TY - JOUR

T1 - Cell Model for Testing Pharmaceuticals Targeting Human PD-L1

AU - Shashkova, O.A.

AU - Terekhina, L.A.

AU - Malakhov, I.S.

AU - Pinevich, A.A.

AU - Vartanyan, N.L.

AU - Avrov, K.O.

AU - Krutetskaya, I.Yu.

AU - Gryazeva, I.V.

AU - Berlina, M.A.

AU - Stolbovaya, A.Yu.

AU - Smirnov, I.V.

AU - Fedorenko, S.V.

AU - Krylova, A.A.

AU - Nadporojskii, M.A.

AU - Shatik, S.V.

AU - Stanzhevskii, A.A.

AU - Samoilovich, M.P.

N1 - Export Date: 18 November 2024 Химические вещества/CAS: atezolizumab, 1380723-44-3

PY - 2024/10/30

Y1 - 2024/10/30

N2 - The aim of this study was to create and evaluate a cell model designed for in vitro and in vivo testing of anti-human PD-L1 therapeutic and diagnostic agents’ specificity. Materials and Methods. Genetically modified cells expressing human PD-L1 (strain CT26-PD-L1) were obtained by retroviral transduction of murine CT26 carcinoma cells. PD-L1 gene activity was assessed by real-time PCR, and PD-L1 expression on cells was identified by flow cytometry. Cells were tested using recombinant single-domain human anti-PD-L1 antibodies (nanoantibodies) conjugated with radioisotopes68 Ga or177 Lu. Immunoreactive fraction and cell internalization of the radioconjugates were evaluated in vitro. For in vivo experiments CT26-PD-L1 cells were transplanted into mice, radioimmunoconjugates were injected 9–14 days later, in 1–48 h the tumors were retrieved and subjected to direct radiometry. Intact CT26 cells not expressing the antigen served as a control. Results. CT26-PD-L1 strain of murine tumor cells expressing human membrane PD-L1 was created. When transplanted into intact BALB/c mice or sublethally irradiated F1(DBA×BALB/c) mice, these cells formed tumors. Thus, a significant advantage of the model was the possibility of in vivo testing of human PD-L1-affinity agents using animals under conventional vivarium conditions. When radioimmunoconjugates were administered to tumor bearing mice, radionuclides accumulated in tumors generated from the transplanted CT26-PD-L1 cells, but not CT26 cells. CT26-PD-L1 cells internalized anti-PD-L1 nanobodies in vitro. Due to a high density of target molecules, CT26-PD-L1 cells allowed both to confirm pharmaceuticals’ specificity and to quantify the target-binding fraction of conjugates in a single test. Conclusion. The created cells are the first genetically engineered cells designed to evaluate affinity of anti-human PD-L1 therapeutic and diagnostic agents in Russia. Test results confirmed the model suitability for in vitro and in vivo testing of the specificity of pharmaceuticals targeting human PD-L1. © 2024, Privolzhsky Research Medical University. All rights reserved.

AB - The aim of this study was to create and evaluate a cell model designed for in vitro and in vivo testing of anti-human PD-L1 therapeutic and diagnostic agents’ specificity. Materials and Methods. Genetically modified cells expressing human PD-L1 (strain CT26-PD-L1) were obtained by retroviral transduction of murine CT26 carcinoma cells. PD-L1 gene activity was assessed by real-time PCR, and PD-L1 expression on cells was identified by flow cytometry. Cells were tested using recombinant single-domain human anti-PD-L1 antibodies (nanoantibodies) conjugated with radioisotopes68 Ga or177 Lu. Immunoreactive fraction and cell internalization of the radioconjugates were evaluated in vitro. For in vivo experiments CT26-PD-L1 cells were transplanted into mice, radioimmunoconjugates were injected 9–14 days later, in 1–48 h the tumors were retrieved and subjected to direct radiometry. Intact CT26 cells not expressing the antigen served as a control. Results. CT26-PD-L1 strain of murine tumor cells expressing human membrane PD-L1 was created. When transplanted into intact BALB/c mice or sublethally irradiated F1(DBA×BALB/c) mice, these cells formed tumors. Thus, a significant advantage of the model was the possibility of in vivo testing of human PD-L1-affinity agents using animals under conventional vivarium conditions. When radioimmunoconjugates were administered to tumor bearing mice, radionuclides accumulated in tumors generated from the transplanted CT26-PD-L1 cells, but not CT26 cells. CT26-PD-L1 cells internalized anti-PD-L1 nanobodies in vitro. Due to a high density of target molecules, CT26-PD-L1 cells allowed both to confirm pharmaceuticals’ specificity and to quantify the target-binding fraction of conjugates in a single test. Conclusion. The created cells are the first genetically engineered cells designed to evaluate affinity of anti-human PD-L1 therapeutic and diagnostic agents in Russia. Test results confirmed the model suitability for in vitro and in vivo testing of the specificity of pharmaceuticals targeting human PD-L1. © 2024, Privolzhsky Research Medical University. All rights reserved.

KW - cell model

KW - CT26

KW - PD-L1

KW - radioconjugate

KW - targeted agent

KW - tumor model

KW - VHH

KW - atezolizumab

KW - bms 936559

KW - nanobody

KW - programmed death 1 ligand 1

KW - radioisotope

KW - animal cell

KW - animal experiment

KW - animal model

KW - animal tissue

KW - Bagg albino mouse

KW - bound fraction

KW - controlled study

KW - drug analysis

KW - drug screening

KW - engineered cell

KW - flow cytometry

KW - gene activity

KW - genetic recombination

KW - human

KW - human cell

KW - in vitro study

KW - internalization (cell)

KW - mouse

KW - nonhuman

KW - radiometry

KW - real time polymerase chain reaction

KW - review

UR - https://www.mendeley.com/catalogue/281860e8-dffd-3948-9f91-eec4a8a8b5e4/

U2 - 10.17691/stm2024.16.5.01

DO - 10.17691/stm2024.16.5.01

M3 - статья

VL - 16

SP - 5

EP - 15

JO - СОВРЕМЕННЫЕ ТЕХНОЛОГИИ В МЕДИЦИНЕ

JF - СОВРЕМЕННЫЕ ТЕХНОЛОГИИ В МЕДИЦИНЕ

SN - 2076-4243

IS - 5

ER -