University of Massachusetts

Children's Medical Center Corporation

Scot Andrew Wolfe

Mehmet Fatih Bolukbasi

Erik J Sontheimer

Nadia Amrani

Ankit Gupta

Christian Mueller

Daniel E Bauer

Thoru Pederson

Shaojie Zhang

Ardalan Naseri

Jae Keith Joung

Hanhui Ma

Kevin Luk

Metewo Selase Kosi Enuameh

Nicola Anne Kearns

Rene Maehr

Manuel Garber

Sneha Suresh

Sukanya Iyer

Jennifer Chen

Xiangdong Meng

Yuxuan Wu

Benjamin Roscoe

Dongsheng Guo

Jing Zeng

Ryan Michael Jude Genga

Charles P Emerson

Oliver King

Pengpeng Liu

The present invention is directed to the filed of gene therapy. In particular, compositions and methods are disclosed that repair gene microduplication mutations by reversion to a wild type sequence. For example, the creation of a double stranded break by a programmable nuclease protein within a microduplication induces the microhomology mediated end joining DNA repair pathway that in the process of DNA repair removes the microduplication mutation and restores the wild type sequence.

Microhomology mediated repair of microduplication gene mutations

The present disclosure relates to methods of and systems for modifying the transcriptional regulation of stem or progenitor cells to promote their differentiation or reprogramming of somatic cells. Further, the labeling and editing of human genomic loci in live cells with three orthogonal CRISPR/Cas9 components allow multicolor detection of genomic loci with high spatial resolution, which provides an avenue for barcoding elements of the human genome in the living state.

Compositions and methods for differential Cas9 gene labeling and/or editing

Provided herein are methods and compositions for increasing fetal hemoglobin levels in a cell by disrupting BCL11A expression at the genomic level. Also provided herein are methods and compositions relating to the treatment of hemoglobinopathies by reinduction of fetal hemoglobin levels.

Targeting BCL11A distal regulatory elements with a CAS9-CAS9 fusion for fetal hemoglobin reinduction

The present invention provides a Cas9 platform to facilitate single-site nuclease gene editing precision within a human genome. For example, a Cas9 nuclease/DNA-targeting unit (Cas9-DTU) fusion protein precisely delivers a Cas9/sgRNA complex to a specific target site within the genome for subsequent sgRNA-dependent cleavage of an adjacent target sequence. Alternatively, attenuating Cas9 binding using mutations to the a protospacer adjacent motif (PAM) recognition domain makes Cas9 target site recognition dependent on the associated DTU, all while retaining Cas9's sgRNA-mediated DNA cleavage fidelity. Cas9-DTU fusion proteins have improved target site binding precision, greater nuclease activity, and a broader sequence targeting range than standard Cas9 systems. Existing Cas9 or sgRNA variants (e.g., truncated sgRNAs (tru-gRNAs), nickases and FokI fusions) are compatible with these improvements to further reduce off-target cleavage. A robust, broadly applicable strategy is disclosed to impart Cas9 genome-editing systems with the single-genomic-site accuracy needed for safe, effective clinical application.

CAS9-CAS9 fusion proteins

Cas9-DNA targeting unit chimeras

The present application relates to the hybrid selection methods in prokaryotes using counterselectable reporter genes.

Bacterial counterselectable hybrid systems

The invention is related to a fusion polypeptide involving transcription regulation of a target gene. Said fusion polypeptide consists of (i) a zinc finger domain that recognizes and binds to the corresponding nucleic acid recognition sequence in the vicinity of a promoter of the target gene; (ii) a dimerization domain of a tetracycline (tet) repressor which contains a drug (e.g., tetracycline)-dependent switch, and (iii) an accessory domain which alters the expression of the target gene (such as an transcriptional activation or repression domain). This fusion polypeptide can be administered to, or expressed in, a cell harboring the target gene to be regulated.

Zinc finger-based drug-dependent gene regulation system

Growing community of inventors

Winchester, MA, United States of America

Scot Andrew Wolfe