Safe lentiviral vectors for targeted delivery of multiple therapeutic molecules

Inventors

Lai, Zhennan • Galvin, Jeffrey A.

Assignees

American Gene Technologies International Inc

Abstract

The present application discloses a lentiviral transfer system which includes: (i) a self-inactivating transfer vector comprising: multiple gene units, wherein each gene unit includes a heterologous nucleic acid sequence operably linked to a regulatory nucleic acid sequence; and (ii) a helper construct which lacks a 5′ LTR, wherein the 5′ LTR has been replaced with a heterologous promoter, in which the helper construct further comprises: a lentiviral env nucleic acid sequence containing a deletion, wherein the deleted env nucleic acid sequence does not produce functional env protein; and a packaging signal contains a deletion, wherein the deleted packaging signal is nonfunctional.

Core Innovation

The invention relates to a self-inactivating lentiviral transfer system derived from HIV-1 for targeted delivery and long-term expression of multiple therapeutic molecules. The system includes a self-inactivating transfer vector that carries multiple gene units, where each gene unit comprises a heterologous nucleic acid sequence operably linked to a regulatory nucleic acid sequence. The vector further includes a mammalian insulator sequence and splice acceptor and splice donor sites, and the self-inactivating transfer vector is free of wPRE (wood-chuck hepatitis virus post-transcriptional element).

In the described system, helper construct features and deletions render env nucleic acid nonfunctional and disable the packaging signal by deletion, and the functional 5′ LTR is replaced by a heterologous promoter. Multiple heterologous gene units allow combined genetic payloads, including RNAi agents and tumor suppressor P53. Optional regulatory and control elements and encoded functional additions are described, including trafficking-signal fusions and translation initiation elements such as IRES, as well as enzyme-cleavage control elements.

The disclosed payloads include RNAi targeting BCL2 (Bcl-2) and P53, and the system is described as supporting combinations such as P53–BCL2 RNAi, with tumor reduction and tumor marker changes reported in in vitro and in vivo examples. Cell/tissue-specific promoter usage is described in the context of prostate cancer, including human prostate-specific antigen-related outcomes. Broader therapeutic indications described in the document include other cancers and multiple genetic or neurological disorders via replacement enzymes and/or RNAi.

Claims Coverage

The independent claims cover a lentiviral transfer system with a specific self-inactivating multi-gene-unit vector architecture and a pharmaceutical composition comprising lentiviral particles produced using that system. Across the independent claims, the inventive structure centers on multiple gene units with heterologous nucleic acids linked to regulatory nucleic acids, inclusion of a mammalian insulator and splice acceptor/donor sites, and the vector being free of wPRE.

Overall, claim coverage emphasizes a wPRE-free self-inactivating lentiviral transfer vector architecture with multiple gene units, heterologous nucleic acids linked to regulatory nucleic acids, and a mammalian insulator plus splice acceptor/donor sites. The independent pharmaceutical-composition claim ties a lentiviral particle product to that same vector architecture, with dependent claims adding optional envelope functionality, promoter selection for cell/tissue specificity, trafficking-signal encoding, and payloads including RNAi agents and Bcl-2 expression inhibition.

Stated Advantages

Documented Applications