A431 Squamous Cell Carcinoma Research Articles

Potential Antitumor Mechanism of Propolis Against Skin Squamous Cell Carcinoma A431 Cells Based on Untargeted Metabolomics.

Propolis is a sticky substance produced by honeybees (Apis mellifera) through the collection of plant resins, which they mix with secretions from their palate and wax glands. Propolis can inhibit tumor invasion and metastasis, thereby reducing the proliferation of tumor cells and inducing cell apoptosis. Previous research has shown that propolis has an inhibitory effect on skin squamous cell carcinoma A431 cells. Nevertheless, its inhibitory mechanism is unclear because of many significantly different Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways between the ethanol extract of the propolis (EEP) group and the control group of cells. In this study, the main components of EEP and the antitumor mechanism at an IC50 of 29.04 μg/mL EEP were determined via untargeted metabolomics determined using ultra high-performance liquid chromatography tandem mass spectrometry (UHPLC‒MS/MS), respectively. The results revealed 43 polyphenolic components in the EEP and 1052 metabolites, with 160 significantly upregulated and 143 significantly downregulated metabolites between cells treated with EEP and solvent. The KEGG enrichment results revealed that EEP significantly inhibited A431 cell proliferation via the steroid hormone biosynthesis and linoleic acid metabolism pathways. These findings may provide valuable insights for the development of targeted therapies for the treatment of cutaneous squamous cell carcinoma.

Cold plasma irradiation inhibits skin cancer via ferroptosis

Cold atmospheric plasma (CAP) has been extensively utilized in medical treatment, particularly in cancer therapy. However, the underlying mechanism of CAP in skin cancer treatment remains elusive. In this study, we established a skin cancer model using CAP treatmentin vitro. Also, we established the Xenograft experiment modelin vivo. The results demonstrated that treatment with CAP induced ferroptosis, resulting in a significant reduction in the viability, migration, and invasive capacities of A431 squamous cell carcinoma, a type of skin cancer. Mechanistically, the significant production of reactive oxygen species (ROS) by CAP induces DNA damage, which then activates Ataxia-telangiectasia mutated (ATM) and p53 through acetylation, while simultaneously suppressing the expression of Solute Carrier Family 7 Member 11 (SLC7A11). Consequently, this cascade led to the down-regulation of intracellular Glutathione peroxidase 4 (GPX4), ultimately resulting in ferroptosis. CAP exhibits a favorable impact on skin cancer treatment, suggesting its potential medical application in skin cancer therapy.

VDR and PDIA3 Are Essential for Activation of Calcium Signaling and Membrane Response to 1,25(OH)2D3 in Squamous Cell Carcinoma Cells.

The genomic activity of 1,25(OH)2D3 is mediated by vitamin D receptor (VDR), whilst non-genomic is associated with protein disulfide isomerase family A member 3 (PDIA3). Interestingly, our recent studies documented that PDIA3 is also involved, directly or indirectly, in the modulation of genomic response to 1,25(OH)2D3. Moreover, PDIA3 was also shown to regulate cellular bioenergetics, possibly through the modulation of STAT signaling. Here, the role of VDR and PDIA3 proteins in membrane response to 1,25(OH)2D3 and calcium signaling was investigated in squamous cell carcinoma A431 cell line with or without the deletion of VDR and PDIA3 genes. Calcium influx was assayed by Fura-2AM or Fluo-4AM, while calcium-regulated element (NFAT) activation was measured using a dual luciferase assay. Further, the levels of proteins involved in membrane response to 1,25(OH)2D3 in A431 cell lines were analyzed via Western blot analysis. The deletion of either PDIA3 or VDR resulted in the decreased baseline levels of Ca2+ and its responsiveness to 1,25(OH)2D3; however, the effect was more pronounced in A431∆PDIA3. Furthermore, the knockout of either of these genes disrupted 1,25(OH)2D3-elicited membrane signaling. The data presented here indicated that the VDR is essential for the activation of calcium/calmodulin-dependent protein kinase II alpha (CAMK2A), while PDIA3 is required for 1,25(OH)2D3-induced calcium mobilization in A431 cells. Taken together, those results suggest that both VDR and PDIA3 are essential for non-genomic response to this powerful secosteroid.

Dissection of an impact of VDR and RXRA on the genomic activity of 1,25(OH)2D3 in A431 squamous cell carcinoma

BackgroundHuman skin is the natural source, place of metabolism, and target for vitamin D3. The classical active form of vitamin D3, 1,25(OH)2D3, expresses pluripotent properties and is intensively studied in cancer prevention and therapy. To define the specific role of vitamin D3 receptor (VDR) and its co-receptor retinoid X receptor alpha (RXRA) in genomic regulation, VDR or RXRA genes were silenced in the squamous cell carcinoma cell line A431 and treated with 1,25(OH)2D3 at long incubation time points 24 h/72 h. Extending the incubation time of A431 WT (wild-type) cells with 1,25(OH)2D3 resulted in a two-fold increase in DEGs (differentially expressed genes) and a change in the amount of downregulated from 37% to 53%. VDR knockout led to a complete loss of 1,25(OH)2D3-induced genome-wide gene regulation at 24 h time point, but after 72 h, 20 DEGs were found, of which 75% were downregulated, and most of them belonged to the gene ontology group “immune response”. This may indicate the existence of an alternative, secondary response to 1,25(OH)2D3. In contrast, treatment of A431 ΔRXRA cells with 1,25(OH)2D3 for 24 h only partially affected DEGs, suggesting RXRA-independent regulation. Interestingly, overexpression of classic 1,25(OH)2D3 targets, like CYP24A1 (family 24 of subfamily A of cytochrome P450 member 1) or CAMP (cathelicidin antimicrobial peptide) was found to be RXRA-independent. Also, immunofluorescence staining of A431 WT cells revealed partial VDR/RXRA colocalization after 24 h and 72 h 1,25(OH)2D3 treatment.Comparison of transcriptome changes induced by 1,25(OH)2D3 in normal keratinocytes vs. cancer cells showed high cell type specific expression pattern with only a few genes commonly regulated by 1,25(OH)2D3. Activation of the genomic pathway at least partially reversed the expression of cancer-related genes, forming a basis for anti-cancer activates of 1,25(OH)2D3. In summary, VDR or RXRA independent genomic activities of 1,25(OH)2D3 suggest the involvement of alternative factors, opening new challenges in this field.

Multifaceted approach toward mapping out the anticancer properties of small molecules via invitro evaluation on melanoma and nonmelanoma skin cancer cells, and in silico target fishing.

Melanoma and nonmelanoma skin cancers are among the most prevalent and most lethal forms of skin cancers. To identify new lead compounds with potential anticancer properties for further optimization, invitro assays combined with in-silico target fishing and docking have been used to identify and further map out the antiproliferative and potential mode of action of molecules from a small library of compounds previously prepared in our laboratory. From screening these compounds invitro against A375, SK-MEL-28, A431, and SCC-12 skin cancer cell lines, 35 displayed antiproliferative activities at the micromolar level, with the majority being primarily potent against the A431 and SCC-12 squamous carcinoma cell lines. The most active compounds 11 (A431: IC50 = 5.0 μM, SCC-12: IC50 = 2.9 μM, SKMEL-28: IC50 = 4.9 μM, A375: IC50 = 6.7 μM) and 13 (A431: IC50 = 5.0 μM, SCC-12: IC50 = 3.3 μM, SKMEL-28: IC50 = 13.8 μM, A375: IC50 = 17.1 μM), significantly and dose-dependently induced apoptosis of SCC-12 and SK-MEL-28 cells, as evidenced by the suppression of Bcl-2 and upregulation of Bax, cleaved caspase-3, caspase-9, and PARP protein expression levels. Both agents significantly reduced scratch wound healing, colony formation, and expression levels of deregulated cancer molecular targets including RSK/Akt/ERK1/2 and S6K1. In silico target prediction and docking studies using the SwissTargetPrediction web-based tool suggested that CDK8, CLK4, nuclear receptor ROR, tyrosine protein-kinase Fyn/LCK, ROCK1/2, and PARP, all of which are dysregulated in skin cancers, might be prospective targets for the two most active compounds. Further validation of these targets by western blot analyses, revealed that ROCK/Fyn and its associated Hedgehog (Hh) pathways were downregulated or modulated by the two lead compounds. In aggregate, these results provide a strong framework for further validation of the observed activities and the development of a more comprehensive structure-activity relationship through the preparation and biological evaluation of analogs.

Antitumor Effect of Poplar Propolis on Human Cutaneous Squamous Cell Carcinoma A431 Cells

Propolis is a gelatinous substance processed by western worker bees from the resin of plant buds and mixed with the secretions of the maxillary glands and beeswax. Propolis has extensive biological activities and antitumor effects. There have been few reports about the antitumor effect of propolis against human cutaneous squamous cell carcinoma (CSCC) A431 cells and its potential mechanism. CCK-8 assays, label-free proteomics, RT–PCR, and a xenograft tumor model were employed to explore this possibility. The results showed that the inhibition rate of A431 cell proliferation by the ethanol extract of propolis (EEP) was dose-dependent, with an IC50 of 39.17 μg/mL. There were 193 differentially expressed proteins in the EEP group compared with the control group (p < 0.05), of which 103 proteins (53.37%) were upregulated, and 90 proteins (46.63%) were downregulated. The main three activated and suppressed Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were extracellular matrix (ECM)-receptor interaction, amoebiasis, cell adhesion molecules (CAMs), nonalcoholic fatty liver disease (NAFLD), retrograde endocannabinoid signaling, and Alzheimer’s disease. The tumor volume of the 100 mg/kg EEP group was significantly different from that of the control group (p < 0.05). These results provide a theoretical basis for the potential treatment of human CSCC A431 cell tumors using propolis.

Human Blood Serum Can Diminish EGFR-Targeted Inhibition of Squamous Carcinoma Cell Growth through Reactivation of MAPK and EGFR Pathways.

Regardless of the presence or absence of specific diagnostic mutations, many cancer patients fail to respond to EGFR-targeted therapeutics, and a personalized approach is needed to identify putative (non)responders. We found previously that human peripheral blood and EGF can modulate the activities of EGFR-specific drugs on inhibiting clonogenity in model EGFR-positive A431 squamous carcinoma cells. Here, we report that human serum can dramatically abolish the cell growth rate inhibition by EGFR-specific drugs cetuximab and erlotinib. We show that this phenomenon is linked with derepression of drug-induced G1S cell cycle transition arrest. Furthermore, A431 cell growth inhibition by cetuximab, erlotinib, and EGF correlates with a decreased activity of ERK1/2 proteins. In turn, the EGF- and human serum-mediated rescue of drug-treated A431 cells restores ERK1/2 activity in functional tests. RNA sequencing revealed 1271 and 1566 differentially expressed genes (DEGs) in the presence of cetuximab and erlotinib, respectively. Erlotinib- and cetuximab-specific DEGs significantly overlapped. Interestingly, the expression of 100% and 75% of these DEGs restores to the no-drug level when EGF or a mixed human serum sample, respectively, is added along with cetuximab. In the case of erlotinib, EGF and human serum restore the expression of 39% and 83% of DEGs, respectively. We further assessed differential molecular pathway activation levels and propose that EGF/human serum-mediated A431 resistance to EGFR drugs can be largely explained by reactivation of the MAPK signaling cascade.

The Association of Propranolol with 2-Deoxy-D-Glucose Reduces the Metabolism and the Proliferation of Cutaneous Squamous Carcinoma A431 Cell Line

Non-selective β-blocking (±)-Propranolol Hydrochloride was demonstrated to improve the progression-free survival of oncological patients. Since the expression of β-adrenoceptors in the epidermal squamous cell carcinoma was described, we hypothesized that the topical application of a β- adrenoceptors-blockers over the tumor lesion may decrease its extension before the surgical excision, becoming an adjuvant therapy against cutaneous squamous cell carcinoma. However, it is known that β-AR-blocker anti-cancer activity as a single agent is limited. Hence, we suggested that the combination of propranolol with the glucose analog 2-Deoxy-D-glucose could improve its antiproliferative effect through the induction of metabolic stress. Propranolol and 2-Deoxy-D-glucose effect on A431 squamous cell carcinoma and normal keratinocytes evaluating metabolic activity, proliferation and apoptosis through MTT, immunofluorescence Ki-67 and AnnexinV assays, cell cycle analysis and migration assay. Our results demonstrated that the addition of 2-Deoxy-D-glucose low dose to propranolol improve its effect on reduction of A431 cells metabolism and proliferation, similar effect was observed on HaCaT viability and mobility. However, the HaCaT migration ability is not completely compromised. The combination of propranolol with a low dose of 2-Deoxy-D-glucose could be a promising treatment to be topically applied avoiding systemic adverse effects in patients with cutaneous squamous cell carcinoma.

Effects of photodynamic therapy using Red LED-light combined with hypocrellin B on apoptotic signaling in cutaneous squamous cell carcinoma A431 cells

BackgroundThe incidence of cutaneous squamous cell carcinoma (cSCC) has been demonstrating yearly increases. cSCC is a malignant cancer and exerts a major impact on patients' health and quality of life. Thus, the development and use of novel therapies in the treatment of cSCC are needed. It has been reported that LED photodynamic therapy (LED PDT) mediated by Hypocrellin B and its derivatives, a second-generation photosensitizer, can induce apoptosis in a variety of tumor cells, However, its potential pro-apoptotic effects on cSCC have yet to be investigated. ObjectiveThis study aims to investigate the pro-apoptotic effects and molecular mechanisms of HB-LED PDT in cutaneous squamous cell carcinoma A431 cells (Subsequent abbreviation A431 cells). Such information can provide an important theoretical foundation for the clinical translation of HB-LED PDT in the treatment of cSCC. Methods1. Effects of HB on A431 cells were determined using a Cell Counting Kit-8 assay, which method can indirectly reflect the number of living cells. In this way, this assay can then provide a means to identify the optimal concentrations of HB required for the induction of apoptosis in A431 cells. 2. The effects of HB-LED PDT on the morphology of A431 cells and changes in the nuclei after Hoechst33342 staining as determined using inverted fluorescent microscopy. 3. Use of the Annexin V-FITC test kit to detect levels of apoptosis in A431 cells in response to treatment with HB. Changes in reactive oxygen species and mitochondrial membrane potential following HB-LED PDT treatment in A431 cells were determined using fluorescence activated cell sorting (FACS). 4. Real-time quantitative PCR and Western Blot were applied to assess changes in several key factors involved in apoptosis including Bax, Bcl-2, and Caspase-3, at both transcription and translation levels. With these assays, it was possible to investigate the apoptotic signaling pathway in A431 cells in response to HB-LED PDT. ResultsHB-LED PDT inhibited proliferation activity and promoted nuclear fragmentation within these A431 cells. HB-LED PDT inhibited mitochondrial activity, increased reactive oxygen species production, and promoted apoptosis of A431 cells. In addition, several key factors in the apoptotic signaling pathway were increased at both the transcriptional and translational levels in A431 cells in response to the HB-LED PDT, indicating that the apoptotic signaling pathway was activated by HB-LED PDT. ConclusionHB-LED PDT induces apoptosis in A431 cells through a mitochondria-mediated apoptotic pathway. Such findings serve as an important foundation for the development of new approaches in the treatment of cSCC.

Abstract LB111: Epigenetic mechanism underlying pathogenesis of skin photo-damage and actinic keratoses

Abstract Chronic skin exposure to solar ultraviolet B (UVB) radiation causes cutaneous inflammation, sun-burns and pre-malignant lesions known as Actinic Keratosis (AK). About 20% of AKs may progress to highly invasive malignant lesions known as Squamous Cell Carcinoma (SCC). These lesions carry multiple mutations in oncogenes and/or tumor suppressor genes. However, the molecular mechanism underlying AK is not incompletely defined. Here, we demonstrate that cutaneous UVB exposure leads to activation of bromodomain-4 (BRD4), an epigenetic modifier. We observed enhanced expression of BRD4 and its hyper-acetylation marks, H3K18ac and H3K27ac in AK, accompanied by elevated transcription of BRD4-target genes: MMP13, CXCL9, IL33, Apol19b, IL19 and Oasl2. Using confocal microscopy, we observed enhanced nuclear accumulation of BRD4 and H3K27ac in the sun-exposed lesional skin, AK and in SCC tissues from patients. Ptch+/−/SKH1 mice irradiated with to chronic UVB manifest similar molecular alterations in highly photo-damaged skin. By manipulating BRD4 expression in human skin SCC A431 cells, we identified two important BRD4-regulated signaling pathways, which may be involved in the molecular pathogenesis of AKs and SCCs. BRD4-mediated inflammatory cytokine signaling is regulated by EZH2-SOCS3-NFkB/STAT3 axis while Myc-Survivin-CDK4-E2F1 axis drives tumor cell hyper-proliferation. Co-localization of BRD4/H3K27ac and Survivin-E2F1 was contiguous from sun-burn skin to AK and SCC. We observed enhanced copy number of Myc in human AK lesional skin and associated SCCs. We also show that Myc-Survivin-CDK4-E2F1 axis is critical in driving the progression of tumor keratinocytes to highly invasive and fast growing SCCs. This signaling acts by enhancing the expression epithelial-mesenchymal transition transcription factors and other proteins involved in UVB-induced SCC development. 9cUAB30, a RXR agonist developed by us blocked BRD4 activation and its downstream signaling in A431 and SCC13 cells. Oral administration of 9cUAB30 abrogates UVB-induced inflammation and keratinocyte hyper-proliferation in murine skin in BRD4 dependent manner. 9cUAB30 treatment attenuates growth of human SCC xenograft tumors in highly immune-suppressed mice, a response which was significantly more profound when the drug was administered with a survivin inhibitor, LQZ-7I. These translational studies demonstrate that UVB-induced cutaneous BRD4 drives AK pathogenesis and their progression to invasive SCCs. Citation Format: Mahendra P. Kashyap, Rajesh Sinha, Rubina Shafi, Venkatram R. Atigadda, Craig A. Elmets, Mohammad Athar. Epigenetic mechanism underlying pathogenesis of skin photo-damage and actinic keratoses [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr LB111.

The Activation of PPARγ by (2Z,4E,6E)-2-methoxyocta-2,4,6-trienoic Acid Counteracts the Epithelial-Mesenchymal Transition Process in Skin Carcinogenesis.

Cutaneous squamous cell carcinoma (cSCC) is the most common UV-induced keratinocyte-derived cancer, and its progression is characterized by the epithelial-mesenchymal transition (EMT) process. We previously demonstrated that PPARγ activation by 2,4,6-octatrienoic acid (Octa) prevents cutaneous UV damage. We investigated the possible role of the PPARγ activators Octa and the new compound (2Z,4E,6E)-2-methoxyocta-2,4,6-trienoic acid (A02) in targeting keratinocyte-derived skin cancer. Like Octa, A02 exerted a protective effect against UVB-induced oxidative stress and DNA damage in NHKs. In the squamous cell carcinoma A431 cells, A02 inhibited cell proliferation and increased differentiation markers' expression. Moreover, Octa and even more A02 counteracted the TGF-β1-dependent increase in mesenchymal markers, intracellular ROS, the activation of EMT-related signal transduction pathways, and cells' migratory capacity. Both compounds, especially A02, counterbalanced the TGF-β1-induced cell membrane lipid remodeling and the release of bioactive lipids involved in EMT. In vivo experiments on a murine model useful to study cell proliferation in adult animals showed the reduction of areas characterized by active cell proliferation in response to A02 topical treatment. In conclusion, targeting PPARγ may be useful for the prevention and treatment of keratinocyte-derived skin cancer.

Understanding of black salve toxicity by multi-compound cytotoxicity assays

BackgroundBlack salve is a controversial complementary and alternative medicine (CAM) associated with skin toxicity and skin cancer treatment failures. Black salve formulations vary between manufacturers and contain a number of botanical and synthetic constituents. The skin cancer cytotoxicity of a number of these constituents has not been assessed to date. The alkaloids from the rhizomes of Sanguinaria canadensis, a key black salve ingredient, have had their single compound cytotoxicity assessed; however, whether they possess synergistic cytotoxicity with other compounds has not been studied and is of direct clinical relevance. This research aimed to improve our understanding of the skin cancer cytotoxicity of black salve constituents.MethodsThe cytotoxicity of individual and combination black salve constituents were assessed against the A375 melanoma and A431 squamous cell carcinoma cell lines. Cytotoxicity was determined using the Resazurin assay with fluorescence measured using a Tecan Infinite 200 Pro Microplate reader, compound cytotoxicity being compared to that of the topical cancer therapeutic agent, 5- fluouracil. Docetaxal was used as a positive control. Dunnetts p value was used to determine whether significant synergistic cytotoxicity was present.ResultsSanguinarine was the most cytotoxic compound tested with a 24-hour IC50 of 2.1 μM against the A375 Melanoma cell line and 3.14 μM against the A431 SCC cell line. All black salve constituents showed greater cytotoxicity against the two skin cancer cell lines tested than the skin cancer therapeutic 5-Fluouracil with 24 hours of compound exposure. Chelerythrine and minor Quaternary Benzophenanthridine Alkaloids (QBAs) present in black salve, at concentrations not having a cytotoxic effect by themselves, boosted the cytotoxic effects of sanguinarine. This could be a synergistic rather than additive cytotoxic effect although the synergistic effect was cell line and concentration dependent.ConclusionsBlack salve contains several cytotoxic compounds, a number of which have been found to possess synergistic cytotoxicity for the first time against skin cancer cell lines. In addition, these compounds together increase the overall cytotoxic effect. Assessing multi-compound cytotoxicity in herbal medicine can provide additional information about both their therapeutic and toxicity potential. As black salve is currently being used by patients, further cytotoxicity work should be undertaken to assess whether synergistic cytotoxicity exists when tested in normal skin cells.

Annexin A6 and NPC1 regulate LDL-inducible cell migration and distribution of focal adhesions

Cholesterol is considered indispensable for cell motility, but how physiological cholesterol pools enable cells to move forward remains to be clarified. The majority of cells obtain cholesterol from the uptake of Low-Density lipoproteins (LDL) and here we demonstrate that LDL stimulates A431 squamous epithelial carcinoma and Chinese hamster ovary (CHO) cell migration and invasion. LDL also potentiated epidermal growth factor (EGF) -stimulated A431 cell migration as well as A431 invasion in 3-dimensional environments, using organotypic assays. Blocking cholesterol export from late endosomes (LE), using Niemann Pick Type C1 (NPC1) mutant cells, pharmacological NPC1 inhibition or overexpression of the annexin A6 (AnxA6) scaffold protein, compromised LDL-inducible migration and invasion. Nevertheless, NPC1 mutant cells established focal adhesions (FA) that contain activated focal adhesion kinase (pY397FAK, pY861FAK), vinculin and paxillin. Compared to controls, NPC1 mutants display increased FA numbers throughout the cell body, but lack LDL-inducible FA formation at cell edges. Strikingly, AnxA6 depletion in NPC1 mutant cells, which restores late endosomal cholesterol export in these cells, increases their cell motility and association of the cholesterol biosensor D4H with active FAK at cell edges, indicating that AnxA6-regulated transport routes contribute to cholesterol delivery to FA structures, thereby improving NPC1 mutant cell migratory behaviour.

PINCH-1 promotes IGF-1 receptor expression and skin cancer progression through inhibition of the GRB10-NEDD4 complex.

Background: Insulin-like growth factor 1 receptor (IGF-1R) expression and signaling play important roles in promotion of skin cancer progression. Identification of signaling pathways that regulate IGF-1R is crucial for understanding the pathogenesis and therapeutic treatment of skin cancer.Methods: Molecular, cellular and genetic approaches were used to investigate the function of PINCH-1 in regulation of IGF-1R expression and skin cell behavior. Furthermore, conditional PINCH-1 knockout mouse and carcinogen (7, 12-dimethylbenz[a]anthracene (DMBA)/12-O-tetradecanoylphorbol-13-acetate (TPA))-induced skin cancer model were employed to determine the function of PINCH-1 in regulation of IGF-1R expression and skin carcinogenesis in vivo.Results: Knockdown of PINCH-1 from HaCaT keratinocytes or A431 squamous carcinoma cells diminished IGF-1R levels, suppressed cell proliferation and increased apoptosis. Re-expression of PINCH-1 in PINCH-1 knockdown cells restored IGF-1R expression, cell proliferation and survival. Furthermore, depletion of NEDD4 effectively reversed PINCH-1 deficiency-induced down-regulation of IGF-1R expression, cell proliferation and survival. Conditional knockout of PINCH-1 from keratin 5 (K5) positive keratinocytes in mice, like depletion of PINCH-1 from keratinocytes in culture, reduced the IGF-1R level. Using a mouse model of DMBA/TPA-induced skin cancer, we show that the levels of both PINCH-1 and IGF-1R were significantly increased in response to treatment with the carcinogens. Genetic ablation of PINCH-1 from the epidermis markedly reduced the IGF-1R expression and cell proliferation despite stimulation with DMBA/TPA, resulting in resistance to chemical carcinogen-induced skin cancer initiation and progression.Conclusions: Our results reveal a PINCH-1-NEDD4-IGF-1R signaling axis that is critical for promotion of skin tumorigenesis and suggest a new strategy for therapeutic control of skin cancer progression.

Lycium barbarum polysaccharides and ferroptosis: jumping into the era of novel regulated cell death.

Lycium barbarum polysaccharides and ferroptosis: jumping into the era of novel regulated cell death.

In vitro Antiproliferative Effects of Cotinus coggygria Scop. on human non-melanoma and melanoma skin cancer cells

Skin cancer is one of the most frequently diagnosed malignancies worldwide and its incidence constantly increases. The disease divides in two major subtypes: non-melanoma and melanoma skin cancer. The main drawbacks of the traditional skin cancer therapy are primary and acquired drug resistance and serious side effects due to the nonspecific treatments targeting. Despite advances in therapy strategies there is a need of new affordable natural anti-skin cancer agents, which to possess higher efficiency without causing detrimental side effects. Medicinal plants provide great possibility for the discovery of new anticancer therapeutics with preventive and treatment potential. Cotinus coggygria Scop. is a plant species widely applied in phytotherapy predominantly against disorders of the skin and mucosal tissues. The herb has a large range of valuable biological activities but its anticancer properties have not been thoroughly studied. The aim of the present research was to assess the antiproliferative properties of the crude leaf aqueous ethanolic extract from Bulgarian herb C. coggygria and its chloroformic and aqueous fractions on a panel of human skin cancer cell lines: basal cell carcinoma (TE 354.T), squamous cell carcinoma (A431) and malignant melanoma (A375) and to compare them to the cell growth inhibitory potential on normal dermal cell line (BJ). The antiproliferative capacity of the plant substances was investigated using MTT assay and microscopy cell morphology observation after 72 h cell treatment in a wide scale of concentrations. The obtained results showed that the crude extract and both fractions inhibit significant proliferation of A431 squamous cell carcinoma and A375 melanoma cells with the highest cytostatic effect registered for the aqueous fraction on A375 cells with a half maximal inhibitory concentration value of 44.33 ?g/ml. C. coggygria exhibited no cytostatic activity towards TE 354.T basal cell carcinoma cells. The established marked slight

Human blood serum can donor-specifically antagonize effects of EGFR-targeted drugs on squamous carcinoma cell growth

Many patients fail to respond to EGFR-targeted therapeutics, and personalized diagnostics is needed to identify putative responders. We investigated 1630 colorectal and lung squamous carcinomas and 1357 normal lung and colon samples and observed huge variation in EGFR pathway activation in both cancerous and healthy tissues, irrespectively on EGFR gene mutation status. We investigated whether human blood serum can affect squamous carcinoma cell growth and EGFR drug response. We demonstrate that human serum antagonizes the effects of EGFR-targeted drugs erlotinib and cetuximab on A431 squamous carcinoma cells by increasing IC50 by about 2- and 20-fold, respectively. The effects on clonogenicity varied significantly across the individual serum samples in every experiment, with up to 100% differences. EGF concentration could explain many effects of blood serum samples, and EGFR ligands-depleted serum showed lesser effect on drug sensitivity.

Cytotoxic, Antimitotic, DNA Binding, Photocatalytic, H2O2 Sensing, and Antioxidant Properties of Biofabricated Silver Nanoparticles Using Leaf Extract of Bryophyllum pinnatum (Lam.) Oken.

Bryophyllum pinnatum is a perennial herb traditionally used in ethnomedicine. In the present report, silver nanoparticles (AgNPs) were synthesized using B. pinnatum leaf extract. BP-AgNPs were confirmed following UV-Vis spectroscopy with SPR peak at 412 nm and further characterized by FTIR, XRD, SEM-EDX, and TEM. Microscopic images confirmed the spherical shape and ~15 nm average size of nanostructures. BP-AgNPs were evaluated for photocatalytic degradation of hazardous dyes (methylene blue and Rhodamine-B) and showed their complete reduction within 100 and 110 min., respectively. BP-AgNPs have emerged as a unique SPR-based novel sensor for the detection of H2O2, which may deliver exciting prospects in clinical and industrial areas. DPPH and ABTS free radical scavenging activity were studied with respective IC50 values of 89 and 259 μg/mL. A strong intercalating interaction of CT-DNA with BP-AgNPs was investigated. Observed chromosomal abnormalities confirm the antimitotic potential of BP-AgNPs in the meristematic root tip. The cytotoxicity of BP-AgNPs against B16F10 (melanoma cell line) and A431 (squamous cell carcinoma cell line), was assessed with respective IC50 values of 59.5 and 96.61 μg/ml after 24 h of treatment. The presented green synthetic approach provides a novel and new door for environmental, industrial, and biomedical applications.

Vandetanib inhibits cell growth in EGFR-expressing cutaneous squamous cell carcinoma

Advanced cutaneous squamous cell carcinoma (SCC) responds poorly to chemotherapy, leading to significant morbidity or death. Overexpression of epidermal growth factor receptor (EGFR) is frequently observed in advanced cutaneous SCC. Vandetanib is a multiple tyrosine kinase targeting vascular endothelial growth factor receptor-2 (VEGFR2), EGFR, and the rearranged during transfection (RET) proto-oncogene. Vandetanib has been reported to inhibit tumor growth in head and neck SCC. However, the efficacy of vandetanib against cutaneous SCC has not been thoroughly investigated. The aim of this study is to evaluate the efficacy of vandetanib against cutaneous SCC in vitro and in vivo. Vandetanib is found to inhibit the proliferation of cutaneous SCC cells as assessed by cell viability and clonogenic assay. Cell death analysis indicates that vandetanib induces cell death in SCC cells but not in normal human keratinocytes or fibroblasts. The in vivo anti-tumor effect of vandetanib is shown in xenograft tumor models using A431 SCC cells. Mechanistically, vandetanib suppresses the phosphorylation of EGFR in SCC cells. Clinically, EGFR expression levels are elevated in cutaneous SCC specimens, relative to normal epidermis. In conclusion, we identified vandetanib as a novel therapeutic option for cutaneous SCC, especially in tumors with high EGFR expression.

Higher Anti-Tumor Efficacy of the Dual HER3-EGFR Antibody MEHD7945a Combined with Ionizing Irradiation in Cervical Cancer Cells

The outcome of locally advanced cervical cancer (LACC) is dismal. Biomarkers are needed to individualize treatments and to improve patient outcomes. Here, we investigated whether coexpression of epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 3 (HER3) could be an outcome prognostic biomarker, and whether targeting both EGFR and HER3 with a dual antibody (MEHD7945A) enhanced ionizing radiation (IR) efficacy. Expression of EGFR and HER3 was evaluated by immunohistochemistry in cancer biopsies (n = 72 patients with LACC). The antitumor effects of the MEHD7945A and IR combotherapy were assessed in 2 EGFR- and HER3-positive cervical cancer cell lines (A431 and CaSki) and in A431 cell xenografts. The mechanisms involved in tumor cell radiosensitization were also studied. The interaction of MEHD7945A, IR, and cisplatin was evaluated using dose-response matrix data. EGFR and HER3 were coexpressed in only in 7 of the 22 biopsies of FIGO IVB cervix cancer. The median overall survival was 14.6 months and 23.1 months in patients with FIGO IVB tumors that coexpressed or did not coexpress EGFR and HER3, respectively. In mice xenografted with A431 (squamous cell carcinoma) cells, MEHD7945A significantly increased IR response by reducing tumor growth and increasing cleaved caspase-3 expression. In A431 and CaSki cells, the combotherapy increased DNA damage and cell death, particularly immunogenic cell death, and decreased survival by inhibiting the MAPK and AKT pathways. An additive effect was observed when IR, MEHD7945A, and cisplatin were combined. Targeting EGFR and HER3 with a specific dual antibody enhanced IR efficacy. These preliminary results and the prognostic value of EGFR and HER3 coexpression should be confirmed in a larger sample.