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14-02-2017 | Image

Figure 6: Biomarker discovery for combination immunotherapy and proposed new concepts for clinical management with immunotherapy based on biomarkers.

a | For biomarker studies, excisional or needle biopsy samples are taken from patients together with blood samples. Tumour tissue is processed either by enzymatic digestion followed by dimethyl sulfoxide freezing, by snap-freezing or by being formaldehyde-fixated and paraffin-embedded (FFPE). Blood samples undergo Ficoll density centrifugation for peripheral blood mononuclear cell (PBMC) isolation. Serum and granulocytes will be frozen. The digested tumour material will be used to grow patient-derived xenografts (PDXs) in non-obese diabetic (NOD) mice that have severe combined immunodeficiency and are deficient for the common γ-chain (also known as IL-2RG) to grow tumour cell lines and to isolate tumour-infiltrating lymphocytes (TILs). TILs will be subjected to flow cytometry studies, co-culture with tumour cell lines or tumour tissue digest, or used for adoptive transfer in PDX models. The fresh-frozen tumour tissue will be used for deep sequencing (both whole-exome DNA and whole-exome RNA sequencing, and for the development of gene expression signatures), and the FFPE material for analyses by immunohistochemistry (IHC) of the tumour and the tumour microenvironment (which involves staining for the receptors that are shown on the T cell in Fig. 1 and other cells in the microenvironment). PBMCs will be used for flow cytometry analyses. Results obtained from these analyses will be correlated with patient outcomes to find predictive biomarkers. b | Proposed new concepts for clinical management with immunotherapy. As we progress with immunotherapy, we face paradigm shifts regarding clinical management. Patients experiencing durable responses that are sustained even off treatment require new concepts in risk management and mitigation, while making the most of the clinical benefit. Overall, a phased approach can be envisioned in which aggressive combination regimens should achieve frequent clinical responses, to be followed by maintenance with less aggressive and safer regimens, reaching the point of weaning patients off treatment. Immunotherapy has the peculiarity of keeping patients in response off treatment134, 135 and the possibility of re-induction136, even with the same agents that were used in induction, with an important proportion of patients responding again. Identifying biomarkers will be crucial for optimal clinical management. BTLA, B lymphocyte and T lymphocyte attenuator; CTLA4, cytotoxic T lymphocyte-associated antigen 4; IDO1, indoleamine-2,3-dioxygenase 1; IL-10, interleukin-10; LAG3, lymphocyte activation gene 3 protein; MDSC, myeloid-derived suppressor cell; PD1, programmed cell death protein 1; PDL1, PD1 ligand; TAM, tumour-associated macrophages; TGFβ, transforming growth factor-β; TIM3, T cell immunoglobulin and mucin domain-containing 3; TReg cell, regulatory T cell.