Very High vs. High Tumor Mutational Burden Across Tumors: Real-World Associations with MSI, Pathway Features, and Immunotherapy Outcomes

Teixeira, Maria Fernanda; Tomaz, Victoria; Barbosa e Silva, Lucas Campos; Donizete, Uelson; Tustumi, Francisco; Nakaya, Helder Imoto; Beal, Juliana Rodrigues; Moura, Fernando; Borad, Mitesh J.; Campregher, Paulo Vidal; Uson Junior, Pedro Luiz Serrano

doi:10.3390/biomedicines14030593

Open AccessArticle

Very High vs. High Tumor Mutational Burden Across Tumors: Real-World Associations with MSI, Pathway Features, and Immunotherapy Outcomes

by

Maria Fernanda Teixeira

¹

,

Victoria Tomaz

¹,

Lucas Campos Barbosa e Silva

¹,

Uelson Donizete

¹,

Francisco Tustumi

^1,*

,

Helder Imoto Nakaya

¹

,

Juliana Rodrigues Beal

¹,

Fernando Moura

¹,

Mitesh J. Borad

²,

Paulo Vidal Campregher

^1,3 and

Pedro Luiz Serrano Uson Junior

¹

Center for Personalized Medicine, Hospital Israelita Albert Einstein, São Paulo 05652900, Brazil

²

Mayo Clinic Cancer Center, Phoenix, AZ 85054, USA

³

Genesis Genomics, São Paulo 04703901, Brazil

^*

Author to whom correspondence should be addressed.

Biomedicines 2026, 14(3), 593; https://doi.org/10.3390/biomedicines14030593

Submission received: 26 January 2026 / Revised: 22 February 2026 / Accepted: 3 March 2026 / Published: 6 March 2026

(This article belongs to the Special Issue Gastrointestinal Diseases: From Pathophysiology to Innovative Therapeutic Approaches)

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Abstract

Background: Tumor mutational burden (TMB) is an FDA-approved biomarker for immune checkpoint inhibitor (ICI) therapy. However, its predictive value varies among tumor types and molecular contexts. We investigated whether a very high TMB identifies a biologically distinct subset and whether a higher cutoff provides additional clinical insights beyond the conventional high TMB threshold. Methods: We analyzed 133 patients with advanced solid tumors and TMB ≥ 10 mutations/Mb (mut/Mb) who underwent tumor genomic profiling using a 523-gene DNA/RNA next-generation sequencing panel. Tumors were stratified into prespecified TMB categories: 10–20 mut/Mb (TMB-H) and >20 mut/Mb (TMB-VH). The clinical characteristics, ICI outcomes (in the treated subset), and pathway-level genomic features were compared between groups. Results: TMB-VH was observed in 42/133 (31.6%) patients and spanned more than 20 tumor types. MSI was markedly more prevalent in TMB-VH than in TMB-H tumors (38.1% vs. 2.2%; Fisher’s exact p = 8.9 × 10⁻⁸). Pathway-level comparisons did not identify statistically significant differences after false discovery rate correction (all q > 0.05), and the observed patterns were descriptive in nature. In the ICI-treated subset with complete follow-up, objective response did not differ according to the TMB group. Overall survival (OS) was also similar between groups, whether measured from metastatic diagnosis (log-rank p = 0.937) or from ICI initiation (log-rank p = 0.814), although OS was numerically longer in the TMB-VH group in both analyses without reaching statistical significance. Conclusions: In this cohort study, TMB-VH was strongly associated with MSI but not independently associated with improved ICI outcomes. Larger multicenter cohorts are needed to validate pathway-oriented patterns and clarify the clinical utility of extreme TMB thresholds across various histologies. Integrating the functional context (e.g., MSI status, gene-level context, and pathway-level features) with TMB magnitude may enable more robust, tumor-aware biomarker models for immunotherapy selection.

Keywords: tumor mutational burden; immune checkpoint inhibitors; microsatellite instability; biomarkers; pathway analysis; precision oncology

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MDPI and ACS Style

Teixeira, M.F.; Tomaz, V.; Barbosa e Silva, L.C.; Donizete, U.; Tustumi, F.; Nakaya, H.I.; Beal, J.R.; Moura, F.; Borad, M.J.; Campregher, P.V.; et al. Very High vs. High Tumor Mutational Burden Across Tumors: Real-World Associations with MSI, Pathway Features, and Immunotherapy Outcomes. Biomedicines 2026, 14, 593. https://doi.org/10.3390/biomedicines14030593

AMA Style

Teixeira MF, Tomaz V, Barbosa e Silva LC, Donizete U, Tustumi F, Nakaya HI, Beal JR, Moura F, Borad MJ, Campregher PV, et al. Very High vs. High Tumor Mutational Burden Across Tumors: Real-World Associations with MSI, Pathway Features, and Immunotherapy Outcomes. Biomedicines. 2026; 14(3):593. https://doi.org/10.3390/biomedicines14030593

Chicago/Turabian Style

Teixeira, Maria Fernanda, Victoria Tomaz, Lucas Campos Barbosa e Silva, Uelson Donizete, Francisco Tustumi, Helder Imoto Nakaya, Juliana Rodrigues Beal, Fernando Moura, Mitesh J. Borad, Paulo Vidal Campregher, and et al. 2026. "Very High vs. High Tumor Mutational Burden Across Tumors: Real-World Associations with MSI, Pathway Features, and Immunotherapy Outcomes" Biomedicines 14, no. 3: 593. https://doi.org/10.3390/biomedicines14030593

APA Style

Teixeira, M. F., Tomaz, V., Barbosa e Silva, L. C., Donizete, U., Tustumi, F., Nakaya, H. I., Beal, J. R., Moura, F., Borad, M. J., Campregher, P. V., & Uson Junior, P. L. S. (2026). Very High vs. High Tumor Mutational Burden Across Tumors: Real-World Associations with MSI, Pathway Features, and Immunotherapy Outcomes. Biomedicines, 14(3), 593. https://doi.org/10.3390/biomedicines14030593

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Very High vs. High Tumor Mutational Burden Across Tumors: Real-World Associations with MSI, Pathway Features, and Immunotherapy Outcomes

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