Abstract
In gene therapy, effective and selective suicide gene expression is crucial. We exploited the endogenous Long INterspersed Element-1 (L1) machinery often reactivated in human cancers to integrate the Herpes Simplex Virus Thymidine Kinase (HSV-TK) suicide gene selectively into the genome of cancer cells. We developed a plasmid-based system directing HSV-TK expression only when reverse transcribed and integrated in the host genome via the endogenous L1 ORF1/2 proteins and an Alu element. Delivery of these new constructs into cells followed by Ganciclovir (GCV) treatment selectively induced mortality of L1 ORF1/2 protein expressing cancer cells, but had no effect on primary cells that do not express L1 ORF1/2. This novel strategy for selective targeting of tumour cells provides high tolerability as the HSV-TK gene cannot be expressed without reverse transcription and integration, and high selectivity as these processes take place only in cancer cells expressing high levels of functional L1 ORF1/2.
Highlights
Suicide gene therapy for cancer treatment, referred to as gene-directed enzyme pro-drug therapy (GDEPT), aims at selectively targeting cancer cells as an alternative or complementary adjuvant to classical chemotherapies [1]
We describe a novel strategy for efficient and selective expression of the Herpes Simplex Virus Thymidine Kinase (HSV-TK) suicide gene in cancer cells based on their Long INterspersed Element-1 (L1) ORF1/2p expression
We show that transfection with Alu-driven vectors or infection with the corresponding Associated Virus vectors (AAVs) renders the tumour cells sensitive to efficient GCV mediated growth inhibition both as monolayers and spheroid cultures
Summary
Suicide gene therapy for cancer treatment, referred to as gene-directed enzyme pro-drug therapy (GDEPT), aims at selectively targeting cancer cells as an alternative or complementary adjuvant to classical chemotherapies [1]. GDEPT is based on introducing into tumour cells a viral or a bacterial “suicide” gene encoding an enzyme able to activate a non- or mildly toxic prodrug leading to tumour cell death. The suicide gene has to be selectively active in tumour cells and the pro-drug is delivered either by local or systemic administration and is only activated if the cells express the suicide gene. The most extensively studied suicide gene system is the combination of Herpes Simplex Virus Thymidine Kinase (HSV-TK) and the pro-drug Ganciclovir (GCV). DNA polymerase incorporates GCV-triphosphate into replicating DNA leading to cell death by polymerase inhibition [2]. To be effective GDEPT approaches have to target tumour cells
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