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Whole-exome sequencing and clinical interpretation of formalin-fixed, paraffin-embedded tumor samples to guide precision cancer medicine


Translating whole-exome sequencing (WES) for prospective clinical use may have an impact on the care of patients with cancer; however, multiple innovations are necessary for clinical implementation. These include rapid and robust WES of DNA derived from formalin-fixed, paraffin-embedded tumor tissue, analytical output similar to data from frozen samples and clinical interpretation of WES data for prospective use. Here, we describe a prospective clinical WES platform for archival formalin-fixed, paraffin-embedded tumor samples. The platform employs computational methods for effective clinical analysis and interpretation of WES data. When applied retrospectively to 511 exomes, the interpretative framework revealed a 'long tail' of somatic alterations in clinically important genes. Prospective application of this approach identified clinically relevant alterations in 15 out of 16 patients. In one patient, previously undetected findings guided clinical trial enrollment, leading to an objective clinical response. Overall, this methodology may inform the widespread implementation of precision cancer medicine.

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Figure 1: FFPE and frozen sample sequencing metrics.
Figure 2: FFPE and frozen sample data yield comparable alteration data.
Figure 3: PHIAL reveals the long tail of clinically relevant events.
Figure 4: Clinically relevant findings from individual patients.
Figure 5: Clinical sequencing informs clinical trial enrollment and experimental discovery.

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We thank the patients and clinicians for their participation in this project. We thank the Broad Genomics Platform (specifically K. Anderka, A. Cheney, E. Wheeler, T. Mason and C. Crawford). We thank C. Sougnez for facilitating data deposition. We also thank A. Lane and A. Yoda (Dana-Farber Cancer Institute), for their contributions to the JAK3 experimental work: A. Lane provided Ba/F3 cells, and A. Yoda provided murine stem cell virus–puromycin vector. G.G. is partially funded by a Paul C. Zamecnik, MD, Chair in Oncology at Massachusetts General Hospital. This work was supported by the Starr Cancer Foundation (L.A.G.), the Prostate Cancer Foundation (E.M.V.A. and L.A.G.), US National Institutes of Health (NIH) NHGRI Clinical Sequencing Exploratory Research grant 1U01HG006492 (L.A.G.), the NIH National Cancer Institute grant 1U24CA126546 (L.A.G. and E.S.L.), the US Department of Defense (L.A.G.), NIH U24CA143845 grant (G.G.) and the Dana-Farber Leadership Council (E.M.V.A.).

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Authors and Affiliations



All authors contributed extensively to the work presented in this paper. E.M.V.A. and N.W. contributed equally to this work. S.G., G.G. and L.A.G. contributed equally to this work. D.L.P., D.C.F., J.F., E.S.L., S.A.F., E.M.V.A. and S.G. contributed to FFPE sample sequencing protocols and analysis of sequencing metrics. E.M.V.A., P.S., D.F., K.C., G.K., S.L.C., A.M., A.S., A.K., D.V., M.L., L.T.L., J.G.G., M.R. and G.G. contributed to computational analyses for FFPE versus frozen sample comparisons and analysis of WES data generally. E.M.V.A., N.W., G.K., F.W.H., S.W.G., S.J., P.J., J.G., L.M., N.L., B.R., P.K. and L.A.G. contributed to clinical analysis and interpretation methods and application. N.W., S.M., J.J.-V. and L.A.G. contributed to experimental follow-up of the JAK3 mutation. D.B. and L.G. contributed clinical input for the patient case. All authors discussed the results and implications and commented on the manuscript at all stages.

Corresponding authors

Correspondence to Gad Getz or Levi A Garraway.

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Competing interests

L.A.G. and N.W. are equity holders and consultants in Foundation Medicine. L.A.G. is a consultant to Novartis, Millenium/Takeda and Boehringer Ingelheim and a recipient of a grant from Novartis. G.G. is a consultant to The Kew Group. D.B. is a consultant to N-of-1.

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Van Allen, E., Wagle, N., Stojanov, P. et al. Whole-exome sequencing and clinical interpretation of formalin-fixed, paraffin-embedded tumor samples to guide precision cancer medicine. Nat Med 20, 682–688 (2014).

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