Does Detection of Circulating ONYX-015 Genome by Polymerase Chain Reaction Indicate Vector Replication?
Authors: Antoine Yver, John J. Nemunaitis, Casey Cunningham
Journal: Journal of Clinical Oncology (2001)

Article Tools SPECIAL DEPARTMENTS Article Tools OPTIONS & TOOLS Export Citation Track Citation Add To Favorites Rights & Permissions COMPANION ARTICLES No companion articles ARTICLE CITATION DOI: 10.1200/JCO.2001.19.12.3155 Journal of Clinical Oncology - published online before print September 21, 2016 PMID: 11408514 Does Detection of Circulating ONYX-015 Genome by Polymerase Chain Reaction Indicate Vector Replication? Antoine YverxAntoine YverSearch for articles by this author John J. NemunaitisxJohn J. NemunaitisSearch for articles by this author , Casey CunninghamxCasey CunninghamSearch for articles by this author Show More Aventis Pharmaceuticals Inc, Bridgewater, NJUS Oncology, Dallas, TX https://doi.org/10.1200/JCO.2001.19.12.3155 First Page Full Text PDF Figures and Tables © 2001 by American Society of Clinical OncologyjcoJ Clin OncolJournal of Clinical OncologyJCO0732-183X1527-7755American Society of Clinical OncologyResponse15062001In Reply:We appreciate the additional unpublished data provided by Dr Yver, particularly as published literature has not identified detection (> 1 × 103 plaque forming units/0.5 mL) of RPR/ING201 in blood for more than 90 minutes.1-3 Nevertheless, although we agree with Yver that injection of replication-defective adenoviral vectors may be associated with transient circulation of viral genome, we still maintain that the prolonged detectable circulation of Onyx-015 genome as detailed in our article is suggestive of intratumoral replication.4 We agree that our data does not prove replication. However, much of our argument is based on the consideration that, given the assay's sensitivity, judgements regarding the significance of polymerase chain reaction detection of nucleic acids always hinge strongly on the precise definition of what constitutes a threshold detection limit. Our assay was validated for a detection limit sensitivity to 1.05 × 104 Ad particles/mL, with a qualification sensitivity of 4.2 × 104 Ad particles/mL. Our reported results are all above the sensitivity level (ie, > 4.2 × 104 particles/mL), and the assay was validated to be specific for Onyx-015 and was performed independently by Althea Technologies (San Diego, CA). Because Yver has not published his results, it is difficult to interpret his statement. Once his data are published, the scientific community can rigorously evaluate the sensitivity and specificity of his assays and data in the context of ours. Further, although Yver's methods of demonstrating that the viral genome recovered was intact virus are elegant, we also note that E1-deleted vectors such as RPR/ING201 can, and do, replicate in some cancer cells.5,6 This is a critical issue to determine whether or not Yver's observation represents viral replication in his system and one that cannot be evaluated on unpublished data.In addition, in one of our subsequent trials where we administered intravenous Onyx-015, we demonstrated plasma levels of Onyx-015 genome between 5.6 and 890 × 104 genomes/mL 7 days after a single infusion (2 × 1010 to 2 × 1013 particles/mL). And further, we were able to demonstrate an increase in circulating viral genome at 48 hours 2.5- to 10-fold above those seen 6 hours after initial Onyx-015 infusion. Such an increase with already significant levels of circulating viral genome is most easily explained by continued viral replication. Finally, other evidence provided by electron micrographs of biopsied malignant tissue reveal bulging nuclei filled with Onyx-015 during the period when viral genome is detected in circulation, whereas adjacent nonmalignant tissue shows no evidence of viral presence.7,8In summary, we reported the first peer-reviewed data of persistently detectable viral genome after administration of replication-competent virus, which does suggest viral replication. This result is based on rigorously validated assays in the context of peer-reviewed publication. However, we appreciate Yver's point that further research is needed. We are in the process of development of a quantitative plaque assay to potentially measure functional Onyx-015 from stored plasma samples of previously treated patients with detectable circulating genome. We thank Dr Yver and the editors for the opportunity to expand here the discussion of this important area.1. Clayman G: Adenovirus-mediated p53 gene transfer in patients with advanced recurrent head and neck squamous cell carcinoma. J Clin Oncol 16:: 2221,1998-2232, Link, Google Scholar2. Nemunaitis J, Swisher G, Timmons T, et al: Adenovirus-mediated p53 gene transfer in sequence with cisplatin to tumors of patients with non-small-cell lung cancer. J Clin Oncol 18:: 609,2000-622, Link, Google Scholar3. Swisher SG, Roth JA, Nemunaitis J, et al: Adenovirus-mediated p53 gene transfer in advanced non-small-cell lung cancer. J Natl Cancer Inst 91:: 763,1999-771, Crossref, Medline, Google Scholar4. Nemunaitis J, Khuri F, Ganly I, et al: Phase II trial of intratumoral injection of ONYX-015, a replication-selective adenovirus, in patients with refractory head and neck cancer. J Clin Oncol 19:: 289,2001-298, Link, Google Scholar5. Steinwaerder D, Carlson C, Otto D, et al: Tumor-specific gene expression in hepatic metastases by a replication-activated adenovirus vector. Nat Med 7:: 240,2001-244, Crossref, Medline, Google Scholar6. Steinwaerder D, Carlson C, Lieber A: DNA replication of first-generation adenovirus vectors in tumor cells. Gene Therapy (in press) Google Scholar7. Nemunaitis J, Cunningham C, Buchanan A, et al: Intravenous infusion of a replication-selective adenovirus (ONYX-015) in cancer patients: Safety, feasibility and biological activity. Gene Therapy (in press) Google Scholar8. Nemunaitis J, Ganly I, Khuri F, et al: Selective replication and oncolysis in p53 mutant tumors with ONYX-015, an E1B-55kD gene-deleted adenovirus, in patients with advanced head and neck cancer: A Phase II trial. Cancer Res 60:: 6359,2000-6366, Medline, Google Scholar