Introduction. Microtubules are highly dynamic polymers of α, β-tubulin dimers involves in a broad spectrum of the processes, such as intracellular transport and cell proliferation. This makes them an attractive molecular target for anti-cancer therapies. Substances that affect the dynamic state of tubulin microtubules are known as the mitotic poisons that are effectiveand widely used in the chemotherapy of various tumors. Mitotic poisons are able to interfere with polymerization (stabilization) or depolymerization of tubulin, which in turn leads to the arrest of cells in the M-phase (named as a mitotic catastrophe) and their subsequent death via activation of apoptotic mechanisms. However, the effectiveness of MP-based therapies is gradually decreasing over the time due to development of multiple drug resistance mechanisms in cancer cells. Thus, development of novel compounds selectively targeting tubulin and effectively overcoming multiple drugresistance phenotype in cancer is an urgent need in current oncology. Aim. To examine the cytotoxic and antitumor activities of several pyrrole-containing heterocyclic compounds (EPC-91, EPC-92 and PCA-93) against cancer cell lines with epithelial and mesenchymal origin, including those with multiple drug resistance phenotype. Materials and methods. Studies were performed on parental human cancer cell lines – triple-negative breast cancer HCC1806, gastrointestinal stromal tumor GIST T-1, osteosarcoma SaOS-2, – sensitive to chemotherapy (paclitaxel, doxorubicin) and their resistant sublines (HCC1806 Tx-R, GIST T-1 Tx-R, SaOS-2 Dox-R), as well as on murine colorectal adenocarcinoma cell line Colon-26, exhibiting primary resistance to the aforementioned chemotherapeutic agents. Results. The cytotoxic activities of EPC-91 and PCA-93 were due to their abilities to depolymerize tubulin. The results of immunofluorescence microscopy and Western blotting indicated that the compounds disrupt assembly of tubulin microtubules and prevent polymerization of α-tubulin in cancer cells. Inhibition of tubulin polymerizations led to significant increasein number of round-shaped and phospho-histone 3 (e. g. mitotic) cells, followed by their death through apoptosis. PCA-93 also exhibited potent anti-tumor effect against Colon-26 cells due to its anti-proliferative and proapoptotic activities. Conclusion. The data shown here illustrates potent cytotoxic activities of EPC-91 and PCA-93 against multiple cancer cell lines in vitro including those with multiple drug resistance phenotype. Similarly, PCA-93 was found to be highly effective against Colon-26 cell in vivo, thereby illustrating the attractive platform for the development of novel pyrrole-based agents exhibiting potent anti-tumor activities.

Background: Hypoxia-inducible factor-1α (HIF-1α) is a well-known transcriptional regulator that mediates a broad spectrum of cellular responses to hypoxia, including angiogenesis, extracellular matrix remodeling, and metabolic reprogramming. These activities can be achieved by upregulation of numerous genes, such as vascular endothelial growth factors, fibroblast growth factors, and platelet-derived growth factors, which are involved in the growth regulation of normal tissues and solid tumors. Notably, HIF-1α-mediated regulation of the solid tumor’s microenvironment effectively modulates tumor sensitivity to anticancer therapies and thereby can contribute to disease progression. Methods: The study was performed on breast, lung and prostate cancer cell lines. Protein expression was examined by western blotting. Antitumor activity of 2-ANPC was measured by syngeneic 4T1 breast cancer mouse model. Results: We show here that a 2-aminopyrrole derivative (2-amino-1-benzamido-5-(2-(naphthalene-2-yl)-2-oxoethylidene)-4-oxo-4,5-dihydro-1-H-pyrrole-3-carboxamide—2-ANPC), previously shown as a potent microtubule-targeting agent, effectively downregulates HIF-1α expression in a broad spectrum of cancer cell lines, including breast, lung, and prostate cancer. The downregulation of HIF-1α expression in 2-ANPC-treated cancer cells was due to enhanced proteasome-mediated degradation, whereas the proteasome inhibitor MG-132 effectively reversed this downregulation. 2-ANPC’s potency in downregulating HIF-1α was also shown in vivo by using the 4T1 breast cancer syngraft model. Importantly, this 2-aminopyrrole derivative also downregulated the expression of vascular endothelial growth factor receptors 1 and 3 (VEGFR1 and 3) in 4T1 tumors, which correlated with decreased tumor weight and size. As expected, an increase in apoptotic (i.e., cleaved caspase-3-positive) cells was detected in 4T1 tumors treated with 2-aminopyrrole derivative. Lastly, using various computational tools, we identified four potential binding sites for 2-ANPC to interact with HIF-1α, HIF-1β, and the p300 complex. Conclusions: Collectively, we show here, for the first time, that HIF-1α is a novel molecular target for the 2-aminopyrrole derivative (2-ANPC), thereby illustrating it as a potential scaffold for the development of potent chemotherapeutic agents with anti-angiogenic activity.

Background. Local therapy aimed to suppress infection of the osteoarticular apparatus is based on the use of coated spacers. As a rule, the coating surfaces are composed of bone cement supplemented with bioactive substances, e.g., antibiotics. However, current approaches aimed to stimulate the release of antibiotics from bone cement are not always effective and may even decrease their properties and biological activities. The aim of the study — to examine the effect of the lattice structures of base implants on the dynamics and amount of antibiotics released from bone cement in vitro. Methods. A novel type of implants with a lattice structure and traditional reinforced spacer filled with bone cement and antibiotic were used in present study. The release of vancomycin and cefazolin was measured spectrophotometrically for up to 30 days. Calibration curves were made according to the regression lines based on the values of the stock solutions with the well-known concentrations of the antibiotics. Results. For all tested samples, the elution profile of antibiotics was determined and illustrated the significant release of the drugs during the initial time-period followed by the decrease (day 7) and reaching a uniform elution plateau (day 15). In all samples, the amount of released antibiotic did not exceed 1% of total amount of the drug present in the implant. Notably, the higher release of the antibiotics was found for the samples with lattice structure when compared with control group, despite the relatively higher initial amount of bone cement and antibiotics in control samples. A difference in the amount of antibiotics was also detected between the tested samples of two types of lattice structures. Conclusion. The lattice structure of the implants increases the amount of antibiotic release from the bone cement and also facilitates the antibiotic dynamics release when compared to the traditional bone cement used as the reinforced spacers.

Introduction The established treatments for purulent infection in the bone and joint involve one- or two‑stage local effect on the biofilm with use of bone cement and an active substance including an antibiotic in addition to systemic therapy.The objective was to evaluate experimental qualitative and quantitative antibiotic release from bone cement introduced into a new type of lattice-structured spacer.Material and methods A new type of lattice-structured implant/spacer manufactured using additive technologies and a comparison sample simulating a traditional reinforced spacer made of bone cement + antibiotic were used. Vancomycin release was measured by spectrophotometry for periods of 30 days. A regression line was used to plot calibration curves based on data obtained from mother solutions.Results An effective profile of antibiotic release from bone cement was obtained in the first days of the experiment, followed by a decrease at the end of the first week and an exit to a uniform plateau. The amount of fixed antibiotic in solutions did not exceed 1 % of the total mass of bone cement and active substance. The amount of antibiotic released from the lattice-structured samples was higher than that in the comparison samples.Discussion Antibiotic release is a superficial process and is not dependent on the total volume of bone cement. A possible increase in the volume of the medicinal composition does not lead to a proportional increase in the amount of the active substance released. The findings showed that the antibiotic release is more intense even with a smaller volume of material in the lattice structures compared to the control samples, which emphasizes the importance of optimizing the geometry and structure of the material to achieve maximum efficiency of the release of active substances.Conclusion The lattice structure of implants quantitatively affects the release of antibiotic from bone cement into the environment.

Hypoxia-inducible factor-1α (HIF-1α) is a well-known transcriptional regulator that me-diates a broad spectrum of cellular responses to hypoxia, including angiogenesis, extra-cellular matrix remodeling, and metabolic reprogramming. These activities can be achieved by up-regulation of numerous genes, such as vascular endothelial growth fac-tors, fibroblast growth factors, and platelet-derived growth factors, which are involved in the growth regulation of normal tissues and solid tumors. Of note, HIF-1α-mediated regu-lation of the solid tumor’s microenvironment effectively modulates tumor sensitivity to anticancer therapies and thereby can contribute to disease progression. We show here that a 2-aminopyrrole derivative (2-amino-1-benzamido-5-(2-(naphthalene-2-yl)-2-oxoethylidene)-4-oxo-4,5-dihydro-1-H-pyrrole-3-carboxamide – 2-ANPC), previously shown as a potent microtubule-targeting agent, effectively down-regulates HIF-1α expression in a broad spectrum of cancer cell lines, including breast, lung, and prostate cancer. The downregulation of HIF-1α expression in 2-ANPC-treated cancer cells was due to en-hanced proteasome-mediated degradation, whereas the proteasome inhibitor MG-132 ef-fectively reversed this downregulation. 2-ANPC’s potency to down-regulate HIF-1α was also shown in vivo by using the 4T1 breast cancer syngraft model. Important, this 2-aminopyrrole derivative also down-regulated the expression of vascular endothelial growth factor receptors 1 and 3 (VEGFR1 and 3) in 4T1 tumors, which correlated with de-creased tumor weight and size. As expected, an increase of apoptotic (i.e., cleaved caspa-se-3-positive) cells was detected in 4T1 tumors treated with 2-aminopyrrole derivative. Lastly, using various computational tools, we identified four potential binding sites for 2-ANPC to interact with HIF-1α, HIF-1β, and the p300 complex. Collectively, we show here, for the first time, that HIF-1α is a novel molecular target for the 2-aminopyrrole de-rivative (2-ANPC), thereby illustrating it as a potential scaffold for the development of po-tent chemotherapeutic agents with anti-angiogenic activity.