Glyceraldehyde-3-phosphate dehydrogenase as a target for small-molecule disease-modifying therapies in human neurodegenerative disorders

Abstract

Recent articles have highlighted numerous additional functions of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) that are independent of its well-documented glycolytic function. One of the most intriguing of these functions is as an initiator of programmed cell death cascades. This activity involves a nuclear appearance of GAPDH, a considerable proportion of which requires synthesis of new GAPDH protein and has characteristics suggesting the involvement of a novel isozyme. The relevance of such findings to human neurodegenerative conditions is emphasized by the increased nuclear GAPDH observed in postmortem samples from patients with Parkinson's disease, Alzheimer's disease, Huntington's disease and glaucoma, among others. A number of small-molecule compounds have now been identified that show anti-apoptotic activity because of their ability to interact with GAPDH and prevent its nuclear accumulation. These compounds, one of which is currently being tested in late-stage Phase II clinical trials as a disease-modifying therapy for Parkinson's disease, have potential utility in the treatment of human neurodegenerative conditions.

Fig. 1: Appearance of acidic isoforms of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) after treatment with 6-hydroxydopamine (6-OHDA). Human neuroblastoma (SH-SY5Y) cells were induced to undergo programmed cell death (PCD) by treatment with 6-OHDA. Cells were subcellular fractionated and nuclear extracts subjected to 2-dimensional gel electrophoresis. Control cultures were treated identically with the exception of vehicle treatment in place of 6-OHDA. The cultures treated with 6-OHDA show a pronounced appearance of acidic GAPDH bands in the nuclear fractions. Similar bands are absent from the control culture nuclear fractions.

Fig. 2: Changes in nuclear GAPDH after treatment with 6-OHDA. SH-SY5Y cells were induced to undergo PCD by the addition of 6-OHDA. Cells were subcellular fractionated and GAPDH in the nuclear fractions examined. Upper panel: 6-OHDA treatment was associated with a pronounced increase in nuclear GAPDH levels. Lower panel: under control conditions, most (> 80%) of the low levels of nuclear GAPDH present were readily extracted as a soluble fraction after centrifugation. Less than 5% of nuclear GAPDH was inextractable by RNase/DNase/2M salt treatment under control conditions. After treatment with 6-OHDA, only 35% of the increased nuclear GAPDH was extractable as a soluble fraction by centrifugation. In excess of 55% of nuclear GAPDH was resistant to extraction by RNase/DNase/2M salt after 6-OHDA treatment. SEM = standard error of the mean, S420 = the soluble fraction from a 420-mmol/L salt extraction, SRNase = the soluble fraction following RNase treatment, SDNase = the soluble fraction following DNase treatment, S2M = the soluble fraction following 2-mol/L salt extraction, P2M = the insoluble fraction following 2-mol/L salt extraction.

Fig. 3: A schematic representation of the GAPDH- mediated PCD cascade. A minor fraction of GAPDH is initially released from cytoplasmic binding sites. This minor fraction translocates to the nucleus and initiates synthesis of a novel nuclear-targeted GAPDH isoform. Transcription of this novel, nuclear GAPDH could also conceivably be induced by other factors. The novel, nuclear GAPDH then initiates PCD cascades through regulation of transcription, leading to loss of mitochondrial membrane potential and cell death. Small-molecule anti-apoptotic compounds have been identified that selectively bind to a fraction of GAPDH, preventing the increase in nuclear GAPDH and downstream events. Tetrahydroaminoacridine (THA) and donepezil have been reported to act at the level of new GAPDH transcription by binding to promoter regions. Aliphatic compounds, which like THA and donepezil have been shown to prevent increases in GAPDH mRNA, could also act at the level of transcription, although this has yet to be studied. Binding of small molecules to the newly synthesized GAPDH, thereby preventing its expression in the nucleus, is also possible in addition to binding to GAPDH released from cytoplasmic binding sites. The question marks above arrows indicate possible sites of action that have yet to be studied.