0301: Novel HMGCS1 Deficiency Expands Proinflammatory Monocytes and Cytotoxic Populations with Increased Cytokine Release

Background/Purpose: The mevalonate (MVK) pathway synthesizes isoprenoids that mediate post-translational protein modifications via prenylation. Deficient protein prenylation due to decreased mevalonate kinase (MK) activity leads to innate immune activation and autoinflammatory disorders. Patients using statins, HMG-CoA reductase inhibitors, may present with myopathy. However, the precise pathogenesis of prenylation deficiency-associated inflammation and myotoxicity remains unclear. We identified a rare homozygous pathogenic variant in the HMGCS1 gene (c.265C >T, p. Arg89Trp), encoding for HMG-CoA synthase, the first enzyme in the MVK pathway, in two siblings presenting with necrotizing myopathy, arthropathy/arthritis, and susceptibility to infections.

Methods: Single-cell RNA sequencing (scRNA-seq) was conducted on PBMCs obtained from affected siblings, an unaffected sibling (wild-type), the mother (heterozygous) and unrelated healthy control. Muscle biopsies from affected siblings were used for bulk RNA sequencing. Using spectral flow cytometry, whole blood and/or PBMCs from affected individuals were assessed for IFN-g, IL-1b and IL-6 expression in monocytes, while IFN-g, granzyme B and perforin were assessed on NK and T cells.

Results: ScRNA-seq data evidenced an expansion of activated monocyte and cytotoxic populations in the affected but not in the unaffected brother, suggesting monocytes, T and NK cells as drivers of inflammation. Since muscle biopsies of the affected brothers revealed a strong IFN-g signature and IL-1 and IL-6 are crucial for inflammasome formation, we investigated these cytokines and inflammatory markers in LPS-stimulated myeloid subsets, and IL-12-stimulated NK cells and T cells of affected and unaffected controls. Percentage of IL-1b⁺ and IL-6⁺ non-classical monocytes (NCM) were significantly elevated in the affected siblings. IFN-g levels were increased in all monocyte subpopulations of the affected siblings, consistent with the IFN-g signature observed in the muscle biopsies. Monocytes expressing inflammatory markers, such as CD64, Siglec-1, and CD11c, were increased with a marked expression in NCM. Classical monocytes (CM), intermediate monocytes (IM), and NCM of the affected siblings presented lower PSGL-1 levels, aligned with the observed inflammatory responses, suggesting an infiltration of activated monocytes into muscle tissues. We observed a decrease in NK cells in affected donors, while T cells, specifically Th1 and Th17 cells, were expanded. Further, T cells presented elevated IFN-g expression in affected siblings, correlating with T cells infiltration seen in muscle tissue. Notably, T cells of the affected donors showed higher expression of granzyme B and perforin but decreased CD107a, indicating these cells were activated but with diminished degranulation activity.

Conclusion: Our data provide strong evidence that monocytes and T cells may play a pivotal role in the pathogenesis of inflammation and necrotizing myositis in patients with HMGCS1 deficiency, with NCM and cytotoxic T cells presenting the most proinflammatory profile suggesting an important role of these cells in sustaining HMGCS1 mutation-associated autoinflammatory disorder.