Cell CTL (BiAT) with bispecific antibody as well as preparation method and application thereof

A bispecific antibody and specific technology, applied in animal cells, vertebrate cells, blood/immune system cells, etc., can solve problems such as immune recognition barriers, limit the efficacy of tumor immunotherapy, and unclear targeting, and achieve Increase the number of targets, kill efficiently, and improve the effect of targeting

Active Publication Date: 2015-03-25
SHENZHEN BEIKE BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The tumor killing effect of CIK is low, and the targeting is not obvious. The combination of DC and DC-CTL technology can present tumor-associated antigens (tumor-associated antigens, TAAs) to a certain extent, but the multi-target specific tumor killing effect is still limited , and the multi-target effect can specifically solve the immune escape of tumors by down-regulating the expression of some TAAs to reduce their immunogenicity; although CTL has a specific and relatively high-ef...

Method used

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  • Cell CTL (BiAT) with bispecific antibody as well as preparation method and application thereof
  • Cell CTL (BiAT) with bispecific antibody as well as preparation method and application thereof
  • Cell CTL (BiAT) with bispecific antibody as well as preparation method and application thereof

Examples

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Effect test

Embodiment 1

[0055] Example 1. Preparation of cell CTL (BiAT) with bispecific antibody

[0056] Preparation of cell CTL (BiAT) with bispecific antibody comprises the following steps:

[0057] 1) Mix the CD3 monoclonal antibody and one or more monoclonal antibodies against TAAs in an equimolar ratio, and combine them to produce a bispecific antibody BiAb;

[0058] 2) Add specific cytotoxic T lymphocytes (CTL) to the bispecific antibody BiAb and incubate them for equipping to obtain cell CTL (BiAT) with bispecific antibody.

[0059] 1. Preparation of specific cytotoxic T lymphocytes (CTL)

[0060] The method for obtaining specific cytotoxic T lymphocytes (CTL) in the step 2) may include the following steps:

[0061] 2.1. DC culture and antigen loading

[0062] Mononuclear cells (PBMC) were collected by leukocyte apheresis and resuspended in serum-free medium (purchased from Nipro) at 37°C, 5% CO 2The concentration was incubated in an incubator for 3 hours, the adherent cells (DC precurso...

Embodiment 2

[0085] Example 2. Antitumor effect of cell CTL (BiAT) with bispecific antibody

[0086] 1. In vitro tumor cell-targeted killing effect of cellular CTL (BiAT) with bispecific antibody

[0087] HER2(+) breast adenocarcinoma SK-BR-3, human malignant glioma cell A-172, human gastric cancer cell SGC-7901 and HER2(-) hepatocellular carcinoma cell Hep3B were used as target cells, CTL, CTL (BiAT-HER2), CTL (BiAT-HER2 / EGFR), CIK and Bi-CIK (HER2) were used as effector cells, and T lymphocytes and PBS were used as controls. Target cells 4×10 3 Inoculate / well in a 96-well plate at 37°C, 5% CO 2 Cultivate overnight in the incubator, add various effector cells at the effect-to-target ratio of 10:1, 20:1, 40:1, and 60:1 on the second day, and store at 37°C, 5% CO 2 Incubate together in the incubator for 24h, add MTS 20μl / well, place at 37°C, 5% CO 2 Incubated in the incubator for 4h, the optical density (OD) value of the enzyme label detector detection wavelength 495nm, killing rate (%)...

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Abstract

The invention discloses cell CTL (BiAT) with a bispecific antibody as well as a preparation method and an application thereof. The cytotoxicity T lymphocyte (CTL) with the bispecific antibody is prepared by assembling CTL cells by combining a CD3 monoclonal antibody and any antibody aiming at tumor-associated antigens (TAAs), and the CTL is named as CTL (BiAT) cells. The target binding of targeting specificity CTL and one tumor-associated antigen or more tumor-associated antigens is generated due to combined use of multiple tumor-associated antigen epitope peptides to stimulate DC induction, the targeting action of the CTL cells is ensured due to dual mechanisms, the number of the targets acting on the CTL cells is increased, the targeting property of the CTL cellular immune effect is further improved, and tumor cells can be efficiently killed.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to targeted immune cell therapy drugs for malignant tumors, in particular to a cell CTL (BiAT) with bispecific antibodies and a preparation method thereof, and its use in the preparation of enhanced specific cytotoxic T lymphocytes (CTL) in the application of the medicine of immune killing effect. Background technique [0002] Immune cell therapy is currently the mainstream treatment method for malignant tumor immunotherapy. It collects a certain amount of mononuclear cells from the peripheral blood of tumor patients, cultures them in vitro in a GMP laboratory, expands and activates them to obtain immune activity with anti-tumor effects. cells, or induce targeted treatment with tumor antigens, and then reinfuse them into the patient's body through intravenous, intradermal injection, intervention, etc. ability. [0003] At present, adoptive tumor immune cell therapy technology is commonl...

Claims

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Application Information

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IPC IPC(8): C12N5/0783C12N5/0784A61K35/17A61K35/15A61P35/00
Inventor 胡祥孙平刘沐芸丁长才张芸
Owner SHENZHEN BEIKE BIOTECH
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