Screening chimeric GAA variants in preclinical study results in hematopoietic stem cell gene therapy candidate vectors for Pompe disease.

Pompe disease is a rare genetic neuromuscular disorder caused by acid α-glucosidase (GAA) deficiency resulting in lysosomal glycogen accumulation and progressive myopathy. Enzyme replacement therapy, the current standard of care, penetrates poorly into the skeletal muscles and the peripheral and central nervous system (CNS), risks recombinant enzyme immunogenicity, and requires high doses and frequent infusions. Lentiviral vector-mediated hematopoietic stem and progenitor cell (HSPC) gene therapy was investigated in a Pompe mouse model using a clinically relevant promoter driving nine engineered GAA coding sequences incorporating distinct peptide tags and codon optimizations.

Gene Therapy Developments for Pompe Disease.

Pompe disease is an inherited neuromuscular disorder caused by deficiency of the lysosomal enzyme acid alpha-glucosidase (GAA). The most severe form is infantile-onset Pompe disease, presenting shortly after birth with symptoms of cardiomyopathy, respiratory failure and skeletal muscle weakness. Late-onset Pompe disease is characterized by a slower disease progression, primarily affecting skeletal muscles.

Inducible histone K-to-M mutations are dynamic tools to probe the physiological role of site-specific histone methylation in vitro and in vivo.

Development and differentiation are associated with profound changes to histone modifications, yet their in vivo function remains incompletely understood. Here, we generated mouse models expressing inducible histone H3 lysine-to-methionine (K-to-M) mutants, which globally inhibit methylation at specific sites. Mice expressing H3K36M developed severe anaemia with arrested erythropoiesis, a marked haematopoietic stem cell defect, and rapid lethality.

The Hippo pathway target, YAP, promotes metastasis through its TEAD-interaction domain.

The transcriptional coactivator Yes-associated protein (YAP) is a major regulator of organ size and proliferation in vertebrates. As such, YAP can act as an oncogene in several tissue types if its activity is increased aberrantly. Although no activating mutations in the yap1 gene have been identified in human cancer, yap1 is located on the 11q22 amplicon, which is amplified in several human tumors.

TEL-AML1 corrupts hematopoietic stem cells to persist in the bone marrow and initiate leukemia.

The initial steps in the pathogenesis of acute leukemia remain incompletely understood. The TEL-AML1 gene fusion, the hallmark translocation in Childhood Acute Lymphoblastic Leukemia and the first hit, occurs years before the clinical disease, most often in utero. We have generated mice in which TEL-AML1 expression is driven from the endogenous promoter and can be targeted to specific populations.

Analysis of histone 2B-GFP retention reveals slowly cycling hematopoietic stem cells.

Hematopoietic stem cells (HSCs) are thought to divide infrequently based on their resistance to cytotoxic injury targeted at rapidly cycling cells and have been presumed to retain labels such as the thymidine analog 5-bromodeoxyuridine (BrdU). However, BrdU retention is neither a sensitive nor specific marker for HSCs. Here we show that transient, transgenic expression of a histone 2B (H2B)-green fluorescent protein (GFP) fusion protein in mice has several advantages for label-retention studies over BrdU, including rapid induction of H2B-GFP in virtually all HSCs, higher labeling intensity and the ability to prospectively study label-retaining cells, which together permit a more precise analysis of division history.

Gfi-1 restricts proliferation and preserves functional integrity of haematopoietic stem cells.

Haematopoietic stem cells (HSCs) sustain blood production throughout life. HSCs are capable of extensive proliferative expansion, as a single HSC may reconstitute lethally irradiated hosts. In steady-state, HSCs remain largely quiescent and self-renew at a constant low rate, forestalling their exhaustion during adult life.

Tel/Etv6 is an essential and selective regulator of adult hematopoietic stem cell survival.

Hematopoietic stem cells (HSCs) sustain blood formation throughout life. Pathways regulating maintenance of adult HSCs are largely unknown. Here we report that the Ets-related transcription factor Tel/Etv6, the product of a locus frequently involved in translocations in leukemia, is a selective regulator of HSC survival.