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A comprehensive characterization of cell cultures and xenografts derived from a human verrucous penile carcinoma

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Tumor Biology

Abstract

This study aimed to establish and characterize primary cell cultures and xenografts derived from penile carcinoma (PeCa) in order to provide experimental models for cellular processes and efficacy of new treatments. A verrucous squamous cell carcinoma (VSCC) was macrodissected, dissociated, and cultivated in KSFM/DF12 medium. Cell cultures were evaluated at passage 5 (P5) using migration and invasion assays and were serially propagated, in vivo, in BALB/c nude mice until passage 3 (X1–X3). Immunophenotypic characterization of cultures and xenografts was performed. Genomic (CytoScan HD, Affymetrix) and transcriptomic profiles (HTA 2.0 platform, Affymetrix) for VSCC, cell cultures, and xenografts were assessed. P5 cells were able to migrate, invade the Matrigel, and produce tumors in immunodeficient mice, demonstrating their malignant potential. The xenografts unexpectedly presented a sarcomatoid-like carcinoma phenotype. Genomic analysis revealed a high similarity between the VSCC and tumor-derived xenograft, confirming its xenograft origin. Interestingly, a subpopulation of P5 cells presented stem cell-related markers (CD44+CD24− and ALDH1high) and sphere-forming capacity, suggesting their potential xenograft origin. Cell cultures and xenografts retained the genomic alterations present in the parental tumor. Compared to VSCC, differentially expressed transcripts detected in all experimental conditions were associated with cellular morphology, movement, and metabolism and organization pathways. Malignant cell cultures and xenografts derived from a verrucous penile carcinoma were established and fully characterized. Nevertheless, xenograft PeCa models must be used with caution, taking into consideration the selection of specific cell populations and anatomical sites for cell/tumor implantation.

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Acknowledgments

The authors are grateful for the assistance given by MSc. Rainer Marco Lopes Lapa and Dr Vilma R Martins for the scientific support with the xenograft model.

Funding

This study was supported by grants from the National Council of Technological and Scientific Development (CNPq 573589/08-9) and São Paulo Research Foundation (FAPESP 2009/52088-3, 2010/51601-6, and 2009/14027-2).

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    Authors and Affiliations

    1. International Research Center—CIPE, A. C. Camargo Cancer Center, São Paulo, SP, Brazil

      Juan J. Muñoz, Hellen Kuasne, Rolando A. R. Villacis, Fabio A. Marchi, Tiago G. Santos & Silvia R. Rogatto

    2. Genetic Graduation Program, Institute of Biosciences, São Paulo State University—UNESP, Botucatu, SP, Brazil

      Juan J. Muñoz

    3. NeoGene Laboratory, Department of Urology, Medical School, São Paulo State University—UNESP, CEP: 18.618-000, Botucatu, SP, Brazil

      Sandra A. Drigo, Hellen Kuasne & Silvia R. Rogatto

    4. Department of Pathology, Medical School, São Paulo State University—UNESP, Botucatu, SP, Brazil

      Maria A. C. Domingues

    5. Department of Pelvic Surgery, A. C. Camargo Cancer Center, São Paulo, SP, Brazil

      Ademar Lopes

    Authors
    1. Juan J. Muñoz
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    2. Sandra A. Drigo
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    3. Hellen Kuasne
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    4. Rolando A. R. Villacis
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    5. Fabio A. Marchi
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    6. Maria A. C. Domingues
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    7. Ademar Lopes
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    Corresponding authors

    Correspondence to Sandra A. Drigo or Silvia R. Rogatto.

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    Additional information

    Juan J. Muñoz and Sandra A. Drigo contributed equally to this work.

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    Fig. S1

    Schematic flow chart of the study design including the establishment and characterization of verrucous penile carcinoma cell cultures and xenografts. IHC: immunohistochemistry; P1-P5: cell culture passage from first to fifth (in vitro), X1, X2 and X3: xenograft in the first, second and third passages (in vivo). Immunophenotyping of xenograft cell cultures (X1, X2 and X3) was done (rectangle with dashed lines). (TIF 945 kb)

    Table S1

    Antibodies used for phenotypic characterization assays. (DOC 41 kb)

    Table S2

    Phenotypic characterization of cell cultures and xenografts. (DOC 39 kb)

    Table S3

    Genomic alterations detected in cell cultures (P1 and P5) and xenograft on passage 1 (X1) in comparison with the parental penile carcinoma (VSCC). (DOC 106 kb)

    Table S4

    Top 20 differentially expressed genes detected in cell cultures (P1 and P5) and xenografts (X1-X3) in comparison with the parental penile carcinoma (VSCC). (DOC 54 kb)

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    Muñoz, J.J., Drigo, S.A., Kuasne, H. et al. A comprehensive characterization of cell cultures and xenografts derived from a human verrucous penile carcinoma. Tumor Biol. 37, 11375–11384 (2016). https://doi.org/10.1007/s13277-016-4951-z

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    • DOI: https://doi.org/10.1007/s13277-016-4951-z

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