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T Cell Exhaustion in the Cervical Cancer Tumor Microenvironment: PD-1 Overexpression and Co-Expression with TIGIT, Tim-3, LAG-3, and NKG2A
by
, , , , ,
Nadia Tatiana García-Barrientos
1
,
Fabiola Solorzano-Ibarra
1,
Ksenia Klimov-Kravtchenko
1,
Jose Manuel Rojas-Diaz
1,
Marcela Sofia Guitron-Aviña
1,
Francisco Javier Ceja-Flores
2,
Jose Alfonso Cruz-Ramos
3,
Pablo Cesar Ortiz-Lazareno
4,
Felipe de Jesus Bustos-Rodriguez
5,
Juan Carlos Vazquez-Limon
5,
Miriam Ruth Bueno-Topete
1,
Martha Cecilia Tellez-Bañuelos
2,
Jesse Haramati
2,*,† and
Susana del Toro-Arreola
1,6,*,† 1
Instituto de Investigación en Enfermedades Crónico Degenerativas, Departamento de Biología Molecular y Genómica, Centro Universitario de Ciencias de la Salud, Universidad de Guadalajara, Colonia Independencia, Guadalajara 44340, Mexico
2
Laboratorio de Inmunología Traslacional, Departamento de Biología Celular y Molecular, Centro Universitario de Ciencias Biológicas y Agropecuarias, Universidad de Guadalajara, Zapopan 45200, Mexico
3
Coordinación de Investigación, Subdirección de Desarrollo Institucional, Instituto Jalisciense de Cancerología, Guadalajara 44200, Mexico
4
Centro de Investigación Biomédica de Occidente, División de Inmunología, Instituto Mexicano del Seguro Social (IMSS), Guadalajara 44340, Mexico
5
Departamento de Oncología del Hospital Civil de Guadalajara “Dr. Juan I. Menchaca”, Guadalajara 44340, Mexico
6
Laboratorio de Inmunología, Departamento de Fisiología, Centro Universitario de Ciencias de la Salud, Universidad de Guadalajara, Guadalajara 44340, Mexico
*
Authors to whom correspondence should be addressed.
†
These authors contributed equally to this work.
Cancers 2025, 17(22), 3627; https://doi.org/10.3390/cancers17223627
Submission received: 24 September 2025
/
Revised: 30 October 2025
/
Accepted: 5 November 2025
/
Published: 11 November 2025
(This article belongs to the Special Issue Recent Advances and Strategies for the Management of CIN and HPV Eradication Starategies for the Prevention of Uterine Cervical Cancer)
Simple Summary
Immune checkpoint inhibitor therapy is of growing importance for the treatment and management of cervical cancer. In this analysis of known and emerging inhibitory checkpoint receptors in patients with cervical cancer, we observed that in addition to an increase in PD-1 and TIGIT, the emerging receptors Tim-3, LAG-3 and NKG2A were likewise upregulated. The co-expression of multiple immune checkpoints indicates an interplay of exhaustion pathways and possible future therapeutic targets. The tumor microenvironment was highly immunosuppressive: T cells expressing multiple checkpoints were more likely to be found in the tumor than in peripheral blood mononuclear cells (PBMC)s, up to 6-fold higher in the case of PD-1+ NKG2A+ cells, highlighting the importance of examining biopsies, not only peripheral blood, for exhausted T cell signatures that may be key to cancer diagnosis and management. Understanding how treatment alters these pathways could support the development of rational combination immunotherapies to restore CD8+ T cell function in cervical cancer.
Abstract
Objective: T cell exhaustion is a major mechanism of immune evasion in cancer, characterized by the sustained expression of multiple inhibitory receptors. This study aimed to evaluate the expression of immune checkpoints in peripheral and tumor-infiltrating CD8+ T cells from cervical cancer patients. Methods: We enrolled 104 participants: 37 treatment-naïve patients, 36 treated patients, and 31 age-matched healthy donors. Peripheral blood mononuclear cells (PBMCs) were isolated from all participants. Ten cervical biopsies were collected for tumor-infiltrating lymphocyte (TIL) isolation and paraffin fixation. Immune checkpoint expression was analyzed by multiparametric flow cytometry and immunohistochemistry. Results: In peripheral CD8+ T cells, we found a significant upregulation of exhaustion-associated markers PD-1, TIGIT, Tim-3, and LAG-3. In the tumor infiltrating lymphocytes, these same molecules, with the addition of NKG2A, were notably upregulated further. While BTLA and NKG2A showed no systemic changes, NKG2A increased in TILs and BTLA decreased in TILs. The co-expression of PD-1 with TIGIT, Tim-3, LAG-3, and NKG2A was notably enriched between 2- and 6-fold in TILs compared with patient PBMCs. The tumor microenvironment was highly immunosuppressive, characterized by enrichment with PD-1, PD-L1, and TIGIT; TIGIT was notably upregulated in locally advanced versus early-stage tumors. Conclusions: Our findings highlight the strongly immunosuppressive environment of cervical tumors in treatment-naïve patients and the presence of elevated inhibitory checkpoint expression in peripheral blood of both pre- and post-treatment patients. These results underscore the importance of investigating immune regulation within the tumor site itself and suggest that immune checkpoint co-expression may serve as a biomarker of T cell exhaustion and therapeutic resistance. Understanding how treatment alters these pathways could guide rational combination immunotherapies to restore CD8+ T cell function in cervical cancer.
Keywords:
cervical cancer; checkpoint therapy; ICI; CIN; cervical dysplasia; chemoradiotherapy; TIGIT; LAG-3; PD-1; PD-L1
