Figure S2 shows clinical biomarker changes including body composition, healthy eating index, paraprotein, LDL and IGF1.

Figure S4 shows change in peripheral blood CRP and inflammatory biomarkers, bone marrow cell proportions and differentially expressed genes in paired samples by BMI and microbiome diversity as well as cell interactions.

TET2 is a commonly mutated gene in hematologic malignancies, including as an initiating event in clonal hematopoiesis (CH). Its mutation alters hematopoietic self-renewal, differentiation, and systemic inflammation responses. TP53 mutations co-occur with TET2 mutations and are also observed in patients with high-risk clonal hematopoiesis and hematologic malignancies. Using a murine model, we found that HSPCs with both mutations initially promoted a myeloproliferative phenotype. Over time these double mutant HSPCs acquire additional genomic alternations, leading to disease progression to acute leukemias including B-ALL. We observed enhanced inflammatory signatures at transformation and identified NLRP1 as a target of TP53 activation. Decreased response to an inflammatory cell death pathway in the setting of TP53 mutation allows cells to tolerate inflammatory stress. This pathway also modifies response to chemotherapies that induce protein translational stalling. Our results identify a hematopoietic stem cell stress response pathway with implications on adaptation to inflammation and chemotherapy tolerance. Significance:TET2 and TP53 mutations co-operate leading to advanced hematologic malignancy. TET2 mutations promote an inflammatory environment and TP53 mutation supports tolerance to this inflammatory stress.

Figure S6 shows the effect of high-fiber diet on the gut microbiota composition and immune response in Vk*MYC mice.

Consumption of a Western diet and high body mass index (BMI) are risk factors for progression from premalignant phenotypes to multiple myeloma, a hematologic cancer. In the NUTRIVENTION trial (NCT04920084), we administered a high-fiber, plant-based diet (HFPBD; meals for 12 weeks, coaching for 24 weeks) to 23 participants with myeloma precursor states and elevated BMI. The intervention was feasible and improved quality of life and modifiable risk factors: metabolic (BMI, insulin resistance), microbiome (diversity, composition), and immune (inflammation, monocyte subsets). Disease progression trajectory improved (n = 2) or was stable. Findings were translated to Vk*MYC mice modeling the myeloma precursor state, in which a high-fiber diet (HFD) delayed disease progression through improved metabolism and microbiome composition, leading to increased short-chain fatty acid production that reinvigorated antitumor immunity and inhibited tumor growth. These effects from fiber consumption were independent of calorie restriction and weight loss. A HFD is a low-risk intervention that may delay progression to myeloma. SIGNIFICANCE:A HFPBD in participants with precursor plasma cell disorders and an elevated BMI improved metabolic, microbiome, and immune biomarkers of disease. In a subset, it may delay progression to myeloma. In mouse models, a HFD delayed progression to myeloma independent of calorie restriction. See related commentary by Fairfield and Reagan, p. 623.

Figure S8 reports flow cytometry gating strategy related to fig. 4, 5, S6.

Abstract Background: Mucinous breast cancer (MucBC) is an uncommon histologic subtype of estrogen receptor (ER)-positive/HER2-negative breast cancer (BC) characterized by tumor cells floating in pools of mucin. In contrast to ER-positive/HER2-negative invasive ductal carcinoma of no special type (IDC-NST), MucBCs harbor fewer PIK3CA mutations and typically lack concurrent 1q gains/16q losses. Yet, no pathognomonic genetic alterations have been identified to explain their mucinous phenotype. Here, we sought to determine whether MucBCs might be driven by distinctive epigenetic alterations. Methods: We analyzed 40 pure MucBCs using genome-wide DNA methylation profiling (n=40) and/or RNA-sequencing (n=27). We evaluated DNA methylation age, epigenetic mitotic score, and integrated methylation-transcriptomic profiles, comparing MucBCs with IDC-NSTs from The Cancer Genome Atlas (TCGA) lacking PIK3CA mutations and concurrent 1q gains/16q losses to minimize genetic confounding factors. IDC-NSTs were matched at a 1:1 ratio by menopausal status, ER/HER2 status and histologic grade. The ER-positive/HER2-negative CAMA1, ZR-75-1 and MCF7 breast cancer cell lines were used to interrogate candidate pathways and transcriptomic changes. Results: Compared to matched IDC-NSTs, MucBCs exhibited accelerated DNA methylation age relative to patient chronological age (p=0.015), indicative of epigenetic dysregulation, lower epigenetic mitotic scores (p=0.0071), and pronounced global enhancer hypomethylation (p< 0.001). Integrated methylation-transcriptome analyses identified ZBTB20 and ZNF133 as key transcriptional regulators in MucBCs (FDR < 0.05), as well as significant dysregulation of the TGF-β and estrogen-response pathways (FDR<0.05). In CAMA1 and ZR-75-1 BC cells, pharmacologic inhibition of the TGF-β pathway led to increased expression of mucin-encoding genes (MUC2, MUC20, MUCL1, MUC5B), while silencing of ZBTB20 or ZNF133 resulted in reduced mucin gene expression. In MCF7 cells, overexpression of ZBTB20 or ZNF133 similarly resulted in upregulation of mucin encoding genes and induced transcriptional changes in the TGF-β pathway, including downregulation of TGFBR1, TGFB1, SMAD3 and TRIM33 (TIF1γ) and upregulation of ID1, mirroring patterns observed in MucBCs. Conclusions: MucBC display accelerated DNA methylation age and widespread enhancer hypomethylation, underscoring profound epigenomic dysregulation. Key transcription factors, including ZBTB20 and ZNF133, appear to orchestrate mucin gene expression as well as TGF-β pathway reprogramming, contributing to the mucinous phenotype of MucBC. Citation Format: Lorenzo Ferrando, Higinio Dopeso, Laxmi Gusain, Edaise M da Silva, Thais Basili de Oliveira, Lounes Djerroudi, Hannah Y. Wen, Hong Zhang, Edi Brogi, Richard P. Koche, Pierre-Jacques Hamard, Larry Norton, Jorge S. Reis-Filho, Britta Weigelt, Fresia Pareja. Epigenetic and transcriptional drivers of mucinous breast carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 5296.

Table S1, Plant-Based Dietary Intervention Reporting Checklist Table S2, Baseline Characteristics Table S3, Individual Participant Data Table S4, BMI, Dietary Intake and Activity Changes Table S5, Body Composition Change Table S6, Meal Examples Table S7, Adverse Events Table S8, Quality of Life by EORTC QLQ C30 Table S9, Post-Intervention Qualitative Feedback Survey Results Table S10, Feedback Comments Table S11, Case Examples of Patients with Improved Disease Trajectory Table S12, Metabolic Changes Table S13, Microbiome Changes Table S14, Immune Changes Table S15, Immune Flow Cytometry Changes Table S16, scRNAseq global cell numbers and patients with each identified cell type at baseline and W52. Table S17, Purified Diet Formulations Table S18, Standard Diet Table S19, Vk*MYC mice individual data Table S20, Human Flow Cytometry Leukocyte Panel Antibodies Table S21, Pan Leukocyte Panel Subsets Table S22, Mouse Flow Cytometry Antibodies

Figure S3 shows change in microbiome diversity, composition, top genera, top species and top butyrate producer species.

Figure S5 shows the effect of high fiber diet on Vk*MYC mice and t-VkMYC mice.

Role of 2-HG and KDM3B catalytic activity in KDM3B dependency

Figure S7 shows the effect of SCFAs on myeloma cells and immunodeficient mice.

Figure S1 shows the meals provided, CONSORT Diagram and sample timepoints analyzed.

Plasticity transitions during carcinoma progression generate fetal-like progenitor states with metastatic capacity. How these progenitors emerge and persist during tumor progression remains unclear. Here, we elucidate a process that drives the emergence of SOX2+ metastatic progenitors in lung adenocarcinomas (LUAD). LUAD cells at the tumor invasive front and distant metastases express the cell adhesion molecule L1CAM, a marker of regenerative epithelial progenitors and a mediator of cell-basement membrane and cell-cell interactions, as well as the proliferation of extravasated micrometastatic cells. We now identify a distinct and broader role of L1CAM as promoter of the SOX2+ LUAD progenitor state. We show that L1CAM at cell-cell interfaces promotes the assembly of the planar cell polarity (PCP) complex in metastatic LUAD progenitors. L1CAM-dependent PCP acting through a non-canonical WNT signaling activates c-Jun, which cooperates with the chromatin remodeling factor CHD1 to drive SOX2 expression and metastatic activity. This axis sustains the tumor-initiating and regenerative capacity of LUAD progenitor cells. By illuminating the role of L1CAM and PCP signaling in the generation of SOX2+ LUAD progenitors, our findings identify potential new targets to treat metastatic cancer.

Recurrent mosaic somatic mutations in circulating leukocytes can be frequently found in the aging population. These mutations frequently arise in epigenetic modifier genes like TET2. In the absence of signs of hematologic malignancies this condition is termed clonal hematopoiesis (CH). Although CH has been associated with increased incidence and adverse outcomes in patients with solid tumors, the disease-specific effects and mechanisms by which CH alters solid tumor biology have not been delineated. To develop insights into the interplay between mutant CH clones and epithelial tumor cells we analyzed a cohort of over forty-seven thousand patients who underwent paired blood and tumor sequencing. After correcting for age, sex, ancestry, stage, smoking, and previous treatment history, we identify unique patterns of poor survival associated with specific CH genotypes and solid tumor histologies. Given that the detection of tumor-infiltrating TET2-CH clones is especially associated with poorer outcomes in thyroid cancer patients (HR 2.18, 95%CI 1.18-4.05, p=0.013), we focused on studying the role of CH in thyroid cancer biology. Compared to other CH alleles, TET2-mutant CH is enriched in the tumor microenvironment (TME) across solid tumors and is associated with adverse prognosis specifically in Anaplastic thyroid cancer (ATC) patients. ATC is a clinically aggressive malignancy with a dismal prognosis. Combined BRAF/MEK inhibition offers significant therapeutic benefit in patients with BRAFV600E-mutant ATCs. However, relapses are common and overall survival remains poor. A hallmark of ATC is significant infiltration with myeloid cells, particularly macrophages. ATCs are most common in the aging population, which also has an increased incidence of TET2-mutant CH. These mutant macrophages have been shown to accelerate CH-associated pathophysiology including atherosclerosis. However, the clinical and mechanistic contribution of TET2-mutant clones to the prognosis and/or treatment response in solid tumors has not been elucidated. Using subclonal murine models of Tet2-mutant CH we confirm that mutant macrophages selectively infiltrate mouse BrafV600E-mutant ATC models (58% higher enrichment of Tet2-/- cells in the TME compared to the WT counterparts) and confer resistance to BRAF/MEK inhibition (68-days median survival compared to no mortality after 147 days). Using single-cell CITEseq, we identify that the overexpression of Tgfβ-family ligands by CH macrophages is the driver of this resistance. Importantly, inhibition of Tgfβ signaling or the depletion of mutant-macrophages completely restored the sensitivity to MAPK pathway inhibition. In summary, this work identified a novel actionable resistance mechanism mediated by a clonally driven process within the tumor-immune microenvironment. Pablo Sanchez Vela, Vera Tiedje, Julie L. Yang, Brian R. Untch, Laura Boucai, Aaron J. Stonestrom, Alberto Bueno Costa, Sebastià Franch Expósito, Sebastià Franch Expósito, Avi Srivastava, Marina Kerpelev, Jillian Greenberg, Matthew Wereski, Amanda Kulick, Kevin Chen, Tianyue Qin, Soo-Yeon Im, Anthony R. Martinez Benitez, Raquel Pluvinet, Merve Sahin, Kamal Menghrajani, Gnana P. Krishnamoorthy, Elisa de Stanchina, Ahmet Zehir, Rahul Satija, Jeffrey Knauf, Robert L. Bowman, Manel Esteller, Sean Devlin, Michael F. Berger, Richard P. Koche, Ross L. Levine, James A. Fagin. TET2-mutant clonal hematopoiesis enrichment in the tumor microenvironment is an actionable driver of treatment resistance in solid tumors: Thyroid cancer as a paradigm [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 2552.

Small cell lung cancer (SCLC) is known for its high metastatic potential, with most patients demonstrating clinically evident metastases in multiple organs at diagnosis. The factors contributing to this exceptional metastatic capacity have not been defined. To bridge this gap, we compare gene expression in SCLC patient samples who never experienced metastasis or relapse throughout their clinical course, versus primary SCLC patient samples from more typical patients who had metastatic disease at diagnosis. This analysis identifies FOXA2 as a transcription factor strongly associated with SCLC metastasis. Subsequent analyses in experimental models demonstrates that FOXA2 induces a fetal neuroendocrine gene expression program and promotes multi-site metastasis. Moreover, we identify ASCL1, a transcription factor known for its initiating role in SCLC tumorigenesis, as a direct binder of the FOXA2 promoter and regulator of FOXA2 expression. Taken together, these data define the ASCL1-FOXA2 axis as a critical driver of multiorgan SCLC metastasis.

Phenotype switching is a form of cellular plasticity in which cancer cells reversibly move between two opposite extremes: proliferative versus invasive states1,2. Although it has long been hypothesized that such switching is triggered by external cues, the identity of these cues remains unclear. Here we demonstrate that mechanical confinement mediates phenotype switching through chromatin remodelling. Using a zebrafish model of melanoma coupled with human samples, we profiled tumour cells at the interface between the tumour and surrounding microenvironment. Morphological analysis of interface cells showed elliptical nuclei, suggestive of mechanical confinement by the adjacent tissue. Spatial and single-cell transcriptomics demonstrated that interface cells adopted a gene program of neuronal invasion, including the acquisition of an acetylated tubulin cage that protects the nucleus during migration. We identified the DNA-bending protein HMGB2 as a confinement-induced mediator of the neuronal state. HMGB2 is upregulated in confined cells, and quantitative modelling revealed that confinement prolongs the contact time between HMGB2 and chromatin, leading to changes in chromatin configuration that favour the neuronal phenotype. Genetic disruption of HMGB2 showed that it regulates the trade-off between proliferative and invasive states, in which confined HMGB2high tumour cells are less proliferative but more drug-resistant. Our results implicate the mechanical microenvironment as a mechanism that drives phenotype switching in melanoma.

There is a crucial need for strategies that stimulate repair in the adult brain. The neonatal mouse cerebellum can regenerate via the adaptive reprogramming of nestin (Nes)-expressing progenitors (NEPs). However, analysis of Nes + cells of the adult cerebellum is limited. Using reporter lines and genetic inducible fate mapping, we show that adult Nes + cells are mainly Bergmann glia ( Nes + Bg) that have in vitro sphere-forming ability. Following injury, Nes + Bg increase in number due to upregulation of Nes in Hopx -expressing Bg, but the cells exhibit limited regeneration. ATAC-seq of Nes + Bg reveals that silencing of developmental genes compared to neonatal NEPs contributes to the impaired regeneration. Activating sonic hedgehog signalling augments the number of Nes + Bg after injury but not neurogenesis, showing additional cues are required. Our results demonstrate an age-dependent decline in the regenerative potential of NEPs and highlight Nes+ Bg as potential injury-responsive cells that could facilitate regeneration.

Cancer stem cells are essential for initiation and therapy resistance of many cancers, including acute myeloid leukemias (AML). Here, we apply functional genomic profiling to diverse human leukemias, including high-risk MLL- and NUP98-rearranged specimens, using label tracing in vivo. Human leukemia propagation is mediated by a rare quiescent label-retaining cell (LRC) population undetectable by current immunophenotypic markers. AML quiescence is reversible, preserving genetic clonal competition and epigenetic inheritance. LRC quiescence is defined by distinct promoter-centered chromatin and gene expression dynamics controlled by an AP-1/ETS transcription factor network, where JUN is necessary and sufficient for LRC quiescence and associated with persistence and chemotherapy resistance in diverse patients. This enables prospective isolation and manipulation of immunophenotypically-varied leukemia stem cells, establishing the functions of epigenetic plasticity in leukemia development and therapy resistance. These findings offer insights into leukemia stem cell quiescence and the design of therapeutic strategies for their clinical identification and control.