Usage of an Alternative Transcription Start Site in PRKN mutation carriers: Key to Later-Onset Parkinson's Disease and a Novel Therapeutic Target

Biallelic pathogenic variants of PRKN, encoding the Parkin RBR E3 ubiquitin protein ligase, are the most common known cause of autosomal recessive Parkinsons disease (PD). PARK-PRKN is characterized by an early median age at onset (AAO) of 31 years with a wide range (3-81 years). When evaluating the 16 previously published carriers of a homozygous deletion of Exon 2 (PRKNdelEx2) from the MDSGene database, the median AAO is later (39.5 years; range: 25-75 years) than in carriers of other PRKN pathogenic variants. Here, we investigated 26 homozygous PRKNdelEx2 patients, including 20 from additional sources, and confirmed the later median AAO (37 years, range: 13-60 years). Furthermore, we investigated one carrier who was still unaffected in her 60s. To elucidate the functional basis for this observation, we used induced pluripotent stem cell (iPSC)-derived dopaminergic neurons (iDN) from this unaffected carrier as well as genome-edited iDNs from two control lines and neuroblastoma cell lines (SH-SY5Y) with the introduction of a homozygous PRKNdelEx2. We observed elevated levels of an N-terminally truncated form of Parkin (Parkin{Delta}1-79), starting at an internal translation initiation site (TIS) in Exon 3 (p.Met80 in the full-length protein) in cells with the Exon 2 deletion. Furthermore, in silico prediction and Parkin quantification in isogenic neuroblastoma cell lines suggested the presence of residual Parkin{Delta}1-79 also for other PRKN variants upstream of the alternative TIS. iDNs from PRKNdelEx2 carriers partially retained Parkin E3 ubiquitin ligase activity in contrast to carriers of other exonic deletions in PRKN (PRKNdelEx3, PRKNdelEx7) via the expression of Parkin{Delta}1-79. Importantly, endogenous Parkin E3 ubiquitin ligase activity of Parkin{Delta}1-79 was enhanced in neurons derived from a homozygous PRKNdelEx2 variant carrier upon treatment with a small molecule allosteric modulator of Parkin (BIO-2007817). In summary, the later AAO in patients with homozygous PRKNdelEx2 is associated with partially retained Parkin function. The residual ligase activity can be further increased pharmacologically, providing mutation-specific personalized counseling opportunities and a potential novel therapy for selected patients with PARK-PRKN.