Evaluation of an AAV9-mini-dystrophin gene therapy candidate in a rat model of Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is an X-linked disease caused by loss-of-function mutations in the dystrophin gene and is characterized by muscle wasting and early mortality. Adeno-associated virus-mediated gene therapy is being investigated as a treatment for DMD. In the nonclinical study documented here, we determined the effective dose of fordadistrogene movaparvovec, a clinical candidate adeno-associated virus serotype 9 vector carrying a human mini-dystrophin transgene, after single intravenous injection in a dystrophin-deficient (DMDmdx) rat model of DMD. Overall, we found that transduction efficiency, number of muscle fibers expressing the human mini-dystrophin polypeptide, improvement of the skeletal and cardiac muscle tissue architecture, correction of muscle strength and fatigability, and improvement of diastolic and systolic cardiac function were directly correlated with the amount of vector administered. The effective dose was then tested in older DMDmdx rats with a more dystrophic phenotype similar to the pathology observed in older patients with DMD. Except for a less complete rescue of muscle function in the oldest cohort, fordadistrogene movaparvovec was also found to be therapeutically effective in older DMDmdx rats, suggesting that this product may be appropriate for evaluation in patients with DMD at all stages of disease.

Protein Phosphatase Inhibitor-1 Gene Therapy in a Swine Model of Nonischemic Heart Failure.

BACKGROUND: Increased protein phosphatase-1 in heart failure (HF) induces molecular changes deleterious to the cardiac cell. Inhibiting protein phosphatase-1 through the overexpression of a constitutively active inhibitor-1 (I-1c) has been shown to reverse cardiac dysfunction in a model of ischemic HF. OBJECTIVES: This study sought to determine the therapeutic efficacy of a re-engineered adenoassociated viral vector carrying I-1c (BNP116.I-1c) in a preclinical model of nonischemic HF, and to assess thoroughly the safety of BNP116.I-1c gene therapy. METHODS: Volume-overload HF was created in Yorkshire swine by inducing severe mitral regurgitation. One month after mitral regurgitation induction, pigs were randomized to intracoronary delivery of either BNP116.I-1c (n = 6) or saline (n = 7). Therapeutic efficacy and safety were evaluated 2 months after gene delivery. Additionally, 24 naive pigs received different doses of BNP116.I-1c for safety evaluation. RESULTS: At 1 month after mitral regurgitation induction, pigs developed HF as evidenced by increased left ventricular end-diastolic pressure and left ventricular volume indexes. Treatment with BNP116.I-1c resulted in improved left ventricular ejection fraction (-5.9 ± 4.2% vs. 5.5 ± 4.0%; p < 0.001) and adjusted dP/dt maximum (-3.39 ± 2.44 s-1 vs. 1.30 ± 2.39 s-1; p = 0.007). Moreover, BNP116.I-1c-treated pigs also exhibited a significant increase in left atrial ejection fraction at 2 months after gene delivery (-4.3 ± 3.1% vs. 7.5 ± 3.1%; p = 0.02). In vitro I-1c gene transfer in isolated left atrial myocytes from both pigs and rats increased calcium transient amplitude, consistent with its positive impact on left atrial contraction. We found no evidence of adverse electrical remodeling, arrhythmogenicity, activation of a cellular immune response, or off-target organ damage by BNP116.I-1c gene therapy in pigs. CONCLUSIONS: Intracoronary delivery of BNP116.I-1c was safe and improved contractility of the left ventricle and atrium in a large animal model of nonischemic HF.

Detection of porcine endogenous retrovirus in cultures of freshly isolated porcine bone marrow cells.

Pigs are under consideration as possible sources of organs for xenotransplantation in humans. The induction of hematopoietic microchimerism through xenotransplantation of source animal hematopoietic cells has been suggested as a means to induce tolerance in potential recipients. Because all porcine cells contain genetic information for porcine endogenous retrovirus (PERV), coculture techniques, reverse transcriptase (RT) and reverse transcriptase-polymerase chain reaction assays were used to determine whether infectious PERV is released from fresh porcine bone marrow cells cultured in the presence or absence of porcine cytokines. Human embryonic kidney cell line, HEK-293 cells cocultured with porcine bone marrow cells were positive for PERV RNA but never became positive for viral RT activity, suggesting the PERV infection was not productive. In contrast, high levels of RT activity was detected in porcine ST-IOWA cells after coculture, demonstrating that these cells became productively infected. PERV was released from cultured porcine bone marrow cells without stimulation, and combinations of the porcine hematopoietic cytokines, interleukin-3, granulocyte macrophage-colony stimulating factor and stem cell factor had no additional effect on the infectivity or in vitro tropism of released PERV virions.