Despite advances in therapy, glioblastoma remains an incurable disease with a dismal prognosis. Recent studies have implicated cancer stem cells within glioblastoma (glioma stem cells, GSCs) as mediators of therapeutic resistance and tumor progression. In this study, we investigated the role of the transforming growth factor-β (TGF-β) superfamily, which has been found to play an integral role in the maintenance of stem cell homeostasis within multiple stem cell systems, as a mediator of stem-like cells in glioblastoma. We find that BMP and TGF-β signaling define divergent molecular and functional identities in glioblastoma, and mark relatively quiescent and proliferative GSCs, respectively. Treatment of GSCs with BMP inhibits cell proliferation, but does not abrogate their stem-ness, as measured by self-renewal and tumorigencity. Further, BMP pathway activation confers relative resistance to radiation and temozolomide chemotherapy. Our findings define a quiescent cancer stem cell population in glioblastoma that may be a cellular reservoir for tumor recurrence following cytotoxic therapy.

Purpose: In estrogen receptor–positive (ER+)/HER2− breast cancer, multiple measures of intratumor heterogeneity are associated with a worse response to endocrine therapy. We sought to develop a novel experimental model to measure heterogeneity in response to tamoxifen treatment in primary breast tumors. Experimental Design: To investigate heterogeneity in response to treatment, we developed an operating room-to-laboratory pipeline for the collection of live normal breast specimens and human tumors immediately after surgical resection for processing into single-cell workflows for experimentation and genomic analyses. Live primary cell suspensions were treated ex vivo with tamoxifen (10 µmol/L) or control media for 12 hours, and single-cell RNA libraries were generated using the 10X Genomics droplet-based kit. Results: In total, we obtained and processed normal breast tissue from two women undergoing reduction mammoplasty and tumor tissue from 10 women with ER+/HER2− invasive breast carcinoma. We demonstrate differences in tamoxifen response by cell type and identify distinctly responsive and resistant subpopulations within the malignant cell compartment of human tumors. Tamoxifen resistance signatures from resistant subpopulations predict poor outcomes in two large cohorts of ER+ breast cancer patients and are enriched in endocrine therapy–resistant tumors. Conclusions: This novel ex vivo model system now provides the foundation to define responsive and resistant subpopulations within heterogeneous human tumors, which can be used to develop precise single cell–based predictors of response to therapy and to identify genes and pathways driving therapeutic resistance.

The senescence-associated secretory phenotype (SASP) is a hallmark of senescent cells and plays a critical role in the development and progression of various age-related diseases, including cancer, cardiovascular disorders, and neurodegenerative diseases. In this study, we characterize SASP heterogeneity using single-cell RNA sequencing (scRNA-seq) data, focusing on the transcriptional signatures associated with elevated expression of individual SASP genes in mature senescent cells, as well as time-dependent variation in SASP expression across the early and mature senescent states in the WI-38 human lung fibroblast cell line. We generated multiple gene sets, each representing the transcriptional landscape linked to high expression of a specific SASP gene, and integrated them into an ensemble that reflects the temporal dynamics of SASP gene expression. Applying SASP scores derived from this ensemble of gene sets (SASP scores/EGS) to publicly available scRNA-seq datasets from human lung, skin, and eye tissues enabled the identification of senescent fibroblasts and revealed IGFBP7 as a consistently upregulated marker in p21⁺ or p16⁺ fibroblasts across diverse human tissues. Our framework supports improved detection of both early and mature fibroblast replicative senescent cells, offering valuable insights into aging and age-related disease research.

In ER+/HER2- breast cancer, multiple measures of intra-tumor heterogeneity are associated with worse response to endocrine therapy. To investigate heterogeneity in response to treatment, we developed an operating room-to-laboratory pipeline for the collection of live human tumors and normal breast specimens immediately after surgical resection for processing into single-cell workflows for experimentation and genomic analyses. We demonstrate differences in tamoxifen response by cell type and identify distinctly responsive and resistant subpopulations within the malignant cell compartment of human tumors. Tamoxifen resistance signatures from 3 distinct resistant subpopulations are prognostic in large cohorts of ER+ breast cancer patients and enriched in endocrine therapy resistant tumors. This novel ex vivo model system now provides a foundation to define responsive and resistant sub-populations within heterogeneous tumors, to develop precise single cell-based predictors of response to therapy, and to identify genes and pathways driving resistance to therapy.

Comparison of tamoxifen and control treated epithelial subpopulations from normal samples

Supplementary Table S4 from Tamoxifen Response at Single-Cell Resolution in Estrogen Receptor–Positive Primary Human Breast Tumors