Anti-HER2/anti-PD-L1 bifunctional antibody and application thereof

A bifunctional antibody, PD-L1 technology, applied in the field of molecular immunology, can solve problems such as low production efficiency, poor pharmacokinetic performance, and difficulties in the development of bispecific antibodies

Active Publication Date: 2022-01-18
DARTSBIO PHARM LTD +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there are more than 40 types of bifunctional antibody forms that have been developed, but due to problems such as low production efficiency and poor pharmacokinetic performance, the development of bispecific antibodies has always been difficult
At present, there is no anti-HER2 / anti-PD-L1 bifunctional antibody at home and abroad

Method used

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  • Anti-HER2/anti-PD-L1 bifunctional antibody and application thereof
  • Anti-HER2/anti-PD-L1 bifunctional antibody and application thereof
  • Anti-HER2/anti-PD-L1 bifunctional antibody and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0058] Example 1. Thermal stability screening

[0059] The concentration of anti-HER2 / anti-PD-L1 bifunctional antibody was adjusted to 10mg / ml. Incubate for 1 h at room temperature, 60°C, and 70°C in a water bath, and then evaluate the anti-HER2 / anti-PD-L1 bifunctional antibody (hereinafter also referred to as "HER2 / PD-L1 bifunctional antibody" or "bifunctional antibody") in a binding ELISA. Antibody (bsAb)") for binding human PD-L1 and human HER2 antigens. A 96-well immunoplate was coated with 2 μg / ml His-tagged antigen overnight at 4°C. Three-fold serial dilutions of the HER2 / PD-L1 diabody and trastuzumab were added to the wells. After one hour of incubation, the bound HER2 / PD-L1 diabody was detected with horseradish peroxidase (HRP)-conjugated goat anti-human IgG (H+L), and 3,3′,5,5′-tetramethyl benzidine substrate for development. It was then detected at 450 nm with a SpectraMaxM5e (Molecular Devices) microplate reader. The result is as Figure 4 As shown, the bindin...

Embodiment 2

[0060] Embodiment 2. Light stability screening

[0061] The photodegradation of HER2 / PD-L1 bifunctional antibody was carried out according to the ICH guidelines (Q1B, 1997) in Zuocheng SHH-3SDT series photostability test chamber (1 cycle-1.2 million lux hours of visible light (400-700nm), 200Wh Determination of UVA light (320-400nm) per square meter. After the dark control group and the samples exposed to light were compressed with rubber stoppers, the 15ml glass vials were stored in a photostability chamber. HER2 / PD-L1 bifunctional antibody Concentration is 10mg / ml.Then, carries out the activity measurement of sample in conjunction with ELISA.Result is as Figure 5 As shown, the binding activity between 77-1#bsAb and hu-PD-L1 is almost undetectable at 100nM antibody concentration; the binding EC50 value of 97-1#bsAb to hu-PD-L1 after light irradiation is 4.825nM and untreated group Compared with the EC50 value of 0.2761nM, it decreased by 17 times; the EC50 values ​​of 262-2...

Embodiment 3

[0062] Example 3. Biofilm Interferometry (BLI)

[0063] Dilute the HER2 / PD-L1 bifunctional antibody to 10 μg / ml in the sample buffer (0.02% Tween 20 and 0.1% BSA in PBS), and dilute the antigens hu-HER2 and hu-PD-L1 in the sample buffer Diluted to 100nM. Binding of HER2 / PD-L1 diabodies to specific hu-HER2 and hu-PD-L1 antigens was analyzed by BLI (forteBIO; Pall). All measurements were performed in sample buffer at room temperature. The bsAb (10 μg / ml) was immobilized on the proteinA biosensor and the antigen was titrated from 100 nM to obtain rate constants and affinities. KD values ​​were obtained by applying a 1:1 binding isotherm using the software provided by the vendor. The result is as Figure 6 As shown, the binding affinity of 262-2#bsAb to hu-PD-L1 is stronger than that of the positive control antibody Avelumab; the binding affinity of 262-2#bsAb to hu-HER2 is consistent with that of the positive control antibody trastuzumab.

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Abstract

The invention relates to an anti-HER2/anti-PD-L1 bifunctional antibody and an application thereof. The anti-HER2/anti-PD-L1 bifunctional antibody comprises: a monoclonal antibody unit, which aims at HER2 and comprises two heavy chains and two light chains; a single-chain antibody unit aims at PD-L1 and comprises two single-chain antibodies, each single-chain antibody comprises a heavy chain variable region and a light chain variable region, the heavy chain variable region and the light chain variable region are connected through a connecting peptide, and N ends or C ends of the two single-chain antibodies are respectively connected with C ends of Fc fragments of two heavy chains of the monoclonal antibody unit through linker. The anti-HER2/anti-PD-L1 bifunctional antibody disclosed by the invention can be combined with PD-L1 and also can be combined with HER2, so that the anti-HER2/anti-PD-L1 bifunctional antibody is a novel antitumor drug.

Description

technical field [0001] The invention belongs to the technical field of molecular immunology, and specifically relates to an anti-HER2 / anti-PD-L1 bifunctional antibody and its application. Background technique [0002] HER2 overexpression has been detected in a variety of solid tumors, including breast cancer (BC), colorectal cancer (CRC), gastric and gastroesophageal junction (GEJ) cancer, non-small cell lung cancer (NSCLC), biliary tract cancer (BTC) and bladder cancer. Trastuzumab It is a humanized monoclonal antibody (mAb) against HER2 and is an effective therapy against HER2+ tumors. Treatment of HER2+ BC patients has been significantly improved through the use of targeted therapeutics. However, only 50–80% of HER2+BC patients benefit from targeted therapeutics, while 20–50% of patients are unresponsive from treatment initiation or develop resistance after treatment. In recent years, immune checkpoint inhibitors have become a mainstay of treatment for a variety of c...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/46C12N15/62C12N15/85C12N5/10A61K39/395A61P35/00
CPCC07K16/32C07K16/2827C12N15/85A61K39/3955A61K39/39533A61P35/00C07K2317/24C07K2317/31C07K2317/56C07K2317/565C07K2317/51C07K2317/515C07K2317/622C07K2317/92A61K2039/507
Inventor 陈艺丽王春河刘国键刘鑫园谭杰陈淦均唐磊丁华平吴委涛洪一峯
Owner DARTSBIO PHARM LTD
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