1. ¹Department of Microbiology and Immunology, University of Illinois College of Medicine, Chicago, IL, USA
2. ²School of Life Sciences and Biotechnology, Korea University, Seoul, Korea
3. ³GenUs BioSystems Inc., Northbrook, IL, USA

Correspondence: Dr EP Cohen, Department of Microbiology and Immunology (m/c 790), 835 South Wolcott Ave, Chicago, IL 60615, USA. E-mail: EPCohen@UIC.EDU

Received 22 July 2006; Revised 16 October 2006; Accepted 17 November 2006; Published online 2 February 2007.

Abstract

This study describes a new strategy for the identification of squamous carcinoma antigens tumor-associated antigens (TAA). The antigens were discovered by comparing microarrays of squamous carcinoma vaccines highly enriched for immunotherapeutic cells with non-enriched vaccines. The vaccines were prepared by transferring sheared genomic DNA fragments (25 kb) from KLN205 cells, a squamous carcinoma cell line (DBA/2 mouse origin (H-2^d) into LM fibroblasts (C3H/He origin, H-2^k). The transferred tumor DNA segments integrate spontaneously into the genome of the recipient cells, replicate as the cells divide and are expressed. As only a small proportion of the transfected cell population was expected to have incorporated DNA segments that included genes specifying TAA (the vast majority specify normal cellular constituents), a novel strategy was employed to enrich the vaccine for TAA-positive cells. Microarrays were used to compare genes expressed by enriched and non-enriched vaccines. Seventy-five genes were overexpressed in cells from the enriched vaccine. One, the gene for Cytochrome P450 (family 2, subfamily e, polypeptide 1) (Cyp2e1), was overexpressed in the enriched but not the non-enriched vaccine. A vaccine for squamous carcinoma was prepared by transfer of a 357 bp fragment of the gene for Cyp2e1 into the fibroblast cell line. Robust immunity, sufficient to result in indefinite survival, was induced in tumor-bearing mice immunized with cells transfected with this gene fragment.