Tumor reduction in vivo after adenoviral mediated gene transfer of the herpes simplex virus thymidine kinase gene and ganciclovir treatment in human head and neck squamous cell carcinoma

Abstract

Gene transfer offers the possibility of novel therapies for head and neck squamous cell carcinoma (HNSCC). To this end, we demonstrate that a replication deficient adenovirus vector (Ad.RSVlacZ) can efficiently transduce foreign genes into human HNSCC cell lines in vitro, and that adenoviral mediated transfer of herpes simplex virus thymidine kinase (Ad.RSVtk) followed by exposure to ganciclovir results in tumor cell killing in vitro and in vivo. Exposure to Ad.RSVlacZ resulted in lacZ expression at multiplicities of infection (MOIs) of 10 and 100 for the cell lines HEp-2 and FaDu, respectively. This increased to 97% (HEp-2) and 49% (FaDu) at an MOI of 10,000. For HEp-2, maximum expression occurred during the first 48 hours after exposure (52% at 24 hours, 48% at 48 hours; MOI 500), then declined by 40% per day. This rapid decline may be caused by dilution of the gene through cell proliferation, because normalizing for the increase in total protein shows that the total number of cells expressing lacZ is stable from days 1 to 4. FaDu and HEp-2 were then transduced by AD.RSVtk and exposed to 20 μM ganciclovir for 24 hours. Significant tumor cell killing, as measured by a colony forming assay, occurred at an MOI of 2 for HEp-2 and 20 for FaDu. At an MOI of 200, 100% of HEp-2 and 97% of FaDu cells were killed. Next, subcutaneous tumor nodules derived from FaDu and HEp-2 were established in the flanks of SCID mice. Direct intratumoral injection of Ad.RSVtk followed by 7 days of ganciclovir therapy resulted in an adenovirus dose dependent reduction of tumor growth, and an actual size reduction of established tumor nodules at the highest does (10 10 plaque forming units). In conclusion, an adenovirus vector can efficiently transduce HNSCC cell lines in vitro. Maximum marker gene expression occurred during the first 48 hours after transduction. Transduction by Ad.RSVtk followed by exposure to ganciclovir resulted in tumor cell killing in vitro and in vivo.