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JP-7856432-B2 - Method for manufacturing eribulin-based antibody-drug conjugates

JP7856432B2JP 7856432 B2JP7856432 B2JP 7856432B2JP-7856432-B2

Inventors

  • 松尾 公博
  • 中村 太樹
  • 宮下 祐輔

Assignees

  • エーザイ・アール・アンド・ディー・マネジメント株式会社

Dates

Publication Date
20260511
Application Date
20201221
Priority Date
20191223

Claims (13)

  1. A method for producing a compound represented by formula (B), [In equation (B), m is an integer between 1 and 10.] (i) The compound represented by formula (1) [In equation (1), m is an integer between 1 and 10.] By reacting with the compound represented by formula (2), To obtain the compound represented by formula (3), [In equation (3), m is an integer between 1 and 10.] (ii) Producing a compound represented by formula (A) by acyling a hydroxyl group in a compound represented by formula (3) using an acyling agent, wherein the acyling agent contains a phenoxy group or a nitrophenoxy group, (iii) Compound represented by formula (A) [In formula (A), m is an integer between 1 and 10, and X is a phenoxy group or a nitrophenoxy group.] A method comprising reacting eribulin or a salt thereof with a compound represented by formula (B).
  2. A method for producing a compound represented by formula (B), [In equation (B), m is an integer between 1 and 10.] (a) an acylating agent containing a phenoxy group or a nitrophenoxy group, (b) a compound represented by formula (3). [In equation (3), m is an integer between 1 and 10.] , and (c) preparing a mixture containing eribulin or a salt thereof, Using the aforementioned acylating agent, the hydroxyl group in the compound represented by formula (3) is acylated to obtain the compound represented by formula (A). [In formula (A), m is an integer between 1 and 10, and X is a phenoxy group or a nitrophenoxy group.] To obtain, A method comprising reacting a compound represented by formula (A) with eribulin or a salt thereof to produce a compound represented by formula (B).
  3. The method according to claim 1 or 2 , wherein eribulin or a salt thereof is a methanesulfonate of eribulin.
  4. The method according to any one of claims 1 to 3 , wherein the reaction of the compound represented by formula (A) with eribulin or a salt thereof is carried out in the presence of a base.
  5. The method according to claim 4 , wherein the base is a tertiary amine or a nitrogen-containing aromatic compound.
  6. The method according to claim 5, wherein the tertiary amine is triethylamine or N,N-diisopropylethylamine.
  7. The method according to claim 5 or 6, wherein the nitrogen-containing aromatic compound is pyridine or 2,6-lutidine.
  8. The method according to any one of claims 1 to 7, wherein the acylating agent is diphenyl carbonate, bis(4-nitrophenyl) carbonate, bis(2-nitrophenyl) carbonate, bis(3-nitrophenyl) carbonate, phenyl chloroformate, 4-nitrophenyl chloroformate, 2 -nitrophenyl chloroformate, or 3-nitrophenyl chloroformate.
  9. A method for producing an antibody-drug conjugate represented by formula (I), [In the formula, Ab is an antibody or its antigen-binding fragment, D is eribulin, m is an integer from 1 to 10, and p is an integer from 1 to 8.] (i) The compound represented by formula (1) [In equation (1), m is an integer between 1 and 10.] It is reacted with the compound represented by formula (2), To obtain the compound represented by formula (3), [In equation (3), m is an integer between 1 and 10.] (ii) Using an acylating agent to acylate a hydroxyl group in a compound represented by formula (3), wherein the acylating agent contains a phenoxy group or a nitrophenoxy group, thereby acylating a compound represented by formula (A). [In formula (A), m is an integer between 1 and 10, and X is a phenoxy group or a nitrophenoxy group.] To manufacture, (iii) Reacting eribulin or a salt thereof with a compound represented by formula (A) to obtain a compound represented by formula (B), [In equation (B), m is an integer between 1 and 10.] (iv) A method comprising reacting a compound represented by formula (B) with an antibody or an antigen-binding fragment thereof to obtain an antibody-drug conjugate represented by formula (I).
  10. (A) The Ab comprises an antibody or antigen-binding fragment that binds to folate receptor alpha (FRA), and the antibody or antigen-binding fragment is (i) Three heavy chain CDR amino acid sequences represented by SEQ ID NO: 2 (heavy chain CDR1), SEQ ID NO: 3 (heavy chain CDR2), and SEQ ID NO: 4 (heavy chain CDR3), as defined by the Kabat numbering system, and three light chain CDR amino acid sequences represented by SEQ ID NO: 7 (light chain CDR1), SEQ ID NO: 8 (light chain CDR2), and SEQ ID NO: 9 (light chain CDR3), (ii) Three heavy chain CDR amino acid sequences represented by SEQ ID NO: 13 (heavy chain CDR1), SEQ ID NO: 14 (heavy chain CDR2), and SEQ ID NO: 15 (heavy chain CDR3), as defined by the IMGT numbering system, and three light chain CDR amino acid sequences represented by SEQ ID NO: 16 (light chain CDR1), SEQ ID NO: 17 (light chain CDR2), and SEQ ID NO: 18 (light chain CDR3), (iii) A heavy chain region containing the amino acid sequence represented by SEQ ID NO: 1, and a light chain region containing the amino acid sequence represented by SEQ ID NO: 6, or (iv) A heavy chain variable region containing the amino acid sequence represented by SEQ ID NO: 23, and a light chain variable region containing the amino acid sequence represented by SEQ ID NO: 24, Does it include, (B) The above Ab comprises an antibody or antigen-binding fragment that binds to human epidermal growth factor receptor 2 (HER2), and the above antibody or antigen-binding fragment is (v) Three heavy chain CDR amino acid sequences represented by SEQ ID NO: 71 (heavy chain CDR1), SEQ ID NO: 72 (heavy chain CDR2), and SEQ ID NO: 73 (heavy chain CDR3), as defined by the Kabat numbering system, and three light chain CDR amino acid sequences represented by SEQ ID NO: 74 (light chain CDR1), SEQ ID NO: 75 (light chain CDR2), and SEQ ID NO: 76 (light chain CDR3), (vi) Three heavy chain CDR amino acid sequences represented by SEQ ID NO: 191 ( heavy chain CDR1), SEQ ID NO: 192 (heavy chain CDR2) , and SEQ ID NO: 193 (heavy chain CDR3) , as defined by the IMGT numbering system, and three light chain CDR amino acid sequences represented by SEQ ID NO: 194 (light chain CDR1), SEQ ID NO: 195 (light chain CDR2), and SEQ ID NO: 196 (light chain CDR3), (vii) A heavy chain variable region containing the amino acid sequence represented by SEQ ID NO: 27, and a light chain variable region containing the amino acid sequence represented by SEQ ID NO: 28, or (viiii) A heavy chain region containing the amino acid sequence represented by SEQ ID NO : 307 or 347, and a light chain region containing the amino acid sequence represented by SEQ ID NO: 308, including, or (C) The Ab comprises an antibody or antigen-binding fragment that binds to mesothelin, and the antibody or antigen-binding fragment is (ix) Three heavy chain CDR amino acid sequences represented by SEQ ID NO: 65 (heavy chain CDR1), SEQ ID NO: 66 (heavy chain CDR2), and SEQ ID NO: 67 (heavy chain CDR3), as defined by the Kabat numbering system, and three light chain CDR amino acid sequences represented by SEQ ID NO: 68 (light chain CDR1), SEQ ID NO: 69 (light chain CDR2), and SEQ ID NO: 70 (light chain CDR3), (x) Three heavy chain CDR amino acid sequences represented by SEQ ID NO: 185 (heavy chain CDR1), SEQ ID NO: 186 (heavy chain CDR2), and SEQ ID NO: 187 (heavy chain CDR3), as defined by the IMGT numbering system, and three light chain CDR amino acid sequences represented by SEQ ID NO: 188 (light chain CDR1), SEQ ID NO: 189 (light chain CDR2), and SEQ ID NO: 190 (light chain CDR3), The method according to claim 9, comprising (xi) a heavy chain variable region containing the amino acid sequence represented by SEQ ID NO: 25, and a light chain variable region containing the amino acid sequence represented by SEQ ID NO: 26, and (xi) a heavy chain region containing the amino acid sequence represented by SEQ ID NO: 305, and a light chain region containing the amino acid sequence represented by SEQ ID NO: 306 .
  11. The method according to claim 9 or 10 , wherein p is 3 or 4.
  12. The method according to any one of claims 9 to 11 , wherein eribulin or a salt thereof is a methanesulfonate of eribulin.
  13. The method according to any one of claims 9 to 12, wherein the acylating agent is diphenyl carbonate, bis(4-nitrophenyl) carbonate, bis(2-nitrophenyl) carbonate, bis(3-nitrophenyl) carbonate, phenyl chloroformate, 4-nitrophenyl chloroformate, 2 -nitrophenyl chloroformate, or 3-nitrophenyl chloroformate.

Description

This invention relates to a method for producing an eribulin-based antibody-drug conjugate (ADC). Cancer is one of the leading causes of disease and death worldwide, with approximately 14 million new cases and 8.2 million cancer-related deaths in 2012. The most common causes of cancer death are lung cancer (1.59 million deaths), liver cancer (745,000 deaths), stomach cancer (723,000 deaths), colorectal cancer (694,000 deaths), breast cancer (521,000 deaths), and esophageal cancer (400,000 deaths). The number of new cancer cases is projected to increase by approximately 70% in the next 20 years, reaching approximately 22 million new cases per year (Non-Patent Literature 1). Microtubules are dynamic, fibrous cytoskeletal proteins involved in a variety of cellular functions, including intracellular migration and transport, cell signaling, and maintenance of cell shape. Microtubules also play a crucial role in mitotic cell division by forming the mitotic spindle, which is necessary for separating chromosomes into two daughter cells. The biological function of microtubules in all cells is largely regulated by their polymerization dynamics, which result from the reversible non-covalent addition of α and β tubulin dimers at both ends of the microtubule. This dynamic behavior and the resulting control over microtubule length are critical to proper spindle function. Even slight changes in microtubule dynamics can trigger spindle checkpoints, halting cell cycle progression in mitosis and subsequently leading to cell death (Non-Patent Literature 2). Due to their rapid cell division, cancer cells are generally more sensitive than normal cells to compounds that bind to tubulin and disrupt its normal function. For this reason, tubulin inhibitors and other microtubule-targeting drugs represent a promising class of drugs for treating cancer (Non-Patent Literature 3). Folate receptor alpha (FRA) is a glycophosphatidylinositol (GPI)-binding membrane protein that binds to folate. While the role of FRA in the biology of normal and cancerous tissues is not fully understood, it is highly overexpressed in a high percentage of epithelial ovarian cancers (Non-Patent Literature 4) and also highly overexpressed in a certain percentage of non-small cell lung cancers (Non-Patent Literature 5). FRA expression is also restricted in normal tissues. These characteristics make FRA an attractive target for cancer immunotherapy. The proto-oncogene human epidermal growth factor receptor 2 (HER2) encodes a transmembrane tyrosine kinase receptor belonging to the human epidermal growth factor receptor (EGFR) family (Non-Patent Literature 6). Overexpression of HER2 enables constitutive activation of growth factor signaling pathways such as the PI3K-AKT-mTOR pathway, thereby serving as an oncogenic driver in several cancers, including approximately 20% of invasive breast cancers (Non-Patent Literature 7-8). Because HER2 amplification mediates a transformative phenotype, HER2 is another promising target for cancer treatment. International Publication No. 2017/151979 World Cancer Report 2014.Mukhtar et al. (2014) Mol. Cancer Ther. 13:275-84.Dumontet and Jordan (2010) Nat. Rev. Drug Discov. 9:790-803.O’Shannessy et al. (2013) Int. J. Gynecol. Pathol. 32(3):258-68.Christoph et al. (2014) Clin. Lung Cancer 15(5):320-30.King et al. (1985) Science 229:974-6.Slamon et al. (1989) Science 244:707-12.Gajria and Chandarlapaty (2011) Expert Rev. Anticancer Ther.11:263-75.O’Shannessy et al., (2011) Oncotarget 2: 1227-43. Embodiments of the present invention will be described in detail below. One embodiment of the present invention relates to a method for producing an antibody-drug conjugate (ADC) represented by formula (I). Therefore, let's first explain the ADC represented by equation (I). ADCs can bind to tumor cells (e.g., FRA-expressing tumor cells), internalize them, and kill them. Furthermore, the antibody portion (Ab) used in the ADC is preferably an antibody or its antigen-binding fragment, which targets tumor cells. The antibody or its antigen-binding fragment may be, for example, (a) comprising three heavy chain CDRs containing the amino acid sequences of heavy chain complementarity determining region (heavy chain CDR) 1 represented by SEQ ID NO: 2, heavy chain CDR 2 represented by SEQ ID NO: 3, and heavy chain CDR 3 represented by SEQ ID NO: 4, as defined by the Kabat numbering system (Kabat, Sequences of Proteins of Immunological Interest (National Institutes of Health, Bethesda, Md. (1987 and 1991))), and three light chain CDRs containing the amino acid sequences of light chain complementarity determining region (light chain CDR) 1 represented by SEQ ID NO: 7, light chain CDR 2 represented by SEQ ID NO: 8, and light chain CDR 3 represented by SEQ ID NO: 9, or, (b) comprising three heavy chain CDRs including the amino acid sequences of heavy chain CDR1 represented by SEQ ID NO: 13, heavy chain CDR2 represented by SEQ ID NO: 14, and heavy chain CDR3 r