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Intramuscular Transplantation Improves Engraftment Rates for Esophageal Patient-Derived Tumor Xenografts

  • Translational Research and Biomarkers
  • Published:
Annals of Surgical Oncology Aims and scope Submit manuscript

Abstract

Background

Recently, there has been an increase in the availability of targeted molecular therapies for cancer treatment. The application of these approaches to esophageal cancer, however, has been hampered by the relative lack of appropriate models for preclinical testing. Patient-derived tumor xenograft (PDTX) models are gaining popularity for studying many cancers. Unfortunately, it has proven difficult to generate xenografts from esophageal cancer using these models. The purpose of this study was to improve the engraftment efficiency of esophageal PDTXs.

Methods

Fresh pieces of esophageal tumors obtained from endoscopic biopsies or resected specimens were collected from 23 patients. The tumors were then coated in Matrigel and transplanted in immunocompromised mice subcutaneously (n = 6) and/or using a novel implantation technique whereby the tumor is placed in a dorsal intramuscular pocket (n = 18). They are then monitored for engraftment.

Results

With the novel intramuscular technique, successful engraftment was achieved for all 18 patient tumors. Among these PDTXs, 13 recapitulated the original patient tumors with respect to degree of differentiation, molecular and genetic profiles, and chemotherapeutic response. Lymphomatous transformation was observed in the other five PDTXs. Successful engraftment was achieved for only one of six patient tumors using the classic subcutaneous approach.

Discussion

We achieved a much higher engraftment rate of PDTXs using our novel intramuscular transplant technique than has been reported in other published studies. It is hoped that this advancement will help expedite the development and testing of new therapies for esophageal cancer.

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Acknowledgment

This work was supported by a NHMRC of Australia Centres for Research Excellence Grant (1040947). MR received the Thornell-Shore Memorial scholarship from the Royal Australasian College of Surgeons (RACS) and the Sir Thomas Naghten Fitzgerald scholarship from The University of Melbourne. DL received an NHMRC Australian Postgraduate Research award and Foundation for a Surgery Scholarship from the RACS. JS is supported by the Victorian Cancer Agency/Snowdome Foundation “Eva & Les Erdi Fellowship.” Some tissue samples used in this Project were provided by the Victorian Cancer Biobank with appropriate ethics approval. The Victorian Cancer Biobank is supported by the Victorian Government, Australia.

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Author information

Authors and Affiliations

  1. Surgical Oncology Research Laboratory, Peter MacCallum Cancer Centre, Melbourne, VIC, Australia

    Matthew Read MBBS, David Liu MBBS, BMedSci, Cuong P. Duong MBBS, FRACS, PhD, Carleen Cullinane BSc(Hons), PhD, Christina M. Fennell BBiomedSc(Hons), Nicholas J. Clemons BSc(Hons), PhD & Wayne A. Phillips BSc(Hons), PhD

  2. Sir Peter MacCallum Department of Oncology, University of Melbourne, Melbourne, VIC, Australia

    Matthew Read MBBS, David Liu MBBS, BMedSci, Carleen Cullinane BSc(Hons), PhD, Christina M. Fennell BBiomedSc(Hons), Jake Shortt BMedSc, MBChB, FRACP, FRCPA, PhD, Nicholas J. Clemons BSc(Hons), PhD & Wayne A. Phillips BSc(Hons), PhD

  3. Division of Cancer Surgery, Peter MacCallum Cancer Centre, Melbourne, VIC, Australia

    Cuong P. Duong MBBS, FRACS, PhD & Wayne A. Phillips BSc(Hons), PhD

  4. Department of Surgery (St. Vincent’s Hospital), University of Melbourne, Melbourne, VIC, Australia

    Nicholas J. Clemons BSc(Hons), PhD & Wayne A. Phillips BSc(Hons), PhD

  5. Department of Pathology, Peter MacCallum Cancer Centre, Melbourne, Australia

    William K. Murray MBBS, FRCPA, FRCPath & David Westerman MBBS, FRACP, FRCPA, FFSc

  6. School of Clinical Sciences at Monash Health, Melbourne, Australia

    Jake Shortt BMedSc, MBChB, FRACP, FRCPA, PhD

  7. Faculty of Medicine, Nursing & Health Sciences, Monash University, Melbourne, Australia

    Jake Shortt BMedSc, MBChB, FRACP, FRCPA, PhD

  8. University of Melbourne, Melbourne, Australia

    David Westerman MBBS, FRACP, FRCPA, FFSc

  9. Monash University Centre for Obesity Research and Education, Alfred Hospital, Melbourne, Australia

    Paul Burton MBBS, FRACS, PhD

  10. Cabrini Hospital, Melbourne, VIC, Australia

    Paul Burton MBBS, FRACS, PhD

Authors
  1. Matthew Read MBBS
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  2. David Liu MBBS, BMedSci
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  3. Cuong P. Duong MBBS, FRACS, PhD
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  4. Carleen Cullinane BSc(Hons), PhD
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  5. William K. Murray MBBS, FRCPA, FRCPath
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  6. Christina M. Fennell BBiomedSc(Hons)
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  7. Jake Shortt BMedSc, MBChB, FRACP, FRCPA, PhD
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  8. David Westerman MBBS, FRACP, FRCPA, FFSc
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  10. Nicholas J. Clemons BSc(Hons), PhD
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  11. Wayne A. Phillips BSc(Hons), PhD
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Corresponding author

Correspondence to Wayne A. Phillips BSc(Hons), PhD.

Additional information

Nicholas J. Clemons and Wayne A. Phillips are co-senior authors.

Electronic supplementary material

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10434_2015_4425_MOESM1_ESM.tif

Supplementary Fig. 1 Intramuscular (IM) implantation technique and PDTX formation. a After placement of a stay suture, a small IM pocket is created on the dorsum of the mouse that is just large enough to accommodate the tumor piece. b After implantation, the IM pocket is sutured closed with the implant evident immediately below the outer muscle fibers. c Gross view of an IM PDTX at the time of harvest (TIFF 2961 kb)

10434_2015_4425_MOESM2_ESM.tif

Supplementary Fig. 2 Identification and characterization of lymphomagenic transformation within a subset of established PDTXs. Immunohistochemistry using pan-cytokeratin AE1/AE3 (1:100) (NCL-L-AE1/AE3; Novocastra/Leica Microsystems, Newcastle, UK), CD20 (1:200) (M7055, DakoCytomation; Dako, Glostrup, Denmark), and CD45 (1:200) (M0701, DakoCytomation) antibodies on the original patient tumor and the corresponding PDTX. The PDTX of patient 9 (a) is negative for the pan-cytokeratin marker AE1/AE3 but positive for the common leukocyte antigen CD45 and the B-lymphocyte restricted antigen CD20, indicating that it is predominantly composed of human B cells. In comparison, the PDTX of patient 16 (b) exhibits a combination of epithelial cells (nests of AE1/AE3-positive cells) and lymphocytes. c Further analysis via flow cytometry was performed as previously described (Craig FE, Foon KA. Blood 2008;222:3941–67) of single cells derived from PDTX 14 and confirmed that the lymphoid cells were CD19+ B cells (left plot). The abnormal B-cell population was then gated and is depicted on the FSC vs. SSC plot in blue (right plot), indicating that these cells are also large. d Right In situ hybridization for the Epstein Barr virus (EBV) (blue staining) was performed on sections of formalin-fixed paraffin-embedded PDTX using the INFORM EBER Probe and ISH iVIEW Blue Detection Kit (Ventana/Roche, Tucson, AZ, USA) as per the manufacturer’s protocol and counterstained with eosin (pink staining). Left EBV-positive nasopharyngeal carcinoma (positive control). Right Section from PDTX 9 (TIFF 3402 kb)

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Read, M., Liu, D., Duong, C.P. et al. Intramuscular Transplantation Improves Engraftment Rates for Esophageal Patient-Derived Tumor Xenografts. Ann Surg Oncol 23, 305–311 (2016). https://doi.org/10.1245/s10434-015-4425-3

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