Nicole D. Vincelette

Myelodysplastic syndrome (MDS) is characterized by S100A9-mediated constitutive innate immune activation that leads to the inflammatory death of hematopoietic stem/progenitor cells (HSPCs). This suppressive microenvironment includes the secretion of TGFβ and is phenocopied by the S100A9-transgenic (Tg) mice. However, the role of the different ALK/TGFB receptors in hematopoiesis or in S100A9-mediated inflammation is not understood. Primary low-risk MDS bone marrow mononuclear cells (BM-MNCs) were analyzed in vitro for hematopoietic restoration by an ALK4/ALK5 inhibitor (Vactosertib), an ALK2 inhibitor (TP-0184/Itacnosertib) and an ALK5/TGFBR1-specific inhibitor (TP-6379). Specimens significantly responded to TP-6379 treatment restoring hematopoiesis, with a positive bias for specimens harboring spliceosomal mutations. We previously shown that S100A9 drives phenotypic pathology of MDS, and it has been known to induce and cooperate with TGFβ in this disease. Hence, we treated healthy BM-MNC with recombinant human (rh) S100A9 where TP-6379 rescued the S100A9-reduced hematopoiesis. The effect of TP-6379 was validated in vivo in S100A9-Tg mice. Similar results were obtained with K562 cells, with or without the knock-in (KI) mutation in SF3B1, with strong sensitivity to rhS100A9 that was overcome by TP-6379. Hence, directly targeting TGFBR1/ALK5 with TP-6379 restores healthy hematopoiesis in low-risk MDS preclinical models.

Mosaic chromosomal alterations (mCAs) are a prevalent but poorly understood form of clonal hematopoiesis (CH). Whether mCAs contribute to disease independently of CHIP, and whether their large-scale genomic effects can be resolved to actionable targets, remain unknown. In 452,594 UK Biobank participants, we show that mCAs confer multimorbidity and mortality risk independent of CHIP. Notably, mCA-CHIP co-occurrence defines a very high-risk clonal state with synergistically elevated mortality, identifying a population not captured by CHIP screening alone. To resolve large mCAs to specific disease mechanisms, a cytoband-level mapping framework was developed that links mCAs to discrete genomic loci and candidate effector genes. Functional validation using single-cell transcriptomics and mouse models prioritized MYC (chr8 gain) and S100A9 (chr1 gain) as key drivers of systemic inflammation and multiorgan pathology. These findings establish mCAs as independent, synergistic, and genetically-resolvable drivers of age-related disease, with immediate implications for screening, risk stratification, and therapeutic development. SIGNIFICANCE: Hematopoietic mCAs drive age-related multimorbidity and mortality independently of CHIP, while co-occurrence defines a synergistically high-risk clonal state undetectable by standard screening. Integrative cytoband-level mapping and functional validation resolve large chromosomal alterations to discrete effector genes, enabling mechanistic risk stratification and informing precision surveillance and targeted therapeutic strategies.

Figure S5. Changes in signaling in BM of trisomy 8+ TN-MF patient and MYC MF mice. Figure S6. Effects of silencing S100a9 in MYC-driven MF or overexpression of S100a9. Figure S7. Effects of inhibition of S100a9 or MYC in MYC-driven MF. Figure S8. Synthesis and Characterization of Tasquinimod. Figure S9. Synthesis and Characterization of MYCi975.

Supplementary Table 7. Targeted Next Generation Sequencing (NGS) panels used for the Moffitt Cancer Center, Nice University Hospital, and University of Maryland Cohorts.

Supplementary Table 2. Predictors of complete response with (CR) or without (CRi) count recovery.

Supplementary Figure 10. Nested Bootstrap Analysis.

Supplementary Figure 1. Study Flow and Model Development Pipeline.

Supplementary Figure 12. Comparison of Risk Group Classifications Between the Moffitt Model vs. Other Prognostic Models.

Supplementary Table 3. Predictors of Overall Survival.

Comparison of gene expression profiles of Mx1-Cre+/-;Rosa26LSL-MYC/LSL-MYC vs. Mx1-Cre+/-;Rosa26+/+ mouse.