We addressed the therapeutic shortcomings of human arginase-I (hArgI) using bioinorganic and protein engineering strategies, resulting in an enzyme with a 10-fold higher catalytic efficiency for L-Arg hydrolysis, 12-15 fold reduction in the IC50 towards a variety of malignant cell lines and, importantly a 5 fold improvement of half-life in serum. PEGylation resulted in a circulatory t1/2 in balb/c mice of 63 ± 10 hrs with a 9-fold increase in pharmacodynamic efficacy relative to similarly prepared native hArgI and effectively controlled HCC, metastatic melanoma, and SCLC tumors in mouse xenograft models.
Using public funding from State of Texas and NIH we are carrying out late stage preclinical development of engineered hArgI-5 Kda PEG in a manner that satisfies industry standards. A high cell density E.coli fermentation process yielding >3.8 g/L culture of hArgI was developed; a four column purification process and PEGylation were used to produce preclinical enzyme at >98% purity with very low endotoxin level (<0.1 EU/mg). GMP production and GLP toxicology studies are under way with a physician-sponsored IND expected to be filed in Jan 2012 followed by a Phase I/IIa clinical trial of metastatic melanoma patients.
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