Microsatellite instability (MSI) and the tumor mutation burden (TMB) as biomarkers of response to immune checkpoint inhibitors in prostate cancer

Both microsatellite instability (MSI) and the tumor mutation burden (TMB) have obtained a tumor agnostic approval as biomarkers of response to the immune checkpoint inhibitor pembrolizumab (1). The two biomarkers have a common underlying mechanism that relates to the presence of a high number of tumor neo-antigens for presentation to immune effector cells (2). Immune cells are disinhibited by the immune checkpoint inhibitor treatment and may facilitate a robust anti-tumor effect targeting these neo-antigens. The two biomarkers have, though, significant differences that require careful considerations in their use. High MSI is produced by one underlying mechanism, which is mismatch repair (MMR) deficiency (3). MMR is one of the cellular mechanisms that cells employ to repair nucleotide mismatches in the two complementary DNA chromatid strands produced during replication (3). Four proteins, MSH2, MSH6, MLH1 and PMS2, participate in MMR and their genetic or somatic mutation or epigenetic loss produces MSI. Microsatellites are stretches of single nucleotide or oligonucleotide repeats that are particularly sensitive to mismatching during replication and therefore are prone to mutations that remain unrepaired and can be detected in cells with MMR deficiency. Mutations in MMR genes are more prevalent in certain cancers such as colorectal, endometrial and gastric, but are also present with a lower prevalence in other cancer types. Independently of the primary site, MMR deficiency-associated high neo-antigen burden has the potential to trigger an immune response in cancer cells with intact DNA sensing mechanisms (4). On the other hand a high TMB has a varying mechanistic origin and may be produced by several mechanisms besides MMR deficiency (2). Among cancers across primary types with high TMB, defined as more than 20 mutations/Mb, only 16% of cases had MMR deficiency/high MSI (5). A well characterized additional mechanism is mutations in the exonuclease domain of the gene encoding for the proofreading polymerase epsilon (POLE), but other cancers have a high TMB without MMR or POLE defects (6). Another difference between MMR deficiency and high TMB as biomarkers of immune checkpoint inhibitor immunotherapy is that, whereas MMR deficiency is a concrete state produced by specific repair protein function loss, TMB is a continuous variable which has artificially been set at 10 mutations /Mb as a cut-off to define high TMB for the purpose of regulatory approval for immune checkpoint inhibitor treatment (1). This cut-off may be sub-optimal as a predictor of immunotherapy response for many cancers. Higher TMB values are associated with higher probability of response to immunotherapy, while cancers with TMB just above the cut-off are technically candidates for these treatments but have realistically low response rates (7). For example in colorectal cancer the cut-off of 10 mutations/Mb did not define microsatellite stable (MSS) cancers benefiting from immune checkpoint inhibitors (8). A small OS benefit was observed, though, with the combination of durvalumab and tremelimumab versus best supportive care in a phase 2 study of late line colorectal cancer patients, which was more pronounced in patients with a TMB more than 28 variants/Mb in a liquid biopsy based assay (9).

The group of patients with MSS/high TMB from various primary sites has attracted interest as potential targets for immunotherapy with immune checkpoint inhibitors (10-12). In the Foundation Medicine pan-cancer database with over 148,000 samples, MSS/high TMB (defined as ≥20 mutations/Mb) cancers represented the 6.6% of cases (10). The prostate cancer sub-cohort of this database with 3,326 samples displayed a high TMB prevalence of 5.1% (13). A few MSS/high TMB patients in the Foundation Medicine pan-cancer database were treated with immune checkpoint inhibitors and derived a significant benefit with 67% benefit rate and a median PFS of 26.8 months. In colorectal cancers the group of MSS/high TMB, defined as >10 mutations/Mb, represented 9.8% of all MSS patients in the cohort from The Cancer Genome Atlas (TCGA), while among gastroesophageal adenocarcinomas 3% to 6% of cases were MSS/TMB high in the respective gastroesophageal cancer cohort of TCGA (11,12). The pathophysiologic foundation of high TMB creation in these MSS cases appears to be heterogeneous. However, although no individual gene was dominant, both colorectal and gastroesophageal MSS cancers with high TMB displayed a high prevalence of mutations in genes involved in DNA damage response and repair and in epigenetic modifiers. Mutagenic patterns have been examined as a means to discover underlying causes of hypermutability (14). This effort has identified, in addition to MMR deficiency and proofreading polymerase deficiency associated signatures, several other signatures associated with UV light, tobacco smoke and alkylating agents exposures, as well as a signature associated with defects in APOBEC (apolipoprotein B mRNA editing catalytic polypeptide-like) enzyme family members. However, these exposures are also observed in many cancer cases without hypermutability and therefore the underlying pathophysiology and additional required events remain ill-defined for most cancers with high TMB and may be divergent dependent on primary sites and additional mutagenic exposures.

In prostate cancer, a retrospective genomic study based on the MSK-IMPACT targeted panel, MSI high/MMR deficient cancers constituted 2.8% of the 2,257 patients in the cohort, while 1.5% of the patients were MSS with a high TMB, above 10 mutations/Mb (15). The median TMB in patients with MSI high/MMR deficient cancers was 41 mutations/Mb (interquartile range, 26–57 mutations/Mb), while the median TMB in patients with MSS/high TMB was 15 mutations/Mb (interquartile range, 11–27 mutations/Mb). In the group of 33 patients with MSS/high TMB, 3 patients had mutations in the exonuclease domain of POLE producing TMB of 34, 169 and 183 mutations/Mb (15). The rest of the patients in the group had other alterations that may have contributed to the high TMB observed, albeit lower than the respective TMB of MSI high/MMR deficient cases. These included alterations in BRCA2/BRCA1 genes (21% of patients with MSS/high TMB prostate cancers in the series), ATM (12%) and MUTYH (15%). The prognosis of patients with metastatic prostate cancer in the three groups with MSI high/MMR deficiency, MSS/high TMB and MSS/low TMB was not significantly different (median overall survival 45, 39 and 48 months, respectively, log-rank test P=0.36). Patients with MSI high/MMR deficiency who received checkpoint inhibitor immunotherapy and had evaluable disease showed an overall response rate of 45%. In the MSS/high TMB group of patients only 5 patients who received immunotherapy had evaluable disease and the best response of all 5 patients was stable disease (15). In another small series of 22 patients with metastatic castration resistant prostate cancer and MSI high/MMR deficiency or high TMB, defined as more than 10 mutations/Mb, treated with pembrolizumab, the overall response rate was 50%, including 27.3% complete response rate (16). However among the 6 patients without high MSI or CDK12 alterations, one patient had stable disease and the remaining 5 patients had no response. Median OS was 5.1 months for patients with a TMB between 10 and 14.9 mutations/Mb, 20.5 months for patients with a TMB between 15 and 24.9 mutations/Mb and not reached for patients with a TMB above 24.9 mutations/Mb (16). These results suggest that prostate cancer patients in the lower range of high TMB do not derive benefit from PD-1 inhibitor immunotherapy in the absence of MSI.

The timing of acquisition of the defect leading to MMR deficiency or other cause of high TMB in the life span of a cancer has also implications for the use of resulting phenotypes as markers for immune checkpoint inhibitors effectiveness (17). Although the mutation patterns did not differ between cancers in patients with genetic defects in MMR genes (Lynch syndrome) and those with somatic origin of MMR deficiency, the clonality of the defect produced higher heterogeneity in the latter cases. Divergent clones in cancers that have acquired defects producing hypermutability later in their natural history would be expected to be differentially responsive to immune checkpoint inhibitors, leading to immunoediting and tumor treatment escape with predominance of surviving clones with low TMB. This concept has been observed in other targeted therapies such as poly(ADP-ribose) polymerase (PARP) inhibitors, which are more effective in cancers with germline BRCA1/BRCA2 mutations compared with somatic mutations (18).

Besides the zygosity and resulting clonality of the defects leading to hypermutability, the number and type of mutations in a particular high TMB cancer is of critical importance for determining response to immune checkpoint inhibitors (19). An increasing benefit with increasing number of mutations has been documented in some primary cancer types, including non-small cell lung cancer, melanoma, bladder carcinoma and colorectal cancer (termed category I cancers), but not in other types of primary cancers, including breast, head and neck, gastroesophageal, prostate and renal carcinomas, termed category II cancers (19). In this latter category increasing TMB has no statistically significant effect in OS (19). Moreover, the response to immune checkpoint inhibitors of MSS/high TMB (above 10 mutations/Mb) category I tumors was 22.6%, while the response of MSS/high TMB category II tumors was 5% (odds ratio =0.18, P=0.03) (20). With a more conservative cut-off of 16 mutations/Mb the response of category I and category II tumors were 42.9% and 16.6%, respectively. Tumors in category I group showed a correlation between neoantigen load and CD8⁺ infiltrating lymphocyte levels (21). Therefore, the cut-off of high TMB as a clinical predictive biomarker for immunotherapy should be individualized according to the type of the primary tumor. Different cut-offs may also need to be set according to the specific immunotherapy type and whether treatment is given as monotherapy or combination with other immunotherapies or chemotherapy. The source of the tumor tissue where the evaluation for TMB was performed is of potential importance as primary sites tend to have a lower TMB than metastatic sites (22).

In conclusion, TMB is a pan-cancer biomarker of response to immune checkpoint inhibitors. However the cut-off of 10 mutations/Mb appears to be inadequate for predicting clinically meaningful responses in prostate cancer patients, as in several other cancers. Prostate cancer patients with TMB above 10 mutations/Mb as a group do not derive significant benefit from immune checkpoint inhibitor therapy and therefore the optimal higher cut-off requires further study. This is of significant clinical importance as immune checkpoint inhibitors are not without adverse effects, which can be severe or even lethal in some patients (23). Moreover, some patients appear not only to derive no benefit for immunotherapy treatments but also to suffer an accelerated worsening of their disease (hyper-progression) (24). Although this adverse outcome is rare and even debatable in its existence, it makes the accurate prediction of benefit from immunotherapy an even more pressing clinical problem (24).

Acknowledgments

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Footnote

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References

1. Thein KZ, Myat YM, Park BS, et al. Target-Driven Tissue-Agnostic Drug Approvals-A New Path of Drug Development. Cancers (Basel) 2024;16:2529. [Crossref] [PubMed]
2. Jardim DL, Goodman A, de Melo Gagliato D, et al. The Challenges of Tumor Mutational Burden as an Immunotherapy Biomarker. Cancer Cell 2021;39:154-73. [Crossref] [PubMed]
3. Németh E, Szüts D. The mutagenic consequences of defective DNA repair. DNA Repair (Amst) 2024;139:103694. [Crossref] [PubMed]
4. Lu C, Guan J, Lu S, et al. DNA Sensing in Mismatch Repair-Deficient Tumor Cells Is Essential for Anti-tumor Immunity. Cancer Cell 2021;39:96-108.e6. [Crossref] [PubMed]
5. Chalmers ZR, Connelly CF, Fabrizio D, et al. Analysis of 100,000 human cancer genomes reveals the landscape of tumor mutational burden. Genome Med 2017;9:34. [Crossref] [PubMed]
6. Voutsadakis IA. Polymerase epsilon mutations and concomitant β2-microglobulin mutations in cancer. Gene 2018;647:31-8. [Crossref] [PubMed]
7. Marques A, Cavaco P, Torre C, et al. Tumor mutational burden in colorectal cancer: Implications for treatment. Crit Rev Oncol Hematol 2024;197:104342. [Crossref] [PubMed]
8. Vegivinti CTR, Gonzales Gomez C, Syed M, et al. The role of immune checkpoint inhibitors for patients with advanced stage microsatellite stable colorectal cancer and high tumor mutation burden: quantity or quality? Expert Opin Biol Ther 2023;23:595-601. [Crossref] [PubMed]
9. Chen EX, Jonker DJ, Loree JM, et al. Effect of Combined Immune Checkpoint Inhibition vs Best Supportive Care Alone in Patients With Advanced Colorectal Cancer: The Canadian Cancer Trials Group CO.26 Study. JAMA Oncol 2020;6:831-8. [Crossref] [PubMed]
10. Goodman AM, Sokol ES, Frampton GM, et al. Microsatellite-Stable Tumors with High Mutational Burden Benefit from Immunotherapy. Cancer Immunol Res 2019;7:1570-3. [Crossref] [PubMed]
11. Voutsadakis IA. High tumor mutation burden (TMB) in microsatellite stable (MSS) colorectal cancers: Diverse molecular associations point to variable pathophysiology. Cancer Treat Res Commun 2023;36:100746. [Crossref] [PubMed]
12. Voutsadakis IA. The Landscape and Prognosis of Microsatellite Stable (MSS) Esophageal, Gastro-Esophageal Junction and Gastric Adenocarcinomas with High Tumor Mutation Burden (TMB). Cancer Invest 2024;42:697-709. [Crossref] [PubMed]
13. Chung JH, Dewal N, Sokol E, et al. Prospective Comprehensive Genomic Profiling of Primary and Metastatic Prostate Tumors. JCO Precis Oncol 2019;3:PO.18.00283.
14. Campbell BB, Light N, Fabrizio D, et al. Comprehensive Analysis of Hypermutation in Human Cancer. Cell 2017;171:1042-1056.e10. [Crossref] [PubMed]
15. Lenis AT, Ravichandran V, Brown S, et al. Microsatellite Instability, Tumor Mutational Burden, and Response to Immune Checkpoint Blockade in Patients with Prostate Cancer. Clin Cancer Res 2024;30:3894-903. [Crossref] [PubMed]
16. Mosalem O, Tan W, Bryce AH, et al. A real-world experience of pembrolizumab monotherapy in microsatellite instability-high and/or tumor mutation burden-high metastatic castration-resistant prostate cancer: outcome analysis. Prostate Cancer Prostatic Dis 2025;28:138-44. [Crossref] [PubMed]
17. Martin S, Katainen R, Taira A, et al. Lynch syndrome-associated and sporadic microsatellite unstable colorectal cancers: different patterns of clonal evolution yield highly similar tumours. Hum Mol Genet 2024;33:1858-72. [Crossref] [PubMed]
18. Stravodimou A, Voutsadakis IA. Neo-adjuvant therapies for ER positive/HER2 negative breast cancers: from chemotherapy to hormonal therapy, CDK inhibitors, and beyond. Expert Rev Anticancer Ther 2024;24:117-35. [Crossref] [PubMed]
19. Zheng M. Dose-Dependent Effect of Tumor Mutation Burden on Cancer Prognosis Following Immune Checkpoint Blockade: Causal Implications. Front Immunol 2022;13:853300. [Crossref] [PubMed]
20. McGrail DJ, Pilié PG, Rashid NU, et al. Validation of cancer-type-dependent benefit from immune checkpoint blockade in TMB-H tumors identified by the FoundationOne CDx assay. Ann Oncol 2022;33:1204-6. [Crossref] [PubMed]
21. McGrail DJ, Pilié PG, Rashid NU, et al. High tumor mutation burden fails to predict immune checkpoint blockade response across all cancer types. Ann Oncol 2021;32:661-72. [Crossref] [PubMed]
22. Schnidrig D, Turajlic S, Litchfield K. Tumour mutational burden: primary versus metastatic tissue creates systematic bias. Immunooncol Technol 2019;4:8-14. [Crossref] [PubMed]
23. Schneider BJ, Naidoo J, Santomasso BD, et al. Management of Immune-Related Adverse Events in Patients Treated With Immune Checkpoint Inhibitor Therapy: ASCO Guideline Update. J Clin Oncol 2021;39:4073-126. [Crossref] [PubMed]
24. Adashek JJ, Subbiah IM, Matos I, et al. Hyperprogression and Immunotherapy: Fact, Fiction, or Alternative Fact? Trends Cancer 2020;6:181-91. [Crossref] [PubMed]