In situ forming implants (ISFIs) have shown promise as a sustained, local drug delivery system for therapeutics in a variety of applications. However, development of ISFIs has been hindered by poor correlation between in vitro study results and in vivo performance. In contrast to oral dosage forms, there is currently no clear consensus on a standard for in vitro drug dissolution studies for parenteral formulations. Recent studies have suggested that the disparity between in vivo and in vitro behavior of phase-inverting ISFIs may be, in part, due to differences in injection site stiffness. Accordingly, this study aimed to create acrylamide-based hydrogel phantoms of various porosities and stiffness, which we hypothesized would better predict in vivo performance. Implant microstructure and shape were found to be dependent on the stiffness of the phantoms, while drug release was found to be dependent on both phantom porosity and stiffness. Specifically, SEM analysis revealed that implant porosity and interconnectivity decreased with increasing phantom stiffness and better mimicked the microstructure seen in vivo. Burst release of drug increased from 31% to 43% when in standard acrylamide phantoms vs macroporous phantoms (10 kPa), improving the correlation to the burst release seen in vivo. Implants in 30 kPa macroporous phantoms had the best correlation with in vivo burst release, significantly improving (p < 0.05) the burst release relative to in vivo from 64%, using a standard PBS dissolution method, to 92%. These findings confirm that implant behavior is affected by injection site stiffness. Importantly, with appropriate optimization and validation, hydrogel phantoms such as the one investigated here could be used to improve the in vitro-in vivo correlation of in situ forming implant formulations and potentially augment their advancement to clinical use.

The increase of a skin malignant melanoma (melanoma malignum) incidence in the world has been observed in recent years. The tumour, especially in advanced stadium with metastases, is highly resistant to conventional treatment. One of the strategies is to modulate melanogenesis using chemical compounds. In this study, the processes of differentiation and melanogenesis induced by dimethylsulfoxide (DMSO) in human melanoma cells (A-375) were investigated. Natural melanin isolated from A-375 melanoma cell line treated with 0.3% DMSO was analyzed by pyrolysis-gas chromatography-mass spectrometry (Py-GC/MS) method. The products derived from pheomelanin have not been stated in the pyrolytic profile of analyzed melanin. Within all products derived from eumelanins, 1,2-benzenediol has been predominated. It has been shown that in the melanoma cells stimulated with 0.3% and 1% DMSO, the increase of transcriptional activity of the tyrosinase gene took place. It was accompanied by the rise of tyrosinase activity and an accumulation of melanin in the cells. The better knowledge about the structure of melanins can contribute to establish the uniform criteria of malignant melanoma morbidity risk.

Malignant melanoma (melanoma malignum) is one of the most dangerous types of tumor. It is very difficult to cure. In recent years, a lot of attention has been given to chemoprevention. This method uses natural and synthetic compounds to interfere with and inhibit the process of carcinogenesis. In this study, a new treatment strategy was proposed consisting of a combination of 5,7-dimethoxycoumarin (DMC), an activator of melanogenesis, and valproic acid (VPA), a well-known drug that is one of the histone deacetylase inhibitors (HDACis). In conjunction with 1 mM VPA, all of the tested concentrations of DMC (10–150 μM) significantly decreased the proliferation of A-375 cells. VPA and DMC also induced the synthesis of melanin and the formation of dendrite and star-shaped cells. Tyrosinase gene expression and tyrosinase activity significantly increased in response to VPA treatment. Pyrolysis with gas chromatography and mass spectrometry (Py-GC/MS) was used to investigate the structure of the isolated melanin. This showed that the quantitative and qualitative components of melanin degradation products are dependent on the type of applied melanogenesis inductor. Products derived from eumelanin were detected in the pyrolytic profile of melanin isolated from A-375 cells stimulated with DMC. Thermal degradation of melanin isolated from melanoma cells after exposure to VPA or a mixture of VPA and DMC revealed the additional presence of products derived from pheomelanin.

Aim of the studyAbnormalities in signaling as well as altered gene expression have been identified in numerous diseases, including cancer. The biological functions of signal transducer and activator of transcription 3 (STAT3) are very broad. It is thought that STAT3 can also contribute to oncogenesis. RNA interference (RNAi) is one of the most efficient tools for silencing gene expression within cells. The main goal of the study was to verify the effectiveness of STAT3 gene silencing and its influence on cell proliferation and activation of apoptosis in bladder cancer cells.Material and methodsThe study was conducted on cellular material, which was the stable human bladder cancer cell line T24. The synthesis of shRNA (short hairpin RNA) interfering with the STAT3 gene was based on pSUPER. neo expression vector. The gene expression at the mRNA level was determined by the real-time PCR method. The influence of STAT3 gene silencing on apoptosis induced in cells with modulated STAT3 expression was evaluated using parallel quantification of mono- and oligonucleosomal DNA degradation of genomic DNA.ResultsIn transfected T24 cells, the STAT3 mRNA expression decreased to the level of 68.3% compared to the scrambled (SCR) control. Silencing the STAT3 gene induced changes in the phenotype of T24 cells. Statistically significant differences in cell proliferation (p = 0.0318) and apoptosis induction (p = 0.0376) were observed.ConclusionsApplication of the designed shRNA for the STAT3 gene contributed to a decrease of expression of the examined gene. It also decreased the proliferation and increased the susceptibility to apoptosis in T24 bladder cancer cells.