Researchers have implicated a member of the cellular death squad known as the “necrosome” in the pathogenesis of Gaucher’s disease. Receptor-interacting kinase 3 (RIPK3), a protein involved in a type of cell suicide called necroptosis, ramped up neuroinflammation in mice with the disease, according to a report in the January 19 Nature Medicine. Knocking out the kinase calmed Gaucher's symptoms. Led by Anthony Futerman at the Weizmann Institute of Science in Rehovot, Israel, the investigators propose RIPK3 as a therapeutic target for Gaucher’s. This lysosomal storage disorder is relevant to the understanding of Parkinson’s disease and dementia with Lewy bodies, as well.
“It’s exciting that this paper demonstrates a role for necroptosis in this disease,” said Junying Yuan at Harvard Medical School, who was not involved in the study. Yuan, who discovered the process and coined the term “necroptosis” (see Degterev et al., 2005), is developing inhibitors of the death cascade in hopes of treating a variety of diseases.
Lysosomal storage disorders are characterized by the buildup of lipids in lysosomes, the cell’s trash processors. People with Gaucher’s have mutations in the glucocerebrosidase (GBA) gene, which encodes a lysosomal enzyme that chops up lipids. Of the three known forms of the disease, only types 2 and 3 affect the central nervous system. In those cases, the accumulating lipids trigger the destruction of neurons, leading to loss of motor coordination, mental deterioration, and early death.
Scientists have linked the enzyme to other neurodegenerative diseases as well. Heterozygous mutations in GBA are one of the strongest genetic risk factors for Parkinson's and dementia with Lewy bodies (see April 2013 news story and PDGene).
One of the enduring unknowns in the Gaucher’s field is how the disease kills neurons, Futerman told Alzforum. To figure that out, first author Einat Vitner and colleagues examined mice that lacked GBA only in cells of neuronal lineage. In the brains of these nGD mice, the investigators found many dead neurons but, curiously, no markers of apoptosis. Instead, they found more RIPK1 and RIPK3 than the controls had. These two kinases form a complex that triggers necroptosis. Mice treated with the irreversible GBA inhibitor conduritol B epoxide also produced more of these two kinases in the brain, while immunofluorescence revealed expression of RIPK3 in neurons and activated microglia, but not in astrocytes. The findings suggest that neurons in Gaucher’s die through necroptosis rather than apoptosis, the authors concluded.
Interestingly, microglia in nGD mice—which do express functional GBA—contained more RIPK3 than wild-type microglia. While the authors are unsure why this is, they noted that RIPK3 can turn on inflammation as well as cell death. This raised the possibility that the kinase triggers neuronal death indirectly by way of activating microglia, as well as by setting off necroptosis within neurons.
Futerman and colleagues next explored RIPK3’s part in Gaucher’s disease pathogenesis by pharmacologically inducing the disease in RIPK3-knockout mice. Starting with eight-day-old mice, the researchers injected the animals daily with the GBA inhibitor. Ripk3+/- mice stopped gaining weight at 20 days of age, whereas their Ripk3-/- littermates continued to grow. By 31 days of age, Ripk3+/- mice fell off a rotating rod more readily than did Ripk3-/- mice. Ripk3+/- mice died prior to 40 days of age, whereas Ripk3-/- mice lived longer than 100 days. The work suggests that RIPK3 facilitates toxicity due to loss of GBA.
The work also hinted at a role for inflammation in Gaucher’s. When the GBA inhibitor-treated mice were 27 days old, the researchers detected fewer activated microglia in the brains of Ripk3 homozygous knockout mice compared to heterozygotes. In the latter, microglia became activated before there were detectable signs of neuronal death. This suggests that inflammation may trigger the demise of the neurons, though the researchers did not show this in the current study. They did find that mice lacking tumor necrosis factor, a known inflammatory cytokine and inducer of cell death, succumbed to GBA inhibition just as wild-type mice did. This suggests that other inflammatory triggers may be responsible for inducing the deadly cascade, Futerman said.
The precise chain of events that destroy neurons in GD is still unclear. Even so, Peter Vandenabeele at Ghent University in Belgium, who was not involved in the study, said that targeting RIPK3 could be a strategy in treating the disease. “I think RIPK3 kinase activity is an important hub in cell death in neuronal cells and in inflammation in microglia cells,” Vandenabeele wrote to Alzforum. “By targeting both processes, synergistic and devastating interaction between cell death and inflammation might be efficiently targeted.”
Krishnaraj Rajalingam at Goethe University in Frankfurt, Germany, called for a deeper understanding of the death complex. “Every day the dogma about how this pathway works is changing,” Rajalingam told Alzforum. “The more we understand about the contents of this complex, we will possibly have more avenues to explore to quench necroptosis in pathological conditions.”
Lifelong enzyme replacement therapy with Cerezyme, a recombinant form of glucocerebrosidase manufactured by the biotech company Genzyme in Cambridge, Massachusetts, is the only available treatment for people with Gaucher’s. The costly therapy poorly penetrates the brain, leaving people with the neurological forms of the disease undertreated.
That a RIPK3-targeting strategy could work as a stand-alone treatment for Gaucher’s is doubtful, Pablo Sardi of Genzyme told Alzforum; he was not involved in the study. “The question is whether this will affect the basis of the disease, which is the accumulation of lipids,” Sardi said. However, he noted that targeting RIPK3 could give Gaucher's patients a leg up when taken in combination with therapies that ameliorate the lipid storage. However, Sardi agreed that RIPK3 could have broader relevance, since GBA mutations have been linked to buildup of α-synuclein in people with Parkinson's and dementia with Lewy bodies (see June 2011 news story). “[Targeting RIPK3] might work better for PD, where the progression of the disease is slower,” Sardi said.
Whether necroptosis plays a role in neuronal cell death in Alzheimer’s disease remains to be seen. Both necrotic and apoptotic forms of death have been implicated in AD, along with malfunction in lysosomal processing (see Yamashima, 2013, and Nixon and Yang, 2012). Oddly, scientists recently found that RIPK3 itself forms amyloid fibrils in cultured cells, which initiate necroptotic signaling (see July 2012 news story).—Jessica Shugart
- Parkinson’s Gene Increases Risk of Dementia
- Feedback Loop—Molecular Mechanism for PD, Gaucher’s Connection
- Amyloid Fibrils as Signaling Complexes? Role in Cell Death Pathway
- Degterev A, Huang Z, Boyce M, Li Y, Jagtap P, Mizushima N, Cuny GD, Mitchison TJ, Moskowitz MA, Yuan J. Chemical inhibitor of nonapoptotic cell death with therapeutic potential for ischemic brain injury. Nat Chem Biol. 2005 Jul;1(2):112-9. PubMed.
- Yamashima T. Reconsider Alzheimer's disease by the 'calpain-cathepsin hypothesis'-A perspective review. Prog Neurobiol. 2013 Jun;105:1-23. PubMed.
- Nixon RA, Yang DS. Autophagy and neuronal cell death in neurological disorders. Cold Spring Harb Perspect Biol. 2012;4(10) PubMed.
- Shachar T, Lo Bianco C, Recchia A, Wiessner C, Raas-Rothschild A, Futerman AH. Lysosomal storage disorders and Parkinson's disease: Gaucher disease and beyond. Mov Disord. 2011 Aug 1;26(9):1593-604. Epub 2011 May 26 PubMed.
- Yang NY, Lee YN, Lee HJ, Kim YS, Lee SJ. Glucocerebrosidase, a new player changing the old rules in Lewy body diseases. Biol Chem. 2013 Feb 22; PubMed.
- Ofengeim D, Yuan J. Regulation of RIP1 kinase signalling at the crossroads of inflammation and cell death. Nat Rev Mol Cell Biol. 2013 Nov;14(11) Epub 2013 Oct 16 PubMed.
- Vitner EB, Salomon R, Farfel-Becker T, Meshcheriakova A, Ali M, Klein AD, Platt FM, Cox TM, Futerman AH. RIPK3 as a potential therapeutic target for Gaucher's disease. Nat Med. 2014 Feb;20(2) Epub 2014 Jan 19 PubMed.