Mehdi Farshchian

Serpin Peptidase Inhibitor Clade A Member 1 (SerpinA1) Is a Novel Biomarker for Progression of Cutaneous Squamous Cell Carcinoma

The incidence of keratinocyte-derived nonmelanoma skin cancers is increasing worldwide because of cumulative recreational exposure to sunlight. At present, no specific molecular markers are available for assessing the progression of premalignant actinic keratoses to invasive cutaneous squamous cell carcinoma (SCC). We examined the role of the Serpin family in skin SCCs. Expression profiling of cutaneous SCC cell lines (n = 8) revealed up-regulation of SerpinA1 compared with normal epidermal keratinocytes (n = 5). Analysis with quantitative RT-PCR showed that the mean level of SerpinA1 mRNA was markedly up-regulated in cutaneous SCC cell lines (n = 8) compared with in normal keratinocytes. SerpinA1 production by SCC cells was dependent on p38 mitogen-activated protein kinase activity and was up-regulated by epidermal growth factor, tumor necrosis factor-α, interferon-γ, and IL-1β. Immunostaining of tissue arrays with 148 human tissue samples revealed tumor cell-associated expression of SerpinA1 in 19 of 36 actinic keratoses, 22 of 29 Bowens disease samples, 67 of 71 sporadic SCCs, and all 12 recessive dystrophic epidermolysis bullosa-associated SCCs examined. Moreover, tumor cell-associated SerpinA1 staining was detected in all chemically induced mouse skin SCCs studied (n = 17). Overexpression of SerpinA1 mRNA was also detected by quantitative RT-PCR in chemically induced mouse skin SCCs (n = 14) compared with control tissues (n = 14). These data identify SerpinA1 as a novel tumor cell-associated biomarker for progression of cutaneous SCCs.

Complement Factor I Promotes Progression of Cutaneous Squamous Cell Carcinoma

The incidence of cutaneous squamous cell carcinoma (cSCC) is rising worldwide. We have examined the role of complement components in the progression of cSCC. Analysis of cSCC cell lines (n=8) and normal human epidermal keratinocytes (n=11) with whole transcriptome profiling (SOLiD), quantitative real-time reverse transcriptase-PCR, and western blotting revealed marked overexpression of complement factor I (CFI) in cSCC cells. Immunohistochemical analysis for CFI in vivo showed stronger tumor cell-specific labeling intensity in invasive sporadic cSCCs (n=83) and recessive dystrophic epidermolysis bullosa-associated cSCCs (n=7) than in cSCC in situ (n=65), premalignant epidermal lesions (actinic keratoses, n=64), benign epidermal papillomas (seborrheic keratoses, n=39), and normal skin (n=9). The expression of CFI was higher in the aggressive Ha-ras-transformed cell line (RT3) than in less tumorigenic HaCaT cell lines (HaCaT, A5, and II-4). The expression of CFI by cSCC cells was upregulated by IFN-γ and IL-1β. Knockdown of CFI expression inhibited proliferation and migration of cSCC cells and resulted in inhibition of basal extracellular signal-regulated kinase (ERK) 1/2 activation. Knockdown of CFI expression potently inhibited growth of human cSCC xenograft tumors in vivo. These results provide evidence for the role of CFI in the progression of cSCC and identify it as a potential therapeutic target in this nonmelanoma skin cancer.

Treatment of common warts with an intralesional mixture of 5-fluorouracil, lidocaine, and epinephrine: a prospective placebo-controlled, double-blind randomized trial.

BACKGROUND 5-Fluorouracil is an antimetabolite that has been known to be effective for the treatment of common warts. OBJECTIVE The objective was to evaluate the efficacy of a combination of 5-fluorouracil, lidocaine, and epinephrine (5-FU+LE) for the treatment of common warts. MATERIALS AND METHODS Of the 40 patients initially enrolled, 34 patients with at least 68 verruca vulgaris (one pair for each subject) completed the study. The selected warts were randomized into two treatment groups, with one wart on each patient receiving intralesional 5-FU+LE (4 mL of 50 mg/mL 5-fluorouracil and 1 mL of a mixture of 20 mg/mL [2%] lidocaine and 0.0125 mg/mL epinephrine) and the other receiving intralesional normal saline placebo using a Mantoux needle. Patients received up to four injections at weekly intervals and were followed at 1 and 6 months after the initial injection. RESULTS Complete response was observed in 64.7% of the warts treated with 5-FU+LE compared to 35.3% in the warts treated with placebo (p<.05). Systemic reaction or treatment-related adverse medical events and recurrence rate did not differ significantly between two groups. CONCLUSION The results show that intralesional 5-FU+LE is a safe and effective approach for common warts with high success rate.

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.

New perspectives on role of tumor microenvironment in progression of cutaneous squamous cell carcinoma

AbstractEpidermal keratinocyte-derived cutaneous squamous cell carcinoma (cSCC) is the most common metastatic skin cancer, and its incidence is increasing worldwide. Solar UV radiation is an important risk factor for cSCC and leads to genetic and epigenetic changes both in epidermal keratinocytes and dermal cells. Tumor cells in cutaneous cSCCs typically harbor several driver gene mutations, but epidermal keratinocytes in sun-exposed normal skin also contain mutations in these same genes. Therefore, alterations in the microenvironment of premalignant lesions are evidently required for their progression to invasive and metastatic cSCC. For example, alterations in the composition of basement membrane and dermal extracellular matrix are early events in cSCC progression. The presence of microbial structures and the influx of inflammatory cells promote the secretion of proteases, which in turn regulate the availability of growth factors, cytokines, and chemokines and thus influence the growth and invasion of cSCC. Together, these observations emphasize the role of the tumor microenvironment in the progression of cSCC and identify it as a novel therapeutic target in cSCC and other malignant tumors. Graphical abstractTumor–stroma interactions in the progression of cutaneous squamous cell carcinoma (cSCC). Epidermal layer is separated by a well-organized basement membrane (BM) from the dermal layer. UV radiation, other environmental insults, and aging target both epidermal keratinocytes and dermal fibroblasts and lead to genetic and epigenetic changes in these cells. In addition, epidermal keratinocytes in normal sun-exposed skin harbor several mutations in the cSCC driver genes. During transition to premalignant actinic keratosis (AK), the differentiation of keratinocytes is disturbed resulting in a neoplastic epithelium with hyperplastic cells. Expression of proteinases, such as matrix metalloproteinases (MMP) by neoplastic cells and activated stromal fibroblasts and macrophages is induced in AK, and collagen XV and XVIII are lost from the dermal BM. Furthermore, inflammatory cells accumulate at the site of the hyperplastic epithelium. During a later stage of cSCC progression, the number of inflammatory cells increases, and the expression of complement components and inhibitors by tumor cells is induced (CFI complement factor I, CFH complement factor H, FHL-1 Factor H-like protein 1). In addition to MMPs, activated fibroblasts also produce growth factors and promote inflammation, growth, and invasion of tumor cells

Tumor cell-specific AIM2 regulates growth and invasion of cutaneous squamous cell carcinoma.

Cutaneous squamous cell carcinoma (cSCC) is the most common metastatic skin cancer. Inflammation is a typical feature in cSCC progression. Analysis of the expression of inflammasome components in cSCC cell lines and normal human epidermal keratinocytes revealed upregulation of the expression of AIM2 mRNA and protein in cSCC cells. Elevated levels of AIM2 mRNA were noted in cSCCs in vivo compared with normal skin. Strong and moderate tumor cell specific expression of AIM2 was detected with immunohistochemistry (IHC) in sporadic human cSCCs in vivo, whereas expression of AIM2 was moderate in cSCC in situ (cSCCIS) and low or absent in actinic keratosis (AK) and normal skin. IHC of cSCCs, cSCCIS and AKs from organ transplant recipients also revealed strong and moderate tumor cell specific expression of AIM2 in cSCCs. Knockdown of AIM2 resulted in reduction in viability of cSCC cells and onset of apoptosis. RNA-seq and pathway analysis after knockdown of AIM2 in cSCC cells revealed downregulation of the biofunction category Cell cycle and upregulation of the biofunction category Cell Death and Survival. Knockdown of AIM2 also resulted in reduction in invasion of cSCC cells and downregulation in production of invasion proteinases MMP1 and MMP13. Knockdown of AIM2 resulted in suppression of growth and vascularization of cSCC xenografts in vivo. These results provide evidence for the role of AIM2 in the progression of cSCC and identify AIM2 inflammasome function as a potential therapeutic target in these invasive and metastatic tumors.

Dasatinib promotes apoptosis of cutaneous squamous carcinoma cells by regulating activation of ERK1/2

Cutaneous squamous cell carcinoma (cSCC) is the most common metastatic skin cancer among White population.1 Treatment of patients with unresectable and metastatic cSCC is challenging in the absence of targeted therapies, and the metastatic disease is associated with poor prognosis.2,3 Tyrosine kinases have been identified as desirable targets for cancer therapy. Imatinib mesylate, an inhibitor of multiple tyrosine kinases, is indicated for the therapy of chronic myeloid leukaemia (CML) and gastrointestinal stromal tumors.4 A secondgeneration tyrosine kinase inhibitor, dasatinib, is an inhibitor of BCRABL and Src family kinases, indicated for the therapy of imatinibresistant CML.5 Recent studies have demonstrated the effect of dasatinib in the treatment of solid cancers alone or in combination with other antineoplastic drugs, but the efficacy of dasatinib in the treatment of cSCC is not known.6 Eph receptors represent the largest family of receptor tyrosine kinases. Eph receptors and their ligands, ephrins, play an important role in various biological processes involving cell migration, for example organ development, tissue remodelling, axon guidance and angiogenesis, as well as in progression of cancer.7 We have recently reported that EphB2 promotes growth and invasion of cSCC cells in culture and in human cSCC xenografts in vivo, identifying EphB2 as a putative therapeutic target in cSCC.8 EphB2 has been listed as one of putative targets for dasatinib, but the effect of dasatinib on endogenous EphB2 is not known.9

Drug-induced skin reactions: a 2-year study

Background The aim of this study was to analyze the clinical characteristics of patients with adverse cutaneous drug reactions, which occur when a medicinal product results in cutaneous morbidity. Methods The study included 308 patients who were diagnosed as having an adverse cutaneous drug reaction during the study period (2007–2009). In 84 cases, histopathologic examination of skin biopsies were also performed. Results Patients with drug reactions were found to be more commonly female (63%) than male (37%). Beta-lactam antibiotics were found to be the most frequent cause of adverse cutaneous drug reactions (42.7%), followed by non-steroidal anti-inflammatory drugs (16.5%). Acute urticaria was the most common clinical presentation (59.2%) followed by fixed drug eruptions (18.5%), and maculopapular eruptions (14.9%). Conclusion Adverse cutaneous drug reactions in our study population were mainly induced by beta-lactam antibiotics and non-steroidal anti-inflammatory drugs. The most common forms of cutaneous adverse drug reactions were found to be acute urticaria, fixed drug eruptions, and maculopapular rashes.

Expression of claudin-11 by tumor cells in cutaneous squamous cell carcinoma is dependent on the activity of p38δ

The incidence of cutaneous squamous cell carcinoma (cSCC) is rapidly increasing, and the prognosis of patients with metastatic disease is poor. There is an emerging need to identify molecular markers for predicting aggressive behaviour of cSCC. Here, we have examined the role of tight junction (TJ) components in the progression of cSCC. The expression pattern of mRNAs for TJ components was determined with RNA sequencing and oligonucleotide array‐based expression analysis from cSCC cell lines (n=8) and normal human epidermal keratinocytes (NHEK, n=5). The expression of CLDN11 was specifically elevated in primary cSCC cell lines (n=5), but low or absent in metastatic cSCC cell lines (n=3) and NHEKs. Claudin‐11 was detected in cell‐cell contacts of primary cSCC cells in culture by indirect immunofluorescence analysis. Analysis of a large panel of tissue samples from sporadic UV‐induced cSCC (n=65), cSCC in situ (n=56), actinic keratoses (n=31), seborrhoeic keratoses (n=7) and normal skin (n=16) by immunohistochemistry showed specific staining for claudin‐11 in intercellular junctions of keratinizing tumor cells in well and moderately differentiated cSCCs, whereas no staining for claudin‐11 was detected in poorly differentiated tumors. The expression of claudin‐11 in cSCC cells was dependent on the activity of p38δ MAPK and knock‐down of claudin‐11 enhanced cSCC cell invasion. These findings provide evidence for the role of claudin‐11 in regulation of cSCC invasion and suggest loss of claudin‐11 expression in tumor cells as a biomarker for advanced stage of cSCC.

Abstract 3201: Complement component C3 and complement factor B regulate growth of cutaneous squamous cell carcinoma

Cutaneous squamous cell carcinoma (cSCC) is the most common metastatic skin cancer and its incidence is increasing globally. We have examined the role of complement component C3 and complement factor B (CFB) in the progression of cSCC. Significantly elevated mRNA levels for C3 and CFB were detected din cSCC cell lines (n = 8) compared to normal human epidermal keratinocytes (n = 11) by quantitative real-time reverse transcriptase-PCR. Increased production of C3 and CFB by cSCC cell lines was detected by western blotting. The mRNA levels for C3 and CFB were also markedly higher in cSCC tumors (n = 6) than in normal skin (n = 10). Tumor cell specific staining for C3 and CFB was detected in cSCC tumors in vivo by immunohistochemistry and the staining intensity was stronger in invasive cSCCs (n = 71) than in cSCC in situ (n = 69), premalignant epidermal lesions (actinic keratoses, n = 65) and normal skin (n = 5). Significant upregulation of C3 and CFB mRNA expression was also noted in chemically induced mouse skin cSCCs (n = 27) compared to benign papillomas (n = 17). The expression of C3 and CFB at mRNA and protein level was markedly higher in invasive tumorigenic Ha-ras-transformed HaCaT cell line than in benign tumorigenic ras-transformed HaCaT cells and parental non-tumorigenic HaCaT cells. The expression of both C3 and CFB was significantly upregulated by IFN-γ and TNF-α in cSCC cells. Knockdown of CFB and C3 by specific siRNAs inhibited migration of cSCC cells. Knockdown of CFB inhibited proliferation of cSCC cells and this was associated with potent inhibition of ERK1/2 activation. Knockdown of C3 and CFB also significantly inhibited growth of human cSCC xenograft tumors in vivo in SCID mice. These results provide evidence for the role of C3 and CFB in progression of cSCC and identify them as specific biomarkers and putative therapeutic targets for invasive cSCC. Citation Format: Pilvi Riihila, Mehdi Farshchian, Markku Kallajoki, Atte Kivisaari, Seppo Meri, Reidar Grenman, Ritva Heljasvaara, Taina Pihlajaniemi, Juha Peltonen, Sirkku Peltonen, Veli-Matti Kahari. Complement component C3 and complement factor B regulate growth of cutaneous squamous cell carcinoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3201. doi:10.1158/1538-7445.AM2015-3201