CGT and ACD Inhibitors for SRT Treatment of Krabbe Disease

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SUMMARY Krabbe disease (KD) is caused by the deficiency of the ubiquitously expressed lysosomal enzyme galactosylceramidase (GALC) which is responsible for the degradation of galactosylceramides and galactosylsphingosine (psychosine). Because the synthetic pathway conducing to psychosine is not affected in KD, psychosine is continuously produced and accumulated in the Krabbe nervous system. Toxic levels of psychosine are considered the main pathogenic trigger of disease. Currently, the standard of care for KD is hematopoietic stem cell transplantation (HSCT), which is only applicable to asymptomatic or early symptomatic infantile KD cases and only protracts disease. Pre-clinical gene therapy studies using adeno-associated viral (AAV) vectors have shown great promise and in fact, AAV gene therapy applied early in life increases survival, improves quality of life, and decreases neuropathology in twitcher (twi) mice, the natural model for KD. Based on these important successes, AAV-based gene therapy clinical trials are being started only for infantile KD. However, despite the prevention of significant disease-related deficits, HSCT and pre-clinical AAV-gene therapy trials show varied long-term efficacy and resurgence of neurological disease. Thus, the status of gene therapy for KD, the limitations of HSCT to treat primarily presymptomatic infantile KD and the fact that juvenile and adult onset KD patients, which encompass a significant fraction of Krabbe patients, largely remain without any treatment, highlight the need to develop additional strategies to sustain long-term protection for KD patients. The use of substrate reduction therapies (SRT) strategies, singly or combined with current and new therapies for KD, is one potential way to achieve this. In this application we will use two small new compounds which selectively inhibit ceramide galactosyltransferase (CGT) and acid ceramidase (ACD), enzymes that mediate the production of psychosine via galactosylation of ceramides and sphingosine (CGT) and deacylation of galactosylceramide (ACD). Based on the premise that reducing psychosine synthesis will prevent/reduce psychosine-related pathology at early postnatal development of the mammalian brain, we will test the efficacy of SRT of CGT and ACD to enhance HSCT and AAV-GALC gene therapy in the mouse model of infantile KD (twitcher mouse) and the efficacy of single treatment with CGT or ACD inhibitors to ameliorate/prevent disease in a new model of adult-onset KD.