The Influence of Tissue Ischemia Time on RNA Integrity and Patient-Derived Xenografts (PDX) Engraftment Rate in a Non-Small Cell Lung Cancer (NSCLC) Biobank

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Abstract

Introduction

Bio-repositories are invaluable resources to implement translational cancer research and clinical programs. They represent one of the most powerful tools for biomolecular studies of clinically annotated cohorts, but high quality samples are required to generate reliable molecular readouts and functional studies. The objective of our study was to define the impact of cancer tissue ischemia time on RNA and DNA quality, and for the generation of Patient-Derived Xenografts (PDXs).

Methods

One-hundred thirty-five lung cancer specimens were selected among our Institutional BioBank samples. Associations between different warm (surgical) and cold (ex-vivo) ischemia time ranges and RNA quality or PDXs engraftment rates were assessed. RNA quality was determined by RNA integrity number (RINs) values. Fresh viable tissue fragments were implanted subcutaneously in NSG mice and serially transplanted.

Results

RNAs with a RIN>7 were detected in 51% of the sample (70/135), with values of RIN significantly lower (OR 0.08, P = 0.01) in samples preserved for more than 3 hours before cryopreservation. Higher quality DNA samples had a concomitant high RIN. Sixty-three primary tumors (41 adenocarcinoma) were implanted with an overall engraftment rate of 33%. Both prolonged warm (>2 hours) and ex-vivo ischemia time (>10 hours) were associated to a lower engraftment rate (OR 0.09 P = 0.01 and OR 0.04 P = 0.008, respectively).

Conclusion

RNA quality and PDXs engraftment rate were adversely affected by prolonged ischemia times. Proper tissue collection and processing reduce failure rate. Overall, NSCLC BioBanking represents an innovative modality, which can be successfully executed in routine clinical settings, when stringent Standard Operating Procedures are adopted.

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Most cited references 18

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Patient-derived first generation xenografts of non-small cell lung cancers: promising tools for predicting drug responses for personalized chemotherapy.

John C English, Julia Flint, Calum MacAulay … (2010)

Current chemotherapeutic regimens have only modest benefit for non-small cell lung cancer (NSCLC) patients. Cumulative toxicities/drug resistance limit chemotherapy given after the first-line regimen. For personalized chemotherapy, clinically relevant NSCLC models are needed for quickly predicting the most effective regimens for therapy with curative intent. In this study, first generation subrenal capsule xenografts of primary NSCLCs were examined for (a) determining responses to conventional chemotherapeutic regimens and (b) selecting regimens most effective for individual patients. Pieces (1x3x3 mm(3)) of 32 nontreated, completely resected patients' NSCLCs were grafted under renal capsules of nonobese diabetic/severe combined immunodeficient mice and treated with (A) cisplatin+vinorelbine, (B) cisplatin+docetaxel, (C) cisplatin+gemcitabine, and positive responses (treated tumor area

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Gene mutations in primary tumors and corresponding patient-derived xenografts derived from non-small cell lung cancer.

Chuncheng Hao, Li-Li Wang, Shaohua Peng … (2015)

Molecular annotated patient-derived xenograft (PDX) models are useful for the preclinical investigation of anticancer drugs and individualized anticancer therapy. We established 23 PDXs from 88 surgical specimens of lung cancer patients and determined gene mutations in these PDXs and their paired primary tumors by ultradeep exome sequencing on 202 cancer-related genes. The numbers of primary tumors with deleterious mutations in TP53, KRAS, PI3KCA, ALK, STK11, and EGFR were 43.5%, 21.7%, 17.4%, 17.4%, 13.0%, and 8.7%, respectively. Other genes with deleterious mutations in ≥3 (13.0%) primary tumors were MLL3, SETD2, ATM, ARID1A, CRIPAK, HGF, BAI3, EP300, KDR, PDGRRA and RUNX1. Of 315 mutations detected in the primary tumors, 293 (93%) were also detected in their corresponding PDXs, indicating that PDXs have the capacity to recapitulate the mutations in primary tumors. Nevertheless, a substantial number of mutations had higher allele frequencies in the PDXs than in the primary tumors, or were not detectable in the primary tumor, suggesting the possibility of tumor cell enrichment in PDXs or heterogeneity in the primary tumors. The molecularly annotated PDXs generated from this study could be useful for future translational studies.

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Setting up a wide panel of patient-derived tumor xenografts of non–small cell lung cancer by improving the preanalytical steps

Marius Ilie, Manoel Nunes, Lydia Blot … (2014)

With the ongoing need to improve therapy for non–small cell lung cancer (NSCLC) there has been increasing interest in developing reliable preclinical models to test novel therapeutics. Patient-derived tumor xenografts (PDX) are considered to be interesting candidates. However, the establishment of such model systems requires highly specialized research facilities and introduces logistic challenges. We aimed to establish an extensive well-characterized panel of NSCLC xenograft models in the context of a long-distance research network after careful control of the preanalytical steps. One hundred fresh surgically resected NSCLC specimens were shipped in survival medium at room temperature from a hospital-integrated biobank to animal facilities. Within 24 h post-surgery, tumor fragments were subcutaneously xenografted into immunodeficient mice. PDX characterization was performed by histopathological, immunohistochemical, aCGH and next-generation sequencing approaches. For this model system, the tumor take rate was 35%, with higher rates for squamous carcinoma (60%) than for adenocarcinoma (13%). Patients for whom PDX tumors were obtained had a significantly shorter disease-free survival (DFS) compared to patients for whom no PDX tumors (P = 0.039) were obtained. We established a large panel of PDX NSCLC models with a high frequency of mutations (29%) in EGFR, KRAS, NRAS, MEK1, BRAF, PTEN, and PI3KCA genes and with gene amplification (20%) of c-MET and FGFR1. This new patient-derived NSCLC xenograft collection, established regardless of the considerable time required and the distance between the clinic and the animal facilities, recapitulated the histopathology and molecular diversity of NSCLC and provides stable and reliable preclinical models for human lung cancer research.

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Author and article information

Contributors

Rossella Rota: Role: Editor

Journal

Journal ID (nlm-ta): PLoS One

Journal ID (iso-abbrev): PLoS ONE

Journal ID (publisher-id): plos

Journal ID (pmc): plosone

Title: PLoS ONE

Publisher: Public Library of Science (San Francisco, CA USA )

ISSN (Electronic): 1932-6203

Publication date (Electronic): 5 January 2016

Publication date Collection: 2016

Volume: 11

Issue: 1

Electronic Location Identifier: e0145100

Affiliations

[1 ]Department of Thoracic Surgery, University of Torino, 10126, Torino, Italy

[2 ]Department of Molecular Biotechnology and Health Science and Center for Experimental Research and Medical Studies, University of Torino, 10126, Torino, Italy

[3 ]Department of Pathology and Laboratory Medicine, Weill Cornell Medical College, New York, NY, 10021, United States of America

[4 ]Department of Medical Sciences, University of Torino, 10126, Torino, Italy

[5 ]Department of Medical Sciences, Allergology and Immunology, University of Torino, 10126, Torino, Italy

[6 ]Unit of Pulmonology, San Giovanni Battista Hospital, 10126, Torino, Italy

[7 ]Department of Pathology and NYU Cancer Center, New York University School of Medicine, New York, NY, 10016, United States of America

Ospedale Pediatrico Bambino Gesu', ITALY

Author notes

Competing Interests: The authors have declared that no competing interests exist.

Conceived and designed the experiments: FG FT LB ER GI. Performed the experiments: FT LB ER LA MG MT MB EE. Analyzed the data: FM LD PF PS. Contributed reagents/materials/analysis tools: AS LD LA AO GI. Wrote the paper: FG FT GI.

* E-mail: fra.guerrera@ 123456gmail.com

Article

Publisher ID: PONE-D-15-29283

DOI: 10.1371/journal.pone.0145100

PMC ID: 4701130

PubMed ID: 26731692

SO-VID: c900388d-1f36-45d5-8d7c-3010fc45e5cd

License:

This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited

History

Date received : 4 July 2015

Date accepted : 28 November 2015

Page count

Figures: 3, Tables: 3, Pages: 15

Funding

The authors have no support or funding to report.

Custom metadata

Data Availability All relevant data are within the paper and its Supporting Information files.

The Influence of Tissue Ischemia Time on RNA Integrity and Patient-Derived Xenografts (PDX) Engraftment Rate in a Non-Small Cell Lung Cancer (NSCLC) Biobank

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Abstract

Introduction

Methods

Results

Conclusion

Related collections

PLOS Climate

Most cited references 18

Patient-derived first generation xenografts of non-small cell lung cancers: promising tools for predicting drug responses for personalized chemotherapy.

Gene mutations in primary tumors and corresponding patient-derived xenografts derived from non-small cell lung cancer.

Setting up a wide panel of patient-derived tumor xenografts of non–small cell lung cancer by improving the preanalytical steps

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