Epigenetic therapy of Prader-Willi syndrome

Abstract

Prader-Willi syndrome (PWS) is a complex and multisystem neurobehavioral disorder. The molecular mechanism of PWS is deficiency of paternally expressed gene gene or genes from the chromosome 15q11-q13. Due to imprinted gene regulation, the same genes in the maternal chromosome 15q11-q13 are structurally intact but transcriptionally repressed by an epigenetic mechanism. The unique molecular defect underlying PWS renders an exciting opportunity to explore epigenetic-based therapy to reactivate the expression of repressed PWS genes from the maternal chromosome. Inactivation of H3K9m3 methyltransferase SETDB1 and zinc finger protein ZNF274 results in reactivation of SNRPN and SNORD116 cluster from the maternal chromosomes in PWS patient iPSCs and iPSC-derived neurons, respectively. High content screening of small molecule libraries using cells derived from transgenic mice carrying the SNRPN-EGFP fusion protein has discovered that inhibitors of EHMT2/G9a, a histone 3 lysine 9 methyltransferase, are capable of reactivating expression of paternally expressed SNRPN and SNORD116 from the maternal chromosome, both in cultured PWS patient-derived fibroblasts and in a PWS mouse model. Treatment with an EMHT2/G9a inhibitor also rescues perinatal lethality and failure to thrive phenotypes in a PWS mouse model. These findings present the first evidence to support a proof-of-principle for epigenetic-based therapy for the PWS in humans.

Fig 1.

Molecular bases of PWS. Four types of molecular defects found in PWS are diagramed. Chromosome ideogram in pink color represents the maternal origin and in blue color for paternal chromosome. The white strips represent the large or small deletion in the chromosome 15q11-q13. (M, maternal; P, paternal; IC, imprinting center; UPD, uniparental disomy). Type I: Large 6 Mb deletion in the paternal chromosome 15q11-q13. Typ1 II: Maternal UPD of 15. Type III: Imprinting defect that with or without a microdeletion in the PWS—IC. The IC defect causes the maternal epigenotype in the 15q11-q13 region in the paternal chromosome measured by DNA methylation. Type IV: Microdeletion in the region of SNORD116s. For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig 2.

Human chromosome 15q11-q13 imprinting domain and homologous region in the mouse central chromosome 7C. The genes in blue color are expressed exclusively from the paternal chromosome. The gene in pink color is expressed exclusively from the maternal chromosome. The genes in black color are expressed from both maternal and paternal chromosomes. The imprinted expression pattern for PWRN1, NPAP1, and UBE3A is tissue or cell type specific (BP, breakpoint; IC, imprinting center). For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig 3.

Drug screening strategy to discover the small molecule drugs for the epigenetic therapy in the PWS and AS. A, UBE3A-YFP fusion protein transgenic mice. B, Drug screening strategy using cultured neurons derived from UBE3A-YFP fusion protein mice to screen small molecule drug that can reactivate the expression of Ube3a from the paternal chromosome. C, SNRPN-EGFP fusion protein transgenic mice. D, Drug screening strategy using mouse embryonic fibroblasts (MEFs) derived from the maternal SNRPN-EGFP mice to screen small molecule drug that can reactivate the expression of Snrpn from the maternal chromosome.

Fig 4.

A proof-of-principle of epigenetic therapy for PWS by G9a inhibitors. G9a inhibitor of UNC0638 and UNC0642 (block dots) directly reduce H3K9 methylation (red stars), but do not change methylation of PWS—IC (black hexagon). The reduction of H3K9 methylation is shown to be sufficient to activate the expression of PWS genes from the maternal chromosome, thereby offering therapeutic benefits. For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)