Anaplastic lymphoma kinase rearrangement-associated renal cell carcinoma is a rare subtype of kidney cancer that uniquely expresses ALK fusion genes. To elucidate the role of ALK fusion genes in renal tumorigenesis, Noguchi and colleagues engineered human iPSC-derived kidney organoids (HKO) that overexpress the VCL-ALK fusion gene in a doxycycline-inducible manner. Immunohistochemical analysis of VCL-ALK–overexpressing HKO revealed heightened levels of proliferating tubular cells accompanied by reduced glomerular and stromal cells. Long-term culture and hypoxia studies demonstrated that VCL-ALK–overexpressing HKOs maintained their proliferative index under the stress of extensive hypoxic culture while the control organoids dedifferentiated into cartilage. Transplantation of VCL-ALK–overexpressing HKOs into the renal subcapsular space of immunodeficient mice revealed a propensity for invasion into the adjacent renal parenchyma. RNA-sequencing and subsequent GSEA and DEG analysis found a downregulation of nephron development pathways, decreased expression of mature nephron marker NPHS2, and increased expression of immature nephron markers JAG1 and LHX1 in VCL-ALK–overexpressing HKOs compared to the control. Deconvolution analysis of RNA-sequencing data of VCL-ALK–overexpressing HKO identified decreased levels of mature glomerular cells and mature tubular cells, and an increase in immature nephron cells. Together, these results demonstrate that the VCL-ALK fusion gene disrupts nephron maturation and indicates its potential role in early renal tumorigenesis.Methotrexate-packaged tumor cell-derived microparticles (MTX-MP) have demonstrated an ability to recruit neutrophils and activate adaptive immunity, but the mechanism of this antitumor response is unclear. To interrogate the function of MTX-MPs and the pathways implicated in their effect on neutrophils, Zhang and colleagues utilized in vivo and ex vivo models of MTX-MP–activated neutrophils. Tumor growth was significantly disrupted in subcutaneous models injected with MTX-MP–activated neutrophils, and neutrophil-derived microparticles (Neu-MP) in culture medium demonstrated an ability to kill tumor cells in cell culture models. RNA-sequencing and Reactome enrichment analysis of MTX-MP–activated neutrophils showed enrichment of the neutrophil degranulation pathway, and confocal microscopy revealed increased expression of CD11b and Ly-6G on the plasma membrane. Chloroquine-driven inhibition of the lysosome decreased Neu-MP release, and treatment with NAC or DPI also reduced Neu-MPs, indicating Neu-MP release may be dependent on ROS-mediated neutrophil degranulation. The authors also performed flow cytometry analysis of MTX-MP–activated neutrophils and found that pharmacologic inhibition of ROS or Ca2+ flux blocks neutrophil activation and antitumor activity. Overall, the study defines a lysosomal ROS-driven signaling pathway that enables neutrophils to kill tumor cells through the release of cytotoxic microparticles, revealing a therapeutically actionable mechanism for enhancing innate and adaptive antitumor responses.MTHFD2 is highly expressed in tongue squamous cell carcinoma (TSCC), but its mechanism is not well defined. In their study, Tuohuti and colleagues utilized transcriptomic profiling and in vitro and in vivo studies to determine the functional role of MTHFD2. Proliferation and colony formation assays using MTHFD2 knockdown cells demonstrated reduced proliferative and colony formation abilities. Cell migration was also impaired upon MTHFD2 knockdown. Additional studies in nude mice injected with MTHFD2 knockdown cells showed the development of smaller tumors in comparison to the control, accompanied by reduced Ki-67 staining. RNA-sequencing and KEGG pathway enrichment analysis of MTHFD2 knockdown cells revealed an enrichment of differentially expressed genes in the FOXO signaling pathway. Rescue experiments by inhibition of FOXO1 reversed the antiproliferative effects of MTHFD2 knockdown, and restored tumor growth in vivo. Cycloheximide chase assays found that MTHFD2 knockdown reduced FOXO1 degradation, and proteosome inhibition with MG132 also attenuated FOXO1 degradation. Immunoprecipitation demonstrated that MTHFD2 enhances both FOXO1 ubiquitination and methylation. Immunofluorescence assays further showed that MTHFD2 overexpression enhances FOXO1 cytoplasmic translocation. Altogether, this study identifies the MTHFD2-FOXO1 axis and demonstrates its potential utility as a therapeutic target in TSCC.
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