Regulation of herpes simplex virus 1 replication using tumor-associated promoters.
OBJECTIVE
To investigate use of transcriptional regulatory elements (promoters) for tumor-associated antigens to achieve HSV-1 replication preferentially in cells that overexpress the tumor-associated antigens.


BACKGROUND
An important advantage of replicating viruses for cancer therapy is their ability to simultaneously destroy tumor cells by replication and release progeny virion to infect and destroy adjacent cancer cells. This strategy requires regulation of the viral life cycle to obtain robust replication in neoplastic cells and minimize replication in nonneoplastic cells.


METHODS
Promoters for the human carcinoembryonic antigen (CEA) and MUC1/DF3 tumor-associated antigens were characterized and cloned into HSV-1 mutants as heterologous promoters regulating expression of two different HSV-1 genes. Viral replication in tumor cells and cytotoxicity was quantified with in vitro assays. Antineoplastic efficacy was characterized in a flank tumor xenograft model.


RESULTS
Several CEA promoters were cloned and characterized using luciferase reporter assays. The most specific promoter was used to construct and isolate two different HSV-1 mutants in which critical genes are regulated by this promoter (ICP4 and gamma(1) 34.5). Similarly, the promoter for the DF3/MUC1 tumor-associated antigen was cloned into a third HSV-1 mutant such that it regulates expression of gamma(1) 34.5. Regulation of ICP4 expression by the CEA promoter during HSV-1 infection overly attenuates viral replication. Regulation of gamma(1) 34.5 expression by either the CEA promoter or the MUC1/DF3 promoter during HSV-1 infection modulates viral replication, with preferential replication in cells that overexpress the corresponding tumor-associated antigen. A single intratumoral inoculation of an HSV-1 mutant with the MUC1/DF3 promoter regulating gamma(1) 34.5 expression results in significant antineoplastic activity in MUC1-positive pancreatic carcinoma xenografts as compared to mock inoculation.


CONCLUSIONS
Promoters for tumor-associated antigens may be incorporated into the HSV-1 genome to regulate HSV-1 replication. The choices of HSV-1 gene and tumor-associated promoter are important determinants of success of this strategy. Because of its preferential replication in MUC1-positive tumors, an HSV-1 mutant with the MUC1/DF3 promoter regulating gamma(1) 34.5 expression will undergo further examination as a novel cancer therapy agent.
D000818:Animals; D000951:Antigens, Neoplasm / Q000276:immunology* / Q000494:pharmacology*; D002272:Carcinoembryonic Antigen / Q000235:genetics / Q000276:immunology; D002277:Carcinoma / Q000235:genetics* / Q000276:immunology*; D002522:Chlorocebus aethiops; D003110:Colonic Neoplasms / Q000235:genetics* / Q000276:immunology*; D004195:Disease Models, Animal; D015967:Gene Expression Regulation, Viral / Q000187:drug effects* / Q000235:genetics*; D000882:Haplorhini; D018259:Herpesvirus 1, Human / Q000187:drug effects* / Q000235:genetics*; D006801:Humans; D066298:In Vitro Techniques; D007668:Kidney / Q000187:drug effects / Q000276:immunology; D011401:Promoter Regions, Genetic / Q000187:drug effects* / Q000235:genetics*; D014779:Virus Replication / Q000187:drug effects* / Q000235:genetics*
D000951:Antigens, Neoplasm; D002272:Carcinoembryonic Antigen