Table 1.

Comparison of different assays used to detect mismatch repair defects and other predictors of immune therapy response or resistance

Technique or assay	Description	Attributes	Deficiencies
MSI-PCR	PCR-based amplification of known microsatellite loci, gel electrophoresis, and software scoring to detect instability as a multiple-band pattern in amplicons	Focused test with rapid turn-around time	Interpretation difficulties, limited to MSI diagnosis, no information on genetic source of MMRd
dMMR/IHC	Antibody-based staining of FFPE sections from tumor, followed by microscopic examination and scoring to detect MMR proteins (MSH2, MSH6, MLH1, PMS2)	Focused test using a conventional pathology approach, inexpensive, rapid turn-around time	Evaluates the end result of MMR protein depleting alterations only, subject to inter-individual interpretation
CIN/FISH	Hybridization of centromere-specific fluorescent probes to chromosomal spreads, microscopic scoring of centromeric counts to detect aneuploidy	Genome-wide evaluation of chromosomal instability	Counting-based evaluation that is subject to inter-individual interpretation variability
MMR/MSI-NGS panel	NGS of genes for MMR proteins, mutation detection and annotation of pathogenic variants	Focused evaluation of mutations across known MMRd genes	Insensitive to large-scale alterations such as CNVs, insufficient breadth for TMB or neoantigen prediction
NGS/WES	NGS of all known coding exons of genes, mutation detection and annotation of pathogenic variants in genes	Unbiased evaluation of mutations across all coding genes, MSI evaluation (added probes), neoantigen prediction, TMB enabled	Mutational signature calculation may be compromised by lack of breadth
NGS/WGS	NGS of whole genome libraries from cancer and matched normal DNA, comprehensive variant detection, and annotation of variants	Unbiased evaluation of mutations across all coding genes, MSI evaluation, neoantigen prediction, TMB, mutational signature enabled	Expensive to generate sufficient coverage from low cellularity tumors

CIN chromosomal instability, CNV copy number variant, dMMR deficient mismatch repair, FFPE formalin-fixed paraffin-embedded, FISH fluorescence in situ hybridization, IHC immunohistochemistry, MMR mismatch repair, MMRd mismatch repair defect, MSI microsatellite instability, NGS next generation sequencing, TMB tumor mutational burden, WES whole exome sequencing, WGS whole genome sequencing