Allele-Selective Suppression of Mutant Huntingtin in Primary Human Blood Cells

• Published in: Scientific reports Vol. 7; no. 1; p. 46740
• Main Authors: Miller, James R. C., Pfister, Edith L., Liu, Wanzhao, Andre, Ralph, Träger, Ulrike, Kennington, Lori A., Lo, Kimberly, Dijkstra, Sipke, Macdonald, Douglas, Ostroff, Gary, Aronin, Neil, Tabrizi, Sarah J.
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 24.04.2017; Nature Publishing Group
• Subjects: 13/106; 13/109; 13/21; 38/77; 631/250/2504; 631/378/1689; 692/617/375/1558; 82/80; Adult; Aged; Alleles; Blood; Blood cells; Cells, Cultured; Clinical trials; Cohort Studies; Cytokines; Gene silencing; Genotype; Genotyping; Heterozygosity; Humanities and Social Sciences; Humans; Huntingtin; Huntingtin Protein - genetics; Huntington Disease - blood; Huntington Disease - genetics; Huntington's disease; Immune response; Lipopolysaccharides; Middle Aged; multidisciplinary; Mutant Proteins - genetics; Mutants; Myeloid cells; Myeloid Cells - metabolism; Patients; Phagocytosis; Polymorphism, Single Nucleotide; Post-transcription; Proteins; RNA Interference; Science; Single-nucleotide polymorphism; siRNA
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/srep46740

Post-transcriptional gene silencing is a promising therapy for the monogenic, autosomal dominant, Huntington’s disease (HD). However, wild-type huntingtin (HTT) has important cellular functions, so the ideal strategy would selectively lower mutant HTT while sparing wild-type. HD patients were genotyped for heterozygosity at three SNP sites, before phasing each SNP allele to wild-type or mutant HTT . Primary ex vivo myeloid cells were isolated from heterozygous patients and transfected with SNP-targeted siRNA, using glucan particles taken up by phagocytosis. Highly selective mRNA knockdown was achieved when targeting each allele of rs362331 in exon 50 of the HTT transcript; this selectivity was also present on protein studies. However, similar selectivity was not observed when targeting rs362273 or rs362307. Furthermore, HD myeloid cells are hyper-reactive compared to control. Allele-selective suppression of either wild-type or mutant HTT produced a significant, equivalent reduction in the cytokine response of HD myeloid cells to LPS, suggesting that wild-type HTT has a novel immune function. We demonstrate a sequential therapeutic process comprising genotyping and mutant HTT -linkage of SNPs, followed by personalised allele-selective suppression in a small patient cohort. We further show that allele-selectivity in ex vivo patient cells is highly SNP-dependent, with implications for clinical trial target selection.