Summary Mutually exclusive activating mutations in the GNAQ and GNA11 oncogenes, encoding heterotrimeric Gαq family members, have been identified in ~83% and ~6% of uveal and skin melanomas, respectively. However, the molecular events underlying these GNAQ-driven malignancies are not yet defined, thus limiting the ability to develop cancer-targeted therapies. Here, we focused on the transcriptional co-activator YAP, a critical component of the Hippo signaling pathway that controls organ size. We found that Gαq stimulates YAP through a Trio-Rho/Rac signaling circuitry promoting actin polymerization, independently of PLCβ and the canonical Hippo pathway. Furthermore, we show that Gαq promotes the YAP-dependent growth of uveal melanoma cells, thereby identifying YAP as a suitable therapeutic target in uveal melanoma, the first described GNAQ/GNA11-initiated human malignancy.

SUMMARY The integrity of the epidermis and mucosal epithelia is highly dependent on resident self-renewing stem cells, which makes them vulnerable to physical and chemical insults compromising the repopulating capacity of the epithelial stem cell compartment. This is frequently the case in cancer patients receiving radiation or chemotherapy, many of whom develop mucositis, a debilitating condition involving painful and deep mucosal ulcerations. Here, we show that inhibiting the mammalian target of rapamycin (mTOR) with rapamycin increases the clonogenic capacity of primary human oral keratinocytes and their resident self-renewing cells by preventing stem cell senescence. This protective effect of rapamycin is mediated by the increase expression of mitochondrial superoxide dismutase (MnSOD), and the consequent inhibition of ROS formation and oxidative stress. mTOR inhibition also protects from the loss of proliferative basal epithelial stem cells upon ionizing radiation in vivo, thereby preserving the integrity of the oral mucosa and protecting from radiation-induced mucositis.

Oral mucosal wound healing has long been regarded as an ideal system of wound resolution. However, the intrinsic characteristics that mediate optimal healing at mucosal surfaces are poorly understood, particularly in humans. We present a unique comparative analysis between human oral and cutaneous wound healing using paired and sequential biopsies during the repair process. Using molecular profiling, we determined that wound-activated transcriptional networks are present at basal state in the oral mucosa, priming the epithelium for wound repair. We show that oral mucosal wound-related networks control epithelial cell differentiation and regulate inflammatory responses, highlighting fundamental global mechanisms of repair and inflammatory responses in humans. The paired comparative analysis allowed for the identification of differentially expressed SOX2 (sex-determining region Y-box 2) and PITX1 (paired-like homeodomain 1) transcriptional regulators in oral versus skin keratinocytes, conferring a unique identity to oral keratinocytes. We show that SOX2 and PITX1 transcriptional function has the potential to reprogram skin keratinocytes to increase cell migration and improve wound resolution in vivo. Our data provide insights into therapeutic targeting of chronic and nonhealing wounds based on greater understanding of the biology of healing in human mucosal and cutaneous environments.

Genomic alterations in GNAS, the gene coding for the Gαs heterotrimeric G-protein, are associated with a large number human of diseases. Here, we explored the role of Gαs on stem cell fate decisions by using the mouse epidermis as a model system. Conditional epidermal deletion of Gnas or repression of PKA signaling caused a remarkable expansion of the stem cell compartment, resulting in rapid basal cell carcinoma formation. In contrast, inducible expression of active Gαs in the epidermis caused hair follicle stem cell exhaustion and hair loss. Mechanistically, we found that Gαs-PKA disruption promotes the cell autonomous Sonic Hedgehog pathway stimulation and Hippo signaling inhibition, resulting in the non-canonical activation of GLI and YAP1. Our study highlights an important tumor suppressive function of Gαs-PKA, limiting the proliferation of epithelial stem cells and maintaining proper hair follicle homeostasis. These findings can have broad implications in multiple pathophysiological conditions, including cancer.

Activating mutations in GNAQ and GNA11, encoding members of the Gαq family of G protein α subunits, are the driver uveal melanoma oncogenes, while mutations in Gq-linked G proteincoupled receptors (GPCRs) have been identified recently in numerous human malignancies. How Gαq and its coupled receptors transduce mitogenic signals is still unclear, due to the complexity of signaling events perturbed upon Gq activation. Using of a synthetic biology approach and a genome-wide RNAi screen, we found that a highly conserved guanine nucleotide exchange factor, Trio, is essential to activate Rho- and Rac-regulated signaling pathways acting on JNK and p38, thereby transducing proliferative signals from Gαq to the nucleus independently of PLC-β. Indeed, while many biological responses elicited by Gq depend on the transient activation of second messenger system, Gq utilizes a hardwired protein-protein interaction-based signaling circuitry to achieve the sustained stimulation of proliferative pathways, thereby controlling normal and aberrant cell growth.

Dysregulation of the Hippo signaling pathway and the consequent YAP1 activation is a frequent event in human malignancies, yet the underlying molecular mechanisms are still poorly understood. A pancancer analysis of core Hippo kinases and their candidate regulating molecules revealed few alterations in the canonical Hippo pathway, but very frequent genetic alterations in the FAT family of atypical cadherins. By focusing on head and neck squamous cell carcinoma (HNSCC), which displays frequent FAT1 alterations (29.8%), we provide evidence that FAT1 functional loss results in YAP1 activation. Mechanistically, we found that FAT1 assembles a multimeric Hippo signaling complex (signalome), resulting in activation of core Hippo kinases by TAOKs and consequent YAP1 inactivation. We also show that unrestrained YAP1 acts as an oncogenic driver in HNSCC, and that targeting YAP1 may represent an attractive precision therapeutic option for cancers harboring genomic alterations in the FAT1 tumor suppressor genes.

Summary Two studies published in this issue of Cancer Discovery describe the emerging mutational landscape of head and neck squamous cell carcinomas (HNSCC) and their genomic and epigenetic alterations, thus identifying novel actionable cancer drivers and predictive biomarkers for targeted therapies. Most genomic alterations in HNSCC converge in a handful of molecular pathways resulting in cell cycle deregulation, genomic instability, cell differentiation defects, and persistent mitogenic signaling, the latter involving aberrant PI3K/mTOR pathway activation thereby rendering HNSCC responsive to PI3K/mTOR inhibitors.

BackgroundCetuximab, a monoclonal blocking antibody against the epidermal growth factor receptor EGFR, has been approved for the treatment of squamous cell carcinomas of the head and neck (HNSCC). However, only few patients display long-term responses, prompting the search for cetuximab resistance mechanisms and new therapeutic options enhancing cetuximab effectiveness.MethodsCetuximab-sensitive HNSCC cells were retro-engineered to express PIK3CA and RAS oncogenes. These cells and HNSCC cells harboring endogenous PIK3CA and RAS oncogenes were xenografted into mice (n = 10 per group) and studied for their biochemical, antitumor, antiangiogenic, and antilymphangiogenic responses to cetuximab and mTOR targeting agents. All P values are two-sided.ResultsCetuximab treatment of PIK3CA- and RAS-expressing HNSCC xenografts promoted an initial antitumor response, but all tumors relapsed within few weeks. In these tumors, cetuximab did not decrease the activity of mTOR, a downstream signaling target of EGFR, PIK3CA, and RAS. The combined administration of cetuximab and mTOR inhibitors exerted a remarkably increased antitumor activity, particularly in HNSCC cells that are resistant to cetuximab as a single agent. Indeed, cotargeting mTOR together with cetuximab caused a rapid tumor collapse of both PIK3CA- and RAS-expressing HNSCC xenografts (P < .001), concomitant with reduced proliferation (P < .001) and lymphangiogenesis (P < .001).ConclusionThe presence of PIK3CA and RAS mutations and other alterations affecting the mTOR pathway activity in HNSCC could be exploited to predict the potential resistance to cetuximab, and to select the patients that may benefit the most from the concomitant administration of cetuximab and PI3K and/or mTOR inhibitors as a precision molecular therapeutic option for HNSCC patients.