Harnessing the power of a photoinitiated thiol-ene "click" reaction for the efficient synthesis of S-lipidated collagen model peptide amphiphiles.

A photoinitiated thiol-ene "click" reaction was used to synthesize S-lipidated collagen model peptide amphiphiles. Use of 2-iminothiolane provided an epimerization-free thiol handle required for thiol-ene based incorporation of lipid moieties onto collagen-based peptide sequences. This approach not only led to improvements in the triple helical characteristics of the resulting collagen model peptides but also increased the aqueous solubility of the peptide amphiphiles. As a result, this methodology holds significant potential for the design and advancement of functional peptide amphiphiles, offering enhanced capabilities across a wide range of applications.

A pilot study using unique targeted testing of the urogenital microbiome has potential as a predictive test during IVF for implantation outcome.

PURPOSE: This pilot study aimed to develop a methodology characterising the urogenital microbiome as a predictive test in the IVF workup. METHODS: Using unique custom qPCRs, we tested for the presence of specific microbial species from vaginal samples and First Catch Urines from the male. The test panel included a range of potential urogenital pathogens, STIs, 'favourable bacteria' (Lactobacillus spp.) and 'unfavourable bacteria' (anaerobes) reported to influence implantation rates. We tested couples attending Fertility Associates, Christchurch, New Zealand for their first round of IVF. RESULTS: We found that some microbial species affected implantation. The qPCR result was interpreted qualitatively using the Z proportionality test. Samples from women at the time of Embryo Transfer who did not achieve implantation had significantly higher percent of samples that were positive for Prevotella bivia and Staphylococcus aureus compared to women who did achieve implantation. DISCUSSION: The results provide evidence that most other microbial species chosen for testing had little functional effect on implantation rates. The addition of further microbial targets (yet to be determined) could be combined in this predictive test for vaginal preparedness on the day of embryo transfer. This methodology has a substantial advantage of being affordable and easily performed in any routine molecular laboratory. This methodology is most suitable as a foundation on which to develop a timely test of microbiome profiling. Using the indicators detected to have a significant influence, these results can be extrapolated. CONCLUSION: Using a rapid antigen test, a woman can self-sample prior to embryo transfer and obtain an indication of microbial species present which could influence implantation outcome.

Collagen I dysregulation is pivotal for ovarian cancer progression.

As a principal matrisomal protein, collagen is involved in the regulation of the structural framework of extracellular matrix (ECM) and therefore is potentially crucial in determining the biophysical character of the ECM. It has been suggested that collagen architecture plays a role in ovarian cancer development, progression and therapeutic responses which led us to examine the collagen morphology in normal and cancerous ovarian tissue. Also, the behaviour of ovarian cancer cells cultured in four qualitatively different collagen gels was investigated. The results here provide evidence that collagen I morphology in the cancerous ovary is distinct from that in the normal ovary. Tumour-associated collagen I showed streams or channels of thick elongated collagen I fibrils. Moreover, fibril alignment was significantly more prevalent in endometrioid and clear cell cancers than other ovarian cancer subtypes. In this work, for the first-time collagen I architecture profiling (CAP) was introduced using histochemical staining, which distinguished between the collagen I morphologies of ovarian cancer subtypes. Immunohistochemical examination of ovarian normal and cancerous tissues also supported the notion that focal adhesion and Rho signalling are upregulated in ovarian cancers, especially in the high-grade serous tumours, as indicated by higher expression of p-FAK and p190RhoGEF. The results also support the concept that collagen I architecture, which might be collagen I concentration-dependent, influences proliferation in ovarian cancer cells. The study provides evidence that modification of collagen I architecture integrity is associated with ovarian cancer development and therapeutic responses.

Deciphering Biophysical Modulation in Ovarian Cancer Cells.

It has been long known that the oncogenic extracellular environment plays an indispensable role in developing and nurturing cancer cell progression and in resistance to standard treatments. However, by how much the biophysical components of tumour extracellular environment contribute to these processes is uncertain. In particular, the topographic environment is scarcely explored. The biophysical modulation of cell behaviour is primarily facilitated through mechanotransduction-associated mechanisms, including focal adhesion and Rho/ROCK signalling. Dysregulation of these pathways is commonly observed in ovarian cancer and, therefore, biophysical modulation of these mechanisms may be of great importance to ovarian cancer development and progression. In this work, aspects of the biophysical environment were explored using a bioimprinting technique. The study showed that topography-mediated substrate sensing delayed cell attachment, however, cell-cell interactions overrode the effect of topography in some cell lines, such as OVCAR-5. Also, 3D topographical cues were shown to modulate the inhibition of focal adhesion and Rho signalling, which resulted in higher MAPK activity in cells on the bioimprints. It was revealed that c-Src is vital to the biophysical modulation of cell proliferation and inhibition of c-Src could downregulate biophysically modulated MAPK activity. This study provides evidence that the biophysical extracellular environment affects key intracellular mechanisms associated with tumourigenicity in ovarian cancer cells.

Extracellular biophysical environment: Guilty of being a modulator of drug sensitivity in ovarian cancer cells.

The roles of the extracellular biophysical environment in cancer are barely studied. This study considers the possibility that cell-like topography of a cancer cell environment may influence chemo-responses. Here, a novel bioimprinting technique was employed to produce cell-like topography on the polystyrene substrates used for cell culture. In this work, we have shown that extracellular biophysical cues generated from the topography alter the chemosensitivity of ovarian cancer cells. The three-dimensionality of the bioimprinted surface altered the cell-surface interaction, which consequently modulated intracellular signalling and chemoresponses. Sensitivity to platinum was altered more than that to paclitaxel. The effect was largely mediated through the integrin/focal adhesion system and the Rho/ROCK pathway. Moreover, the results provided evidence that biophysical cues also modulate MAPK signalling associated with chemo-resistance in ovarian cancer. Therefore, the novel findings from of this study revealed for the first time that the effects of the biophysical environment, such as substrate topography, influences ovarian cancer cell responses to clinical drugs. These observations suggest that a full clinical understanding of ovarian cancer will include biophysical aspects of tumour microenvironment.

The extracellular topographical environment influences ovarian cancer cell behaviour.

The importance of the biophysical cellular environment in cancer development has been increasingly recognised but so far has been only superficially studied. Here we investigated the effect of cell-like substrate topography on ovarian cancer cell behaviour and potential underlying signalling pathways. We observed changes in cell morphology in response to substrate topography, which implies modification of structure-function associations. Differences in focal adhesion signalling and Rho/ROCK activity suggested their involvement in the biomechanically-driven cellular responses. Cell-like topography was also shown to modulate the MAPK pathway and hence potentially regulate cell proliferation. The selective regulation of the cells by the mechanotransduction pathways that we noted has wide ranging implications for understanding cancer development. We established that the physical architecture of cell culture substrate is sufficient to influence cancer cell behaviour, independent of genetic composition or biochemical milieu.

NRAS and EPHB6 mutation rates differ in metastatic melanomas of patients in the North Island versus South Island of New Zealand.

Melanoma, the most aggressive skin cancer type, is responsible for 75% of skin cancer related deaths worldwide. Given that New Zealand (NZ) has the world's highest melanoma incidence, we sought to determine the frequency of mutations in NZ melanomas in recurrently mutated genes. NZ melanomas were from localities distributed between North (35°S-42°S) and South Islands (41°S-47°S). A total of 529 melanomas were analyzed for BRAF exon 15 mutations by Sanger sequencing, and also by Sequenom MelaCarta MassARRAY. While, a relatively low incidence of BRAFV600E mutations (23.4%) was observed overall in NZ melanomas, the incidence of NRAS mutations in South Island melanomas was high compared to North Island melanomas (38.3% vs. 21.9%, P=0.0005), and to The Cancer Genome Atlas database (TCGA) (38.3% vs. 22%, P=0.0004). In contrast, the incidence of EPHB6G404S mutations was 0% in South Island melanomas, and was 7.8% in North Island (P=0.0002). Overall, these data suggest that melanomas from geographically different regions in NZ have markedly different mutation frequencies, in particular in the NRAS and EPHB6 genes, when compared to TCGA or other populations. These data have implications for the causation and treatment of malignant melanoma in NZ.