The present disclosure encompasses engineered meganucleases that bind and cleave recognition sequences within a dystrophin gene. The present disclosure also encompasses methods of using such engineered meganucleases to make genetically modified cells. Further, the disclosure encompasses pharmaceutical compositions comprising engineered meganuclease proteins, or polynucleotides encoding engineered meganucleases of the disclosure, and the use of such compositions for the modification of a dystrophin gene in a subject, or for treatment of Duchenne Muscular Dystrophy.

The present disclosure provides, among other things, a recombinant adeno-associated viral (rAAV) vector encoding an agent that inhibits the proteolytic activity of plasma kallikrein. The disclosure also provides, a recombinant adeno-associated viral (rAAV) vector encoding an anti/plasma kallikrein antibody heavy drain and an anti-plasma kallikrein antibody light chain.

The present disclosure provides, among other things, a recombinant adeno-associated virus (rAAV) vector comprising an AAV8 capsid and a codon-optimized SERPING1 sequence encoding a human C1-esterase inhibitor. The disclosure also provides a method of treating a subject having Hereditary angioedema (HAE), comprising administering to the subject in need thereof a recombinant adeno-associated virus (rAAV) vector comprising an AAV8 capsid, and codon-optimized SERPING1 sequences encoding a human C1-esterase inhibitor.

The invention involves inducing 3-50 or more mutations (e.g., any whole number between 3 and 50 of mutations, with it noted that in some embodiments there can be up to 16 different RNA(s), e.g., sgRNAs each having its own a promoter, in a vector, such as AAV, and that when each sgRNA does not have its own promoter, there can be twice to thrice that amount of different RNA(s), e.g., sgRNAs, e.g., 32 or even 48 different guides delivered by one vector) in transgenic Cas9 eukaryotes to model genetic disease, e.g. cancer. The invention comprehends testing putative treatments with such models, e.g., testing putative chemical compounds that may be pharmaceutically relevant for treatment or gene therapy that may be relevant for treatment, or combinations thereof. The invention allows for the study of genetic diseases and putative treatments to better understand and alleviate a genetic disease or a condition, e.g., cancer.

Provided are compositions and methods for inducing expression of human beta-globin in erythrocytes for use in prophylaxis and/or therapy of a hemoglobinopathy in an individual. The method generally entails introducing into CD34+ cells a polynucleotide encoding: i) a 5′ long terminal repeat (LTR) and a self-inactivating 3′ LTR; ii) at least one polyadenylation signal; iii) at least one promoter; iv) a globin gene locus control region (LCR); v) an ankyrin insulator element (Ank); vi) a Woodchuck Post-Regulatory Element (WPRE) configured such that the WPRE does not integrate into a target genome; and vii) a sequence that is a reverse complement of a sequence encoding human beta-globin, and can include beta-globin that has a PT87Q mutation. Intron 2 of the beta globin gene can be a complete intron. Modified erythrocyte progenitor cells, recombinant vectors and virions comprising recombinant polynucleotides, and methods of making the vectors and virions are included.

In alternative embodiments, provided are compositions, including recombinant expression systems and vectors, products of manufacture and kits, and methods, for remotely-controlled and non-invasive manipulation of intracellular nucleic acid expression, genetic processes, function and activity in live cells such as T cells in vivo, for example, activating, adding functions or changing or adding specificities for immune cells, for monitoring physiologic processes, for the correction of pathological processes and for the control of therapeutic outcomes. In alternative embodiments, provided are blue-light-mediated light-inducible nuclear translocation and dimerization (LINTAD) systems for gene regulation to control cell activation based on the integration of light-sensitive LOV2-based nuclear localization, light-induced active transportation via the biLINuS motif, and CRY2-CIB1 dimerization that feature high spatiotemporal control to control or alter cell activities in vivo, for example, to limit CAR T cell activity to the tumor site for immunotherapy applications.

Methods are provided for treating osteoarthritis by administering αKlotho protein and sTGFβ-R2 protein to a site within a mammal exhibiting symptoms of osteoarthritis, such as a knee joint. The αKlotho protein and the sTGFβ-R2 protein are both present at the osteoarthritic site.

Disclosed herein include methods, compositions, and kits enabling expression of multiple proteins from a single mRNA with a predetermined stoichiometry. There are provided, in some embodiments, nucleic acid compositions comprising a promoter operably linked to a polynucleotide comprising a first nucleic acid unit encoding first unit payload protein(s) and a second nucleic acid unit encoding second unit payload protein(s). The first nucleic acid unit and the second nucleic acid unit can each comprise a 3′ engineered translation initiation site (eTIS) comprising a three-nucleotide tunable element immediately upstream of a start codon. The eTIS of each of the first nucleic acid unit and the second nucleic acid unit can be configured to achieve a predetermined stoichiometry of the first unit payload protein(s) and the second unit payload protein(s) in a cell or cell-like environment.

The present invention relates to a product comprising (i) an anti-VEGF entity; and (ii) a negative complement regulator, or nucleotide sequences encoding therefor, as a combined preparation for simultaneous, separate or sequential use in therapy. In particular, the anti-VEGF entity is an anti-VEGF antibody, preferably aflibercept and the negative complement regulator is Complement Factor I (CFI) or Complement Factor H Like Protein 1 (FHL1). The main uses are for the treatment of eye diseases, in particular age-related macular degeneration (AMD).

The present disclosure provides, in part, nucleic acid loaded flowable hydrogels and compositions, systems and methods related thereto, to effectively deliver nucleic acids to cells that contact the flowable hydrogels.