Pharmacologically induced transgene ablation system

摘要

The present invention relates to gene therapy systems designed for the delivery of a therapeutic product to a subject using replication-defective virus composition(s) engineered with a built-in safety mechanism for ablating the therapeutic gene product, either permanently or temporarily, in response to a pharmacological agent—preferably an oral formulation, e.g., a pill. The invention is based, in part, on the applicants' development of an integrated approach, referred to herein as “PITA” (Pharmacologically Induced Transgene Ablation), for ablating a transgene or negatively regulating transgene expression. In this approach, replication-deficient viruses are used to deliver a transgene encoding a therapeutic product (an RNA or a protein) so that it is expressed in the subject, but can be reversibly or irreversibly turned off by administering the pharmacological agent; e.g., by administration of a small molecule that induces expression of an ablator specific for the transgene or its RNA transcript.

主权项

1. A composition for AAV-mediated delivery of a therapeutic product, said composition comprising:
(a) a first recombinant AAV vector containing a nucleic acid molecule comprising:
(i) a nucleic acid sequence encoding a therapeutic product operably linked to a promoter that controls transcription of the therapeutic product; and(ii) at least one endonuclease ablation site which comprises a sequence of at least 30nucleic acid base pairs which are specifically recognized by at least ten (10×) zinc fingers, said at least one endonuclease ablation site being located at least 5′ to the sequence encoding the therapeutic product; and (b) a second recombinant AAV vector containing the coding sequence for at least one ablator which is a chimeric endonuclease comprising at least ten copies of a zinc finger domain linked to a functional endonuclease catalytic domain in operative association with a promoter, wherein transcription of the at least one ablator and/or ablation activity is induced in response to a pharmacological agent and said ablator recognizes at least one endonuclease ablation site in the first recombinant AAV vector; wherein the nucleic acid sequence in (a)(i) consists of at least one endonuclease ablation site on the first strand of the nucleic acid sequence and at least one second endonuclease ablation is located on the second strand of the nucleic acid sequence, wherein said second endonuclease ablation site is distinct from said endonuclease ablation site on the first strand and is specifically and selectively recognized by different zinc fingers.