cancer, kidney cancer, such as renal carcinoma, hematopoietic neoplasms.
  • head and neck cancer e.g squamous cell carcinoma of the head and neck (SCCHN), sarcoma, melanoma
  • lung cancer such as non-small cell lung cancer, breast cancer, ovarian cancer, cervical cancer, prostate cancer, retinoblastoma, melanoma
  • the HER3-associated cancer is head and neck cancer, breast cancer, ovarian cancer, prostate cancer, liver cancer, kidney cancer, bladder cancer, pancreatic cancer, brain cancer, hematopoietic neoplasms, retinoblastoma, melanoma, colorectal cancer, gastric cancers, or lung cancer.
  • the HER3- associated cancer is head and neck cancer.
  • the HER3- associated cancer is squamous cell carcinoma of the head and neck.
  • the primary tumor site is oral cavity.
  • HER3 -expressing cell is meant a cell expressing HER3.
  • the methods for treating a cancer provided herein are methods for treating a HER3- associated cancer, wherein treatment with a HER3 inhibitor increases the expression of HER3 in the cancer.
  • the methods for treating a HER3- associated cancer provided herein are methods for treating squamous cell carcinoma of the head and neck.
  • the methods for treating a HER3- associated cancer provided herein are methods for treating squamous cell carcinoma of the head and neck that expresses HER3.
  • the methods for treating a cancer provided herein are methods for treating a cancer associated with B-Raf, wherein treatment with a B-Raf inhibitor increases the expression of HER3 in the cancer.
  • the methods for treating a cancer provided herein are methods for treating a cancer associated with MEK, wherein treatment with a MEK inhibitor increases the expression of HER3 in the cancer.
  • the methods for treating a cancer provided herein are methods for treating a cancer associated with MEK and B-Raf, wherein treatment with a MEK inhibitor and a B-Raf inhibitor increases the expression of HER3 in the cancer.
  • the methods for treating a cancer provided herein are methods for treating thyroid cancer.
  • the methods for treating a cancer provided herein are methods for treating melanoma. In even more specific embodiments, the methods for treating a cancer provided herein are methods for treating thyroid cancer or melanoma that expresses HER3 and/or B-Raf.
  • the cancer is characterized by a BRAF mutation (e.g, V600E or V600K). In specific embodiments, the BRAF mutation is the BRAF V600E mutation. In specific embodiments, the BRAF mutation is the BRAF V600K mutation. In certain embodiments, the cancer is resistant to treatment with a BRAF inhibitor (e.g, vemurafenib or dabrafenib).
  • the cancer is resistant to treatment with a MEK inhibitor (e.g, trametinib).
  • a MEK inhibitor e.g, trametinib
  • the cancer is resistant to treatment with a BRAF inhibitor (e.g, vemurafenib or dabrafenib) and a MEK inhibitor (e.g, trametinib).
  • methods of treating a HER3 -associated cancer described in this disclosure can comprise administering a HER3 inhibitor in combination with a second agent.
  • the second agent is an EGFR inhibitor, a B-Raf inhibitor, or a MEK inhibitor.
  • the second agent can be administered concurrently with the HER3 inhibitor.
  • the second agent can be
  • the second agent can be administered before the HER3 inhibitor, and the second agent can be administered after the HER3 inhibitor.
  • One or both of the HER3 inhibitor and the second agent can each be administered once, or one or both of the HER3 inhibitor and the second agent can each be administered more than once, e.g. , in an alternating manner.
  • a method of treating a HER3- associated cancer comprising administering to a patient diagnosed with said cancer a therapeutically effective amount of a HER3 inhibitor and a second agent, e.g. , an EGFR inhibitor (for example, an anti -EGFR antibody, such as cetuximab), a B-Raf inhibitor, or a MEK inhibitor, wherein the cancer is characterized in that cells from the cancer comprise a mutation in the FAT1 gene.
  • the mutation in the FAT1 gene is a loss- of-function mutation.
  • a method for treating a patient with a HER3 inhibitor and a second agent e.g., an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab), a B-Raf inhibitor, or a MEK inhibitor, wherein the patient is suffering from a HER3 -associated cancer, the method comprising the steps of:
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • a method of treating a FIER3 -associated cancer in a patient diagnosed therewith comprising the steps of: determining whether cells of the cancer comprise a mutation in the FAT1 gene; and if the ceils of the cancer comprise a mutation in the FAT1 gene, then administering a therapeutically effective amount of a HER3 inhibitor and a second agent, e.g, an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab), a B-Raf inhibitor, or a MEK inhibitor, to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • a method of treating a HER3- associated cancer comprising administering to a patient diagnosed with said cancer a therapeutically effective amount of a HER3 inhibitor and a second agent, e.g, an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab), a B-Raf inhibitor, or a MEK inhibitor, wherein the cancer is characterized in that cells from the cancer comprise at least one mutation in at least one NOTCH gene.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • a FIER3 inhibitor for example, an anti-EGFR antibody, such as cetuximab
  • a B-Raf inhibitor for example, a B-Raf inhibitor
  • MEK inhibitor for example, an anti-EGFR antibody, such as cetuximab
  • the patient is suffering from a HER3 -associated cancer, the method comprising the steps of:
  • the at least one mutation in at least one NOTCH gene is one or more loss- of-function mutations.
  • a method of treating a HER3 -associated cancer in a patient diagnosed therewith comprising the steps of: determining whether cells of the cancer comprise at least one mutation in at least one NOTCH gene; and if the cells of the cancer comprise at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor and a second agent, e.g, an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab), a B-Raf inhibitor, or a MEK inhibitor, to the patient.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • a method of treating a HER3- associated cancer comprising administering to a patient diagnosed with said cancer a therapeutically effective amount of a HER3 inhibitor and a second agent, e.g, an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab), a B-Raf inhibitor, or a MEK inhibitor, wherein the cancer is characterized in that cells from the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene.
  • a second agent e.g, an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab), a B-Raf inhibitor, or a MEK inhibitor
  • a method of treating a HER3 -associated cancer comprising administering to a patient diagnosed with said cancer a therapeutically effective amount of a HER3 inhibitor and a second agent, e.g., an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab), a B-Raf inhibitor, or a MEK inhibitor, wherein the cancer is characterized in that tissue from the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • a method for treating a patient with a HER3 inhibitor and a second agent e.g., an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab), a B-Raf inhibitor, or a MEK inhibitor, wherein the patient is suffering from a HER3 -associated cancer, the method comprising the steps of:
  • a method for treating a patient with a HER3 inhibitor and a second agent e.g, an EGFR inhibitor (for example, an anti-EGFR antibody, such as cetuximab), a B-Raf inhibitor, or a MEK inhibitor, wherein the patient is suffering from a HER3 -associated cancer, the method comprising the steps of: determining whether tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one
  • the tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • a method of treating a HER3 -associated cancer in a patient diagnosed therewith comprising the steps of: determining whether cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and if the cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor and a second agent, e.g, an EGFR inhibitor (for example, an anti -EGFR antibody, such as cetuximab), a B-Raf inhibitor, or a MEK inhibitor, to the patient.
  • an EGFR inhibitor for example, an anti -EGFR antibody, such as cetuximab
  • a B-Raf inhibitor such as cetuximab
  • MEK inhibitor e.g, MEK inhibitor
  • a method of treating a TIER 3 -associated cancer in a patient diagnosed therewith comprising the steps of: determining whether tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and if the tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations
  • a HER3 inhibitor used in the methods herein may be any HER3 inhibitor known in the art.
  • the HER3 inhibitor may be an anti-HER3 antibody or antigen-binding fragment thereof, for example, 2C2-YTE, or another anti-HER3 antibody or antigen-binding fragment thereof disclosed herein.
  • the HER3 inhibitor may also be a small molecule inhibitor.
  • anti-HER3 antibodies include, but are not limited to, AMG-888 (U3-1287; Amgen and Daiichi Sankyo), MM-121, (Merrimack and Sanofi-Aventis), GE-huMaB-HER3 (Roche), TK-A3 (Takis and University of Cantanzaro (Italy)), TK-A4 (Takis and University of
  • Cantanzaro (Italy)), AV-203 (Aveo Pharmaceuticals), MP-RM-1 (Mediapharma), LJM716 (Novartis and Sanofi Aventis), REGN1400 (Regeneron), MEHD7945A (a bispecific antibody against HER3 and EGFR; Genentech), Pertuzumab (a bispecific antibody against HER3 and HER2; Genentech), MM-111 (a bispecific antibody against HER3 and HER2; Merrimack), and MM-141 (blocks binding of NRG to HER3 and IGF-1 to IGFR; Merrimack).
  • HER3 small molecule inhibitors include, but are not limited to MP-470 (Amuvatinib; Astex Pharmaceuticals) and AZD 8931 (AstraZeneca).
  • a EGFR inhibitor used in the methods herein may be any EGFR inhibitor known in the art.
  • the EGFR inhibitor may be an anti-EGFR antibody or antigen-binding fragment thereof, for example, cetuximab (ERBITUX®; Bristol-Myers Squibb/Lilly), panitumumab (Amgen), or zalutumumab (Genmab).
  • the EGFR inhibitor may also be a small molecule inhibitor.
  • EGFR inhibitors include, but are not limited to, reversible and irreversible inhibitors, such as erlotinib (TARCEVA®; Genentech/Astellas Oncology), AZD9291 (AstraZeneca), gefitinib (IRESSA®; AstraZeneca), icotinib (BPI-2009H; Beta Pharma), rociletinib (CO-1686, AVL-301; Clovis Oncology), poziotinib (NOV120101, HM781-36B; Hanmi Pharmaceuticals/Spectrum Pharmaceuticals), afatinib (BIBW2292;
  • a B-Raf inhibitor used in the methods herein may be any B-Raf inhibitor known in the art.
  • B-Raf inhibitors include, but are not limited to, vemurafenib
  • a MEK inhibitor used in the methods herein may be any MEK inhibitor known in the art.
  • MEK inhibitors include, but are not limited to, such as selumetinib (AZD6244, ARRY- 142866, AstraZeneca), WX-554 (Wilex), trametinib (MEKINIST®;
  • a MEK inhibitor used in the methods herein may be administered with a B-Raf inhibitor used in the methods herein.
  • HER3 binding molecules e.g., antibodies and antigen-binding fragments thereof that specifically bind HER3 (e.g, CL 16, 2C2 or 2C2-YTE antibodies).
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16) for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, comprising: determining whether cells of the cancer comprise a mutation in the FAT1 gene; and if the cells of the cancer comprise a mutation in the FAT1 gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16) for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the cancer is characterized in that cells from the cancer comprise a mutation in the FAT1 gene, said method comprising administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16) for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, comprising: determining whether cells of the cancer comprise at least one mutation in at least one NOTCH gene; and if the cells of the cancer comprise at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16) for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the cancer is characterized in that cells from the cancer comprise at least one mutation in at least one NOTCH gene, said method comprising administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of- function mutations.
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g ., 2C2-YTE or CL16) for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, comprising: determining whether cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and if the cells of the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • an anti-HER3 antibody e.g ., 2C2-YTE or CL16
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g., 2C2-YTE or CL16) for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, comprising: determining whether tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene; and if the tissue of the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, then administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16) for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the cancer is characterized in that cells from the cancer comprise a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, said method comprising administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • an anti-HER3 antibody e.g, 2C2-YTE or CL16
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g, 2C2-YTE or CL16) for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the cancer is characterized in that tissue from the cancer comprises a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene, said method comprising administering a therapeutically effective amount of a HER3 inhibitor to the patient.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • a HER3 inhibitor for example, an anti-HER3 antibody (e.g 2C2-YTE or CL16), for use in a method of treating a HER3 -associated cancer in a patient diagnosed therewith, wherein the HER3 -associated cancer is head and neck cancer, and wherein the primary tumor site is oral cavity.
  • an anti-HER3 antibody e.g 2C2-YTE or CL16
  • the HER3 inhibitor is an anti-HER3 antibody or antigen binding fragment thereof.
  • the anti-HER3 antibody or antigen-binding fragment thereof specifically binds to the same HER3 epitope as an antibody or antigen binding fragment thereof comprising the heavy chain variable region (VH) and light chain variable region (VL) of CL16 or 2C2.
  • VH and VL of CL16 comprise SEQ ID NOs: 2 and 1, respectively
  • the VH and VL of 2C2 comprise SEQ ID NOs: 2 and 3, respectively.
  • anti-HER3 antibodies and antigen binding fragments thereof may be found in International Application Publication No. WO 2013/078191.
  • the full-length amino acid (aa) and nucleotide (nt) sequences for HER3 are known in the art (see, e.g., UniProt Acc. No. P2186 for human HER3, or UniProt Acc. No. 088458 for mouse HER3).
  • the anti-HER3 binding molecules are human antibodies.
  • the HER3 binding molecules are antibodies or antigen-binding fragments thereof.
  • HER3 binding molecules e.g, antibodies or antigen-binding fragments thereof comprise a Fab, a Fab', a F(ab')2, a Fd, a single chain Fv or scFv, a disulfide linked Fv, a V-NAR domain, an IgNar, an intrabody, an IgGACH2, a minibody, a F(ab') 3 , a tetrabody, a triabody, a diabody, a single-domain antibody, DVD-Ig, Fcab, mAb 2 , a (scFv)2, or a scFv-Fc.
  • the antibody is of the IgGl subtype and comprises the triple mutant YTE, as disclosed supra in the Definitions section.
  • the anti-HER3 antibody or antigen-binding fragment thereof is affinity matured.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises an antibody VL and an antibody VH
  • the VL comprises the amino acid sequence: [FW1]X I GSX 2 SNIGLNYVS (SEQ ID NO: 49), [FW2]RNNQRPS (SEQ ID NO: 21), [FW3]AAWDDX X 4 X 5 GEX 6 (SEQ IDNO: 50) [FW4], wherein [FW1], [FW2], [FW3] and [FW4] represent VL framework regions, and wherein (a) Xi represents amino acid residues Arginine (R) or Serine (S), (b) X 2 represents amino acid residues Serine (S) or Leucine (L), (c) X 3 represents amino acid residues Serine (S) or Glycine (G), (d) X 4 represents amino acid residues Leucine (L) or Proline (P), (e) X represents amino acid residues Arginine (R), Isole
  • FW1 comprises SEQ ID NO: 40 or 44
  • FW2 comprises SEQ ID NO: 41
  • FW3 comprises SEQ ID NO: 42
  • FW4 comprises SEQ ID NO: 43
  • FW5 comprises SEQ ID NO: 36
  • FW6 comprises SEQ ID NO: 37
  • FW7 comprises SEQ ID NO: 38
  • FW8 comprises SEQ ID NO: 39.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises a VL and a VH comprising VL-CDR1, VL-CRD2, VL-CDR3, VH-CDR1, VH-CDR2, and VH-CDR3 amino acid sequences identical or identical except for four, three, two, or one amino acid substitutions in one or more CDRs to: SEQ ID NOs: 18, 21, 22, 31, 32, and 35, SEQ ID NOs: 18, 21, 26, 31, 32 and 35, SEQ ID NOs: 18, 21, 27, 31, 32 and 35, SEQ ID NOs: 20, 21, 22, 31, 32 and 35, SEQ ID NOs: 19, 21, 22, 31, 32 and 35, SEQ ID NOs: 18, 21, 25, 31, 32 and 35, SEQ ID NOs: 18, 21, 28, 31, 32 and 35, SEQ ID NOs: 18, 21, 29, 31, 32 and 35, SEQ ID NOs: 18, 21, 30, 31, 32 and 35, SEQ ID NOs: 18, 21, 23, 31, 32 and 35, SEQ ID NOs: 19, 21, 23, 31, 32 and 35, SEQ ID NOs:
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises an antibody VL and an antibody VH, wherein the VL comprises an amino acid sequence at least about 90% to about 100% identical to a reference amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, and SEQ ID NO: 17.
  • a reference amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, and SEQ ID NO: 17.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises an antibody VL and an antibody VH, wherein the VH comprises an amino acid sequence at least about 90% to about 100% identical to a reference amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 12 and SEQ ID NO: 13.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises a VL comprising a sequence at least about 90% to about 100% identical to a reference amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, and SEQ ID NO: 17, and wherein the anti-HER3 antibody or antigen- binding fragment comprises a VH comprising a sequence at least about 90% to about 100% identical to a reference amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 12 and SEQ ID NO: 13.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises a VL-CDR1, a VL-CDR2 and a VL-CDR3 comprising SEQ ID NO: 19, SEQ ID NO: 21 and SEQ ID NO: 23, respectively, and a VH-CDR1, a VH-CDR2, and a VH-CDR3 comprising SEQ ID NO: 31, SEQ ID NO: 32, and SEQ ID NO: 35, respectively.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises a VL comprising SEQ ID NO: 3 and a VH comprising SEQ ID NO: 2.
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises a human heavy chain constant region or fragment thereof.
  • the heavy chain constant region or fragment thereof is an IgG constant region.
  • the IgG constant region is selected from an IgGl constant region, an IgG2 constant region, an IgG3 constant region and an IgG4 constant region.
  • the IgG constant region is an IgGl constant region.
  • the IgG constant domain comprises one or more amino acid substitutions relative to a wild-type IgG constant domain wherein the modified IgG has an increased half-life compared to the half-life of an IgG having the wild- type IgG constant domain.
  • the IgG constant domain comprises one or more amino acid substitutions of amino acid residues at positions 251-257, 285-290, 308-314, 385-389, and 428-436, wherein the numbering is according to the EU index as set forth in Kabat.
  • At least one IgG constant domain amino acid substitution is selected from the group consisting of: (a) substitution of the amino acid at position 252 with Tyrosine (Y), Phenylalanine (F), Tryptophan (W), or Threonine (T), (b) substitution of the amino acid at position 254 with Threonine (T), (c) substitution of the amino acid at position 256 with Serine (S), Arginine (R), Glutamine (Q), Glutamic acid (E), Aspartic acid (D), or Threonine (T), (d) substitution of the amino acid at position 257 with Leucine (L), (e) substitution of the amino acid at position 309 with Proline (P), (f) substitution of the amino acid at position 311 with Serine (S), (g) substitution of the amino acid at position 428 with
  • the human IgG constant domain comprises amino acid substitutions relative to a wild-type human IgG constant domain at positions 252, 254, and 256, wherein (a) the amino acid at position 252 is substituted with Tyrosine (Y), (b) the amino acid at position 254 is substituted with Threonine (T), and (c) the amino acid at position 256 is substituted with Glutamic acid (E), wherein the numbering is according to the EU index as set forth in Kabat.
  • the amino acid at position 434 is substituted with an amino acid selected from the group consisting of Tryptophan (W), Methionine (M), Tyrosine (Y), and Serine (S), and wherein the numbering is according to the EU index as set forth in Kabat.
  • the amino acid at position 428 is substituted with an amino acid selected from the group consisting of Threonine (T), Leucine (L), Phenylalanine (F), and Serine (S), and wherein the numbering is according to the EU index as set forth in Kabat.
  • the amino acid at position 257 is substituted with Leucine (L), and the amino acid at Kabat position 434 is substituted with Tyrosine (Y), and wherein the numbering is according to the EU index as set forth in Kabat.
  • the amino acid at Kabat position 428 is substituted with Leucine (L), and the amino acid at Kabat position 434 is substituted with Serine (S).
  • the anti-HER3 antibody or antigen-binding fragment comprises a light chain constant region selected from the group consisting of a human kappa constant region and a human lambda constant region. In some embodiments, the anti-HER3 antibody or antigen-binding fragment comprises a human lambda constant region. In some embodiments, the anti-HER3 antibody or antigen-binding fragment thereof comprises an antibody VL of SEQ ID NO:3, an antibody VH of SEQ ID NO:2, and an IgGl constant region of SEQ ID NO:46. In some embodiments, the anti-HER3 antibody or antigen-binding fragment thereof comprises a human IgGl constant region and a human lambda constant region.
  • the anti-HER3 antibody is a human antibody, a humanized antibody, a chimeric antibody, a monoclonal antibody, a polyclonal antibody, a recombinant antibody, a multispecific antibody, or an antigen-binding fragment thereof.
  • the anti-HER3 antibody is a human antibody.
  • the antigen-binding fragment is a Fv, Fab, F(ab')2, Fab', dsFv, scFv, or sc(Fv)2.
  • anti-HER3 antibodies or antigen-binding fragments thereof described herein are modified compared to the parent Clone 16 (CL16) antibody.
  • the modifications can include mutations in the CDR regions and/or in the FW regions as compared to CL16.
  • an anti-HER3 antibody described herein comprises modifications to CDR1 and/or CDR3 of the light chain of CL16, including, but not limited to:
  • AAWDDX 3 X 4 X 5 GEX 6 (SEQ IDNO:50), wherein X 3 is selected from S or G, X 4 is selected from L or P, X 5 is selected from R, I, P or S, and Xr, is selected from V or A.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises modifications to CDR2 of the heavy chain of CL16, including, but not limited to a heavy chain CDR1 comprising the consensus sequence
  • an anti-HER3 antibody or antigen binding fragment thereof comprises a VL region comprising the consensus amino acid sequence:
  • an anti-HER3 antibody or antigen binding fragment thereof comprises a VH region comprises the consensus amino acid sequence:
  • an anti-HER3 antibody or antigen binding fragment thereof comprises a VL region comprising the consensus amino acid sequence:
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VL-CDR1 consisting of sequence selected from the group consisting of SEQ ID NOs: 18, 19 and 20.
  • an anti-HER3 antibody or antigen binding fragment thereof described herein comprises a VL-CDR1 comprising a sequence selected from the group consisting of SEQ ID NOs: 18, 19 and 20.
  • an anti- HER3 antibody or antigen-binding fragment thereof described herein comprises a VL-CDR2 consisting of SEQ ID NO: 21.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VL-CDR2 comprising SEQ ID NO: 21.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VL-CDR3 consisting of a sequence selected from the group consisting of SEQ ID NOs: 22, 23, 24, 25, 26, 27, 28, 29, and 30.
  • an anti-HER3 antibody or antigen binding fragment thereof described herein comprises a VL-CDR3 comprising a sequence selected from the group consisting of SEQ ID NOs: 22, 23, 24, 25, 26, 27, 28, 29, and 30.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VH-CDRl consisting of SEQ ID NO: 31. In some aspects, an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VH- CDRl comprising SEQ ID NO: 31. In some aspects, an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VH-CDR2 consisting of a sequence selected from the group consisting of SEQ ID NOs: 32, 33 and 34.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VH-CDR2 comprising a sequence selected from the group consisting of SEQ ID NOs: 32, 33 and 34. In some aspects, an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VH-CDR3 consisting of SEQ ID NO: 35. In some aspects, an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VH-CDR3 comprising SEQ ID NO: 35.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VL-CDR1 consisting of a sequence selected from the group consisting of SEQ ID NOs: 18, 19 and 20, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VL-CDR1 comprising a sequence selected from the group consisting of SEQ ID NOs: 18, 19 and 20, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VL-CDR2 consisting of SEQ ID NO: 21, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen binding fragment thereof described herein comprises a VL-CDR2 comprising SEQ ID NO: 21, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VL-CDR3 consisting of a sequence selected from the group consisting of SEQ ID NOs: 22, 23, 24, 25, 26, 27, 28, 29, and 30, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VL- CDR3 comprising a sequence selected from the group consisting of SEQ ID NOs: 22, 23, 24, 25, 26, 27, 28, 29, and 30, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VH-CDR1 consisting of SEQ ID NO: 31, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen binding fragment thereof described herein comprises a VH-CDR1 comprising SEQ ID NO: 31, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VH-CDR2 consisting of a sequence selected from the group consisting of SEQ ID NOs: 32, 33 and 34, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VH-CDR2 comprising a sequence selected from the group consisting of SEQ ID NOs: 32, 33 and 34, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VH-CDR3 consisting of SEQ ID NO: 35, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VH-CDR3 comprising SEQ ID NO: 35, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VL-CDR1 consisting of a sequence selected from the group consisting of SEQ ID NOs: 18, 19 and 20; a VL-CDR2 consisting of SEQ ID NO: 21; and a VL-CDR3 consisting of a sequence selected from the group consisting of SEQ ID NOs: 22, 23, 24, 25, 26, 27, 28, 29, and 30.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VL-CDR1 comprising a sequence selected from the group consisting of SEQ ID NOs: 18, 19 and 20; a VL-CDR2 comprising SEQ ID NO: 21; and a VL-CDR3 comprising a sequence selected from the group consisting of SEQ ID NOs: 22, 23, 24, 25, 26, 27, 28, 29, and 30.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VH-CDR1 consisting of SEQ ID NO: 31; a VH-CDR2 consisting of a sequence selected from the group consisting of SEQ ID NOs: 32, 33 and 34; and a VH- CDR3 consisting of SEQ ID NO: 35.
  • an anti-HER3 antibody or antigen binding fragment thereof described herein comprises a VH-CDR1 comprising SEQ ID NO: 31; a VH-CDR2 comprising a sequence selected from the group consisting of SEQ ID NOs: 32, 33 and 34; a VH-CDR3 comprising SEQ ID NO: 35.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VL-CDR1 consisting of a sequence selected from the group consisting of SEQ ID NOs: 18, 19 and 20, except for one, two, three or four amino acid substitutions; a VL-CDR2 consisting of SEQ ID NO: 21, except for one, two, three or four amino acid substitutions; and a VL-CDR3 consisting of a sequence selected from the group consisting of SEQ ID NOs: 22, 23, 24, 25, 26, 27, 28, 29, and 30, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VL-CDRl comprising a sequence selected from the group consisting of SEQ ID NOs: 18, 19 and 20, except for one, two, three or four amino acid substitutions; a VL-CDR2 comprising SEQ ID NO: 21, except for one, two, three or four amino acid substitutions; and a VL-CDR3 comprising a sequence selected from the group consisting of SEQ ID NOs: 22, 23, 24, 25, 26, 27, 28, 29, and 30, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VH-CDR1 consisting of SEQ ID NO: 31, except for one, two, three or four amino acid substitutions; a VH-CDR2 consisting of a sequence selected from the group consisting of SEQ ID NOs: 32, 33 and 34, except for one, two, three or four amino acid substitutions; and a VH-CDR3 consisting of SEQ ID NO: 35, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen-binding fragment thereof antibody described herein comprises a VH-CDR1 comprising SEQ ID NO: 31, except for one, two, three or four amino acid substitutions; a VH-CDR2 comprising a sequence selected from the group consisting of SEQ ID NOs: 32, 33 and 34, except for one, two, three or four amino acid substitutions; and VH-CDR3 comprising SEQ ID NO: 35, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises modifications to CDR1, CDR2, and/or CDR3 of the heavy and/or light chain, and further comprises modifications to FW1, FW2, FW3, and/or FW4 of the heavy and/or light chain.
  • FWi comprises SEQ ID NO: 40 or 44
  • FW2 comprises SEQ ID NO: 41
  • FW3 comprises SEQ ID NO: 42
  • FW4 comprises SEQ ID NO: 43
  • FW5 comprises SEQ ID NO: 36
  • FW 6 comprises SEQ ID NO: 37
  • FWv comprises SEQ ID NO: 38
  • FW 8 comprises SEQ ID NO: 39.
  • FWi comprises SEQ ID NO: 40 or 44, except for one, two, three or four amino acid substitutions
  • FW2 comprises SEQ ID NO: 41, except for one, two, three or four amino acid substitutions
  • FW3 comprises SEQ ID NO: 42, except for one, two, three or four amino acid substitutions
  • FW4 comprises SEQ ID NO: 43, except for one, two, three or four amino acid substitutions
  • FW5 comprises SEQ ID NO: 36, except for one, two, three or four amino acid substitutions
  • FW 6 comprises SEQ ID NO: 37, except for one, two, three or four amino acid substitutions
  • FW7 comprises SEQ ID NO: 38, except for one, two, three or four amino acid substitutions
  • FWs comprises SEQ ID NO: 39, except for one, two, three or four amino acid substitutions.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VL and a VH comprising VL-CDR1, VL-CRD2, VL-CDR3, VH- CDR1, VH-CDR2, and VH-CDR3 amino acid sequences identical or identical except for four, three, two, or one amino acid substitutions in one or more CDRs to: SEQ ID NOs: 18, 21, 22, 31, 32, and 35, SEQ ID NOs: 18, 21, 26, 31, 32 and 35, SEQ ID NOs: 18, 21, 27, 31, 32 and 35, SEQ ID NOs: 20, 21, 22, 31, 32 and 35, SEQ ID NOs: 19, 21, 22, 31, 32 and 35, SEQ ID NOs: 18, 21, 25, 31, 32 and 35, SEQ ID NOs: 18, 21, 28, 31, 32 and 35, SEQ ID NOs: 18, 21, 29, 31, 32 and 35, SEQ ID NOs: 18, 21, 30, 31, 32 and 35, SEQ ID NOs: 18, 21, 23, 31, 32 and 35, SEQ ID NOs: 19, 21, 23,
  • Heavy and light chain variable domains of the anti-HER3 antibody or antigen binding fragment thereof described herein include the sequences listed in TABLE 4.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises an antibody VL and an antibody VH, wherein the VL comprises an amino acid sequence at least about 80%, about 85%, about 90%, about 95%, about 96%, about 97%, about 98%, about 99%, or about 100% identical to a reference amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, and SEQ ID NO: 17.
  • the VL comprises VL CDRs identical to those of the VL reference amino acid sequence.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises an antibody VL and an antibody VH, wherein the VH comprises an amino acid sequence at least about 80%, about 85%, about 90%, about 95%, about 96%, about 97%, about 98%, about 99%, or about 100% identical to a reference amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 12 and SEQ ID NO: 13.
  • the VH comprises VH CDRs identical to those of the VL reference amino acid sequence.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises a VL comprising a sequence at least about 80%, about 85%, about 90%, about 95%, about 96%, about 97%, about 98%, about 99%, or about 100% identical to a reference amino acid sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, and SEQ ID NO: 17, and further comprises a VH comprising a sequence at least about 80%, about 85%, about 90%, about 95%, about 96%, about 97%, about 98%, about 99%, or about 100% identical to a reference amino acid sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 12 and SEQ ID NO: 13.
  • an anti-HER3 antibody or antigen-binding fragment provided herein comprises a VL comprising a sequence that is at least 80%, at least 85%, at least 90%, or at least 95% identical to SEQ ID NO: 3 and a VH comprising a sequence that is at least 80%, at least 85%, at least 90%, or at least 95% identical to SEQ ID NO: 2.
  • an anti-HER3 antibody or antigen binding fragment comprises a VL comprising a sequence that is at least 80%, at least 85%, at least 90%, or at least 95% identical SEQ ID NO: 3 and a VH comprising a sequence that is at least 80%, at least 85%, at least 90%, or at least 95% identical to SEQ ID NO: 2, wherein the VL CDRs are identical to those of SEQ ID NO: 3 and the VH CDRs are identical to those of SEQ ID NO: 2.
  • an anti-HER3 antibody or antigen-binding fragment thereof comprises a VH of TABLE 4 and a VL of TABLE 4.
  • Antibodies are designated throughout the specification according to their VL chains.
  • the heavy chains of the specific antibodies disclosed in the present specification correspond to the CL16 original heavy chain (SEQ ID NO: 2).
  • the "CL16 antibody” is an IgGl comprising two original CL16 light chains (SEQ ID NO: 17) and two CL 16 original heavy chains (SEQ ID NO: 2)
  • the "2C2 antibody” is an IgGl comprising two 2C2 light chains (2C2 VL (SEQ ID NO: 3) and two CL16 original heavy chains (SEQ ID NO: 2).
  • the anti-HER3 antibody or antigen-binding fragment thereof comprises a heavy chain constant region or fragment thereof.
  • the heavy chain constant region is an IgG constant region.
  • the IgG constant region can comprise a light chain constant region selected from the group consisting of a kappa constant region and a lambda constant region.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein binds HER3 with substantially the same or better affinity as the CL16 antibody, comprising the CL16 original heavy chain (SEQ ID NO: 2) and the original CL16 light chain (SEQ ID NO: 17).
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein binds HER3 with substantially the same or better affinity as the 2C2 antibody, comprising the 2C2 light chain (2C2 VL (SEQ ID NO: 3) and the CL16 original heavy chain (SEQ ID NO: 2).
  • an anti-HER3 antibody or antigen-binding fragment thereof specifically binds HER3 and antigenic fragments thereof with a dissociation constant of k d (k 0ff /k 0n ) of less than KG 6 M, or of less than KG 7 M, or of less than KG 8 M, or of less than KG 9 M, or of less than KG 10 M, or of less than 1CT 11 M, or of less than KG 12 M, or of less than KG 13 M.
  • an anti-HER3 antibody or antigen binding fragment thereof specifically binds HER3 and antigenic fragments thereof with a dissociation constant between 2 c KG 10 M and 6 c KG 10 M.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein binds to HER3 and/or antigenic fragments thereof with a K 0ff of less than 1 x KG 3 s _1 , or less than 2 c KG 3 s _1 .
  • an anti-HER3 antibody or antigen-binding fragment thereof binds to HER3 and antigenic fragments thereof with a K 0ff of less than KG 3 s _1 , less than 5 c KG 3 s _1 , less than KG 4 s _1 , less than 5 c 1(G 4 s _1 , less than KG 5 s _1 , less than 5x l0 -5 s _1 , less than KG 6 s _1 , less than 5 c KG 6 s _1 , less than less than 5 c 1(G 7 s _1 , less than KG 8 s _1 , less than 5x l(T 8 s _1 , less than KG 9 s _1 , less than 5 c 1(G 9 s _1 , or less than KG 10 s _1 .
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein binds to HER3 and/or antigenic fragments thereof with a K 0ff of between 0.5 x lO -4 s _1 and 2.0x l0 4 s _1 .
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein binds to HER3 and/or antigenic fragments thereof with an association rate constant or k on rate of at least 10 5 M -1 s _1 , at least 5x 10 5 M -1 s _1 , at least 10 6 M -1 s _1 , at least 5x l0 6 M -1 s _1 , at least 10 7 M -1 s _1 , at least 5x l0 7 M -1 s _1 , or at least 10 8 M -1 s _1 , or at least 10 9 M -1 s _1 .
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein binds to HER3 and/or antigenic fragments thereof with an association rate constant or kon rate of between 1 c 10 5 M -1 s _1 and 6x 10 5 M -1 s _1 .
  • VH and VL sequences disclosed in TABLE 4 can be "mixed and matched" to create other anti-HER3 binding molecules described herein.
  • the VH sequences of 15D12.I and 15D12.V are mixed and matched.
  • the VL sequences of 5H6, 8A3, 4H6, 6E.3, 2B11, 2D1, 3A6, 4C4, 1A4, 2C2, 3E.1 can be mixed and matched.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprises mutations that improve the binding to human FcRn and improve the half-life of the anti-HER3 antibody or antigen-binding fragment thereof.
  • mutations are a methionine (M) to tyrosine (Y) mutation in position 252, a serine (S) to threonine (T) mutation in position 254, and a threonine (T) to glutamic acid (E) mutation in position 256, numbered according to the EU index as in Rabat (Rabat, et al. (1991) Sequences of Proteins of Immunological Interest, U.S.
  • an anti-HER3 antibody or antigen-binding fragment thereof comprising an IgG constant domain comprises one or more amino acid substitutions of amino acid residues at positions 251-257, 285-290, 308-314, 385-389, and 428-436, numbered according to the EU index as in Rabat, wherein such mutations increase the serum half-life of the anti-HER3 antibody or antigen-binding fragment thereof.
  • a YTE mutant further comprises a substitution at position 434 of the IgG constant domain, numbered according to the EU index as in Rabat, with an amino acid selected from the group consisting of tryptophan (W), methionine (M), tyrosine (Y), and serine (S).
  • W tryptophan
  • M methionine
  • Y tyrosine
  • S serine
  • a YTE mutant further comprises a substitution at position 434 of the IgG constant domain, numbered according to the EU index as in Rabat, with an amino acid selected from the group consisting of tryptophan (W), methionine (M), tyrosine (Y), and serine (S), and substitution at position 428 of the IgG constant domain, numbered according to the EU index as in Rabat, with an amino acid selected from the group consisting of threonine (T), leucine (L), phenylalanine (F), and serine (S).
  • W tryptophan
  • M methionine
  • Y tyrosine
  • S serine
  • a YTE mutant further comprises a substitution at position 434 of the IgG constant domain, numbered according to the EU index as in Rabat, with tyrosine (Y), and a substitution at position 257 of the IgG constant domain, numbered according to the EU index as in Rabat, with leucine (L).
  • a YTE mutant further comprises a substitution at position 434 of the IgG constant domain, numbered according to the EU index as in Kabat, with serine (S), and a substitution at position 428 of the IgG constant domain, numbered according to the EU index as in Kabat, with leucine (L).
  • an anti-HER3 antibody or antigen-binding fragment thereof comprises a 2C2 light chain variable region (2C2 VL; SEQ ID NO: 3), an original CL16 heavy chain variable region (SEQ ID NO: 2), and an IgGl constant domain comprising a methionine (M) to tyrosine (Y) mutation in position 252, a serine (S) to threonine (T) mutation in position 254, and a threonine (T) to glutamic acid (E) mutation in position 256 of the IgGl constant domain, numbered according to the EU index as in Kabat.
  • M methionine
  • Y tyrosine
  • T serine
  • T threonine
  • E glutamic acid
  • an anti-HER3 antibody or antigen-binding fragment thereof comprises a light chain variable region and a heavy chain variable region as presented in Table 5.
  • TABLE 5 provides the SEQ ID NOs for each clone.
  • an anti-HER3 antibody or antigen-binding fragment thereof comprises a light chain variable region and a heavy chain variable region as described in PCT International Publication No. WO 2013/078191 Al, which is hereby incorporated by reference in its entirety.
  • an anti-HER3 antibody or antigen-binding fragment thereof described herein comprise at least one IgG constant domain amino acid substitution selected from the group consisting of:
  • the positions are numbered according to the EU index as in Rabat, and wherein the modified IgG has an increased serum half-life compared to the serum half-life of an IgG having the wild-type IgG constant domain.
  • the VH and/or VL amino acid sequences can be at least 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to the sequences set forth above, and comprise 1, 2, 3, 4, 5 or more conservative substitutions. In certain aspects, the VH and/or VL amino acid sequences can be at least 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to the sequences set forth above, and comprise 1, 2, 3, 4, 5 or more conservative substitutions which are not within the VH CDRs or VL CDRs.
  • the VH and/or VL amino acid sequences can be at least 80%, 85%, 90%, or 95% identical to the sequences set forth above, and comprise 1, 2, 3, 4, 5 or more conservative substitutions. In certain aspects, the VH and/or VL amino acid sequences can be at least 80%, 85%, 90%, or 95% identical to the sequences set forth above, and comprise 1, 2, 3, 4, 5 or more conservative substitutions which are not within the VH CDRs or VL CDRs.
  • a HER3 antibody having VH and VL regions having high (i.e., 80% or greater) similarity to the VH regions of SEQ ID NOs: 2, 12 or 13 and/or VL regions of SEQ ID NOs: 1, 3, 4, 5, 6, 7, 8, 9, 10, 11, 14, 15, 16, or 17, respectively, can be obtained by mutagenesis (e.g ., site-directed or PCR-mediated mutagenesis) of nucleic acid molecules encoding SEQ ID NOs: 1-17, followed by testing of the encoded altered antibody for retained function using the functional assays described herein.
  • mutagenesis e.g ., site-directed or PCR-mediated mutagenesis
  • the affinity or avidity of an antibody for an antigen can be determined experimentally using any suitable method well known in the art, e.g., flow cytometry, enzyme- linked immunosorbent assay (ELISA), or radioimmunoassay (RIA), or kinetics (e.g,
  • affinity and other antigen-binding parameters are made with standardized solutions of antibody and antigen, and a standardized buffer, as known in the art and such as the buffer described herein.
  • affinity can be measured using a format in which the targeting antibody (e.g, the 2C2 monoclonal antibody) is immobilized onto the chip (referred to as an "IgG down” format) or using a format in which the target protein (e.g, HER3) is immobilized onto the chip (referred to as, e.g, a "HER3 down” format).
  • the targeting antibody e.g, the 2C2 monoclonal antibody
  • HER3 a format in which the target protein
  • epitope refers to a protein determinant capable of binding to an antibody described herein.
  • Epitopes usually consist of chemically active surface groupings of molecules such as amino acids or sugar side chains and usually have specific three dimensional structural characteristics, as well as specific charge characteristics. Conformational and non-conformational epitopes are distinguished in that the binding to the former but not the latter is lost in the presence of denaturing solvents.
  • anti-HER3 antibodies and antigen-binding fragments thereof e.g., human monoclonal antibodies that compete for binding to HER3 with another anti-HER3 antibody or antigen-binding fragment thereof described herein, such as the CL16 antibody or the 2C2 antibody.
  • test antibody to inhibit the binding of, e.g, the CL 16 antibody or the 2C2 antibody
  • the test antibody can compete with that antibody for binding to HER3; such an antibody can, according to non-limiting theory, bind to the same or a related (e.g, a structurally similar or spatially proximal) epitope on HER3 as the anti-HER3 antibody or antigen-binding fragment thereof with which it competes.
  • the anti-HER3 antibody or antigen-binding fragment thereof that binds to the same epitope on HER3 as, e.g, the CL 16 antibody or the 2C2 antibody is a human monoclonal antibody.
  • a method of detecting a mutation in the FAT1 gene in a patient diagnosed with a HER3 -associated cancer comprising: performing an assay on a sample that comprises or is derived from the patient’s cells, e.g, cancerous cells, to determine whether a mutation in the FAT1 gene is present in the sample.
  • cells e.g, cancerous cells
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • a method of detecting a mutation in the FAT1 gene in a patient diagnosed with a HER3 -associated cancer comprising: obtaining a sample, wherein the sample comprises or is derived from the patient’s cells, e.g, cancerous cells; and performing an assay on the sample to determine whether a mutation in the FAT1 gene is present in the sample.
  • the mutation in the FAT1 gene is a loss-of- function mutation.
  • a method of detecting at least one mutation in at least one NOTCH gene in a patient diagnosed with a HER3 -associated cancer comprising: performing an assay on a sample that comprises or is derived from the patient’s cells, e.g, cancerous cells, to determine whether at least one mutation in at least one NOTCH gene is present in the sample.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • a method of detecting at least one mutation in at least one NOTCH gene in a patient diagnosed with a HER3 -associated cancer comprising: obtaining a sample, wherein the sample comprises or is derived from the patient’s cells, e.g. , cancerous cells; and performing an assay on the sample to determine whether at least one mutation in at least one NOTCH gene is present in the sample.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • a method of detecting a mutation in the FAT1 gene in a patient diagnosed with a HER3 -associated cancer comprising: performing an assay on a sample that comprises or is derived from the patient’s cells, e.g. , cancerous cells, to determine whether a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene are present in the sample.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • the mutation in the FAT1 gene is a loss-of-function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • a method of detecting a mutation in the FAT1 gene in a patient diagnosed with a HER3 -associated cancer comprising: obtaining a sample, wherein the sample comprises or is derived from the patient’s cells, e.g. , cancerous cells; and performing an assay on the sample to determine whether a mutation in the FAT1 gene and at least one mutation in at least one NOTCH gene are present in the sample.
  • the mutation in the FAT1 gene is a loss-of-function mutation.
  • the at least one mutation in at least one NOTCH gene is one or more loss-of- function mutations.
  • the mutation in the FAT1 gene is a loss-of- function mutation and the at least one mutation in at least one NOTCH gene is one or more loss-of-function mutations.
  • FAT1 and/or NOTCH mutations may be detected in accordance with any method described in this disclosure in DNA, mRNA or protein from samples by means well known in the art.
  • the sample may be a tumor biopsy sample, a blood sample, a serum sample or a lymph sample, which may contain one or more tumor cells or circulating tumor DNA (ctDNA) from the patient.
  • ctDNA circulating tumor DNA
  • testing comprises detecting mutations in DNA or mRNA from the sample by sequencing or, for example, by use of nucleic acid probes or microarrays.
  • the method further comprises first amplifying the DNA or mRNA molecules from the sample, for example by isothermal amplification or polymerase chain reaction (PCR).
  • PCR polymerase chain reaction
  • FAT1 and/or NOTCH mutations may be assayed by sequencing DNA or RNA by any method known in the art. These methods include, but are not limited to
  • TORRENTTM DNA next generation sequencing ION TORRENTTM RNA next generation sequencing, 454TM pyrosequencing, or Sequencing by Oligo Ligation Detection (SOLIDTM), PCR-based methods, and the like.
  • a change(s) in the amino acid sequence of a protein relative to a non-mutant or wild-type protein may also be used as an indication of the presence of a mutation in the corresponding gene. Therefore, methods of detecting changes in protein sequences may also be used to detect a mutated FAT1 gene and/or a mutated NOTCH gene(s).
  • RNA expression may be assayed by detecting or quantitating mRNA levels by any method known in the art. These methods include, but are not limited to northern blots, ribonuclease protection assays, in situ hybridization, for example, RNAscope ® technology, ILLUMINA® RNASeq, ILLUMINA® next generation sequencing (NGS), ION TORRENTTM RNA next generation sequencing, 454TM pyrosequencing, or Sequencing by Oligo Ligation Detection (SOLIDTM), PCR-based methods, and the like. PCR-based methods include RT-PCR and Real-Time (or quantitative) RT-PCR (qRT-PCR).
  • Protein expression and mutations may be assayed in a particular sample using a variety of methods. Any suitable protein quantification method can be used. In some embodiments, antibody-based/immunospecific methods are used.
  • the immunoassays that can be used include techniques such as immunohistochemistry assays, Western blots, radioimmunoassays, ELISA (enzyme linked immunosorbent assay), "sandwich"
  • ELISA methods include, for example, direct ELISA, indirect ELISA, and sandwich ELISA. Such assays are routine and well known in the art (see, e.g ., Ausubel et al ., eds, (1994) Current Protocols in Molecular Biology (John Wiley & Sons, Inc., NY) Vol. 1, which is incorporated by reference herein in its entirety). Methods such as flow cytometry and cytometric bead array may be used to measure expression of cell-surface proteins. Mass spectroscopic methods may also be used to measure protein expression levels and mutations in a sample.
  • Clinical response can be assessed using screening techniques such as magnetic resonance imaging (MRI) scan, x-radiographic imaging, computed tomographic (CT) scan, flow cytometry or fluorescence-activated cell sorter (FACS) analysis, histology, gross pathology, and blood chemistry, including but not limited to changes detectable by ELISA, RIA, chromatography, and the like.
  • MRI magnetic resonance imaging
  • CT computed tomographic
  • FACS fluorescence-activated cell sorter
  • Example 1 Methods for determining whether a patient diagnosed with a HER3- associated cancer is likely to be responsive to treatment with a Her3 inhibitor, using FAT1 mutation as a biomarker.
  • a patient diagnosed with a HER3 -associated cancer is indicated as likely to be responsive to treatment with a HER3 inhibitor, such as, for example, an anti-HER3 antibody, (e.g., 2C2-YTE or CL16)
  • a patient sample containing a cancer cell is assayed for the presence of a FAT1 gene mutation, e.g. , a loss-of-function mutation in the FAT1 gene.
  • the sample can be, for example, a biopsy, blood or serum sample. After amplification as required, DNA and/or mRNA from the sample is sequenced to establish whether or not a FAT1 gene mutation is present.
  • Example 2 Methods for determining whether a patient diagnosed with a HER3- associated cancer is likely to be responsive to treatment with a Her3 inhibitor, using NOTCH mutation as a biomarker.
  • a patient sample containing a cancer cell is assayed for the presence of one or more mutations in a NOTCH gene (e.g, NOTCH1, NOTCH2, or NOTCH3), e.g, a loss-of-function mutation in the NOTCH gene.
  • a NOTCH gene e.g, NOTCH1, NOTCH2, or NOTCH3
  • mutations in NOTCH2 genes include without limitation: R91*, W16* and splice acceptor mutations (e.g, in exon 6).
  • R91* denotes a mutation that changes R91 of NP 077719.2 to a stop codon (for example, a CGA to TGA mutation at the DNA level).
  • W16* denotes a mutation that changes W16 of NP 077719.2 to a stop codon (for example, a TGG to TGA mutation at the DNA level).
  • the sample can be, for example, a biopsy, blood or serum sample. After amplification as required, DNA and/or mRNA from the sample is sequenced to establish whether or not one or more NOTCH gene mutations (such as a loss-of-function mutation) is present.
  • NOTCH gene mutations such as a loss-of-function mutation
  • Example 3 Methods for determining whether a patient diagnosed with a HER3- associated cancer is likely to be responsive to treatment with a Her3 inhibitor, using the combination of FAT1 mutation and NOTCH mutation as a biomarker.
  • a patient diagnosed with a HER3 -associated cancer is indicated as likely to be responsive to treatment with a HER3 inhibitor, such as, for example, an anti-HER3 antibody, (e.g, 2C2-YTE or CL16)
  • a patient sample containing a cancer cell is assayed for the presence of a FAT1 gene mutation and one or more mutations in a NOTCH gene (e.g, NOTCH1, NOTCH2, or NOTCH3), e.g, a loss-of-function mutation in the FAT1 gene and a loss-of-function mutation in the NOTCH gene.
  • a NOTCH gene e.g, NOTCH1, NOTCH2, or NOTCH3
  • NOTCH2 genes include without limitation: R91*, W16* and splice acceptor mutations (e.g, in exon 6).
  • R91* denotes a mutation that changes R91 of NP 077719.2 to a stop codon (for example, a CGA to TGA mutation at the DNA level).
  • W16* denotes a mutation that changes W16 of NP 077719.2 to a stop codon (for example, a TGG to TGA mutation at the DNA level).
  • the sample can be, for example, a biopsy, blood or serum sample. After amplification as required, DNA and/or mRNA from the sample is sequenced to establish whether or not a FAT1 gene mutation (such as a loss-of-function mutation) and one or more NOTCH gene mutations (such as a loss-of-function mutation) are present.
  • a FAT1 gene mutation such as a loss-of-function mutation
  • NOTCH gene mutations such as a loss-of-function mutation
  • CDX-3379 (a 2C2-YTE antibody) is a fully human anti-HER3 monoclonal IgGlk antibody that is a potent inhibitor of HER3 activation via a unique mechanism of action that locks HER3 in an inactive configuration (Lee el al, 2015, Proc Natl Acad Sci
  • CDX-3379 antitumor activity has been observed in patients with head and neck squamous cell carcinoma (HNSCC) in previous clinical studies.
  • HNSCC head and neck squamous cell carcinoma
  • CDX3379-01 a durable complete response
  • HPV cetuximab-refractory human papillomavirus
  • CDX3379- 02 In a window of opportunity study of CDX-3379 monotherapy (CDX3379- 02), an exceptional clinical response in a patient with HPV(-) HNSCC was observed (Duvvuri et al., 2018, Cancer Research 78(13)suppl:CT058).
  • a phase 2, multi-center, open-label, single-arm clinical trial (CDX3379-04, ClinicalTrials.gov Identifier: NCT03254927) was further conducted to evaluate safety and efficacy of CDX-3379 in combination with cetuximab in patients with advanced HNSCC.
  • One of the objectives of this study was to perform a biomarker analysis to identify biomarkers associated with clinical response to CDX-3379. Fifteen patients were enrolled in the study. They received between 1 and 16 doses of CDX-3379 (median 3 doses) and between 1 and 46 doses of cetuximab (median 6 doses) (see FIG. 2).
  • HNSCC tumor samples from the three clinical trials described above were subjected to next-generation sequencing, targeting more than 1400 cancer-related genes.
  • One sample was from a patient treated in the CDX3379-01 trial (CDX-3379 plus cetuximab), who had a cetuximab-refractory, HPV(-) HNSCC with a durable complete response.
  • Three samples were from three patients treated in the CDX3379-02 trial (CDX-3379 monotherapy), including a patient with an exceptional clinical response (92% tumor shrinkage assessed by physical examination after 2 doses of CDX-3379 once every two weeks) (Duvvuri et al ., 2018, Cancer Research 78(13)suppl:CT058).
  • the remaining samples were from 14 patients treated in the CDX3379-04 trial (CDX-3379 plus cetuximab), including a patient who had an ongoing durable complete response (11+ months) and a patient who had an unconfirmed partial response.
  • the biomarker analysis results are shown in FIG. 3.
  • FAT1 mutations were identified in all patients considered responders in this analysis.
  • inactivating mutations in the FAT1 and NOTCH genes have been identified by the Cancer Genome Atlas Network in 32% and 26% of HPV(-) HNSCC tumors, respectively (The Cancer Genome Atlas Network, Nature 517 (7536):576-582).
  • Example 5 FAT1 knockdown enhanced the anti-proliferative activity of CDX-3379 in head and neck squamous cell carcinoma (HNSCC) cells.
  • HNSCC head and neck squamous cell carcinoma
  • Cal27 cells (American Type Culture Collection (ATCC)) were plated at 4,000 cells/well in a 6-well plate overnight at 37°C in antibiotic free medium.
  • Non-targeting and FATl-specific siRNA (Dharmacon) were diluted to 5 mM using lx siRNA buffer.
  • cells were transfected with diluted siRNAs using DharmaFECT ® reagent (Dharmacon) as recommended, following manufacturer’s protocol.
  • DharmaFECT ® reagent Dharmacon
  • transfection media was removed and cells were treated with either 250 nM human IgGl isotype control or CDX-3379, diluted in MEM (Minimum Essential Medium) with 4% FBS (fetal bovine serum).
  • MEM Minimum Essential Medium
  • FBS fetal bovine serum
  • HNSCC head and neck squamous cell carcinoma

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Abstract

La présente invention concerne des méthodes de traitement de cancers associés à HERS exprimant un biomarqueur spécifique utilisant des inhibiteurs de HERS; ainsi que des biomarqueurs et des utilisations de ces derniers pour déterminer la probabilité d'efficacité d'un traitement anticancéreux avec des inhibiteurs de HERS. L'invention concerne également des méthodes de traitement d'un cancer de la tête et du cou associé à HERS avec des inhibiteurs de HERS, le site tumoral primaire étant la cavité buccale.
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Citations (2)

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WO2010093379A1 (fr) * 2009-02-13 2010-08-19 Fred Hutchinson Cancer Research Center L'établissement de profils d'expression génétique permet d'identifier des gènes prédictifs d'un carcinome malpighien de la cavité buccale et son pronostic
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WO2010093379A1 (fr) * 2009-02-13 2010-08-19 Fred Hutchinson Cancer Research Center L'établissement de profils d'expression génétique permet d'identifier des gènes prédictifs d'un carcinome malpighien de la cavité buccale et son pronostic
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