Related Experiment Videos
Regulation of herpes simplex virus 1 replication using tumor-associated promoters
John T Mullen1, Hideki Kasuya, Sam S Yoon
1Division of Surgical Oncology, Department of Surgery, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 02114, USA.
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.
Summary Background Data:
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.
Insights
This study engineered herpes simplex virus-1 (HSV-1) to replicate specifically in cancer cells using tumor-associated antigen promoters. This targeted replication strategy shows promise for novel cancer therapies by enhancing oncolytic virus efficacy.
Area of Science:
- Oncolytic virotherapy
- Molecular oncology
- Viral gene therapy
Background:
- Replicating viruses offer a dual cancer treatment approach: direct tumor cell lysis and infection of adjacent cancer cells.
- Effective oncolytic virotherapy necessitates viral replication targeted to neoplastic cells while sparing healthy tissues.
Purpose of the Study:
- To engineer herpes simplex virus-1 (HSV-1) for preferential replication in tumor cells overexpressing specific tumor-associated antigens.
- To utilize transcriptional regulatory elements (promoters) from tumor-associated antigens to control HSV-1 gene expression and viral replication.
Main Methods:
- Characterized and cloned promoters for human carcinoembryonic antigen (CEA) and MUC1/DF3 tumor-associated antigens.
- Engineered HSV-1 mutants where viral genes (ICP4, gamma(1) 34.5) are regulated by these tumor-specific promoters.
- Assessed viral replication, tumor cell cytotoxicity in vitro, and antineoplastic efficacy in a pancreatic carcinoma xenograft model.
Main Results:
- CEA and MUC1/DF3 promoters were cloned and characterized; MUC1/DF3 promoter regulated gamma(1) 34.5 expression in an HSV-1 mutant.
- HSV-1 mutants exhibited modulated replication, with preferential replication in cells overexpressing the corresponding tumor-associated antigen.
- An HSV-1 mutant utilizing the MUC1/DF3 promoter demonstrated significant antineoplastic activity in MUC1-positive pancreatic xenografts.
Conclusions:
- Tumor-associated antigen promoters can be integrated into the HSV-1 genome to control viral replication for cancer therapy.
- The selection of the HSV-1 gene and the tumor-associated promoter are critical for therapeutic success.
- An HSV-1 mutant regulated by the MUC1/DF3 promoter warrants further investigation as a novel oncolytic agent due to its tumor-specific replication.