Biospecimen | Method | Tissue/cell types | Pros | Cons | References and recommended reading |
---|---|---|---|---|---|
FFPE | IHC | Tumor cells or tumor infiltrating immune cells | Direct detection; accurately pinpoint cancer cells; highly sensitive; simplicity; low cost | Requirement of trained pathologists; inconsistency for criteria used to score tumors such as PD-L1-positive or negative | |
Multicolor IHC | Tumor cells or tumor infiltrating immune cells | Broad dynamic range; capability for multiplexing using different fluorescence channels; >10 protein targets are identified in the same sample; amenability for co-localization studies | Absence of rigorous quantitative tests; limitation in some biomarker-driven clinical trials; user must select combinations of dyes | Carvajal-Hausdorf DE (2014) [165] | |
T cell receptor deep sequencing | TILs | T cell count information; T cell clonality in tumor | Heterogeneous expression of TIL | Robbins HS (2013) [100] | |
Whole exome sequencing (WES) | Tumor cells | Characterization of tumor mutation load including nucleotide substitutions; structural rearrangements and copy number alterations; identification of the neoantigens and neoepitopes; affordable cost | Require high-performance deep sequencing, computational bioinformatics support; The pipelines are still at early developmental phase | Snyder A (2014) [104]; Rizvi NA (2015) [105]; Bouffet E (2016) [107]; Chen K-H (2016) [166] Hugo W (2016) [106] | |
Blood | ELISPOT assays (IFNγ and granzyme B) | T cells in PBMCs | Detection of tumor antigen-specific CD4+ and CD8+ T cell response with good assay sensitivity; Relatively well validated assay | A poor correlation with clinically relevant immune responses | Shafer-Weaver K (2006) [167]; Janetzki S (2008) [95] Malyguine A (2012) [94]; Janetzki S (2015) [93]; |
Flow cytometry (tetramer, polyfunctional analysis) | T cells in PBMCs | Assessment of tumor antigen-specific CD4+ and CD8+ T cells response; measure multiple functions; detection of neoantigen-specific CD8 + PD-1+ T cells; minimally invasive | Merely in lab research, not as routine clinical monitoring yet | Yuan J (2008) [84]; Attic S (2011) [85]; McNeil LK (2013) [86]; Barrera L (2015) [168]; Gros A (2016) [118] | |
Flow cytometry phenotype staining | Whole blood immune phenotype | Analyses of the frequency and proliferation of different subsets of immune cells; routine operation | Dedicated resource and staff to perform the analyses | ||
RNA-Seq (NGS) | T cells in PBMCs | Identification of genetic variants; a broader dynamic range; detection of more differentially expressed genes; fast and high efficiency | More expensive than microarray; more complex for analysis; bulk signature, not single cell signals; need more validation | Zhao S (2010) [171] | |
qPCR assay | T cells in PBMCs | High specificity; able to detect the reactivity of low-frequency T cells in the peripheral blood of metastatic cancer patients | Bulk signature, not single cell signals; need more validation | Kammula US (2008) [172] | |
Flow cytometry | CTCs | Qualitative analysis at the single cell level in a relatively short period of time; decrease the amount of blood needed; provide valuable information regarding the frequency, phenotype and/or the functionality of T cells | Expensive; need more validation | Zaritskaya L (2010) [83] | |
Cell sieve microfiltration assay and QUASR technique | CTCs | PD-L1 levels from CTCs or CAMLs serves as a surrogate for PD-L1 expression in tumor; as a marker for immunotherapy response | Limited in lab research; need more validation |