Resistance to conventional anti-tumor drugs is one of the significant challenges in oncology, responsible for treatment failure and patient death. Introduction of the targeted drugs (e.g., small molecule tyrosine kinase inhibitors (TKIs) and monoclonal antibodies) in cancer therapy significantly improved overall survival (OS) and progression-free survival (PFS) rates for selected groups of cancer patients and delayed the progression of advanced forms of human malignancies. However, the development of secondary resistance to the targeted drugs remains an unbeatable obstacle to a successful outcome in the long run, thereby making prognosis unfavorable for cancer patients with advanced, recurrent, and metastatic forms of disease. The review focuses on several mechanisms that regulate cancer resistance to conventional chemotherapies. This includes the upregulation of main types of ABC transporters (e.g., ABCB1, ABCC1, and ABCG2), which provides the efflux of chemotherapeutic agents from cancer cells. Additionally, the activation of diverse DNA damage repair (DDR) pathways, epithelial-to-mesenchymal transition (EMT), and the population of cancer stem cells (CSCs) are also discussed in detail, thereby illustrating the diverse molecular mechanisms of cancer sensitivity to chemotherapies. Recently, several TKIs, including those that were initially developed to specifically target FGFR and VEGFR pathways, have also been reported to exhibit “off-target” effects by interacting with ABC transporters and inhibiting their function. This, in turn, illustrates their potency in retaining chemotherapeutic agents within cancer cells and possessing a chemosensitizing function. Of note, FGFR and VEGFR inhibitors may behave as inhibitors or substrates of ABC transporters, depending on the expression of specific pumps and affinity for them, concentrations, and types of co-administered agents, thereby disclosing the complexity of this scenario. Additionally, the aforementioned RTKI can interfere with the other molecular mechanisms regulating tumor sensitivity to conventional chemotherapies, including the regulation of diverse DDR pathways, EMT, and the population of CSCs. Thereby, the aforementioned “off-target” functions of FGFR and VEGFR inhibitors can open novel approaches towards anti-cancer therapies and strategies aimed at counteracting cancer multidrug resistance (MDR), which is important especially as second- or third-line treatments in patients who have progressed on modern chemotherapeutic regimens. Notably, the strategy of using TKIs to potentiate the clinical efficacy of chemotherapies can extend beyond inhibitors of FGFR and VEGFR signaling pathways, thereby providing a rationale for repurposing existing TKIs as an attractive therapeutic approach to overcome cancer chemoresistance.

ABC transporters (ATP Binding Cassette (ABC) transporters) are proteins, which play a dual role in the substances transport across the membrane. On the one hand, they transport nutrients and other molecules inside the cell to supply the necessary nutrients, on the other hand, these proteins excrete some endogenous and exogenous substrates from the cell to maintain their homeostasis in the body and prevent from effects of aggressive environment. ABC transporters play a role in the pathogenesis of various metabolic disorders. In addition, a large amount of evidence has been accumulated about the participation of these proteins in oncogenesis because of their involvement into initiation, progression, invasion and metastasis of tumors, as well as development of multidrug resistance phenotype. Currently, these proteins are attractive therapeutic targets, influence on which can significantly increase the effectiveness of anticancer therapy and improve the prognosis of patients with oncological diseases, including recurrent, metastatic and inoperable forms.The review provides information on drugs that affect the functional activity of ABC transporters and the mechanisms of their action, and also presents the results of clinical trials of these inhibitors.

Introduction. ABC transporters (ATP-binding cassette transporters) are a class of membrane transporter proteins that play a crucial role in various physiological processes, including nutrient uptake, the secretion of signaling molecules, and the elimination of toxins. In addition to the above physiological functions, these proteins play a crucial role in the development of multidrug resistance in malignant neoplasms, thereby contributing to the progression of the disease. The main types of ABC transporters, for which their clinical significance in the development of drug resistance to chemotherapy and targeted therapy in many malignant neoplasms has been proven, include P-glycoprotein (P-gp, MDR1), BCRP and MRP1. However, the role of these proteins in the pathogenesis of gastrointestinal stromal tumors (GIST) and the formation of their resistance to chemotherapy is poorly understood. Aim. To assess the expression of ABC transporters (ABCB1/MDR1, ABCC1/MRP1, and ABCG2/BCRP), the intracellular content of their fluorescent substrates, and stemness markers in imatinib mesylate (IM) sensitive and resistant GIST cell lines, and their role in resistance to chemotherapy. Materials and methods. The study was conducted using GIST cell lines sensitive (GIST T-1) and resistant (GIST T-1R and GIST 430) to IM. As a positive control for the overexpression of ABC transporters in tumor cell lines, we utilized the previously obtained subline of triple-negative breast cancer HCC 1806 Tx-R, which has been proven to exhibit overexpression of ABC transporters. Results. We observed the increased expression of some ABC transporters (MRP1, ABCG2, and MDR1) in the IM-resistant GIST 430 cell line. This resulted in a significant increase in the excretion of their fluorescent substrate rhodamine 123 compared to the IM-resistant GIST T-1R cell line. At the same time, in the presence of P-gp inhibitors (cyclosporine A and tariquidar), as well as MRP1 (MK-571), there was a significant increase in the number of rhodamine-positive cells. A similar pattern was observed for some chemotherapeutic agents exhibiting autofluorescent activity, specifically an increase in the intensity of their fluorescence in the presence of corresponding ABC transporter inhibitors, indicating an increase in their intracellular concentration in GIST 430. Moreover, for doxorubicin, its excretion from GIST cells was proven exclusively by MRP1 protein, while mitoxantrone was “pumped out” from cells mainly through P-gp. The result of hyperexpression of these transporters in GIST 430 cells was their resistance to the drugs mentioned above, as evidenced by a 4- and 5-fold increase in the values of half-maximal inhibitory concentrations (IC50) for doxorubicin and mitoxantrone, respectively, in relation to GIST 430 cells compared to the GIST T-1R cell line. Based on the results of expression of stemness markers (CD44 and CD133) and colony formation, no evidence of the presence of a pool of cells with a stemness phenotype in the GIST 430 cell line was obtained. Conclusion. One of the mechanisms of secondary resistance of GIST 430 cells to chemotherapy is the hyperexpression of ABC transporters (MRP1, ABCG2, and MDR1), which is not associated with the presence of cells with a stemness phenotype in this cell line.

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.

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.