Circulating NK cells are known to convert to a type 1 innate lymphoid cell (ILC1)-like phenotype in response to TGF-b exposure. However, the precise cellular changes defining this process as well as the downstream signaling pathways guiding it remain poorly defined, particularly in humans. We used mass cytometry by time-of-flight (CyTOF) to model this phenotypic shift in vitro and identify a synergistic activity of TGF-b and IL-15 in this cellular conversion. CyTOF profiling identified substantial heterogeneity in the propensity of NK cells to adopt an ILC1-like phenotype in culture, characterized by the step-wise acquisition of various markers, including CD69, CD9, CD103, and CD49a. Activating and inhibitory receptors, including NKG2A, NKG2D, KIR2DL1, KIR3DL1, NKp30, NKp44, and NKp46, were all found to be upregulated exclusively on the cellular subsets that converted most readily in response to TGF-b. An assessment of downstream TGF-b signaling identified TAK1-mediated activation of p38 MAPK as the critical pathway driving conversion. IL-15 enhanced TGF-b-mediated conversion through Ras:RAC1 signaling as well as via the activation of MEK/ERK. Interestingly, the adoption of an ILC1-like phenotype was independent of the effect of IL-15 or TGF-b on mTOR, as the culture of NK cells in the presence of mTOR inhibitors, such as rapamycin or torin1, had minimal impact on the degree of conversion. In conclusion, we have used in vitro human culture systems and CyTOF to define the conversion of circulating NK cells to an ILC1-like phenotype and have clarified the pathways responsible for this process.

Background Vismodegib is a novel Hedgehog pathway inhibitor that has revolutionized the treatment of patients with advanced basal cell carcinoma (BCC) who are poor candidates for surgery or radiation. Few studies have explored the use of vismodegib to facilitate further surgery or radiotherapy, and the optimal treatment duration to balance outcomes with adverse effects. Objectives To characterize the disease response, progression, and recurrence outcomes of BCC patients, and to report the impact of subsequent therapies. Methods We performed a retrospective study of 46 adult patients with advanced basal cell carcinoma (aBCC), including both locally advanced (laBCC) and metastatic (mBCC) disease, treated with vismodegib at a single center from 2012 to 2019. Results Thirty-six had laBCC, and 10 had mBCC. Treatment was given over a mean of 21.9 months. Twenty-three (50%) had a complete response (CR), and 19 (41.3%) achieved partial response (PR). Median time to maximal response was 5.3 months. Eleven (23.9%) had resected disease at median 17.2 months, and 11 patients (23.9%) received radiotherapy. Thirty-two (69.6%) experienced progressive disease after achievement of CR or PR. Among 17 CR patients, who stopped treatment, 14 (82.3%) experienced subsequent relapse; 6 (85%) attained a repeat response. Twenty (43.5%) discontinued treatment at least once due to adverse effects. Conclusions With a response rate of 91%, London Regional Cancer Center’s (LRCP)’s experience with vismodegib supports its effectiveness in treatment of aBCC. Moreover, a significant number of patients treated with vismodegib became amenable to surgery or radiotherapy. Toxicity remained an important factor that limited treatment duration.

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