A method for preparing α-2,3-sialyl glycosyltransferase-mediated site-specific conjugation antibody-drug conjugates and its application.

By using α-2,3-sialyl glycosyltransferase-mediated site-specific glycan conjugation technology, the problems of ADC uniformity and stability were solved, enabling the efficient preparation of antibody-drug conjugates with controllable DAR values, thus improving efficacy and production stability.

CN121271994BActive Publication Date: 2026-05-26WUHAN TANGZHI PHARM CO LTD
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Patent Information

Authority / Receiving Office
CN · China
Patent Type
Patents(China)
Current Assignee / Owner
WUHAN TANGZHI PHARM CO LTD
Filing Date
2025-12-04
Publication Date
2026-05-26

AI Technical Summary

Technical Problem

Existing antibody-drug conjugates (ADCs) suffer from poor uniformity, low stability, and unstable efficacy in traditional conjugation methods. In particular, the site-specific and quantitative conjugation of antibodies with small molecule toxins presents challenges, affecting efficacy and therapeutic window.

Method used

Using α-2,3-sialic acid glycosyltransferase-mediated site-directed glycan coupling technology, human β-1,4-galactosyltransferase and α-2,3-sialic acid glycosyltransferase were used to transfer galactose and azide-modified sialic acid to the glycan ends, respectively, and combined with click chemistry to prepare ADCs with controllable DAR values.

Benefits of technology

It achieves high uniformity, stability and reproducibility of ADCs, improves drug efficacy, and prepares ADCs with DAR values ​​of 2 or 4 through one-pot enzymatic method and three-step method, with higher catalytic activity and better quality control, which is suitable for large-scale production of ADCs.

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Abstract

This invention provides a method for preparing α-2,3-sialyl glycosyltransferase-mediated site-specific conjugation antibody-drug conjugates, comprising the following steps: S1, dissolving the antibody and adding UDP-Gal and Mn. 2+ The process involves several steps: S1) β-1,4-galactosyltransferase reaction, followed by purification to obtain antibody I with galactose-terminated glycans; S2) Dissolving antibody I, adding azide-modified CMP sialic acid derivative, α-2,3-sialic acid glycosyltransferase, and alkaline phosphatase, followed by purification to obtain antibody II with sialic acid derivatives at the glycan ends; S3) Dissolving toxin molecules in DMSO, adding them to antibody II, reacting, and then ultrafiltration to obtain the ADC molecule; or directly preparing the ADC molecule using a one-pot enzymatic method. This invention utilizes the screened ST3Gal3 for site-specific conjugation of ADCs, offering controllable DAR values, batch-to-batch stability, high homogeneity, high reproducibility, better quality control and CMC, thereby improving ADC efficacy.
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