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How basal cell carcinoma (BCC) interacts with its tumor microenvironment to promote growth is unclear. We use singe-cell RNA sequencing to define the human BCC ecosystem and discriminate between normal and malignant epithelial cells. We identify spatial biomarkers of tumors and their surrounding stroma that reinforce the heterogeneity of each tissue type. Combining pseudotime, RNA velocity–PAGA, cellular entropy, and regulon analysis in stromal cells reveals a cancer-specific rewiring of fibroblasts, where STAT1, TGF-β, and inflammatory signals induce a noncanonical WNT5A program that maintains the stromal inflammatory state. Cell-cell communication modeling suggests that tumors respond to the sudden burst of fibroblast-specific inflammatory signaling pathways by producing heat shock proteins, whose expression we validated in situ. Last, dose-dependent treatment with an HSP70 inhibitor suppresses in vitro vismodegib-resistant BCC cell growth, Hedgehog signaling, and in vivo tumor growth in a BCC mouse model, validating HSP70’s essential role in tumor growth and reinforcing the critical nature of tumor microenvironment cross-talk in BCC progression.

Advanced basal cell carcinomas (BCCs) are driven by the Hedgehog (HH) pathway and often possess inherent resistance to SMO inhibitors. Identifying and targeting pathways that bypass SMO could provide alternative treatments. Here, we use a combination of RNA-sequencing analysis of advanced BCC tumor-normal pairs and immunostaining of human and mouse BCC samples to identify an MTOR signature in BCC. Pharmacological inhibition of MTOR activity in BCC cells significantly reduces cell proliferation without affecting HH signaling. Similarly, treatment ofPtch1fl/fl;Gli1-CreERT2mice with everolimus reduces tumor growth and aPKC activity, suggesting that MTOR promotes tumor growth by activating aPKC and demonstrating that suppressing MTOR could be a promising target for BCC patients.

How basal cell carcinoma (BCC) interacts with its tumor microenvironment to promote growth is unclear. Here we use singe-cell RNA sequencing to define the human BCC ecosystem and discriminate between normal and malignant epithelial cells. We identify spatial biomarkers of both tumors and their surrounding stroma that reinforce the heterogeneity of each tissue type. Combining pseudotime, RNA velocity, cellular entropy, and regulon analysis in stromal cells reveal a cancer-specific rewiring of fibroblasts where STAT1, TGF-β, and inflammatory signals induce a non-canonical WNT5A program that maintains the stromal inflammatory state. Cell-cell communication modeling suggests that tumors respond to the sudden burst of fibroblast-specific inflammatory signaling pathways by producing heat shock proteins, which we validated in situ. Finally, dose-dependent treatment with an HSP70 inhibitor suppresses in vitro BCC cell growth and Hedgehog signaling and in vivo tumor growth in a BCC mouse model, validating HSP70’s essential role in tumor growth and reinforcing the critical nature of tumor microenvironment crosstalk in BCC progression.

In RAW 264.7 cells [1], PKC-ɛ regulates FcγR-mediated phagocytosis. BMDM behave similarly; PKC-ɛ concentrates at phagosomes and internalization are reduced in PKC-ɛ−/− cells. Two questions were asked: what is the role of PKC-ɛ? and what domains are necessary for PKC-ɛ concentration? Function was studied using BMDM and frustrated phagocytosis. On IgG surfaces, PKC-ɛ−/− macrophages spread less than WT. Patch-clamping revealed that the spreading defect is a result of the failure of PKC-ɛ−/− macrophages to add membrane. The defect is specific for FcγR ligation and can be reversed by expression of full-length (but not the isolated RD) PKC-ɛ in PKC-ɛ−/− BMDM. Thus, PKC-ɛ function in phagocytosis requires translocation to phagosomes and the catalytic domain. The expression of chimeric PKC molecules in RAW cells identified the ɛPS as necessary for PKC-ɛ targeting. When placed into (nonlocalizing) PKC-δ, ɛPS was sufficient for concentration, albeit to a lesser degree than intact PKC-ɛ. In contrast, translocation of δ(ɛPSC1B) resembled that of WT PKC-ɛ. Thus, ɛPS and ɛC1B cooperate for optimal phagosome targeting. Finally, cells expressing ɛK437W were significantly less phagocytic than their PKC-ɛ-expressing counterparts, blocked at the pseudopod-extension phase. In summary, we have shown that ɛPS and ɛC1B are necessary and sufficient for targeting PKC-ɛ to phagosomes, where its catalytic activity is required for membrane delivery and pseudopod extension.

According to the Centers for Disease Control and Prevention (CDC), breast cancer is currently the 6th leading cause of death of women in the U.S. (CDC, 2007). Currently, estrogen receptor (ER) posi...