ScTrio-seq identifies colon cancer lineages: To better design treatments for cancer, it is important to understand the heterogeneity in tumors and how this contributes to metastasis. To examine this process, Bian et al. used a single-cell triple omics sequencing (scTrio-seq) technique to examine the mutations, transcriptome, and methylome within colorectal cancer tumors and metastases from 10 individual patients. The analysis provided insights into tumor evolution, linked DNA methylation to genetic lineages, and showed that DNA methylation levels are consistent within lineages but can differ substantially among clones. Science , this issue p. 1060

Background: Mononuclear cells in peripheral blood and ascites are important clinical resources commonly used in translational and basic research. However, the impact of different cryopreservation durations and extra freeze-thaw cycles on the number and function of mononuclear cells is unknown. Methods: Peripheral blood samples (n = 21) and ascites samples (n = 8) were collected from healthy volunteers and ovarian cancer patients. Mononuclear cells were isolated, frozen, and thawed at 6 and 12 months. The impact of cryopreservation on cell viability, the phenotype, and the activation and proliferation of T cells were analyzed by flow cytometry. Single-cell sequencing was applied to investigate the underlying mechanism. Results: The cell number and viability of mononuclear cells in peripheral blood and ascites were significantly decreased after cryopreservation. The T lymphocytes, especially CD4⁺ T cells, were affected the most significantly. By contrast, monocytes, natural killer (NK) cells, natural killer T (NKT) cells, and B cells were more tolerant. Meanwhile, T cell proliferation and IL-2 secretion are significantly affected after long-term cryopreservation. Mechanistically, the cell death induced by elevated reactive oxygen species (ROS) was involved in the reduction of CD4⁺ T cells after cryopreservation. Conclusions: Our data indicates that different subtypes of mononuclear cells exhibit different tolerance capacities upon cryopreservation. Thus, our research can provide evidence and support for individuals who are conducting experiments using frozen clinical patient-derived mononuclear cells, for basic research or clinical trials. In addition, extra caution is worthwhile when researchers compare immune cell functionality from peripheral blood or ascites across datasets obtained in different cryopreservation conditions.

Background: Ovarian cancer (OC) remains the most lethal gynaecological malignancy, and acquired resistance to PARP inhibitors such as olaparib represents a major clinical challenge. Although dysregulation of iron metabolism has been implicated in tumorigenesis, the specific role of the iron exporter SLC40A1 (ferroportin, FPN) in mediating olaparib resistance remains poorly understood. The study aimed to elucidate the clinical significance of FPN deficiency and to explore therapeutic strategies for reversing olaparib resistance. Methods: FPN expression was evaluated in OC samples from a clinical cohort of 19 patients, stratified by olaparib responsiveness, to determine its prognostic value. Functional mechanisms were investigated using multiple OC cell lines, patient‐derived organoids (PDOs), and mouse xenograft models. We integrated analyses of iron homeostasis, ROS dynamics, DNA repair capacity, and innate immune signaling, with a focus on the cGAS–STING–STAT3 signaling cascade. The therapeutic potential of combining olaparib with the ferroptosis inducer erastin or the STING inhibitor C‐176 was assessed as a strategy to overcome FPN deficiency‐driven resistance. Results: Clinically, FPN expression was significantly downregulated in olaparib‐resistant tissues compared with sensitive controls ( p < 0.05), and low FPN levels were strongly associated with poor therapeutic outcomes. Mechanistically, FPN deficiency promoted intracellular iron accumulation and ROS generation, which led to increased ferroptosis vulnerability, impaired homologous recombination repair (HRR) and concomitant activation of the cGAS–STING–STAT3 inflammatory axis, thereby driving olaparib resistance. Notably, in vivo studies demonstrated that combining olaparib with either erastin or C‐176 significantly suppressed tumour growth and restored drug sensitivity compared with monotherapy ( p < 0.01), supporting the translational potential of targeting this axis. Conclusions: Our study establishes a clinically relevant mechanistic connection between ferroportin deficiency and cGAS–STING–STAT3‐mediated olaparib resistance in OC. We identify the FPN–ROS–cGAS–STING–STAT3 axis as an actionable vulnerability in resistant OC, indicating that this pathway is a promising therapeutic target to overcome olaparib resistance. Highlights: FPN loss drives olaparib resistance and predicts poor outcomes in ovarian cancer. FPN loss causes iron and ROS accumulation, impairing HRR and activating the cGAS–STING–STAT3 axis. The cGAS–STING–STAT3 pro‐survival program drives olaparib resistance. STING inhibition restores olaparib sensitivity in resistant ovarian cancer.

Single-cell metabolic characteristics are powerful indicators of cellular physiological and pathological states. Flow cytometry enables high-throughput single-cell metabolic profiling and immunophenotyping; however, spectral overlap impedes simultaneous detection of multi-parametric features. This limitation imposes larger sample requirements and increased variabilities due to metabolic dynamics-constraints acutely magnified in studies utilizing clinical microsamples. To overcome these shortcomings, we developed a spectral flow cytometry platform for immunometabolic features, using 13 dual-probe combinations and 11 fluorophore probes. Requiring only 100 μL of whole blood per assay, this platform enables concurrent detection of 4 metabolic biomarkers and 16 immune markers. Application to patients with heart failure revealed heterogeneous metabolic landscapes across 20 immune subpopulations, showing reduced frequencies of naive T cells and NK-like T cells, with a metabolic shift from fatty acid dependence to glucose avidity. Our framework captures metabolic interactions previously inaccessible by sequential detection and will help to enable precision immunometabolism research.

Current histopathological classification systems for ovarian cancer cannot adequately predict platinum chemotherapy response, hindering personalized therapeutic strategies. Here, through integrative analysis of multiple datasets from TCGA, ICGC and GEO databases, we stratify patients into metabolic associated chemotherapy-refractory and chemotherapy-sensitive types (MCRT, MCST for short) with distinct prognosis by 20 genes. Lipidomic profiling of ascites from patients with ovarian cancer further identifies oleic acid as a hallmark metabolite in MCRT cases. Oleic acid treatment facilitates the resistance to cisplatin in ovarian cancer cells and patient-derived organoids. Mechanistically, oleic acid activates and promotes nuclear translocation of YAP through CD36. Inhibition of SCD1—genetically or pharmacologically—synergizes with cisplatin in 26 patient-derived organoids and suppresses tumor growth in xenograft models. Our study proposes a new metabolic classification of ovarian cancer that can predict chemotherapy sensitivity. Moreover, it discovers the role of the oleic acid-CD36-YAP signaling axis in promoting cisplatin resistance. These findings shed light on how oleic acid facilitates the resistance to cisplatin in ovarian cancer and provide a potential therapeutic strategy for treating patients with ovarian cancer.