Preanalytics and Precision Pathology: Pathology Practices to Ensure Molecular Integrity of Cancer Patient Biospecimens for Precision Medicine.

Biospecimens acquired during routine medical practice are the primary sources of molecular information about patients and their diseases that underlies precision medicine and translational research. In cancer care, molecular analysis of biospecimens is especially common because it often determines treatment choices and may be used to monitor therapy in real time. However, patient specimens are collected, handled, and processed according to routine clinical procedures during which they are subjected to factors that may alter their molecular quality and composition. Such artefactual alteration may skew data from molecular analyses, render analysis data uninterpretable, or even preclude analysis altogether if the integrity of a specimen is severely compromised. As a result, patient care and safety may be affected, and medical research dependent on patient samples may be compromised. Despite these issues, there is currently no requirement to control or record preanalytical variables in clinical practice with the single exception of breast cancer tissue handled according to the guideline jointly developed by the American Society of Clinical Oncology and College of American Pathologists (CAP) and enforced through the CAP Laboratory Accreditation Program. Recognizing the importance of molecular data derived from patient specimens, the CAP Personalized Healthcare Committee established the Preanalytics for Precision Medicine Project Team to develop a basic set of evidence-based recommendations for key preanalytics for tissue and blood specimens. If used for biospecimens from patients, these preanalytical recommendations would ensure the fitness of those specimens for molecular analysis and help to assure the quality and reliability of the analysis data.

The Eighth Edition AJCC Cancer Staging Manual: Continuing to build a bridge from a population-based to a more "personalized" approach to cancer staging.

The American Joint Committee on Cancer (AJCC) staging manual has become the benchmark for classifying patients with cancer, defining prognosis, and determining the best treatment approaches. Many view the primary role of the tumor, lymph node, metastasis (TNM) system as that of a standardized classification system for evaluating cancer at a population level in terms of the extent of disease, both at initial presentation and after surgical treatment, and the overall impact of improvements in cancer treatment. The rapid evolution of knowledge in cancer biology and the discovery and validation of biologic factors that predict cancer outcome and response to treatment with better accuracy have led some cancer experts to question the utility of a TNM-based approach in clinical care at an individualized patient level. In the Eighth Edition of the AJCC Cancer Staging Manual, the goal of including relevant, nonanatomic (including molecular) factors has been foremost, although changes are made only when there is strong evidence for inclusion. The editorial board viewed this iteration as a proactive effort to continue to build the important bridge from a "population-based" to a more "personalized" approach to patient classification, one that forms the conceptual framework and foundation of cancer staging in the era of precision molecular oncology. The AJCC promulgates best staging practices through each new edition in an effort to provide cancer care providers with a powerful, knowledge-based resource for the battle against cancer. In this commentary, the authors highlight the overall organizational and structural changes as well as "what's new" in the Eighth Edition. It is hoped that this information will provide the reader with a better understanding of the rationale behind the aggregate proposed changes and the exciting developments in the upcoming edition. CA Cancer J Clin 2017;67:93-99. © 2017 American Cancer Society.

A Study of Thymidylate Synthase Expression as a Biomarker for Resectable Colon Cancer: Alliance (Cancer and Leukemia Group B) 9581 and 89803.

PURPOSE: Tumor levels of thymidylate synthase (TS), a target of 5-fluorouracil (5-FU)-based chemotherapy for colorectal cancer, have been studied as a predictive or prognostic biomarker with mixed results. PATIENTS AND METHODS: Tumor TS levels were prospectively evaluated in two adjuvant therapy trials for patients with resected stage II or III colon cancer. TS expression was determined by standard immunohistochemistry and by automated quantitative analysis. Tumor mismatch repair deficiency (MMR-D) and BRAF c.1799T > A (p.V600E) mutation status were also examined. Relationships between tumor TS, MMR-D, and BRAF mutation status, overall survival (OS), and disease-free survival (DFS) were investigated in the subset of stage III patients. RESULTS: Patients whose tumors demonstrated high TS expression experienced better treatment outcomes, with DFS hazard ratio (HR) = 0.67, 95% confidence interval (CI) = 0.53, 0.84; and OS HR = 0.68, 95% CI = 0.53, 0.88, for high versus low TS expression, respectively. No significant interaction between TS expression and stage was observed (DFS: interaction HR = 0.94; OS: interaction HR = 0.94). Tumors with high TS expression were more likely to demonstrate MMR-D (22.2% vs. 12.8%; p = .0003). Patients whose tumors demonstrated both high TS and MMR-D had a 7-year DFS of 77%, compared with 58% for those whose tumors had low TS and were non-MMR-D (log-rank p = .0006). Tumor TS expression did not predict benefit of a particular therapeutic regimen. CONCLUSION: This large prospective analysis showed that high tumor TS levels were associated with improved DFS and OS following adjuvant therapy for colon cancer, although tumor TS expression did not predict benefit of 5-FU-based chemotherapy. The Oncologist 2017;22:107-114Implications for Practice: This study finds that measurement of tumor levels of thymidylate synthase is not helpful in assigning specific adjuvant treatment for colorectal cancer. It also highlights the importance of using prospective analyses within treatment clinical trials as the optimal method of determining biomarker utility.

The impact of stage, grade, and mucinous histology on the efficacy of systemic chemotherapy in adenocarcinomas of the appendix: Analysis of the National Cancer Data Base.

BACKGROUND: Adenocarcinomas of the appendix represent a heterogeneous disease depending on the presence of mucinous histology, histologic grade, and stage. In the current study, the authors sought to explore the interplay of these factors with systemic chemotherapy in a large population data set. METHODS: Patients in the National Cancer Data Base (NCDB) who were diagnosed with mucinous, nonmucinous, and signet ring cell-type appendiceal neoplasms from 1985 through 2006 were selected. Multivariable Cox proportional hazards regression models were developed. RESULTS: A total of 11,871 patients met the inclusion criteria for the current study: 50.3% had mucinous neoplasms, 40.5% had nonmucinous neoplasms, and 9.2% had signet ring cell-type neoplasms. The 5-year overall survival (OS) stratified by grade was similar among patients with American Joint Committee on Cancer stage I to stage III disease but not for those with stage IV disease. The median OS for patients with stage IV mucinous and nonmucinous tumors was 6.4 years and 2.3 years, respectively, for those with well differentiated histology (P<.0001) and was 1.5 years and 0.8 years, respectively, for those with poorly differentiated histology (P<.0001). In multivariable modeling for stage I to III disease, adjuvant chemotherapy improved OS for both mucinous and nonmucinous histologies, with hazard ratios (HRs) of 0.78 (95% confidence interval [95% CI], 0.68-0.89 [P = .0002]) and 0.83 (95% CI, 0.74-0.94 [P = .002]), respectively. For patients with stage IV disease, systemic chemotherapy significantly improved OS for those with nonmucinous (HR, 0.72; 95% CI, 0.64-0.82 [P<.0001]) but not mucinous (HR, 0.95; 95% CI, 0.86-1.04 [P = .2) histologies, although this was grade-dependent. The median OS for chemotherapy versus no chemotherapy was 6.4 years versus 6.5 years (P value not significant) for patients with mucinous, well-differentiated tumors and 1.6 years versus 1.0 years (P = .0007) for patients with mucinous, poorly differentiated tumors. CONCLUSIONS: Adjuvant chemotherapy demonstrated a significant OS benefit regardless of histology. However, for patients with stage IV disease, the benefit of systemic chemotherapy varied by tumor histology and grade, with patients with well-differentiated, mucinous, appendiceal adenocarcinomas deriving no survival benefit from systemic chemotherapy. Cancer 2016;122:213-221. © 2015 American Cancer Society.

A Novel Approach to High-Quality Postmortem Tissue Procurement: The GTEx Project.

The Genotype-Tissue Expression (GTEx) project, sponsored by the NIH Common Fund, was established to study the correlation between human genetic variation and tissue-specific gene expression in non-diseased individuals. A significant challenge was the collection of high-quality biospecimens for extensive genomic analyses. Here we describe how a successful infrastructure for biospecimen procurement was developed and implemented by multiple research partners to support the prospective collection, annotation, and distribution of blood, tissues, and cell lines for the GTEx project. Other research projects can follow this model and form beneficial partnerships with rapid autopsy and organ procurement organizations to collect high quality biospecimens and associated clinical data for genomic studies. Biospecimens, clinical and genomic data, and Standard Operating Procedures guiding biospecimen collection for the GTEx project are available to the research community.

The national biomarker development alliance: confronting the poor productivity of biomarker research and development.

Making precision (personalized) medicine a routine clinical reality will require a comprehensive inventory of validated biomarkers and molecular diagnostic tests to stratify disease subtypes and improve accuracy in diagnosis and treatment selection. Realization of this promise has been hindered by the poor productivity of biomarker identification and validation. This situation reflects deficiencies that are pervasive across the entire spectrum of biomarker R&D, from discovery to clinical validation and in the failure of regulatory and reimbursement policies to accommodate new classes of biomarkers. The launch of the National Biomarker Development Alliance is the culmination of a 2-year review and consultation process involving diverse stakeholders to advance standards, best practices and guidelines to enhance biomarker discovery and validation by adoption of systems-based approaches and trans-sector collaboration between academia, clinical medicine and relevant private and public sector stakeholders.

Association of TP53 mutational status and gender with survival after adjuvant treatment for stage III colon cancer: results of CALGB 89803.

PURPOSE: The TP53 tumor suppressor is frequently mutated in colon cancer, but the influence of such mutations on survival remains controversial. We investigated whether mutations in the DNA-binding domain of TP53 are associated with survival in stage III colon cancer. EXPERIMENTAL DESIGN: The impact of TP53 genotype was prospectively evaluated in Cancer and Leukemia Group B 89803, a trial that randomized stage III colon cancer patients to receive adjuvant 5-fluorouracil/leucovorin (5FU/LV) or 5FU/LV with irinotecan (IFL). RESULTS: TP53 mutations were identified in 274 of 607 cases. The presence of any TP53 mutation did not predict disease-free survival (DFS) or overall survival with either adjuvant regimen when men and women were considered together or as separate groups. However, outcome differences among women became apparent when tumor TP53 genotype was stratified as wild-type versus zinc- or non-zinc-binding mutations in the TP53 DNA-binding domain. DFS at 5 years was 0.59, 0.52, and 0.78 for women with TP53 wild-type tumors, and tumors with zinc- or non-zinc-binding mutations, respectively. Survival at 5 years for these same women was 0.72, 0.59, and 0.90, respectively. No differences in survival by TP53 genotype were observed in men. CONCLUSIONS: The presence of any TP53 mutation within the DNA-binding domain did not predict survival in stage III colon cancer. However, TP53 genotype was predictive of survival in women following adjuvant therapy. Future colon cancer therapeutic trials, with inclusion of correlative molecular markers, should be designed to permit evaluation of survival and/or response to treatment in women separately from men.

Meeting research needs with postmortem biospecimen donation: summary of recommendations for postmortem recovery of normal human biospecimens for research.

Normal human tissues, bodily fluids, and other biospecimens of known quality are essential for research to understand the development of cancer and other diseases and to develop new diagnostics and therapies. However, obtaining normal biospecimens appropriate for contemporary large-scale molecular and genomic research is one of the most challenging biospecimen acquisition problems for scientists and biospecimen resources that support research. Recognizing this challenge, the U.S. National Cancer Institute recently convened a series of workshops and meetings focused on the acquisition of normal tissues for research and produced an extensive document, Recommendations for Postmortem Recovery of Normal Human Biospecimens for Research. This article summarizes these recommendations, addressing key ethical, operational, and scientific elements for collecting normal reference biospecimens from postmortem donors in the U.S. Awareness of these recommendations can foster more effective collaborations and mitigate potential logistical challenges, while promoting postmortem biospecimen donation options for families and increasing the availability of high quality normal biospecimens for research. The recommendations have been put into practice in the collection of normal human biospecimens for the NIH Genotype-Tissue Expression Program (GTEx), a pilot study of human gene expression and regulation in multiple tissues which will provide valuable insights into the mechanisms of gene regulation and, in the future, its disease-related perturbations (http://commonfund.nih.gov/GTEx/).

Intersection of biobanking and clinical care: should discrepant diagnoses and pathological findings be returned to research participants?

Diagnostic discrepancies occur when the diagnosis made on a biospecimen during the course of review at a biobank differs from the original clinical diagnosis. These diagnostic discrepancies detected during biobanking present unique challenges that are distinct from other types of research results or incidental findings. The proposed process for reporting diagnostic discrepancies or pathological incidental findings identified through a quality assurance evaluation at the biobank includes verification of the biospecimen identity, verification of the diagnosis within the biobank, and re-review of the case by the pathologist at the biospecimen collection site. If the pathologist at the biobank and the original pathologist do not reach agreement, an impartial and knowledgeable third party is consulted. The decision as to whether and how to notify research participants of any confirmed changes in diagnosis would be determined by institutional procedures. Implementation of this proposed process will require clear delineation of the roles and responsibilities of all involved parties in order to promote excellence in patient care and ensure that researchers have access to biospecimens of requisite quality.Genet Med 2012:14(4):417-423.

Biospecimens and biorepositories: from afterthought to science.

Biospecimens are recognized as critical components of biomedical research, from basic studies to clinical trials and epidemiologic investigations. Biorepositories have existed in various forms for more than 150 years, from early small collections in pathology laboratories to modern automated facilities managing millions of samples. As collaborative science has developed, it has been recognized that biospecimens must be of consistent quality. Recent years have seen a proliferation of best practices and the recognition of the field of "biospecimen science." The future of this field will depend on the development of more evidence-based practices in both the research and clinical settings. As the field matures, educating a new generation of biospecimen/biobanking scientists will be an important need.

Documenting the natural history of patients with resected stage II adenocarcinoma of the colon after random assignment to adjuvant treatment with edrecolomab or observation: results from CALGB 9581.

PURPOSE: We conducted a randomized trial comparing adjuvant treatment with edrecolomab versus observation in patients with resected, low-risk, stage II colon cancer. This study also prospectively studied patient- and tumor-specific markers of treatment outcome. PATIENTS AND METHODS: After surgical resection, patients with stage II colon cancer were randomly assigned to either five infusions of edrecolomab at 28-day intervals or observation without adjuvant therapy. RESULTS: Final accrual included 1,738 patients; 865 patients received edrecolomab, and 873 patients were observed without adjuvant treatment. Median follow-up time was 7.9 years. There were no significant outcome differences between study arms (overall survival [OS], P = .71; disease-free survival, P = .64). The combined 5-year all-cause OS was 0.86 (95% CI, 0.84 to 0.88), and the combined 5-year disease-specific OS was 0.93 (95% CI, 0.91 to 0.94). The relationships between demographic and histopathologic factors and survival differed for all-cause and disease-specific survival outcomes, but no combined prognostic factor model was found to adequately classify patients at higher risk of recurrence or death as a result of colon cancer. CONCLUSION: Edrecolomab did not prolong survival. Consequently, this large study with a long duration of follow-up provided unique data concerning the natural history of resected stage II colon cancer. Prognostic factors identified in previous retrospective and pooled analyses were associated with survival outcomes in this stage II patient cohort. Results from ongoing molecular marker studies may enhance our ability to determine the risk profile of these patients.

Microsatellite instability and loss of heterozygosity at chromosomal location 18q: prospective evaluation of biomarkers for stages II and III colon cancer--a study of CALGB 9581 and 89803.

PURPOSE: Colorectal cancer (CRC) develops as a result of a series of accumulated genomic changes that produce oncogene activation and tumor suppressor gene loss. These characteristics may classify CRC into subsets of distinct clinical behaviors. PATIENTS AND METHODS: We studied two of these genomic defects-mismatch repair deficiency (MMR-D) and loss of heterozygosity at chromosomal location 18q (18qLOH)-in patients enrolled onto two phase III cooperative group trials for treatment of potentially curable colon cancer. These trials included prospective secondary analyses to determine the relationship between these markers and treatment outcome. A total of 1,852 patients were tested for MMR status and 955 (excluding patients with MMR-D tumors) for 18qLOH. RESULTS: Compared with stage III, more stage II tumors were MMR-D (21.3% v 14.4%; P < .001) and were intact at 18q (24.2% v 15.1%; P = .001). For the combined cohort, patients with MMR-D tumors had better 5-year disease-free survival (DFS; 0.76 v 0.67; P < .001) and overall survival (OS; 0.81 v 0.78; P = .029) than those with MMR intact (MMR-I) tumors. Among patients with MMR-I tumors, the status of 18q did not affect outcome, with 5-year values for patients with 18q intact versus 18qLOH tumors of 0.74 versus 0.65 (P = .18) for DFS and 0.81 versus 0.77 (P = .18) for OS. CONCLUSION: We conclude that MMR-D tumor status, but not the presence of 18qLOH, has prognostic value for stages II and III colon cancer.

International approaches to advancing biospecimen science.

Biospecimen quality is affected by a number of preanalytical factors that may or may not be obvious to the investigator. These factors are introduced through multiple biospecimen collection, processing, and storage procedures, which can differ dramatically within and between medical institutions and biorepositories. Biospecimen Science is the emerging field of study that is attempting to quantify and control such variability. A variety of efforts are under way around the world to establish research programs, evidence-based biospecimen protocols, and standards to improve the overall quality of biospecimens for research.

Before you analyze a human specimen, think quality, variability, and bias.

Personalized medicine requires capabilities to detect and measure health-associated biomarkers with increasingly specific and sensitive methods, putting analytical chemists at the front lines of translational research. Analytical scientists must be upstream in the experimental design process because the analysis of a biospecimen (tissue, blood, etc.) presents technical and experimental design complexities. (To listen to a podcast about this feature, please go to the Analytical Chemistry multimedia page at pubs.acs.org/page/ancham/audio/index.html.).