Multimodal spatial transcriptomics determines repeat expansion, huntingtin aggregation, and selective cortical neuron loss in Huntington’s Disease

ABSTRACT Huntington’s disease (HD) is caused by CAG expansion in HTT , yet how somatic repeat instability and huntingtin aggregation relate to selective cell loss in the human brain remains unclear. We have developed a multimodal spatial transcriptomics approach that enables defining transcriptional programs with subcellular resolution, somatic CAG repeat lengths, and six other pathology marks including huntingtin aggregates in every cell of intact brain sections. Imaging 428,173 cells in HD cortex revealed selective vulnerability: L5–6 NP and L6b deep-layer excitatory neurons undergo >50% loss, closely linked to very large (>380±55) somatic expansions. Intranuclear aggregation was most prevalent at intermediate somatic repeat expansion (220-300 CAGs) and was accompanied by broader transcriptional changes. In contrast, chandelier and somatostatin+ inhibitory interneurons are lost despite only modest repeat expansion or aggregation. These data provide a comprehensive resource and establish a broadly applicable framework for connecting repeat expansion and protein pathology across diverse cell types. Short Bullet points Development of a novel, multimodal spatial transcriptomics platform enables definition of RNA transcriptomes with subcellular resolution, somatic repeat expansions, and protein accumulation in cells within tissue sections Generation of two complementary datasets for HD and control cortex: 428,173 cells to quantify comprehensively cell-type vulnerability, and a deeper multimodal dataset in which CAG repeat expansion, huntingtin aggregation, six additional pathological readouts, and expression of 1,128 genes were measured in 185,721 cells, providing a comprehensive resource for the neurodegeneration community. Deep layer excitatory neuron loss (L5–6 NP and L6b) was associated with very large somatic CAG expansions (>380±55), while selective inhibitory neuron loss (chandelier and somatostatin interneurons) occurred with modest CAG repeat expansion or huntingtin intranuclear aggregation Intranuclear aggregation, not somatic repeat expansion, was more predictive of transcriptional changes, including chromatin remodeling and RNA export factors