Roles of novel biomarkers in progression of cutaneous squamous cell carcinoma

Roles of novel biomarkers was studied in progression of cutaneous squamous cell carcinoma (cSCC) as the most common metastatic skin cancer. The incidence of cSCC is increasing worldwide due to lifestyle changes such as recreational exposure to sunlight and the aging of the population. Because of an emerging need for molecular markers for the progression of cSCC, we set our goal to characterize three distinct novel markers overexpressed in cSCC cells. Our results identified overexpression of serpin peptidase inhibitor clade A member 1 (SerpinA1), EphB2 and absent in melanoma 2 (AIM2) in cSCC cell lines compared with normal human epidermal keratinocytes (NHEKs). Immunohistochemical analysis of SerpinA1, EphB2 and AIM2 revealed abundant tumor cell-specific expression of cytoplasmic SerpinA1 and AIM2 and cytoplasmic and membranous EphB2 in cSCC tumors in vivo. The staining intensity of SerpinA1, EphB2 and AIM2 was significantly stronger in cSCC as compared with carcinoma in situ (cSCCIS) and actinic keratosis (AK). Tumor cell-associated SerpinA1 and EphB2 was noted in chemically induced mouse skin SCC, and the staining intensity was stronger in mouse cSCCs than in untreated skin. AIM2 staining intensity was significantly more abundant in cSCC of organ transplant recipients (OTR) than in sporadic cSCC in vivo. EphB2 knockdown resulted in inhibition of migration in cSCC cells. In addition, knockdown of EphB2 and AIM2 was found to inhibit the proliferation and invasion of cSCC cells and to delay the growth and vascularization of cSCC xenografts in vivo. Altogether, these findings identify SerpinA1 as a novel biomarker for cSCC. In addition, characterization of the roles of EphB2 and AIM2 in the progression of cSCC was implicated them as possible therapeutic targets for the treatment of cSCC particularly in unresectable and metastatic tumors.Ihon levyepiteelisyövän kehittymiseen liittyvät uudet merkkitekijät Keratinosyyttiperäinen ihon okasolusyöpä on yleisin metastasoituva ihosyöpä. Sen ilmaantuvuus kasvaa maailmanlaajuisesti elintapojen muutoksen kuten auringonvalolle altistumisen sekä väestön ikääntymisen takia. Tarvitaankin uusia merkkitekijöitä tämän syövän etenemisen ennustamiseksi. Työssämme löysimme kolme uutta merkkitekijää ja selvitimme niiden roolia ihon okasolusyövässä. Havaitsimme, että seriiniproteaasin estäjä A1 (SerpinA1), EphB2 ja absent in melanoma 2 (AIM2) ovat voimakkaasti koholla ihon okasolusyövästä eristetyissä solulinjoissa verrattuna ihmisen normaaleihin epidermaalisiin keratinosyytteihin. Tämän ohella havaitsimme immunohistokemiallisessa analyysissä SerpinA1:n, EphB2:n ja AIM2:n ilmentyvän spesifisti kasvaimen soluissa sekä ilmentyvän merkitsevästi enemmän okasolusyövissä verrattuna in situ karsinoomaan ja aktiiniseen keratoosiin in vivo. Lisäksi SerpinA1:n ja EphB2:n havaittiin olevan yliekspressoituja hiiren ihoon kemiallisesti aiheutetussa okasolusyövässä. AIM2:n värjäytymisintensiteetti oli merkitsevästi voimakkaampi elinsiirtopotilaista peräisin olevissa ihon okasolusyövissä kuin sporadisissa okasolusyövissä. EphB2:n hiljentäminen esti merkitsevästi okasolusyövästä eristettyjen solujen migraatiota. Lisäksi EphB2:n ja AIM2:n hiljentäminen syöpäsoluissa vähensi merkitsevästi solujen jakaantumista ja invaasiota sekä tuumorien vaskularisaatiota ja kasvua xenograftimallissa in vivo. Yhdessä nämä havainnot antavat aiheen otaksua, että SerpinA1 voisi toimia merkkitekijänä ihon okasolusyövässä. Lisäksi havaitsimme, että EphB2:lla ja AIM2:lla on tärkeä rooli ihon okasolusyövän kehittymisessä, joten ne voisivat mahdollisesti toimia uusina hoidon kohteina erityisesti metastaattisten ja vaikeasti kirurgisesti poistettavien ihon okasolusyöpien hoidossa.Siirretty Doriast

Complement Component C3 and Complement Factor B Promote Growth of Cutaneous Squamous Cell Carcinoma

Cutaneous squamous cell carcinoma (cSCC) is one of the most common metastatic skin cancers with increasing incidence. We examined the roles of complement component C3 and complement factor B (CFB) in the growth of cSCC. Analysis of cSCC cell lines (n = 8) and normal human epidermal kerati-nocytes (n = 11) with real-time quantitative PCR and Western blotting revealed up-regulation of C3 and CFB expression in cSCC cells. Immunohistochemical staining revealed stronger tumor cell specific Labeling for C3 and CFB in invasive cSCCs (n = 71) and recessive dystrophic epidermolysis bullosa-associated cSCCs (n = 11) than in cSCC in situ (n = 69), actinic keratoses (n = 63), and normal skin (n = 5). Significant up-regulation of C3 and CFB mRNA expression was noted in chemically induced mouse cSCCs, compared to benign papillomas. Knockdown of C3 and CFB expression inhibited migration and proliferation of cSCC cells and resulted in potent inhibition of extracellular signal regulated kinase 1/2 activation. Knockdown of C3 and CFB markedly inhibited growth of human cSCC xenograft tumors in vivo. These results provide evidence for the rotes of C3 and CFB in the development of cSCC and identify them as biomarkers and potential therapeutic targets in this metastatic skin cancer.Peer reviewe

p53-Regulated Long Noncoding RNA PRECSIT Promotes Progression of Cutaneous Squamous Cell Carcinoma via STAT3 Signaling

Long noncoding RNAs (lncRNAs) have emerged as putative biomarkers and therapeutic targets in cancer. The role of lncRNA LINC00346 in cutaneous squamous carcinoma (cSCC) was examined. The expression of LINC00346 was up-regulated in cSCC cells compared with normal human epidermal keratinocytes. Elevated expression of LINC00346 was noted in tumor cells in cSCC tissue sections in vivo, as compared with cSCC in situ, and actinic keratosis by RNA in situ hybridization; and the expression in seborrheic keratosis and normal skin was very low. Immunohistochemical analysis of cSCC tissue sections and functional assays of cSCC cells in culture showed that LINC00346 expression is down-regulated by p53. Knockdown of LINC00346 inhibited invasion of cSCC cells in culture and suppressed growth of human cSCC xenografts in vivo. Knockdown of LINC00346 inhibited expression of activated STAT3 and resulted in down-regulation of the expression of matrix metalloproteinase (MMP)-1, MMP-3, MMP-10, and MMP-13. Based on these observations LINC00346 was named p53 regulated carcinoma-associated STAT3-activating long intergenic non-protein coding transcript (PRECSIT). These results identify PRECSIT as a new p53-regulated lncRNA, which promotes progression of cSCC via STAT3 signaling.</p

Long Noncoding RNA PICSAR Promotes Growth of Cutaneous Squamous Cell Carcinoma by Regulating ERK1/2 Activity

Keratinocyte-derived cutaneous squamous cell carcinoma (cSCC) is the most common metastatic skin cancer, and its incidence is increasing globally. Long noncoding RNAs (lncRNA) are involved in various biological processes, and their role in cancer progression is emerging. Whole transcriptome analysis of cSCC cells (n = 8) and normal human epidermal keratinocytes (n = 4) revealed overexpression of long intergenic ncRNA (LINC00162) in cSCC cells. The expression of LINC00162 in cSCC cells was upregulated by inhibition of the p38α and p38δ mitogen-activated protein kinases. Analysis of tissue sections by RNA in situ hybridization showed that LINC00162 is specifically expressed by tumor cells in cSCCs but not by keratinocytes in normal skin in vivo. Knockdown of LINC00162 inhibited proliferation and migration of cSCC cells, and suppressed the growth of human cSCC xenografts in vivo. Furthermore, knockdown of LINC00162 inhibited extracellular signal-regulated kinase 1/2 activity and upregulated expression of dual specificity phosphatase 6 (DUSP6) in cSCC cells. Based on these observations, LINC00162 was named p38 inhibited cutaneous squamous cell carcinoma associated lincRNA (PICSAR). Our results provide mechanistic evidence for the role of PICSAR in promoting cSCC progression via activation of extracellular signal-regulated kinase 1/2 signaling pathway by downregulating DUSP6 expression. These results also identify PICSAR as a biomarker and putative therapeutic target in cSCC

EphB2 Promotes Progression of Cutaneous Squamous Cell Carcinoma

Keratinocyte-derived skin cancer, cutaneous squamous cell carcinoma (cSCC), is the most common metastatic skin cancer. We have examined the role of Eph/ephrin signaling in the progression of cSCC. Analysis of the expression of EPH and EFN families in cSCC cells and normal epidermal keratinocytes revealed overexpression of EPHB2 mRNA in cSCC cells and cSCC tumors in vivo. Tumor cell–specific overexpression of EphB2 was detected in human cSCCs and in chemically induced mouse cSCCs with immunohistochemistry, whereas the expression of EphB2 was low in premalignant lesions and normal skin. Knockdown of EphB2 expression in cSCC cells suppressed growth and vascularization of cSCC xenografts in vivo and inhibited proliferation, migration, and invasion of cSCC cells in culture. EphB2 knockdown downregulated expression of genes associated with biofunctions cell viability, migration of tumor cells, and invasion of tumor cells. Among the genes most downregulated by EphB2 knockdown were MMP1 and MMP13. Moreover, activation of EphB2 signaling by ephrin-B2-Fc enhanced production of invasion proteinases matrix metalloproteinase-13 (MMP13) and MMP1, and invasion of cSCC cells. These findings provide mechanistic evidence for the role of EphB2 in the early progression of cSCC to the invasive stage and identify EphB2 as a putative therapeutic target in this invasive skin cancer

Identification of Rigosertib for the Treatment of Recessive Dystrophic Epidermolysis Bullosa–Associated Squamous Cell Carcinoma

PURPOSE: Squamous cell carcinoma (SCC) of the skin is the leading cause of death in patients with the severe generalized form of the genetic disease recessive dystrophic epidermolysis bullosa (RDEB). Although emerging data are identifying why patients suffer this fatal complication, therapies for treatment of RDEB SCC are in urgent need. EXPERIMENTAL DESIGN: We previously identified polo-like kinase 1 (PLK1) as a therapeutic target in skin SCC, including RDEB SCC. Here, we undertake a screen of 6 compounds originally designated as PLK1 inhibitors, and detail the efficacy of the lead compound, the multipathway allosteric inhibitor ON-01910, for targeting RDEB SCC in vitro and in vivo. RESULTS: ON-01910 (or rigosertib) exhibited significant specificity for RDEB SCC: in culture rigosertib induced apoptosis in 10 of 10 RDEB SCC keratinocyte populations while only slowing the growth of normal primary skin cells at doses 2 orders of magnitude higher. Furthermore, rigosertib significantly inhibited the growth of two RDEB SCC in murine xenograft studies with no apparent toxicity. Mechanistically, rigosertib has been shown to inhibit multiple signaling pathways. Comparison of PLK1 siRNA with MEK inhibition, AKT inhibition, and the microtubule-disrupting agent vinblastine in RDEB SCC shows that only PLK1 reduction exhibits a similar sensitivity profile to rigosertib. CONCLUSIONS: These data support a “first in RDEB” phase II clinical trial of rigosertib to assess tumor targeting in patients with late stage, metastatic, and/or unresectable SCC

Cutaneous Manifestations in 404 Iranian Patients With Inflammatory Bowel Disease: A Retrospective Study

Background: Cutaneous manifestations of inflammatory bowel disease are relatively common, although they vary widely. Aims: The aim of this study was to determine the prevalence of cutaneous manifestations in inflammatory bowel disease according to their location, age, gender, activity, and type of underlying disease in an Iranian population during a 10-year period. Methods: The medical records of 404 inpatients with inflammatory bowel disease were extracted retrospectively to detect cutaneous manifestations. Results: In this study, the prevalence of cutaneous manifestations was 5.9%. These manifestations were higher in Crohn′s disease (7.29%) than in ulcerative colitis (4.07%), and more frequent in females (52%) than in males (48%). Aphthous stomatitis was observed more frequently in Crohn′s disease; however, pyoderma gangrenosum was seen more often in ulcerative colitis. Erythema nodosum was diagnosed only in female patients with Crohn′s disease. Manifestations secondary to nutritional deficiency or associated conditions including psoriasis and other autoimmune disorders were less frequent. Conclusions: Aphthous stomatitis, pyoderma gangrenosum, and erythema nodosum were the most common skin disorders related to inflammatory bowel disease (IBD), which mainly occurred in women

Inhibition of c-Abl Kinase Activity Renders Cancer Cells Highly Sensitive to Mitoxantrone

<div><p>Although c-Abl has increasingly emerged as a key player in the DNA damage response, its role in this context is far from clear. We studied the effect of inhibition of c-Abl kinase activity by imatinib with chemotherapy drugs and found a striking difference in cell survival after combined mitoxantrone (MX) and imatinib treatment compared to a panel of other chemotherapy drugs. The combinatory treatment induced apoptosis in HeLa cells and other cancer cell lines but not in primary fibroblasts. The difference in MX and doxorubicin was related to significant augmentation of DNA damage. Transcriptionally active p53 accumulated in cells in which human papillomavirus E6 normally degrades p53. The combination treatment resulted in caspase activation and apoptosis, but this effect did not depend on either p53 or p73 activity. Despite increased p53 activity, the cells arrested in G2 phase became defective in this checkpoint, allowing cell cycle progression. The effect after MX treatment depended partially on c-Abl: Short interfering RNA knockdown of c-Abl rendered HeLa cells less sensitive to MX. The effect of imatinib was decreased by c-Abl siRNA suggesting a role for catalytically inactive c-Abl in the death cascade. These findings indicate that MX has a unique cytotoxic effect when the kinase activity of c-Abl is inhibited. The treatment results in increased DNA damage and c-Abl–dependent apoptosis, which may offer new possibilities for potentiation of cancer chemotherapy.</p></div

Collagen VII maintains proteostasis in dermal fibroblasts by scaffolding TANGO1 cargo

Lack of type VII collagen (C7) disrupts cellular proteostasis yet the mechanism remains undescribed. By studying the relationship between C7 and the extracellular matrix (ECM)-associated proteins thrombospondin-1 (TSP1), type XII collagen (C12) and tissue transglutaminase (TGM2) in primary human dermal fibroblasts from multiple donors with or without the genetic disease recessive dystrophic epidermolysis bullosa (RDEB) (n=31), we demonstrate that secretion of each of these proteins is increased in the presence of C7. In dermal fibroblasts isolated from patients with RDEB, where C7 is absent or defective, association with the COPII outer coat protein SEC31 and ultimately secretion of each of these ECM-associated proteins is reduced and intracellular levels are increased. In RDEB fibroblasts, overall collagen secretion (as determined by the levels of hydroxyproline in the media) is unchanged while traffic from the ER to Golgi of TSP1, C12 and TGM2 occurs in a type I collagen (C1) dependent manner. In normal fibroblasts association of TSP1, C12 and TGM2 with the ER exit site transmembrane protein Transport ANd Golgi Organization-1 (TANGO1) as determined by proximity ligation assays, requires C7. In the absence of wild-type C7, or when ECM-associated proteins are overexpressed, C1 proximity and intracellular levels increase resulting in elevated cellular stress responses and elevated TGFβ signaling. Collectively, these data demonstrate a role for C7 in loading COPII vesicle cargo and provides a mechanism for disrupted proteostasis, elevated cellular stress and increased TGFβ signaling in patients with RDEB. Furthermore, our data point to a threshold of cargo loading that can be exceeded with increased protein levels leading to pathological outcomes in otherwise normal cells