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Cutaneous skeletal hypophosphatemia syndrome (CSHS) is a mosaic RA Sopathy characterized by the association of dysplastic skeletal lesions, congenital skin nevi of epidermal and/or melanocytic origin, and FGF23-mediated hypophosphatemia. The primary physiological source of circulating FGF23 is bone cells. However, several reports have suggested skin lesions as the source of excess FGF23 in CSHS. Consequently, without convincing evidence of efficacy, many patients with CSHS have undergone painful removal of cutaneous lesions in an effort to normalize blood phosphate levels. This study aims to elucidate whether the source of FGF23 excess in CSHS is RAS mutation-bearing bone or skin lesions. Toward this end, we analyzed the expression and activity of Fgf23 in two mouse models expressing similar HRAS/Hras activating mutations in a mosaic-like fashion in either bone or epidermal tissue. We found that HRAS hyperactivity in bone, not skin, caused excess of bioactive intact FGF23, hypophosphatemia, and osteomalacia. Our findings support RAS- mutated dysplastic bone as the primary source of physiologically active FGF23 excess in patients with CSHS. This evidence informs the care of patients with CSHS, arguing against the practice of nevi removal to decrease circulating, physiologically active FGF23., Introduction Cutaneous skeletal hypophosphatemia syndrome (CSHS) is characterized by the association of dysplastic skeletal lesions, congenital skin nevi of epidermal and/or melanocytic origin (including epidermal nevi [EN], giant congenital melanocytic [...]

The Ras–mitogen-activated protein kinase (MAPK) pathway is a major target for cancer treatment. To better understand the genetic pathways that modulate cancer cell sensitivity to MAPK pathway inhibitors, we performed a CRISPR knockout screen with MAPK pathway inhibitors on a colorectal cancer (CRC) cell line carrying mutant KRAS. Genetic deletion of the catalytic subunit of protein phosphatase 6 (PP6), encoded by PPP6C, rendered KRAS- and BRAF-mutant CRC and BRAF-mutant melanoma cells more resistant to these inhibitors. In the absence of MAPK pathway inhibition, PPP6C deletion in CRC cells decreased cell proliferation in two-dimensional (2D) adherent cultures but accelerated the growth of tumor spheroids in 3D culture and tumor xenografts in vivo. PPP6C deletion enhanced the activation of nuclear factor κB (NF-κB) signaling in CRC and melanoma cells and circumvented the cell cycle arrest and decreased cyclin D1 abundance induced by MAPK pathway blockade in CRC cells. Inhibiting NF-κB activity by genetic and pharmacological means restored the sensitivity of PPP6C-deficient cells to MAPK pathway inhibition in CRC and melanoma cells in vitro and in CRC cells in vivo. Furthermore, a R264 point mutation in PPP6C conferred loss of function in CRC cells, phenocopying the enhanced NF-κB activation and resistance to MAPK pathway inhibition observed for PPP6C deletion. These findings demonstrate that PP6 constrains the growth of KRAS- and BRAF-mutant cancer cells, implicates the PP6–NF-κB axis as a modulator of MAPK pathway output, and presents a rationale for cotargeting the NF-κB pathway in PPP6C-mutant cancer cells. Editor's summary: Inhibitors of the Ras-MAPK pathway are used to treat cancers carrying RAS and BRAF mutations. Zhang et al. performed a CRISPR-based screen to find modulators of the MAPK pathway and found that protein phosphatase 6 (PP6) increased sensitivity to MAPK pathway inhibitors. Deletion of the catalytic subunit of PP6 promoted the growth of KRAS- and BRAF-mutant cells in spheroid cultures and in mice in an NF-κB–dependent manner and increased their resistance to MAPK pathway inhibitors. These findings indicate that PP6 contributes to MAPK sensitivity in KRAS- and BRAF-mutant cells and suggest that the NF-κB pathway may be cotargeted in PP6C-mutant cells. —Amy E. Baek [ABSTRACT FROM AUTHOR]

Mouse models of breast cancer have revealed that tumour‐bearing hosts must express the oxidoreductase CLIC4 to develop lung metastases. In the absence of host CLIC4, primary tumours grow but the lung premetastatic niche is defective for metastatic seeding. Primary breast cancer cells release EVs that incorporate CLIC4 as cargo and circulate in plasma of wildtype tumour‐bearing hosts. CLIC4‐deficient breast cancer cells also form tumours in wildtype hosts and release EVs in plasma, but these EVs lack CLIC4, suggesting that the tumour is the source of the plasma‐derived EVs that carry CLIC4 as cargo. Paradoxically, circulating EVs are also devoid of CLIC4 when CLIC4‐expressing primary tumours are grown in CLIC4 knockout hosts. Thus, the incorporation of CLIC4 (and perhaps other factors) as EV cargo released from tumours involve specific signals from the surrounding stroma determined by its genetic composition. Since CLIC4 is also detected in circulating EVs from human breast cancer patients, future studies will address its association with disease. [ABSTRACT FROM AUTHOR]

Environmental and molecular carcinogenesis are linked by the discovery that chemical carcinogen induced-mutations in the Hras or Kras genes drives tumor development in mouse skin. Importantly, enhanced expression or allele amplification of the mutant Ras gene contributes to selection of initiated cells, tumor persistence, and progression. To explore the consequences of Ras oncogene signal strength, primary keratinocytes were isolated and cultured from the LSL-Hras G12D and LSL-Kras G12D C57BL/6J mouse models and the mutant allele was activated by adeno-Cre recombinase. Keratinocytes expressing one (H) or two (HH) mutant alleles of Hras G12D , one Kras G12D allele (K), or one of each (HK) were studied. All combinations of activated Ras alleles stimulated proliferation and drove transformation marker expression, but only HH and HK formed tumors. HH, HK, and K sustained long-term keratinocyte growth in vitro , while H and WT could not. RNA-Seq yielded two distinct gene expression profiles; HH, HK, and K formed one cluster while H clustered with WT. Weak MAPK activation was seen in H keratinocytes but treatment with a BRAF inhibitor enhanced MAPK signaling and facilitated tumor formation. K keratinocytes became tumorigenic when they were isolated from mice where the LSL-Kras G12D allele was backcrossed from the C57BL/6 onto the FVB/N background. All tumorigenic keratinocytes but not the non-tumorigenic precursors shared a common remodeling of matrisomal gene expression that is associated with tumor formation. Thus, RAS oncogene signal strength determines cell-autonomous changes in initiated cells that are critical for their tumor-forming potential. [ABSTRACT FROM AUTHOR]

Transgenic mice overexpressing PKC[alpha] in the epidermis (K5-PKC[alpha] mice) exhibit an inducible severe intraepidermal neutrophilic inflammation and systemic neutrophilia when PKC[alpha] is activated by topical 12-O-tetradecanoylphorbol-13-acetate (TPA). This inducible model [...]

The TGF-β-regulated Chloride Intracellular Channel 4 (CLIC4) is an essential participant in the formation of breast cancer stroma. Here, we used data available from the TCGA and METABRIC datasets to show that CLIC4 expression was higher in breast cancers from younger women and those with early-stage metastatic disease. Elevated CLIC4 predicted poor outcome in breast cancer patients and was linked to the TGF-β pathway. However, these associations did not reveal the underlying biological contribution of CLIC4 to breast cancer progression. Constitutive ablation of host Clic4 in two murine metastatic breast cancer models nearly eliminated lung metastases without reducing primary tumor weight, while tumor cells ablated of Clic4 retained metastatic capability in wildtype hosts. Thus, CLIC4 was required for host metastatic competence. Pre- and post-metastatic proteomic analysis identified circulating pro-metastatic soluble factors that differed in tumor-bearing CLIC4-deficient and wildtype hosts. Vascular abnormalities and necrosis increased in primary tumors from CLIC4-deficient hosts. Transcriptional profiles of both primary tumors and pre-metastatic lungs of tumor-bearing CLIC4-deficient hosts were consistent with a microenvironment where inflammatory pathways were elevated. Altogether, CLIC4 expression in human breast cancers may serve as a prognostic biomarker; therapeutic targeting of CLIC4 could reduce primary tumor viability and host metastatic competence. Author summary: CLIC4, a member of a highly conserved CLIC gene family, contributes to the pathogenesis of several human diseases. We show that elevated CLIC4 expression in human breast cancers predicts early invasion and poor outcome in women. Further, in murine models of breast cancer, CLIC4 in the host microenvironment is required for lung metastasis. In the absence of host CLIC4, the microenvironment of the primary tumor and the pre-metastatic lungs is unfavorable for tumor viability and lung colonization. These findings introduce CLIC4 as a potential therapeutic target against a host protein to prevent metastatic spread in breast cancer. [ABSTRACT FROM AUTHOR]