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Table 3 Currently available translational biomarker assays for immune checkpoint inhibitors

From: Current status and perspectives in translational biomarker research for PD-1/PD-L1 immune checkpoint blockade therapy

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

Herbst R (2014) [145]; Loughlin PA (2007) [164]

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

Streitz M (2013) [169]; van Dongen JJ (2012) [170]

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

Steven HL (2015) [173]; Adams DL (2014) [174]

  1. Abbreviations: FFPE formalin-fixed paraffin-embedded, PD-L1 program death-1, TIL tumor-infiltrating lymphocytes, IHC immunohistochemistry, CAMLs cancer-associated macrophage-like cells, ELISPOT enzyme-linked immunospot assay, CTL cytotoxic T-lymphocytes, CTCs circulating tumor cells, PBMC peripheral blood mononuclear cell, WES whole exome sequencing, NGS next-generation sequencing