Anti-cdh6 antibody-pyrrolobenzodiazepine derivative conjugate

An antibody-drug conjugate represented by the following formula (XII):

• wherein, in each structural formula shown above,
• m2 represents an integer of 1 or 2,
• Ab represents an IgG antibody specifically binding to an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO. 4 and having internalization ability that permits cellular uptake, or a functional fragment thereof,
• the N297 glycan of Ab represents any one of N297-(Fuc)MSG1, N297-(Fuc)MSG2, and a mixture thereof, and N297-(Fuc)SG, with N297-(Fuc)MSG1, N297-(Fuc)MSG2, and N297-(Fuc)SG having structures represented by the following formulas,
• wherein each wavy line represents bonding to Asn297 (N297) of the antibody,
• *-L(PEG)- in the N297 glycan represents *—(CH2CH2—O)3—CH2CH2—NH—, wherein
• the amino group at the right end is bound via an amide bond to the carboxylic acid at the 2-position of the sialic acid at the non-reducing terminal in each or either one of the 1-3 and 1-6 branched chains of the β-Man in the N297 glycan, and each asterisk * represents bonding to the nitrogen atom at the 1- or 3-position of the triazole ring in the corresponding structural formula.

An antibody-drug conjugate represented by the following formula (XIII):

• wherein, in each structural formula shown above,
• m2 represents an integer of 1 or 2,
• Ab represents an IgG antibody specifically binding to an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO:
• 4 and having internalization ability that permits cellular uptake, or a functional fragment thereof, and
• the N297 glycan of Ab represents any one of N297-(Fuc)MSG1, N297-(Fuc)MSG2, and a mixture thereof, and N297-(Fuc)SG, with N297-(Fuc)MSG1, N297-(Fuc)MSG2, and N297-(Fuc)SG having structures represented by the following formulas,
• wherein each wavy line represents bonding to Asn297 (N297) of the antibody,
• *-L(PEG)- in the N297 glycan represents *—(CH2CH2—O)3—CH2CH2—NH—, wherein the amino group at the right end is bound via an amide bond to the carboxylic acid at the 2-position of the sialic acid at the non-reducing terminal in each or either one of the 1-3 and 1-6 branched chains of the β-Man in the N297 glycan, and each asterisk * represents bonding to the nitrogen atom at the 1- or 3-position of the triazole ring in the individual structural formulas.

An antibody-drug conjugate represented by the following formula (XIV):

• wherein, in each structural formula shown above,
• m2 represents an integer of 1 or 2,
• Ab represents an IgG antibody specifically binding to an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO:
• 4 and having internalization ability that permits cellular uptake, or a functional fragment thereof, and
• the N297 glycan of Ab represents any one of N297-(Fuc)MSG1, N297-(Fuc)MSG2, and a mixture thereof, and N297-(Fuc)SG, with N297-(Fuc)MSG1, N297-(Fuc)MSG2, and N297-(Fuc)SG having structures represented by the following formulas:
• wherein each wavy line represents bonding to Asn297 (N297) of the antibody,
• *-L(PEG)- in the N297 glycan represents *—(CH2CH2—O)3—CH2CH2—NH—, wherein the amino group at the right end is bound via an amide bond to the carboxylic acid at the 2-position of the sialic acid at the non-reducing terminal in each or either one of the 1-3 and 1-6 branched chains of the β-Man in the N297 glycan, and each asterisk * represents bonding to the nitrogen atom at the 1- or 3-position of the triazole ring in the individual structural formulas.

An antibody-drug conjugate represented by the following formula (XV):

• wherein, in each structural formula shown above,
• m2 represents an integer of 1 or 2,
• Ab represents an IgG antibody specifically binding to an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO:
• 4 and having internalization ability that permits cellular uptake, or a functional fragment thereof, and
• the N297 glycan of Ab represents any one of N297-(Fuc)MSG1, N297-(Fuc)MSG2, and a mixture thereof, and N297-(Fuc)SG, with N297-(Fuc)MSG1, N297-(Fuc)MSG2, and N297-(Fuc)SG having structures represented by the following formulas:
• wherein each wavy line represents bonding to Asn297 (N297) of the antibody,
• *-L(PEG)- in the N297 glycan represents *—(CH2CH2—O)3—CH2CH2—NH—, wherein the amino group at the right end is bound via an amide bond to the carboxylic acid at the 2-position of the sialic acid at the non-reducing terminal in each or either one of the 1-3 and 1-6 branched chains of the β-Man in the N297 glycan, and each asterisk * represents bonding to the nitrogen atom at the 1- or 3-position of the triazole ring in the individual structural formulas.

A method for producing a glycan-remodeled antibody, the method comprising the steps of:

• i) producing an IgG antibody specifically binding to an amino acid sequence comprising the amino acid sequence shown in SEQ ID NO:
• 4 and having internalization ability that permits cellular uptake, or a functional fragment thereof,
• ii) treating the antibody obtained in step i) with hydrolase to produce a (Fucα1,6)GlcNAc-antibody;
• and
• iii)-1 reacting the (Fucα1,6)GlcNAc-antibody and a glycan donor molecule in the presence of a transglycosidase, the glycan donor molecule obtained by introducing a PEG linker having an azide group to the carbonyl group of carboxylic acid at the 2-position of a sialic acid in MSG (9) or SG (10) and oxazolinating the reducing terminal, or
• iii)-2 reacting the (Fucα1,6)GlcNAc-antibody and a glycan donor molecule in the presence of a transglycosidase, the glycan donor molecule obtained by introducing a PEG linker having an azide group to the carbonyl group of carboxylic acid at the 2-position of a sialic acid in (MSG-)Asn or (SG-)Asn with an α-amino group optionally protected and to the carbonyl group of carboxylic acid in the Asn, causing action of hydrolase, and then oxazolinating the reducing terminal.

An antibody or a functional fragment thereof, comprising a light chain variable region, which contains CDRL1 consisting of the amino acid sequence shown in SEQ ID NO:

• 12, CDRL2 consisting of the amino acid sequence shown in SEQ ID NO:
• 13 and CDRL3 consisting of the amino acid sequence shown in SEQ ID NO:
• 14, and a heavy chain variable region, which contains CDRH1 consisting of the amino acid sequence shown in SEQ ID NO:
• 57, CDRH2 consisting of the amino acid sequence shown in SEQ ID NO:
• 58 and CDRH3 consisting of the amino acid sequence shown in SEQ ID NO:
• 59.

An antibody or a functional fragment thereof, comprising a light chain variable region, which contains CDRL1 consisting of the amino acid sequence shown in SEQ ID NO:

• 12, CDRL2 consisting of the amino acid sequence shown in SEQ ID NO:
• 13 and CDRL3 consisting of the amino acid sequence shown in SEQ ID NO:
• 14;
• and a heavy chain variable region, which contains CDRH1 consisting of the amino acid sequence shown in SEQ ID NO:
• 62, CDRH2 consisting of the amino acid sequence shown in SEQ ID NO:
• 63 and CDRH3 consisting of the amino acid sequence shown in SEQ ID NO:
• 64.