Victoria Sherwood

A t-butyloxycarbonyl-modified Wnt5a-derived hexapeptide functions as a potent antagonist of Wnt5a-dependent melanoma cell invasion

The influential role of Wnt5a in tumor progression underscores the requirement for developing molecules that can target Wnt5a-mediated cellular responses. In the aggressive skin cancer, melanoma, elevated Wnt5a expression promotes cell motility and drives metastasis. Two approaches can be used to counteract these effects: inhibition of Wnt5a expression or direct blockade of Wnt5a signaling. We have investigated both options in the melanoma cell lines, A2058 and HTB63. Both express Frizzled-5, which has been implicated as the receptor for Wnt5a in melanoma cells. However, only the HTB63 cell line expresses and secretes Wnt5a. In these cells, the cytokine, TGFβ1, controlled the expression of Wnt5a, but due to the unpredictable effects of TGFβ1 signaling on melanoma cell motility, targeting Wnt5a signaling via TGFβ1 was an unsuitable strategy to pursue. We therefore attempted to target Wnt5a signaling directly. Exogenous Wnt5a stimulation of A2058 cells increased adhesion, migration and invasion, all crucial components of tumor metastasis, and the Wnt5a-derived N-butyloxycarbonyl hexapeptide (Met-Asp-Gly-Cys-Glu-Leu; 0.766 kDa) termed Box5, abolished these responses. Box5 also inhibited the basal migration and invasion of Wnt5a-expressing HTB63 melanoma cells. Box5 antagonized the effects of Wnt5a on melanoma cell migration and invasion by directly inhibiting Wnt5a-induced protein kinase C and Ca2+ signaling, the latter of which we directly demonstrate to be essential for cell invasion. The Box5 peptide directly inhibits Wnt5a signaling, representing an approach to anti-metastatic therapy for otherwise rapidly progressive melanoma, and for other Wnt5a-stimulated invasive cancers.

WNT Signaling: an Emerging Mediator of Cancer Cell Metabolism?

ABSTRACT WNT signaling was discovered in tumor models and has been recognized as a regulator of cancer development and progression for over 3 decades. Recent work has highlighted a critical role for WNT signaling in the metabolic homeostasis of mammals, where its misregulation has been heavily implicated in diabetes. While the majority of WNT metabolism research has focused on nontransformed tissues, the role of WNT in cancer metabolism remains underinvestigated. Cancer is also a metabolic disease where oncogenic signaling pathways regulate energy production and macromolecular synthesis to fuel rapidly proliferating tumors. This review highlights the emerging evidence for WNT signaling in the reprogramming of cancer cell metabolism and examines the role of these signaling pathways as mediators of tumor bioenergetics.

The scope of nanoparticle therapies for future metastatic melanoma treatment

Metastatic melanoma is a highly aggressive malignancy that has traditionally been very difficult to treat. However, after decades of basic research into the signal transduction pathways that promote cancer cell survival, chemoresistance, growth, and crosstalk with the immune system, targeted therapies have now been developed that offer improved survival for patients with metastatic melanoma. Some of the most promising therapies that have been developed include ipilimumab, an anti-cytotoxic T lymphocyte antigen 4 antibody that enhances T-cell activity in the tumour, and selective BRAF inhibitors, such as vemurafenib that blocks tumour cell proliferation in patients with activating BRAF mutations. Although these treatments offer substantial hope for patients, they are not without their drawbacks, which include adverse side-effects, drug resistance, and eventual relapse. Nanotherapeutics holds significant promise to circumvent these shortcomings and has the additional advantage of potentially functioning as a diagnostic device. We will discuss the scope of the use of such multimodal nanoparticles for melanoma treatment and ask whether such particles can offer patients with metastatic melanoma improved prognoses for the future.

Methylation and Loss of Secreted Frizzled-Related Protein 3 Enhances Melanoma Cell Migration and Invasion

Background Wnt signaling is important in development and can also contribute to the initiation and progression of cancer. The Secreted Frizzled Related Proteins (SFRPs) constitute a family of Wnt modulators, crucial for controlling Wnt signaling. Here we investigate the expression and role of SFRP3 in melanoma. Methodology/Principal Findings We show that SFRP3 mRNA is down-regulated in malignant melanoma tumors as compared to normal/benign tissue. Furthermore, we found that SFRP3 expression was lost in the malignant melanoma cell lines, A2058, HTB63 and A375, but not in the non-transformed melanocyte cell line, Hermes 3A. Methylated CpG rich areas were detected in the SFRP3 gene in melanoma cell lines and their SFRP3 expression could be restored using the demethylating agent, 5′aza-deoxycytidine. Addition of recombinant SFRP3 to melanoma cells had no effect on viable cell numbers, but decreased cell migration and invasion. Wnt5a signaling has been shown to increase the migration and invasion of malignant melanoma cells, and high expression of Wnt5a in melanoma tumors has been connected to a poor prognosis. We found that recombinant SFRP3 could inhibit Wnt5a signaling, and that it inhibited melanoma cell migration and invasion in a Wnt5a-dependent manner. Conclusion/Significance We conclude that SFRP3 functions as a melanoma migration and invasion suppressor by interfering with Wnt5a signaling.

WNT5A-mediated β-catenin-independent signalling is a novel regulator of cancer cell metabolism.

WNT5A has been identified as an important ligand in the malignant progression of a number of tumours. Although WNT5A signalling is often altered in cancer, the ligand’s role as either a tumour suppressor or oncogene varies between tumour types and is a contemporary issue for investigators of β-catenin-independent WNT signalling in oncology. Here, we report that one of the initial effects of active WNT5A signalling in malignant melanoma cells is an alteration in cellular energy metabolism and specifically an increase in aerobic glycolysis. This was found to be at least in part due to an increase in active Akt signalling and lactate dehydrogenase (LDH) activity. The clinical relevance of these findings was strengthened by a strong correlation (P < 0.001) between the expression of WNT5A and LDH isoform V in a cohort of melanocytic neoplasms. We also found effects of WNT5A on energy metabolism in breast cancer cells, but rather than promoting aerobic glycolysis as it does in melanoma, WNT5A signalling increased oxidative phosphorylation rates in breast cancer cells. These findings support a new role for WNT5A in the metabolic reprogramming of cancer cells that is a context- dependent event.

Synthesis of Carbohydrate Capped Silicon Nanoparticles and their Reduced Cytotoxicity, In Vivo Toxicity, and Cellular Uptake

The development of smart targeted nanoparticles (NPs) that can identify and deliver drugs at a sustained rate directly to cancer cells may provide better efficacy and lower toxicity for treating primary and advanced metastatic tumors. Obtaining knowledge of the diseases at the molecular level can facilitate the identification of biological targets. In particular, carbohydrate-mediated molecular recognitions using nano-vehicles are likely to increasingly affect cancer treatment methods, opening a new area in biomedical applications. Here, silicon NPs (SiNPs) capped with carbohydrates including galactose, glucose, mannose, and lactose are successfully synthesized from amine terminated SiNPs. The MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] analysis shows an extensive reduction in toxicity of SiNPs by functionalizing with carbohydrate moiety both in vitro and in vivo. Cellular uptake is investigated with flow cytometry and confocal fluorescence microscope. The results show the carbohydrate capped SiNPs can be internalized in the cells within 24 h of incubation, and can be taken up more readily by cancer cells than noncancerous cells. Moreover, these results reinforce the use of carbohydrates for the internalization of a variety of similar compounds into cancer cells.

An early developmental vertebrate model for nanomaterial safety: bridging cell-based and mammalian toxicity assessment

AIM With the rise in production of nanoparticles (NPs) for an ever-increasing number of applications, there is an urgent need to efficiently assess their potential toxicity. We propose a NP hazard assessment protocol that combines mammalian cytotoxicity data with embryonic vertebrate abnormality scoring to determine an overall toxicity index. RESULTS We observed that, after exposure to a range of NPs, Xenopus phenotypic scoring showed a strong correlation with cell based in vitro assays. Magnetite-cored NPs, negative for toxicity in vitro and Xenopus, were further confirmed as nontoxic in mice. CONCLUSION The results highlight the potential of Xenopus embryo analysis as a fast screening approach for toxicity assessment of NPs, which could be introduced for the routine testing of nanomaterials.

WNT Signaling in Cutaneous Squamous Cell Carcinoma: A Future Treatment Strategy?

The molecular mechanisms underlying cutaneous squamous cell carcinoma are less well established than those for other common skin cancers, but recent evidence has highlighted a potentially critical role for WNT signaling in both the development and progression of cutaneous squamous cell carcinoma. WNT pathways are aberrantly regulated in multiple tumor types (albeit in a context-dependent manner), and this has stimulated the development of WNT inhibitory compounds for cancer treatment. In this review, we examine existing evidence for a role of WNT signaling in cutaneous squamous cell carcinoma and discuss if WNT inhibition represents a realistic therapeutic strategy for the future.

Nanoscopic Agents in a Physiological Environment: The Importance of Understanding Their Characteristics

The application of nanotechnology in medicine signifies one of the most exciting developments in science over the last decade. Even though advancement has been made in nanoparticle engineering in terms of size, shape and surface functionalisation, the behaviour in vivo remains poorly characterised and understood. The potential impact of engineered nanomaterials on human health is strictly related to their behaviour in the biological environment. When in contact with biological fluids, nanoparticles spontaneously interact and adsorb proteins to dramatically change their surface properties. Thus, the nanoparticle surface acquires a new biological identity that will influence its stability and interaction with the cellular machinery, thereby affecting the nanoparticle biodistribution in vivo. This protein coating ‘expressed’ at the nanoparticle surface is what is ‘read’ by the cells. Consequently, methods to effectively study the structure and composition of this bio-nano interface have been emerging as key objectives in nanoscience. In this chapter, we discuss the state-of-the-art techniques for the physico-chemical characterisation of nanoparticle-protein complexes in the biological environment with particular emphasis on their impact on the efficiency and safety of a new generation of nanomedicines. We also highlight the barriers faced by nanomedicines for effective targeting and delivery in vivo.

The anti-rheumatic drug, leflunomide, synergizes with MEK inhibition to suppress melanoma growth

Cutaneous melanoma, which develops from the pigment producing cells called melanocytes, is the most deadly form of skin cancer. Unlike the majority of other cancers, the incidence rates of melanoma are still on the rise and the treatment options currently available are being hindered by resistance, limited response rates and adverse toxicity. We have previously shown that an FDA approved drug leflunomide, used for rheumatoid arthritis (RA), also holds potential therapeutic value in treating melanoma especially if used in combination with the mutant BRAF inhibitor, vemurafenib. We have further characterized the function of leflunomide and show that the drug reduces the number of viable cells in both wild-type and BRAFV600E mutant melanoma cell lines. Further experiments have revealed leflunomide reduces cell proliferation and causes cells to arrest in G1 of the cell cycle. Cell death assays show leflunomide causes apoptosis at treatment concentrations of 25 and 50 µM. To determine if leflunomide could be used combinatorialy with other anti-melanoma drugs, it was tested in combination with the MEK inhibitor, selumetinib. This combination showed a synergistic effect in the cell lines tested. This drug combination led to an enhanced decrease in tumor size when tested in vivo compared to either drug alone, demonstrating its potential as a novel combinatorial therapy for melanoma.