Temple University

Drexel University

The Wistar Institute

Wenhui Hu

Kamel Khalili

Yonggang Zhang

Ming H Xiao

Jennifer McCaffrey

Harold C Riethman

Compositions for specifically cleaving target sequences in retroviruses include nucleic acids encoding a Clustered Regularly Interspace Short Palindromic Repeat (CRISPR) associated endonuclease and a guide RNA sequence complementary to one or more target nucleic acid sequences in a retrovirus genome.

Methods and compositions for RNA-guided treatment of HIV infection

Compositions for the in vivo delivery of a gene editing CRISPR/Cas9 complex was developed to eliminate integrated retroviral DNA sequences from latently infected human cells and animal disease models.

Excision of retroviral nucleic acid sequences

A method of treating a subject having a retroviral infection, by administering to the subject a therapeutically effective amount of a composition comprising a vector encoding a CRISPR-associated endonuclease and at least two guide RNAs, wherein the guide RNAs are complementary to two target sequences spanning from the 5'- to 3&#x2032;-LTRs of the sequence in the retrovirus, and eradicating the retroviral infection. A method of immunizing a subject at risk of retroviral infection, by administering a prophylactically effective amount of a composition comprising a Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR)-associated endonuclease, and two or more different guide RNAs (gRNAs), wherein each of the at least two gRNAs is complementary to a different target nucleic acid sequence in a long terminal repeat (LTR) of proviral DNA of a retrovirus to the subject, and preventing retroviral infection in the subject.

A method of preventing transmission of a retrovirus from a mother to her offspring, by administering to the mother a therapeutically effective amount of a composition comprising a Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR)-associated endonuclease, and the two or more different multiplex gRNAs, wherein each of the at least two gRNAs is complementary to a different target nucleic acid sequence in a long terminal repeat (LTR) of proviral DNA of the virus that is unique from the genome of the host cell, cleaving a double strand of the proviral DNA at a first target protospacer sequence with the CRISPR-associated endonuclease, cleaving a double strand of the proviral DNA at a second target protospacer sequence with the CRISPR-associated endonuclease, excising an entire HIV-1 proviral genome, eradicating the HIV-1 proviral DNA from the host cell, and preventing transmission of the proviral DNA to the offspring.

A method of treating a subject at risk for having a virus infection, by administering to the subject a prophylactically effective amount of a composition comprising a vector encoding a CRISPR-associated endonuclease and at least two guide RNAs, wherein the guide RNAs are complementary to two target sequences spanning from the 5'- to 3&#x2032;-LTRs of the sequence in the virus, and preventing a retroviral infection.

Provided herein are methods of detecting a target nucleic acid sequence. In one embodiment, the method includes contacting genomic DNA with a guide RNA having a portion complementary to the target sequence in the genomic DNA and with Cas9 nickase to produce a single-strand break in the genomic DNA at a specific location adjacent to the target sequence. The method further includes contacting the nicked DNA with a polymerase and fluorescently labeled nucleotide. The fluorescently labeled nucleotide is incorporated into the nicked DNA at the specific location and the target nucleic acid sequence is detected via fluorescent label.

Methods of specifically labeling nucleic acids using CRISPR/Cas

A method of inactivating a proviral DNA integrated into the genome of a host cell latently infected with a retrovirus by treating the host cell with a composition comprising a Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR)-associated endonuclease, and two or more different guide RNAs (gRNAs), wherein each of the at least two gRNAs is complementary to a different target nucleic acid sequence in a long terminal repeat (LTR) in the proviral DNA, and inactivating the proviral DNA. A composition for use in inactivating a proviral DNA integrated into the genome of a host cell latently infected with a retrovirus including isolated nucleic acid sequences comprising a CRISPR-associated endonuclease and a guide RNA, wherein the guide RNA is complementary to a target sequence in a human immunodeficiency virus.

Growing community of inventors

Cherry Hill, NJ, United States of America

Wenhui Hu