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Enhanced oncolysis mediated by Coxsackievirus A21 in combination with doxorubicin hydrochloride

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Summary

Virotherapy is an emerging strategy for the treatment of cancer that utilizes both replication-competent and genetically modified viruses to selectively kill tumor cells. We have previously shown that Coxsackievirus A21 (CVA21), a common-cold producing enterovirus, is an effective oncolytic agent against human melanoma, prostate, and breast cancer xenografts in vivo. CVA21 specifically targets and lytically infects susceptible cells expressing the CVA21 cellular receptors, intercellular adhesion molecule-1 (ICAM-1) and decay-accelerating factor (DAF). Herein, the efficacy of CVA21 administered in combination with doxorubicin hydrochloride as a new therapeutic regimen for cancer was investigated. Flow cytometric analysis demonstrated that the human breast, colorectal, and pancreatic cancer cell lines examined expressed moderate levels of surface ICAM-1 and DAF, whilst a normal breast cell line expressed only minimal levels. When CVA21 was combined with doxorubicin hydrochloride, synergistically enhanced cell death was observed when CVA21 was administered both simultaneously or 24 h prior to doxorubicin hydrochloride exposure. Doxorubicin hydrochloride had no effect on CVA21 replication. Through the use of an orthotopic (MDA-MB-231-luc) xenograft SCID mouse model of human breast cancer we showed that a single intravenous injection of CVA21 in combination with an intraperitoneal injection of doxorubicin hydrochloride resulted in significantly greater tumor reduction compared to either agent alone. Overall, these findings highlight the exciting potential of CVA21, administered in combination with doxorubicin hydrochloride, as a new therapeutic regimen for cancer.

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Abbreviations

ANOVA:

One-way analysis of variance

CI:

Combination index

CVA21:

Coxsackievirus A21

DAF:

Decay-accelerating factor

DMSO:

Dimethyl sulfoxide

FBS:

Fetal bovine serum

5-FU:

5-fluorouracil

HSV-1:

Herpes Simplex Virus-1

ICAM-1:

Intercellular adhesion molecule-1

MOI:

Multiplicity of infection

PBS:

Phosphate buffered saline

SCID:

Severe combined immunodeficient

SEM:

Standard error of the mean

SPF:

Specific-pathogen-free

TCID50 :

50% tissue culture infective dose

UV:

Ultraviolet

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Acknowledgements

We gratefully acknowledge those investigators mentioned in the text for the provision of monoclonal antibodies, cell lines and virus that enabled this study to be undertaken. We would like to thank Dr Gough Au, Dr Susanne Johansson, Dr Linda Berry, Dr Erin Haley, and Ms Catherine Delahunty for scientific advice and technical assistance throughout this project. Funding for this work was derived from grants received from the Biotechnology Innovation Fund and Hunter Medical Research Institute. Kathryn Skelding is a National Breast Cancer Foundation Scholar.

Declaration of competing interests

Darren Shafren and Richard Barry hold shares in Viralytics Ltd and are currently conducting research sponsored by this company.

Authors’ contributions

KS participated in the design of the study, carried out the in vitro and in vivo work, and drafted the manuscript. RB participated in the design of the study. DS participated in the design of the study, and was involved in the revision of the manuscript. All authors approved the final manuscript.

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Authors and Affiliations

  1. The Picornaviral Research Unit, Discipline of Immunology and Microbiology, Faculty of Health, The University of Newcastle, David Maddison Clinical Sciences Building, Newcastle, New South Wales, 2300, Australia

    Kathryn A. Skelding, Richard D. Barry & Darren R. Shafren

  2. Viralytics Ltd, Suite 1B, 55-63 Grandview St, Pymble, New South Wales, 2073, Australia

    Darren R. Shafren

Authors
  1. Kathryn A. Skelding
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  2. Richard D. Barry
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  3. Darren R. Shafren
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Corresponding author

Correspondence to Darren R. Shafren.

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Supplementary Table 1

Combination index for MDA-MB-231 cells simultaneously treated with a combination of CVA21 and paclitaxel, tamoxifen, or 5-FU. Combination indices (CI) were calculated using CalcuSyn, as described in the Materials and methods. CI values less than 0.9 were determined to be synergistic, between 0.9 and 1.1 to be additive, and greater than 1.1 to be antagonistic. (GIF 56 kb)

High resolution image (TIFF 9597 kb)

Supplementary Figure 1

Effect of doxorubicin hydrochloride, paclitaxel, 5-FU, and tamoxifen on CVA21 replication. MDA-MB-231 cell monolayers were inoculated with CVA21 (MOI = 10 TCID50/cell). Following the removal of unbound virus, the cell monolayers were incubated with (A) 2.5 or 0.32 μM doxorubicin hydrochloride, (B) 8 or 2 nM paclitaxel, (C) 10 nM or 10 μM 5-FU, or (D) 10 μM or 313 nM tamoxifen for 48 h. At selected time points (0, 2, 4, 6, 8, 10, 24, and 48 h), cell suspensions were freeze-thawed, clarified, and virus yield was determined by viral lytic assay on SK-Mel-28 cells. (GIF 40 kb)

High resolution image (TIFF 2140 kb)

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Skelding, K.A., Barry, R.D. & Shafren, D.R. Enhanced oncolysis mediated by Coxsackievirus A21 in combination with doxorubicin hydrochloride. Invest New Drugs 30, 568–581 (2012). https://doi.org/10.1007/s10637-010-9614-0

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  • DOI: https://doi.org/10.1007/s10637-010-9614-0

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