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

Van Allen EM, Wagle N, Stojanov P, Perrin DL, Cibulskis K, Marlow S, Jane-Valbuena J, Friedrich DC, Kryukov G, Carter SL, McKenna A, Sivachenko A, Rosenberg M, Kiezun A, Voet D, Lawrence M, Lichtenstein LT, Gentry JG, Huang FW, Fostel J, Farlow D, Barbie D, Gandhi L, Lander ES, Gray SW, Joffe S, Janne P, Garber J, MacConaill L, Lindeman N, Rollins B, Kantoff P, Fisher SA, Gabriel S, Getz G, Garraway LA - Nat. Med. (2014)

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Medical Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts, USA. [2] Broad Institute of MIT and Harvard, Cambridge, Massachusetts, USA.

ABSTRACT
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.

Show MeSH

Related in: MedlinePlus

FFPE and Frozen sequencing metricsThe percentage of target bases covered at 20X, percent selected bases, and percent of zero coverage targets in FFPE (n = 99) and non-FFPE tissue (n = 768) (1A–C). Additional quality control metrics for all 867 cases are available in Supplementary Table 1. No statistically significant difference between FFPE and non-FFPE tissue is observed in these three metrics (P > 0.05; two-sided Mann-Whitney test).
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC4048335&req=5

Figure 1: FFPE and Frozen sequencing metricsThe percentage of target bases covered at 20X, percent selected bases, and percent of zero coverage targets in FFPE (n = 99) and non-FFPE tissue (n = 768) (1A–C). Additional quality control metrics for all 867 cases are available in Supplementary Table 1. No statistically significant difference between FFPE and non-FFPE tissue is observed in these three metrics (P > 0.05; two-sided Mann-Whitney test).

Mentions: To produce WES data for clinical use, robust sequencing data must frequently be generated from small quantities of archival FFPE tissue. To test this, DNA was extracted from 99 FFPE samples using the FFPE extraction protocol (Supplementary Table 1, Methods). A comparison of standard WES metrics14 with 768 non-FFPE samples (394 whole blood, 367 frozen, 7 cell lines) sequenced in parallel demonstrated no significant differences independent of input DNA quantity (P > 0.05, Mann-Whitney Test; Fig. 1A–C, Supplementary Table 1). Our lowest successful WES attempts were 13.6ng and 16ng for non-FFPE and FFPE-derived DNA, respectively.


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

Van Allen EM, Wagle N, Stojanov P, Perrin DL, Cibulskis K, Marlow S, Jane-Valbuena J, Friedrich DC, Kryukov G, Carter SL, McKenna A, Sivachenko A, Rosenberg M, Kiezun A, Voet D, Lawrence M, Lichtenstein LT, Gentry JG, Huang FW, Fostel J, Farlow D, Barbie D, Gandhi L, Lander ES, Gray SW, Joffe S, Janne P, Garber J, MacConaill L, Lindeman N, Rollins B, Kantoff P, Fisher SA, Gabriel S, Getz G, Garraway LA - Nat. Med. (2014)

FFPE and Frozen sequencing metricsThe percentage of target bases covered at 20X, percent selected bases, and percent of zero coverage targets in FFPE (n = 99) and non-FFPE tissue (n = 768) (1A–C). Additional quality control metrics for all 867 cases are available in Supplementary Table 1. No statistically significant difference between FFPE and non-FFPE tissue is observed in these three metrics (P > 0.05; two-sided Mann-Whitney test).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4048335&req=5

Figure 1: FFPE and Frozen sequencing metricsThe percentage of target bases covered at 20X, percent selected bases, and percent of zero coverage targets in FFPE (n = 99) and non-FFPE tissue (n = 768) (1A–C). Additional quality control metrics for all 867 cases are available in Supplementary Table 1. No statistically significant difference between FFPE and non-FFPE tissue is observed in these three metrics (P > 0.05; two-sided Mann-Whitney test).
Mentions: To produce WES data for clinical use, robust sequencing data must frequently be generated from small quantities of archival FFPE tissue. To test this, DNA was extracted from 99 FFPE samples using the FFPE extraction protocol (Supplementary Table 1, Methods). A comparison of standard WES metrics14 with 768 non-FFPE samples (394 whole blood, 367 frozen, 7 cell lines) sequenced in parallel demonstrated no significant differences independent of input DNA quantity (P > 0.05, Mann-Whitney Test; Fig. 1A–C, Supplementary Table 1). Our lowest successful WES attempts were 13.6ng and 16ng for non-FFPE and FFPE-derived DNA, respectively.

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Medical Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts, USA. [2] Broad Institute of MIT and Harvard, Cambridge, Massachusetts, USA.

ABSTRACT
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.

Show MeSH
Related in: MedlinePlus