﻿pd-l1 magnetic bead-based pd-1/pd-l1 blocking assay

Flow Analysis of PDL1-Vaxx Antibody Blockade Using Magnetic Fluorescent Beads

In the immune system&#39;s T-cells, B-cells, and intracellular checkpoints, signaling pathways regulate immune activities. Some cancer cells protect themselves from immune attack by stimulating checkpoint targets, which inhibit immune function and promote neoplastic survival and proliferation. Oncologic immunotherapy by checkpoint inhibition uses antibodies to target and block the signaling checkpoints and, thus, restore the anti-neoplastic functions of the immune system1,2,3. Highly effective anti-cancer therapies currently include the monoclonal antibodies nivolumab, which targets programmed death protein 1 (PD-1)4, and atezolizumab, which targets programmed death ligand 1 (PD-L1)5. This approach has shown great clinical success in treating cancer patients. However, the clinical utility of current checkpoint inhibition strategies is mitigated by adverse events and treatment resistance, especially in single-agent therapy6. A combination of immunotherapy and more effective therapeutic strategies with lower toxicity is urgently needed in cancer treatment1,3,6.
Over the past 30 years, Dr. Kaumaya&#39;s laboratory has developed peptide cancer vaccines and peptide mimic-related agents for cancer therapy, some of which are in ongoing clinical trials1,2,7,8,9,10,11,12,13,14. For example, B-Vaxx with HER-2 combination immunotherapy has shown patient benefits against metastatic and/or recurrent solid tumors in clinical trials12. The laboratory&#39;s latest cancer vaccines are PD1-Vaxx2,13 and PDL1-Vaxx14, which have shown great advantages in preclinical studies, especially in combination treatment. The PD1-Vaxx has completed dose-escalation clinical trials in the US and Australia. The PD1-Vaxx will be combined with atezolizumab in the Phase 1b trial to start in May, 2023. This report focuses on evaluating the ability of PDL1-Vaxx-induced antibodies to block the PD-1/PD-L1 interaction.
The PDL1-Vaxx cancer vaccine is a novel B-cell peptide epitope vaccine with PD-L1 amino acids 130-147 linked to the promiscuous T-cell measles virus fusion (MVF) peptide via a GPSL peptide linker. Preclinical studies have shown that PDL1-Vaxx is highly immunogenic in stimulating anti-cancer antibody production in various animal models, prolongs survival, and reduces tumor burden14. These antibodies generated against the PD-L1 peptide can successfully block the PD1/PD-L1 interaction, thus resulting in anti-neoplastic activity. This report introduces an assay that analyzes the blockade of PD1/PD-L1 complex formation by PDL1-Vaxx-induced antibodies using a magnetic bead-based format with a dual-reporter readout on a flow cytometry instrument.

Author Spotlight: Magnetic Fluorescent Bead-Based Dual-Reporter Flow Analysis of PDL1-Vaxx Peptide Vaccine-Induced Antibody Blockade of the PD-1/PD-L1 Interaction 

Magnetic Fluorescent Bead-Based Dual-Reporter Flow Analysis of PDL1-Vaxx Peptide Vaccine-Induced Antibody Blockade of the PD-1/PD-L1 Interaction

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PD-L1 Magnetic Bead-Based PD-1/PD-L1 Blocking Assay  

﻿PD-L1 Magnetic Bead-Based PD-1/PD-L1 Blocking Assay

pd-l1-magnetic-bead-based-pd-1pd-l1-blocking-assay

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JoVE Journal

Cancer Research

310 Views. ﻿PD-L1 Magnetic Bead-Based PD-1/PD-L1 Blocking Assay

Video: PD-L1 Magnetic Bead-Based PD-1/PD-L1 Blocking Assay  

The inhibition of checkpoint receptors (PD-1, PD-L1, and CTLA-4) with monoclonal antibodies has shown great benefit in clinical trials for treating cancer patients and has become a mainstay approach in modern cancer immunotherapy. However, only a subset of patients respond to checkpoint monoclonal antibody immunotherapy. Therefore, it is urgent to develop new therapeutic strategies against cancer. A novel B-cell peptide epitope PDL1 (programmed death ligand 1) cancer vaccine has been developed, with amino acids 130-147 linked to the MVF peptide (&#34;promiscuous&#34; T-cell measles virus fusion protein) via a GPSL linker. Preclinical testing has indicated that this PDL1 vaccine (PDL1-Vaxx) effectively stimulates highly immunogenic antibodies in animals. Animals immunized with PDL1-Vaxx show reduced tumor burden and extended survival rates in various animal cancer models. The mechanisms of action indicate that vaccine-elicited antibodies inhibit tumor cell proliferation, induce apoptosis, and block the PD-1/PD-L1 interaction. This manuscript introduces a magnetic bead-based assay that uses a dual-reporter flow analysis system to evaluate the PD-1/PD-L1 interaction and its blockade by the anti-PDL1 antibodies raised against the PDL1-Vaxx.

Checkpoint inhibitors are important targets in developing therapies for the battle against cancer. This report introduces a novel PDL1 peptide-based cancer vaccine, PDL1-Vaxx, which induces neutralizing polyclonal antibody production that blocks PD-1/PDL1 complex formation. This work also details the development and testing of a fluorescent bead-based assay for analyzing this activity.

The inhibition of checkpoint receptors (PD-1, PD-L1, and CTLA-4) with monoclonal antibodies has shown great benefit in clinical trials for treating cancer ...