Latarsha J. Carithers

Broad Consent for Research on Biospecimens: The Views of Actual Donors at Four U.S. Medical Centers

Commentators are concerned that broad consent may not provide biospecimen donors with sufficient information regarding possible future research uses of their tissue. We surveyed with interviews 302 cancer patients who had recently provided broad consent at four diverse academic medical centers. The majority of donors believed that the consent form provided them with sufficient information regarding future possible uses of their biospecimens. Donors expressed very positive views regarding tissue donation in general and endorsed the use of their biospecimens in future research across a wide range of contexts. Concerns regarding future uses were limited to for-profit research and research by investigators in other countries. These results support the use of broad consent to store and use biological samples in future research.

Development of a consensus approach for return of pathology incidental findings in the Genotype-Tissue Expression (GTEx) project

The active debate about the return of incidental or secondary findings in research has primarily focused on return to research participants, or in some cases, family members. Particular attention has been paid to return of genomic findings. Yet, research may generate other types of findings that warrant consideration for return, including findings related to the pathology of donated biospecimens. In the case of deceased biospecimen donors who are also organ and/or tissue transplant donors, pathology incidental findings may be relevant not to family members, but to potential organ or tissue transplant recipients. This paper will describe the ethical implications of pathology incidental findings in the Genotype-Tissue Expression (GTEx) project, the process for developing a consensus approach as to if/when such findings should be returned, possible implications for other research projects collecting postmortem tissues and how the scenario encountered in GTEx fits into the larger return of results/incidental findings debate.

Small Nucleolar RNA Score: An Assay to Detect Formalin-Overfixed Tissue

Although there are millions of formalin-fixed paraffin-embedded (FFPE) tissue blocks potentially available for scientific research, many are of questionable quality, partly due to unknown fixation conditions. We analyzed FFPE tissue biospecimens as part of the NCI Biospecimen Preanalytical Variables (BPV) program to identify microRNA (miRNA) markers for fixation time. miRNA was extracted from kidney and ovary tumor FFPE blocks (19 patients, cold ischemia ≤2 hours) with 6, 12, 24, and 72 hours fixation times, then analyzed using the WaferGen SmartChip platform (miRNA chip with 1036 miRNA targets). For fixation time, principal component analysis of miRNA chip expression data separated 72 hours fixed samples from 6 to 24 hours fixed samples. A set of small nuclear RNA (snRNA) targets was identified that best determines fixation time and was validated using a second independent cohort of seven different tissue types. A customized assay was then developed, based on a set of 24 miRNA and snRNA targets, and a simple “snoRNA score” defined. This score detects FFPE tissue samples with fixation for 72 hours or more, with 79% sensitivity and 80% specificity. It can therefore be used to assess the fitness-for-purpose of FFPE samples for DNA or RNA-based research or clinical assays, which are known to be of limited robustness to formalin overfixation.

Abstract 1377: Impact of biospecimen pre-analytical factors on molecular analysis

The Biospecimen Pre-analytical Variables (BPV) Program of the Biorepositories and Biospecimen Research Branch (BBRB) at the National Cancer Institute (NCI) is designed to systematically investigate the effects of preanalytical factors on the molecular integrity of biospecimens. Specific parameters related to formalin fixation and paraffin embedding (FFPE) of cancer tissues were examined, including the effects of cold ischemic time (delay to fixation (DTF)) and time in fixative (TIF). Additional preanalytical studies focus on snap freezing method for tissue specimens (dry Ice vs. LN2 vapor), storage temperature (-80°C vs. LN2 vapor), and duration of storage for frozen specimens. Biospecimens for the BPV program were collected at four medical centers, within a tightly regulated infrastructure and strict adherence to SOPs, to enable consistent collection and handling across all sites. Biospecimens were collected from research participants undergoing surgical treatment for renal cell carcinoma; ovarian, fallopian tube, and peritoneal carcinoma; lung adenocarcinoma and squamous cell carcinoma; and colorectal adenocarcinoma. Biospecimens were subjected to specific experimental protocols to systematically vary the preanalytical factors of interest. Extensive annotation was performed with 300+ data elements that include steps in the collection, handling, and processing of the biospecimens, pathological evaluation of the tumor, and clinical information collected from donors. Biospecimen collections concluded in April 2015 with a total of 364 tumor tissue cases collected. The BPV program has conducted multiple, simultaneous molecular analyses to understand the impact of FFPE preanalytical factors on the expression and detection of various molecular analytes. Initial efforts were focused on evaluating the impact of DTF and TIF on the quality of DNA and RNA from FFPE samples using multiple methods and approaches such as NanoDrop 8000 UV-Vis spectrophotometer, Qubit 2.0 Fluorometer, Agilent Bioanalyzer and KAPA Human Genomic DNA Quantification. The QC data from both RIN and Kappa assays showed that FFPE samples have a significant drop in RNA and DNA quality compared with matched frozen samples. 72 hr TIF samples showed a significant drop in both RNA and DNA quality compared to shorter time points (6, 12, or 23 hr TIF), as measured by DV200 and Kappa assays. No significant differences were observed in RNA/DNA quality between the shorter TIF time points or between the DTF time points (1, 2, 3, 12 hr DTF). Further studies are now underway to evaluate additional preanalytical factors. The data from BPV studies will be widely shared with the research community through publication and deposition at a public data repository. The results from these studies will be used to develop evidence-based protocols and best practices for fit-for-purpose collection, processing, and storage of biospecimens. This project is funded by NCI Contract No. HHSN261200800001E. Citation Format: Rachana Agarwal, Ping Guan, Mary Barcus, Jasmin Bavarva, Robin Burges, Philip Branton, Latarsha Carithers, Corinne Camalier, Biswajit Das, Jason Lih, Hana Odeh, Nancy Roche, Dan Rohrer, Michael Sachs, Leslie Sobin, Jewell Scott, Anna Smith, Conrado Soria, Kimberly Valentino, Dana Valley, Mickey Williams, Helen Moore. Impact of biospecimen pre-analytical factors on molecular analysis. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1377.

Abstract 2237: Acquisition of biospecimens to support the Genotype-Tissue Expression (GTEx) project

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA The Genotype-Tissue Expression (GTEx) project is an NIH Common Fund study that is aimed at understanding how genetic variation influences gene expression in normal tissues. Here we describe a biospecimen collection platform that was developed to address the logistical challenges of acquiring a large collection of high-quality tissues from rapid autopsy and organ donors for GTEx. This complex infrastructure is utilized to procure specimens under a rigorous quality management program and a stringent ethical, legal, and regulatory framework. Ultimately, data from the GTEx study will be used to evaluate gene regulatory mechanisms and their contribution to human disease development. Citation Format: Latarsha Carithers, Abhi Rao, Helen Moore. Acquisition of biospecimens to support the Genotype-Tissue Expression (GTEx) project. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2237. doi:10.1158/1538-7445.AM2014-2237