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- Title
Dual Checkpoint Aptamer Immunotherapy: Unveiling Tailored Cancer Treatment Targeting CTLA-4 and NKG2A.
- Authors
Ayass, Mohamad Ammar; Tripathi, Trivendra; Griko, Natalya; Okyay, Tutku; Ramankutty Nair, Ramya; Zhang, Jin; Zhu, Kevin; Melendez, Kristen; Pashkov, Victor; Abi-Mosleh, Lina
- Abstract
Simple Summary: Cancer remains a significant health challenge, with persistent annual increases in new cases and mortality rates. Notably, breast, prostate, and uterine cancers are surging. Economically, cancer care bears an overwhelming burden, with costs varying based on cancer phase and type of treatment. Immunotherapy, including T cell immune-checkpoint inhibitors and CAR-directed T lymphocytes, is pivotal in modern oncology, targeting CTLA-4 and PD-1/PD-L1 pathways. Recent research underscores CTLA-4's diverse role in immune modulation, impacting T cell activation, IDO induction, and T cell proliferation inhibition. Additionally, recognition of the NKG2A axis, interacting with HLA-E on tumor cells, highlights its significance in immune evasion and cancer progression. Current FDA-approved anti-CTLA-4 therapies exhibit promise but face challenges regarding response rates and immune-related adverse events. This study employed computational methods to design aptamers targeting CTLA4 and NKG2A. Aptamer binding specificity was validated through direct and competitive assays using ELISA and flow cytometry. We demonstrated the biological functionality of AYA22T-R2-13, confirming its specific binding to CTLA-4 and NKG2A, and its potential to enhance CD8 T cell and NK cell activity, promoting in vitro tumor cell lysis in human cell lines. Recent strides in immunotherapy have illuminated the crucial role of CTLA-4 and PD-1/PD-L1 pathways in contemporary oncology, presenting both promises and challenges in response rates and adverse effects. This study employs a computational biology tool (in silico approach) to craft aptamers capable of binding to dual receptors, namely, inhibitory CTLA4 and NKG2A, thereby unleashing both T and NK cells and enhancing CD8+ T and NK cell functions for tumor cell lysis. Computational analysis highlighted AYA22T-R2-13 with HADDOCK scores of −78.2 ± 10.2 (with CTLA4), −60.0 ± 4.2 (with NKG2A), and −77.5 ± 5.6 (with CD94/NKG2A). Confirmation of aptamer binding to targeted proteins was attained via ELISA and flow cytometry methods. In vitro biological functionality was assessed using lactate dehydrogenase (LDH) cytotoxicity assay. Direct and competitive assays using ELISA and flow cytometry demonstrated the selective binding of AYA22T-R2-13 to CTLA4 and NKG2A proteins, as well as to the cell surface receptors of IL-2-stimulated T cells and NK cells. This binding was inhibited in the presence of competition from CTLA4 or NKG2A proteins. Remarkably, the blockade of CTLA4 or NKG2A by AYA22T-R2-13 augmented human CD8 T cell- and NK cell-mediated tumor cell lysis in vitro. Our findings highlight the precise binding specificity of AYA22T-R2-13 for CTLA4-B7-1/B7-2 (CD80/CD86) or CD94/NKG2A-HLA-E interactions, positioning it as a valuable tool for immune checkpoint blockade aptamer research in murine tumor models. These in vitro studies establish a promising foundation for further enhancing binding capacity and establishing efficacy and safety in animal models. Consequently, our results underscore the potential of AYA22T-R2-13 in cancer immunotherapy, offering high specificity, low toxicity, and the potential for cost-effective production.
- Subjects
COMBINATION drug therapy; FLOW cytometry; COMPUTER simulation; IN vitro studies; LECTINS; T cells; KILLER cells; CARRIER proteins; COST effectiveness; RESEARCH funding; ANTINEOPLASTIC agents; ENZYME-linked immunosorbent assay; IMMUNE checkpoint inhibitors; NUCLEOTIDES; BIOINFORMATICS; DRUG design; PEPTIDES; CELL lines; TUMORS; INDIVIDUALIZED medicine; BIOLOGICAL assay; CELL receptors; IMMUNE checkpoint proteins; MEDICAL care costs; PHARMACODYNAMICS
- Publication
Cancers, 2024, Vol 16, Issue 5, p1041
- ISSN
2072-6694
- Publication type
Article
- DOI
10.3390/cancers16051041