Is tumor mutational burden predictive of response to immunotherapy?

Reviewer #1 (Public Review):

In their manuscript entitled: "Is tumor mutational burden predictive of response to immunotherapy?", Gurjao and colleagues discuss the use of tumor mutational burden (TMB) as a predictive biomarker for cancer patients to respond to immune checkpoint blockage (ICB). By analyzing a large cohort of 882 patient samples across different tumor types they find either little or no association of TMB to the response of ICB. In addition, they showed that finding the optimal cutoff for patient stratification lead to a severe multiple testing problem. By rigorously addressing this multiple testing problem only non-small cell lung cancer out of 10 cancer types showed a statistically significant association of TMB and response to ICB. Nevertheless, it is clearly shown that in any case the rate of misclassification is too high that TMB alone would qualify as a clinically suitable biomarker for ICB response. Finally, the authors demonstrate with a simple mathematical model that only a few strong immunogenic mutations would be sufficient for an ICB response, thereby showing that also patients with a low TMB score could benefit from immunotherapy. The manuscript is clearly written, the results are well presented and the applied methods are state-of-the-art.

Reviewer #2 (Public Review):

The manuscript points out that TMB cut-offs are not strong predictors of response to immunotherapy or overall survival. By randomly shuffling TMB values within cohorts to simulate a null distribution of log-rank test p-values, they show that under correction, the statistical significance of previously reported TMB cut-offs for predicting outcomes is questionable. There is a clinical need for a better prediction of treatment response than TMB alone can provide. However, no part of the analysis challenges the validity of the well-known pan-cancer correlation between TMB and immunotherapy response. The failure to detect significant TMB cut-offs may be due to insufficient power, as the examined cohorts have relatively low sample sizes. A power analysis would be informative of what cohort sizes are needed to detect small to modest effects of TMB on immune response.

The manuscript provides a simple model of immunogenicity that is tailored to be consistent with a claimed lack of relationship between TMB and response to immunotherapy. Under the model, if each mutation that a tumor has acquired has a relatively high probability of being immunogenic (~10%, they suggest), and if 1-2 immunogenic mutations is enough to induce an immune response, then most tumors produce an immune response, and TMB and response should be uncorrelated except in very low-TMB tumors. The question then becomes whether the response is sufficient to wipe out tumor cells in conjunction with immunotherapy, which is essentially the same question of predicting response that motivated the original analysis. While TMB alone is not an excellent predictor of treatment response, the pan-cancer correlation between TMB and response/survival is highly significant, so the model's only independent prediction is wrong. Additionally, experiments to predict and validate neoepitopes suggest that a much smaller fraction of nonsynonymous mutations produce immune responses1,2.

A key idea that is overlooked in this manuscript is that of survivorship bias: self-evidently, none of the mutations found at the time of sequencing have been immunogenic enough to provoke a response capable of eliminating the tumor. While the authors suggest that immunoediting "is inefficient, allowing tumors to accumulate a high TMB," the alternative explanation fits the neoepitope literature better: most mutations that reach high allele frequency in tumor cells are not immunogenic in typical (or patient-specific) tumor environments. Of course, immunotherapies sometimes succeed in overcoming the evolved immune evasion of tumors. Higher-TMB tumors are likely to continue to have higher mutation rates after sequencing; increased generation of new immunogenic mutations may partially explain their modestly improved responses to therapy.