Somatic gene mutations can alter the vulnerability of cancer cells to T-cell-based immunotherapies. Here we perturbed genes in human melanoma cells to mimic loss-of-function mutations involved in ...resistance to these therapies, by using a genome-scale CRISPR-Cas9 library that consisted of around 123,000 single-guide RNAs, and profiled genes whose loss in tumour cells impaired the effector function of CD8
T cells. The genes that were most enriched in the screen have key roles in antigen presentation and interferon-γ signalling, and correlate with cytolytic activity in patient tumours from The Cancer Genome Atlas. Among the genes validated using different cancer cell lines and antigens, we identified multiple loss-of-function mutations in APLNR, encoding the apelin receptor, in patient tumours that were refractory to immunotherapy. We show that APLNR interacts with JAK1, modulating interferon-γ responses in tumours, and that its functional loss reduces the efficacy of adoptive cell transfer and checkpoint blockade immunotherapies in mouse models. Our results link the loss of essential genes for the effector function of CD8
T cells with the resistance or non-responsiveness of cancer to immunotherapies.
Multiple genetic mutations within melanoma not only cause lesion-specific responses to targeted therapy but also alter the molecular route of resistance to that therapy. Inactivation of PTEN occurs ...in up to 30% of melanomas, frequently with a concurrent activating BRAF mutation. PTEN loss regulates both acquired and intrinsic drug resistance. Here we show that AXL/AKT axis mediated-resistance to BRAF inhibitor (BRAFi) depends upon PTEN status in melanoma. Hyperactivation of both ERK and AKT pathways was associated with BRAFi resistance in melanoma with wildtype PTEN. The PTEN-impaired melanoma cells required only the ERK resistance mechanism. Moreover, we identified AXL as a key upstream effector of AKT pathway-associated resistance to BRAFi in melanoma with wildtype PTEN, but not in melanoma with impaired PTEN. Notably, we confirmed that blocking AXL by shRNA and a small molecular inhibitor could rescue the sensitivity of resistant melanoma cells with wildtype PTEN to BRAFi and inhibit their growth in vitro and in vivo. Our study has uncovered a mechanism by which PTEN status contributes to acquired resistance to BRAFi and offers a rational strategy to guide clinical testing in pre-identified subsets of patients who relapse during treatment with BRAFi. The identified protein AXL represents a promising therapeutic target for BRAF mutant melanoma patients with wildtype PTEN.
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 coactivator 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 phospholipase Cβ 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, a GNAQ/GNA11-initiated human malignancy.
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•The GNAQ oncogene stimulates the transcriptional coactivator YAP in uveal melanoma•A Hippo- and PLCβ-independent Rho GTPase signaling circuitry links GNAQ to YAP•YAP is essential for GNAQ-induced uveal melanoma cell proliferation•YAP represents a suitable therapeutic target for melanomas harboring GNAQ mutations
Feng et al. find that aberrant Gαq/11 activation due to oncogenic mutations leads to YAP-dependent growth in uveal melanoma. In this context, YAP stimulation is independent of PLCβ and the canonical Hippo pathway and instead acts through Trio-Rho/Rac signaling and actin polymerization.
PTEN is the second most highly mutated tumor suppressor in cancer, following only p53. The PTEN protein functions as a phosphatase with lipid- and protein-phosphatase activity. PTEN-lipid-phosphatase ...activity dephosphorylates PIP3 to form PIP2, and it then antagonizes PI3K and blocks the activation of AKT, while its protein-phosphatase activity dephosphorylates different protein substrates and plays various roles in tumorigenesis. Here, we review the PTEN mutations and protein-phosphatase substrates in tumorigenesis and metastasis. Our purpose is to clarify how PTEN protein phosphatase contributes to its tumor-suppressive functions through PI3K-independent activities.
Since their discovery in the early 2000s, microRNAs (miRNAs) and their penchant for RNA interference have taken the scientific community by storm, working their way into virtually every corner of ...biological inquiry. The very nature of their action, the ability to simultaneously extinguish the expression of a multitude of genes and negate their functions, immediately suggested therapeutic promise. In this issue of the JCI, a step toward the realization of this promise is described. Taulli et al. demonstrate that the miRNAs miR-1/miR-206, which are routinely lost in advanced, poorly differentiated rhabdomyosarcoma (RMS) but characteristically expressed in the mature skeletal muscle from which these tumors arise, restore the myogenic differentiation program and block the tumorigenic phenotype (see the related article beginning on page 2366). Their data support the notion that these small RNAs, effectively functioning as "micro-sheriffs" by restoring myogenic law and order, hold substantial clinical potential as differentiation therapy for RMS and perhaps other solid tumors. miRNA reexpression therapy constitutes a novel approach to handcuff oncogenes and arrest tumor development.
Melanoma remains mostly an untreatable fatal disease despite advances in decoding cancer genomics and developing new therapeutic modalities. Progress in patient care would benefit from additional ...predictive models germane for human disease mechanisms, tumor heterogeneity, and therapeutic responses. Toward this aim, this review documents comparative aspects of human and naturally occurring canine melanomas. Clinical presentation, pathology, therapies, and genetic alterations are highlighted in the context of current basic and translational research in comparative oncology. Somewhat distinct from sun exposure-related human cutaneous melanomas, there is growing evidence that a variety of gene copy number alterations and protein structure/function mutations play roles in canine melanomas, in circumstances more analogous to human mucosal melanomas and to some extent other melanomas with murine sarcoma viral oncogene homolog B (
), Neuroblastoma RAS Viral (V-Ras) Oncogene Homolog (
), and neurofibromin 1 tumor suppressor
triple wild-type genotype. Gaps in canine genome annotation, as well as an insufficient number and depth of sequences covered, remain considerable barriers to progress and should be collectively addressed. Preclinical approaches can be designed to include canine clinical trials addressing immune modulation as well as combined-targeted inhibition of Rat Sarcoma Superfamily/Mitogen-activated protein kinase (RAS/MAPK) and/or Phosphatidylinositol-3-Kinase/Protein Kinase B/Mammalian target of rapamycin (PI3K/AKT/mTOR) signal transduction, pathways frequently activated in both human and canine melanomas. Future investment should be aimed towards improving understanding of canine melanoma as a predictive preclinical surrogate for human melanoma and for mutually benefiting these uniquely co-dependent species.
Aberrant HGF/Met signaling promotes tumor migration, invasion, and metastasis through both autocrine and non‐autocrine mechanisms; however, the molecular downstream signaling mechanisms by which ...HGF/Met induces metastasis are incompletely understood. We here report that Ezrin expression is stimulated by HGF and correlates with activated HGF/Met, indicating that HGF/Met signaling regulates the expression of Ezrin. We show that HGF/Met signaling activates the transcription factor Sp1 through the MAPK pathway, and activated Sp1 can in turn directly bind to the promoter of Ezrin gene and regulate its transcription. Notably, knockdown of Ezrin expression by shRNAs inhibits the metastasis induced by either HGF/Met autocrine or non‐autocrine signaling in syngeneic wildtype and HGF transgenic mouse hosts. We also used small molecule drugs in preclinical mouse models to confirm that Ezrin is one of the downstream molecules mediating HGF/Met signaling‐induced metastasis in melanoma. We conclude that Ezrin is a key downstream factor involved in the regulation of HGF/Met signaling‐induced metastasis and demonstrate a link between Ezrin and HGF/Met/MAPK/Sp1 activation in the metastatic process. Our data indicate that Ezrin represents a promising therapeutic target for patients bearing tumors with activated HGF/Met signaling.
What's new?
Aberrant HGF/Met signaling promotes tumor migration, invasion, and metastasis through autocrine and non‐autocrine mechanisms; however, the molecular downstream signaling mechanisms by which HGF/Met induces metastasis remain unclear. Our study provides evidence that the cytoskeletal organizer Ezrin is a key downstream factor involved in the regulation of HGF/Met signaling‐induced metastasis and demonstrates a link between Ezrin and HGF/Met/MAPK/Sp1 activation in the metastatic process. The finding that Ezrin is located downstream of HGF/Met signaling helps explain the high metastatic potential long associated with the Met receptor and highlights Ezrin as a promising therapeutic target for patients bearing tumors with activated HGF/Met signaling.
Metastasis, a complex process, is responsible for most deaths in patients with cancer. Clinically relevant research models are indispensable to advancing our understanding of metastatic mechanisms ...and developing new treatments. We here describe detailed protocols to establish mouse models for melanoma metastasis using the single-cell imaging system and orthotropic footpad injection. The single-cell imaging system permits the tracking and quantification of early metastatic cell survival, while the orthotropic footpad transplantation mimics aspects of the complex metastatic process.
For complete details on the use and execution of this protocol, please refer to Yu et al.1,2
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•The single-cell imaging system for tracking metastatic cell survival in vivo•Tail vein injection for establishing experimental melanoma metastasis model•The orthotropic footpad injection for building up spontaneous melanoma metastasis model
Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.
Metastasis, a complex process, is responsible for most deaths in patients with cancer. Clinically relevant research models are indispensable to advancing our understanding of metastatic mechanisms and developing new treatments. We here describe detailed protocols to establish mouse models for melanoma metastasis using the single-cell imaging system and orthotropic footpad injection. The single-cell imaging system permits the tracking and quantification of early metastatic cell survival, while the orthotropic footpad transplantation mimics aspects of the complex metastatic process.
Cutaneous malignant melanoma is an aggressive cancer of melanocytes with a strong propensity to metastasize. We posit that melanoma cells acquire metastatic capability by adopting an embryonic-like ...phenotype, and that a lineage approach would uncover metastatic melanoma biology. Using a genetically engineered mouse model to generate a rich melanoblast transcriptome dataset, we identify melanoblast-specific genes whose expression contribute to metastatic competence and derive a 43-gene signature that predicts patient survival. We identify a melanoblast gene, KDELR3, whose loss impairs experimental metastasis. In contrast, KDELR1 deficiency enhances metastasis, providing the first example of different disease etiologies within the KDELR-family of retrograde transporters. We show that KDELR3 regulates the metastasis suppressor, KAI1, and report an interaction with the E3 ubiquitin-protein ligase gp78, a regulator of KAI1 degradation. Our work demonstrates that the melanoblast transcriptome can be mined to uncover targetable pathways for melanoma therapy.
Melanoma is the most severe and fatal form of skin cancer, resulting from multiple gene mutations with high intra-tumor and inter-tumor molecular heterogeneity. Treatment options for patients whose ...disease has progressed beyond the ability for surgical resection rely on currently accepted standard therapies, notably immune checkpoint inhibitors and targeted therapies. Acquired resistance to these therapies and treatment-associated toxicity necessitate exploring novel strategies, especially those that can be personalized for specific patients and/or populations. Here, we review the current landscape and progress of standard therapies and explore what personalized oncology techniques may entail in the scope of melanoma. Our purpose is to provide an up-to-date summary of the tools at our disposal that work to circumvent the common barriers faced when battling melanoma.
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