Assessing tumor heterogeneity: integrating tissue and circulating tumor DNA (ctDNA) analysis in the era of immuno-oncology - blood TMB is not the same as tissue TMB

Abstract

Tissue tumor mutational burden (tTMB) is calculated to aid in cancer treatment selection. High tTMB predicts a favorable response to immunotherapy in patients with non-small cell lung cancer. Blood TMB (bTMB) from circulating tumor DNA is reported to have similar predictive power and has been proposed as an alternative to tTMB. Across many studies not only are tTMB and bTMB not concordant but also as reported previously by our group predict conflicting outcomes. This implies that bTMB is not a substitute for tTMB, but rather a composite index that may encompass tumor heterogeneity. Here, we provide a thorough overview of the predictive power of TMB, discuss the use of tumor heterogeneity alongside TMB to predict treatment response and review several methods of tumor heterogeneity assessment. Furthermore, we propose a hypothetical method of estimating tumor heterogeneity and touch on its clinical implications.

Emerging Blood-Based Biomarkers for Predicting Immunotherapy Response in NSCLC

“…Further studies assessing the predictive accuracy of circulating TMB again revealed differences between bTMB and tTMB [ 91 , 92 ]. Zhang et al suggested that, despite the differences between bTMB-tTMB, bTMB could still be predictive of outcomes, underscoring the importance of the quality of included neoantigens rather than the total amount of mutations [ 92 ].…”

“…In the same line, other studies proposed the combination of bTMB with other parameters to improve its predictive capacity [ 91 , 92 , 93 , 94 , 95 ]. For instance, a retrospective analysis of POPLAR and OAK studies again supported that bTMB alone may be insufficient to predict ICI outcomes and advised to use it in combination with Maximum Somatic Allele Frequency (MSAF): a parameter that helps to estimate the fraction of tumor cfDNA in peripheral blood samples [ 95 ].…”

“…In agreement, MSAF in combination with bTMB has been suggested to effectively predict the OS and PFS benefits of ICI vs. chemotherapy [ 95 ]. An additional proposed solution was the inclusion of parameters that assess tumor heterogeneity [ 91 , 92 , 93 , 94 , 95 , 96 ]. In a letter to the editor on the Wang et al study, Liu et al stated that bTMB is likely to carry two sorts of bias: the first one is related to tumor size: larger tumors shed more cells, leading to a higher presence of ctDNA in the bloodstream.…”

“…When Liu et al recalculated the bTMB after adjusting for both factors, the correlation of bTMB with PFS, OS, and OOR was enhanced as compared to the original results. With these claims in mind, Fridland et al proposed an additional index called Tumor Heterogeneity Index (THI), which may well determine whether a given mutation is subclonal, and calculate a score reflecting tumor heterogeneity, consequently giving a less biased vision of tumor mutational landscape than tTMB or bTMB alone [ 91 ].…”

Development of a Novel Reference Material for Tumor Mutational Burden Measurement Based on CRISPR/Cas9 Technology

“…To date, the materials typically used are formalin-fixed and paraffin-embedded (FFPE) surgical specimens or paraffin-embedded human-derived tumor cell lines (with or without matched normal cell lines) ( 21 , 22 ). However, owing to the difficulty of obtaining and mass-producing these materials, intratumor and intertumor heterogeneity ( 23 , 24 ), and variations in the TMB levels in different patients ( 6 , 25 ), FFPE samples from cancer patients display poor reproducibility. Meanwhile, since the validated TMB level in most human-derived tumor-normal matched cell lines that are publicly available and well-characterized is always lower than 10 mutations/megabase (mut/Mb) ( 22 , 26 ), the applicability of tumor-normal cell line-derived samples is hampered by the limited number of cell lines with high TMB values and the cell types.…”