Supplementary Table S3. Mass spectrometry analysis identified two acetylation sites of ATF3

Ischemia-reperfusion injury (IRI) is a major cause of acute kidney injury (AKI) that significantly increases the risk of progression to chronic kidney disease (CKD). Although oxidative stress has been implicated in this transition, the precise mechanisms through which it orchestrates inflammation and fibrosis during IRI-induced AKI-CKD progression remain poorly understood. In this study, we observed sustained reactive oxygen species (ROS) production in post-IRI kidneys. ROS were found to activate AMP-activated protein kinase (AMPK) in macrophages in a calcium-dependent manner. Conditional knockout of AMPKα1 in macrophages (Lyz2-Cre; Prkaa1-fl/fl mice) significantly attenuated renal fibrosis following IRI. Single-cell RNA sequencing analysis further revealed that AMPKα1 deletion reduced the accumulation of Arg1+ MMP12+ macrophages and diminished a profibrotic tubular epithelial cell (TEC) subpopulation marked by persistent expression of PDGFB and VCAM1. These macrophages were shown to interact with PDGFB+ VCAM1+ TECs. Mechanistically, macrophage-derived TWEAK signaling through its receptor Fn14 promoted PDGFB production in TECs, driving maladaptive changes and a fibrogenic phenotype. Importantly, TWEAK neutralization effectively mitigated the AKI-to-CKD transition. Together, our results identify macrophage AMPK as a key redox sensor that, upon activation by oxidative stress, initiates maladaptive macrophage-TEC crosstalk, ultimately promoting renal fibrosis and CKD progression.

Supplementary Table S2. Patient characteristics of tissue microarray

Cancer stem cells (CSCs) are a subpopulation of tumor cells with stem cell-like properties, which are endowed with unique self-renewal capacity and heterogeneity. Specifically, they drive tumor initiation, progression, recurrence, and therapeutic resistance. As the core drivers of tumor development and treatment resistance, CSCs exhibit distinct metabolic characteristics that sustain their self-renewal and differentiation potential. Various cells in the tumor microenvironment (TME) interact with CSCs, thereby creating a favorable niche for maintaining CSCs stemness. Furthermore, the metabolic patterns of CSCs themselves can modulate the functions of other TME-resident cells; collectively, these bidirectional interactions promote tumor recurrence and therapeutic resistance. A growing body of evidence has confirmed the critical role of CSCs in tumor progression, drug resistance, and recurrence, which lays a solid foundation for the development of potential therapeutic targets and intervention strategies. This review systematically summarizes the biological characteristics and targeted therapeutic strategies of CSCs, and explores potential directions to break through current therapeutic bottlenecks in combination with the latest status of clinical research.

Acute myeloid leukemia (AML) featuring retinoic acid receptor-gamma (RARG) rearrangements exhibits morphological features resembling those of acute promyelocytic leukemia but is associated with drug resistance and poor clinical outcomes. However, the mechanisms underlying the role of RARG fusions in leukemogenesis remain elusive. Here, we show that RARG fusions disrupt myeloid differentiation and promote proliferation and self-renewal of hematopoietic stem and progenitor cells (HSPCs) by upregulating BCL2 and ATF3. RARG fusions overexpression leads to preleukemic phenotypes but fails to induce oncogenic transformation. However, the co-occurrence of RARG fusions and heterozygous Wt1 loss induce fully penetrant AML by activating MYC and HOXA9/MEIS1 targets. Leveraging Connectivity Map resources and high-throughput screening, we identify venetoclax, homoharringtonine, and daunorubicin as potential therapeutic options for RARG-AML. Overall, our findings provide pivotal insights into the molecular mechanisms governed by RARG fusions and enhanced by WT1 loss in AML development and propose a rational therapeutic strategy for RARG-AML. Acute Myeloid Leukaemia (AML) with Retinoic acid receptor-gamma (RARG) fusions have similarities to Acute Promyelocytic Leukemia (APL) but are not responsive to standard APL treatments. Here, the authors show the specific molecular mechanisms lead by RARG fusions in AML driving leukemogenesis and as potential therapeutic targets.

SIRPA delivers an anti-phagocytic “don’t-eat-me” signal through its expression on macrophage membrane surface and promotes tumor progression via membrane-independent mechanism. However, current CD47- and SIRPA-targeting agents only disrupt cell-surface inhibitory signaling, highlighting the therapeutic potential of degrading SIRPA. Here, we identified that E3 ubiquitin ligase TRIM2 interacted with SIRPA in vitro. Clinically, elevated TRIM2 expression is associated with prolonged overall and progression-free survival in renal-cell carcinoma (RCC) patients. Mechanistically, TRIM2 catalyzes K48-linked poly-ubiquitination of SIRPA, promoting its proteasomal degradation and reducing SIRPA protein levels. CRISPR/Cas9-mediated deletion of TRIM2 in macrophages upregulated SIRPA, significantly impairing phagocytosis of tumor cells. TRIM2 deficiency also promoted tumor growth by increasing intratumoral infiltration of M2-type macrophage, reducing accumulation of anti-tumor M1-like and antigen-presenting macrophages, and impairing effector CD8 + T-cell recruitment. Importantly, combined TRIM2 overexpression and PD-L1 blockade synergistically enhance anti-tumor immunity. Together, these results suggest that targeting TRIM2 may represent a novel therapeutic strategy against RCC by degrading SIRPA in macrophage and provide a roadmap for clinical application.

Carboxymethyl pachymaran (CMP) was modified by high temperature (HT), high pressure (HP) and gamma irradiation (GI) to obtain HT-CMP, HP-CMP, and GI-CMP samples, respectively. The effects of different modification methods on the immunomodulatory activity of CMPs were evaluated by cell culture in vitro and animal in vivo tests. The modified CMPs showed a stronger immunomodulatory activity than that of CMP in vitro. The modified CMPs improved the proliferative and phagocytic capacities, the ROS release, the TNF-α cytokine secretion of RAW264.7 compared with the CMP. For in vivo test, we found that the CMP and modified CMPs were able to enhance the immune function in mice with cyclophosphamide (CTX) induced by immunosuppression. All the CMPs could increase the cytokine levels (IL-2, IL-6, IFN-γ, TNF-α) and elevate the mRNA expression of NF-κB and TLRs. Furthermore, CMP, HT-CMP and GI-CMP had an ameliorative effect on intestinal flora disorders and could restore the richness and diversity of intestinal flora. Our results provided a theoretical foundation for the potential development of CMP-based immunomodulatory drugs.

Tyrosine kinase inhibitors (TKIs) only partially inhibit the growth of Philadelphia chromosome-positive B-cell acute lymphoblastic leukemia (Ph+ B-ALL) cells, and often lead to rapid relapse. Therefore, it is essential to elucidate the mechanisms of resistance and develop novel treatment strategies. Histone deacetylases (HDACs) are often dysregulated in hematological malignancies, and many HDAC inhibitors have shown potent antitumor activities. In this study, we found that HDAC8 was highly expressed in Ph+ B-ALL patient samples upon TKI treatment. HDAC8 inhibition significantly increased TKI-mediated elimination of leukemia cells and decreased their ability to initiate leukemia. Using two mouse models, we demonstrated that TKIs in combination with targeting HDAC8 effectively inhibited leukemia progression and reduced the frequencies of stem cells. Mechanistically, HDAC8 deacetylated hypoxia inducible factor-1α (HIF-1α) at lysine 19/21, leading to a reduction in PHD2-mediated hydroxylation and subsequent pVHL-mediated ubiquitination, which slowed the degradation of HIF-1α. HIF-1α inhibition induced apoptosis and decreased the initiating capacity of leukemia cells. Importantly, in a Ph+ B-ALL patient–derived xenograft model, targeting HDAC8 or HIF-1α in combination with TKIs significantly inhibited leukemia progression. In conclusion, our study revealed that targeting HDAC8/HIF-1α in combination with TKIs could be a promising strategy for treating Ph+ B-ALL.

Metabolic reprogramming and DNA damage repair are essential in tumorigenesis and chemoresistance, yet their link remains elusive. Here, we show that LDHA deficiency impairs NHEJ and class switch recombination. Additionally, glycolysis-derived lactate promotes XLF lactylation at K288 within its Ku-binding motif (X-KBM) to regulate NHEJ. Mechanistically, DNA damage triggers ATM-mediated GCN5 phosphorylation to increase GCN5-XLF interaction and XLF lactylation, enhancing XLF-Ku80 binding, XLF recruitment to DSBs, and NHEJ efficiency. Cryo-EM structural analysis demonstrates that lactylated X-KBM (laX-KBM) forms a more extensive interface with Ku70/80, inducing conformational changes in the Ku80 vWA domain. XLF lactylation deficiency impairs NHEJ and sensitizes cancer cells to chemotherapy. A specific XLF K288 lactylation peptide inhibitor plus 5-fluorouracil synergistically kills colorectal cancer cells in PDX models with XLF hyperlactylation. These findings highlight that the GCN5-XLF lactylation axis is a critical NHEJ regulator and that targeting XLF lactylation can improve chemotherapy efficiency.

Obesity is a major risk factor for poor breast cancer outcomes, but the impact of obesity-induced tumor microenvironment (TME) metabolites on breast cancer growth and metastasis remains unclear. Here, we performed TME metabolomic analysis in high-fat diet (HFD) mouse models and found that glutathione (GSH) levels were elevated in the TME of obesity-accelerated breast cancer. The deletion of glutamate-cysteine ligase catalytic subunit (GCLC), the rate-limiting enzyme in GSH biosynthesis, in adipocytes but not tumor cells reduced obesity-related tumor progression. Mechanistically, we identified that GSH entered tumor cells and directly bound to lysosomal integral membrane protein-2 (scavenger receptor class B, member 2 [SCARB2]), interfering with the interaction between its N and C termini. This, in turn, recruited mTORC1 to lysosomes through ARF1, leading to the activation of mTOR signaling. Overall, we demonstrated that GSH links obesity and breast cancer progression by acting as an activator of mTOR signaling. Targeting the GSH/SCARB2/mTOR axis could benefit breast cancer patients with obesity.

As the global population ages, the number of patients with osteoporosis is rapidly rising. The existing first-line clinical drugs are bone resorption inhibitors that have difficulty restoring the bone mass of elderly patients to the safe range. The range and period of use of existing peptides and monoclonal antibodies are limited, and small-molecule bone formation–promoting drugs are urgently required. We established an I-9 synthesis route with high yield, simple operation, and low cost that was suitable for future large-scale production. I-9 administration promoted bone formation and increased bone mass in mice with low bone mass in an aged C57 mouse model. Our findings revealed a hitherto undescribed pathway involving the BMP2–ERK–ATF4 axis that promotes osteoblast differentiation; I-9 has favorable biosafety in mice. This study systematically investigated the efficacy, safety, and mechanism of I-9 for treating osteoporosis and positions this drug for preclinical research in the future. Thus, this study has promoted the development of small-molecule bone-promoting drugs.

Abstract Peripheral T-cell lymphoma (PTCL) is a heterogeneous and aggressive disease with a poor prognosis. Histone deacetylase (HDAC) inhibitors have shown inhibitory effects on PTCL. A better understanding of the therapeutic mechanism underlying the effects of HDAC inhibitors could help improve treatment strategies. Herein, we found that high expression of HDAC3 is associated with poor prognosis in PTCL. HDAC3 inhibition suppressed lymphoma growth in immunocompetent mice but not in immunodeficient mice. HDAC3 deletion delayed the progression of lymphoma, reduced the lymphoma burden in the thymus, spleen, and lymph nodes, and prolonged the survival of mice bearing N-methyl-N-nitrosourea–induced lymphoma. Furthermore, inhibiting HDAC3 promoted the infiltration and enhanced the function of natural killer (NK) cells. Mechanistically, HDAC3 mediated ATF3 deacetylation, enhancing its transcriptional inhibitory activity. Targeting HDAC3 enhanced CXCL12 secretion through an ATF3-dependent pathway to stimulate NK-cell recruitment and activation. Finally, HDAC3 suppression improved the response of PTCL to conventional chemotherapy. Collectively, this study provides insights into the mechanism by which HDAC3 regulates ATF3 activity and CXCL12 secretion, leading to immune infiltration and lymphoma suppression. Combining HDAC3 inhibitors with chemotherapy may be a promising strategy for treating PTCL. Significance: Targeting HDAC3 suppresses progression of T‐cell lymphoma by activating ATF3 to induce secretion of CXCL12 and promote infiltration of NK cells, providing an immunostimulatory approach for treating T‐cell lymphoma patients.

The interactions of environmental compartments with epithelial cells are essential for mammary gland development and homeostasis. Currently, the direct crosstalk between the endothelial niche and mammary epithelial cells remains poorly understood. Here, we show that faciogenital dysplasia 5 (FGD5) is enriched in mammary basal cells (BCs) and mediates critical interactions between basal and endothelial cells (ECs) in the mammary gland. Conditional deletion of Fgd5 reduced, whereas conditional knockin of Fgd5 increased, the engraftment and expansion of BCs, regulating ductal morphogenesis in the mammary gland. Mechanistically, murine mammary BC-expressed FGD5 inhibited the transcriptional activity of activating transcription factor 3 (ATF3), leading to subsequent transcriptional activation and secretion of CXCL14. Furthermore, activation of CXCL14/CXCR4/ERK signaling in primary murine mammary stromal ECs enhanced the expression of HIF-1α-regulated hedgehog ligands, which initiated a positive feedback loop to promote the function of BCs. Collectively, these findings identify functionally important interactions between BCs and the endothelial niche that occur through the FGD5/CXCL14/hedgehog axis.

Dysregulated hematopoietic niches remodeled by leukemia cells lead to imbalances in immunological mediators that support leukemogenesis and drug resistance. Targeting immune niches may ameliorate disease progression and tyrosine kinase inhibitor (TKI) resistance in Philadelphia chromosome-positive B-ALL (Ph + B-ALL). Here, we show that T helper type 17 (Th17) cells and IL-17A expression are distinctively elevated in Ph + B-ALL patients. IL-17A promotes the progression of Ph + B-ALL. Mechanistically, IL-17A activates BCR-ABL, IL6/JAK/STAT3, and NF-kB signalling pathways in Ph + B-ALL cells, resulting in robust cell proliferation and survival. In addition, IL-17A-activated Ph + B-ALL cells secrete the chemokine CXCL16, which in turn promotes Th17 differentiation, attracts Th17 cells and forms a positive feedback loop supporting leukemia progression. These data demonstrate an involvement of Th17 cells in Ph + B-ALL progression and suggest potential therapeutic options for Ph + B-ALL with Th17-enriched niches.

CD8 + T cell activation leads to the rapid proliferation and differentiation of effector T cells (T effs ), which mediate antitumor immunity. Although aerobic glycolysis is preferentially activated in CD8 + T effs , the mechanisms that regulate CD8 + T cell glucose uptake in the low-glucose and acidic tumor microenvironment (TME) remain poorly understood. Here, we report that the abundance of the glucose transporter GLUT10 is increased during CD8 + T cell activation and antitumor immunity. Specifically, GLUT10 deficiency inhibited glucose uptake, glycolysis, and antitumor efficiency of tumor-infiltrating CD8 + T cells. Supplementation with glucose alone was insufficient to rescue the antitumor function and glucose uptake of CD8 + T cells in the TME. By analyzing tumor environmental metabolites, we found that high concentrations of lactic acid reduced the glucose uptake, activation, and antitumor effects of CD8 + T cells by directly binding to GLUT10’s intracellular motif. Disrupting the interaction of lactic acid and GLUT10 by the mimic peptide PG10.3 facilitated CD8 + T cell glucose utilization, proliferation, and antitumor functions. The combination of PG10.3 and GLUT1 inhibition or anti–programmed cell death 1 antibody treatment showed synergistic antitumor effects. Together, our data indicate that GLUT10 is selectively required for glucose uptake of CD8 + T cells and identify that TME accumulated lactic acid inhibits CD8 + T cell effector function by directly binding to GLUT10 and reducing its glucose transport capacity. Last, our study suggests disrupting lactate-GLUT10 binding as a promising therapeutic strategy to enhance CD8 + T cell–mediated antitumor effects.

CSCs (Cancer stem cells) with distinct metabolic features are considered to cause HCC (hepatocellular carcinoma) initiation, metastasis and therapeutic resistance. Here, we perform a metabolic gene CRISPR/Cas9 knockout library screen in tumorspheres derived from HCC cells and find that deletion of SCARB2 suppresses the cancer stem cell-like properties of HCC cells. Knockout of Scarb2 in hepatocytes attenuates HCC initiation and progression in both MYC-driven and DEN (diethylnitrosamine)-induced HCC mouse models. Mechanistically, binding of SCARB2 with MYC promotes MYC acetylation by interfering with HDCA3-mediated MYC deacetylation on lysine 148 and subsequently enhances MYC transcriptional activity. Screening of a database of FDA (Food and Drug Administration)-approved drugs shows Polymyxin B displays high binding affinity for SCARB2 protein, disrupts the SCARB2-MYC interaction, decreases MYC activity, and reduces the tumor burden. Our study identifies SCARB2 as a functional driver of HCC and suggests Polymyxin B-based treatment as a targeted therapeutic option for HCC.

Cancer stem cells (CSCs) with distinct metabolic features are considered to cause hepatocellular carcinoma (HCC) initiation, metastasis and therapeutic resistance. Here, we performed a metabolic gene CRISPR/Cas9 knockout screen in tumorspheres derived from HCC cells and found that deletion of the scavenger receptor SCARB2 suppressed the CSC traits of HCC cells. Using lineage tracing of the Scarb2 locus in mice, we demonstrated that Scarb2 positive HCC cells showed tumor-initiating activity. Specifically, Cre-mediated recombination with oncogenic MYC expression in Scarb2 positive cells drove HCC tumor formation, as determined by lineage tracing in Scarb2 mice. Knockout of Scarb2 in hepatocytes attenuated HCC initiation and progression by inhibiting CSC self-renewal. Mechanistically, binding of SCARB2 with MYC promoted MYC acetylation by interfering with HDCA3-mediated MYC deacetylation and subsequently enhanced MYC activity. Screening of a database of FDA-approved drugs showed Polymyxin B displayed high binding affinity for SCARB2 protein, disrupted the SCARB2-MYC interaction, decreased MYC activity, and reduced the tumor burden. Our study identifies SCARB2 as a marker and functional driver of HCC CSCs and suggests a targeted therapeutic option for HCC.

FGD5 promotes basal-like breast cancer tumorigenesis and progression by inducing and maintaining cancer stemness in an EGFR-dependent manner.

The transcription factor MYC is deregulated in almost all human cancers, especially in aggressive lymphomas, through chromosomal translocation, amplification, and transcription hyperactivation. Here, we report that high expression of tribbles homologue 3 (TRIB3) positively correlates with elevated MYC expression in lymphoma specimens; TRIB3 deletion attenuates the initiation and progression of MYC-driven lymphoma by reducing MYC expression. Mechanistically, TRIB3 interacts with MYC to suppress E3 ubiquitin ligase UBE3B-mediated MYC ubiquitination and degradation, which enhances MYC transcriptional activity, causing high proliferation and self-renewal of lymphoma cells. Use of a peptide to disturb the TRIB3-MYC interaction together with doxorubicin reduces the tumor burden in Myc Eμ mice and patient-derived xenografts. The pathophysiological relevance of UBE3B, TRIB3 and MYC is further demonstrated in human lymphoma. Our study highlights a key mechanism for controlling MYC expression and a potential therapeutic option for treating lymphomas with high TRIB3-MYC expression.