Secosteroid diacylhydrazines as novel effective agents against hormone-dependent breast cancer cells.

This research aimed to develop novel selective secosteroids that are highly active against hormone-dependent breast cancer. A simple and convenient approach to N'-acylated 13,17-secoestra-1,3,5(10)-trien-17-oic acid hydrazides was disclosed and these novel types of secosteroids were screened for cytotoxicity against the hormone-dependent human breast cancer cell line MCF7. Most secosteroid N'-benzoyl hydrazides have demonstrated high cytotoxicity against MCF7 cells with IC50 values below 5 µM, which are superior to that of the reference drug cisplatin. Hit compounds 2c, 2e and 2i were characterized by high cytotoxicity (IC50 = 1.6-1.9 µM) and very good selectivity towards MCF7 breast cancer cells. The lead secosteroids 2c, 2e and 2i also exhibit antiestrogenic effects and alter the expression of cell cycle regulating proteins. The effect of selected compounds on PARP (poly(ADP-ribose) polymerase) and Bcl-2 (B-cell CLL/lymphoma 2) indicates their proapoptotic potential. The synthesized secosteroids may be considered as new promising anti-breast cancer agents targeting ERα and apoptosis pathways.

Synthesis and evaluation of sulfonamide derivatives of quinoxaline 1,4-dioxides as carbonic anhydrase inhibitors.

A series of sulfonamide-derived quinoxaline 1,4-dioxides were synthesized and evaluated as inhibitors of carbonic anhydrases (CA) with antiproliferative potency. Overall, the synthesized compounds demonstrated good inhibitory activity against four CA isoforms. Compound 7g exhibited favorable potency in inhibiting a CA IX isozyme with a K i value of 42.2 nM compared to the reference AAZ (K i = 25.7 nM). Nevertheless, most of the synthesized compounds have their highest activity against CA I and CA II isoforms over CA IX and CA XII. A molecular modeling study was used for an estimation of the binding mode of the selected ligand 7g in the active site of CA IX. The most active compounds (7b, 7f, 7h, and 18) exhibited significant antiproliferative activity against MCF-7, Capan-1, DND-41, HL60, and Z138 cell lines, with IC50 values in low micromolar concentrations. Moreover, derivatives 7a, 7e, and 8g showed similar hypoxic cytotoxic activity and selectivity compared to tirapazamine (TPZ) against adenocarcinoma cells MCF-7. The structure-activity relationships analysis revealed that the presence of a halogen atom or a sulfonamide group as substituents in the phenyl ring of quinoxaline-2-carbonitrile 1,4-dioxides was favorable for overall cytotoxicity against most of the tested cancer cell lines. Additionally, the presence of a carbonitrile fragment in position 2 of the heterocycle also had a positive effect on the antitumor properties of such derivatives against the majority of cell lines. The most potent derivative, 3-trifluoromethylquinoxaline 1,4-dioxide 7h, demonstrated higher or close antiproliferative activity compared to the reference agents, such as doxorubicin, and etoposide, with an IC50 range of 1.3-2.1 µM. Analysis of the obtained results revealed important patterns in the structure-activity relationship. Moreover, these findings highlight the potential of selected lead sulfonamides on the quinoxaline 1,4-dioxide scaffold for further in-depth evaluation and development of chemotherapeutic agents targeting carbonic anhydrases.

Target Role of Monocytes as Key Cells of Innate Immunity in Rheumatoid Arthritis.

Rheumatoid arthritis (RA) is a chronic, systemic, and inflammatory autoimmune condition characterized by synovitis, pannus formation (with adjacent bone erosion), and joint destruction. In the perpetuation of RA, fibroblast-like synoviocytes (FLSs), macrophages, B cells, and CD4+ T-cells-specifically Th1 and Th17 cells-play crucial roles. Additionally, dendritic cells, neutrophils, mast cells, and monocytes contribute to the disease progression. Monocytes, circulating cells primarily derived from the bone marrow, participate in RA pathogenesis. Notably, CCR2 interacts with CCL2, and CX3CR1 (expressed by monocytes) cooperates with CX3CL1 (produced by FLSs), facilitating the migration involved in RA. Canonical "classical" monocytes predominantly acquire the phenotype of an "intermediate" subset, which differentially expresses proinflammatory cytokines (IL-1ß, IL-6, and TNF) and surface markers (CD14, CD16, HLA-DR, TLRs, and ß1- and ß2-integrins). However, classical monocytes have greater potential to differentiate into osteoclasts, which contribute to bone resorption in the inflammatory milieu; in RA, Th17 cells stimulate FLSs to produce RANKL, triggering osteoclastogenesis. This review aims to explore the monocyte heterogeneity, plasticity, antigenic expression, and their differentiation into macrophages and osteoclasts. Additionally, we investigate the monocyte migration into the synovium and the role of their cytokines in RA.

Irreversible inhibition of estrogen receptor α signaling and the emergence of hormonal resistance in MCF7 breast cancer cells induced by DNA damage agents.

Combining chemotherapy and hormone therapy is a prevalent approach in breast cancer treatment. While the cytotoxic impact of numerous chemotherapy drugs stems from DNA damage, the exact role of these DNA alterations in modulating estrogen receptor α (ERα) machinery remains elusive. The present study aimed to analyze the impact of DNA damage agents on ERα signaling in breast cancer cells and assess the signaling pathways mediating the influence of DNA damage drugs on the ERα machinery. Cell viability was assessed using the MTT method, while the expression of signaling proteins was analyzed by immunoblotting. ERα activity in the cells treated with various drugs (17ß-estradiol, tamoxifen, 5-fluorouracil) was assessed through reporter gene assays. In vitro experiments were conducted on MCF7 breast cancer cells subjected to varying durations of 5-fluorouracil (5-FU) treatment. Two distinct cell responses to 5-FU were identified based on the duration of the treatment. A singular dose of 5-FU induces pronounced DNA fragmentation, temporally suppressing ERα signaling while concurrently activating AKT phosphorylation. This suppression reverses upon 5-FU withdrawal, restoring normalcy within ten days. However, chronic 5-FU treatment led to the emergence of 5-FU-resistant cells with irreversible alterations in ERα signaling, resulting in partial hormonal resistance. These changes mirror those observed in cells subjected to UV-induced DNA damage, underscoring the pivotal role of DNA damage in shaping estrogen signaling alterations in breast cancer cells. In summary, the results of the present study suggested that the administration of DNA damage agents to cancer cells can trigger irreversible suppression of estrogen signaling, fostering the development of partial hormonal resistance. This outcome may ultimately impede the efficacy of combined or subsequent chemo- and hormone therapy strategies.

Secosteroid thiosemicarbazides and secosteroid-1,2,4-triazoles as antiproliferative agents targeting breast cancer cells: Synthesis and biological evaluation.

A convenient and selective approach to 13,17-secoestra-1,3,5(10)-trien-17-oic acid [N'-arylcarbothioamido]hydrazides and hybrid molecules containing secosteroid and 1,2,4-triazole fragments was disclosed and these novel types of secosteroids were screened for cytotoxicity against hormone-dependent human breast cancer cell line MCF-7. Most of secosteroid-1,2,4-triazole hybrids showed significant cytotoxic effect comparable or superior to that of the reference drug cisplatin. Hit secosteroid-1,2,4-triazole hybrids 4b and 4h were characterized by high cytotoxicity and good selectivity towards MCF-7 breast cancer cells. PARP cleavage (marker of apoptosis) and ERα and cyclin D1 downregulation were discovered in MCF-7 cells treated with lead secosteroid-1,2,4-triazole hybrid 4b. The synthesized secosteroids may be considered as new promising anticancer agents.

Exosomes are involved in the intercellular transfer of rapamycin resistance in the breast cancer cells.

Introduction: Resistance to chemotherapy and/or irradiation remains one of the key features of malignant tumors, which largely limits the efficiency of antitumor therapy. In this work, we studied the progression mechanism of breast cancer cell resistance to target drugs, including mTOR blockers, and in particular, we studied the exosome function in intercellular resistance transfer. Methods: The cell viability was assessed by the MTT assay, exosomes were purified by successive centrifugations, immunoblotting was used to evaluate protein expression, AP-1 activity was analyzed using reporter assay. Results: In experiments on the MCF-7 cell line (breast cancer) and the MCF-7/Rap subline that is resistant to rapamycin, the capability of resistant cell exosomes to trigger a similar rapamycin resistance in the parent MCF-7 cells was demonstrated. Exosome-induced resistance reproduces the changes revealed in MCF-7/Rap resistant cells, including the activation of ERK/AP-1 signaling, and it remains for a long time, for at least several months, after exosome withdrawal. We have shown that both the MCF-7 subline resistant to rapamycin and cells having exosome-triggered resistance demonstrate a stable decrease in the expression of DNMT3A, the key enzyme responsible for DNA methylation. Knockdown of DNMT3A in MCF-7 cells by siRNA leads to partial cell resistance to rapamycin; thus, the DNMT3A suppression is regarded as one of the necessary elements for the development of acquired rapamycin resistance. Conclusion: We propose that DNA demethylation followed by increased expression of key genes may be one of the factors responsible for the progression and maintenance of the resistant cell phenotype that includes exosome-induced resistance.

Novel pentacyclic derivatives and benzylidenes of the progesterone series cause anti-estrogenic and antiproliferative effects and induce apoptosis in breast cancer cells.

The promising antitumor effects of progesterone derivatives have been identified in many studies. However, the specific mechanism of action of this class of compounds has not been fully described. Therefore, in this study, we investigated the antiproliferative and (anti)estrogenic activities of novel pentacyclic derivatives and benzylidenes of the progesterone series. The antiproliferative effects of the compounds were evaluated on hormone-dependent MCF7 breast cancer cells using the MTT test. Estrogen receptor α (ERα) activity was assessed by a luciferase-based reporter assay. Immunoblotting was used to evaluate the expression of signaling proteins. All benzylidenes demonstrated inhibitory effects with IC50 values below 10 µM, whereas pentacyclic derivatives were less active. These patterns may be associated with the lability of the geometry of benzylidene molecules, which contributes to an increase in the affinity of interaction with the receptor. The selected compounds showed significant anti-estrogenic potency. Benzylidene 1d ((8 S,9 S,10R,13 S,14 S,17 S)-17-[(2E)-3-(4-fluorophenyl)prop-2-enoyl]-10,13-dimethyl-1,2,6,7,8,9,11,12,14,15-decahydrocyclopenta[a]phenanthren-3-one) was the most active in antiproliferative and anti-estrogenic assays. Apoptosis induced by compound 1d was accompanied by decreases in CDK4, ERα, and Cyclin D1 expression. Compounds 1d and 3d were characterized by high inhibitory potency against resistant breast cancer cells. Apoptosis induced by the leader compounds was confirmed by PARP cleavage and flow cytometry analysis. Compound 3d caused cell arrest in the G2/M phase. Further analysis of novel derivatives of the progesterone series is of great importance for medicinal chemistry, drug design, and oncology.

Secosteroid-quinoline hybrids as new anticancer agents.

An elegant approach to unknown secosteroid-quinoline hybrids is disclosed. A series of 13,17-secoestra-1,3,5(10)-trien-17-oic acid [N'-(iso)quinolylmethylene]hydrazides was prepared and these novel type of secosteroids was screened for antiproliferative activity against estrogen-responsive human breast cancer cell line MCF-7. Most of the synthesized compounds showed a cytotoxic effect superior to that of reference drug cisplatin; the lead compound exhibits the highest activity with the IC50 value of about 0.8 µM and is 7 times more active than cisplatin. A high selectivity index was observed for the hit 13,17-secoestra-1,3,5(10)-trien-17-oic acid [N'-quinolylmethylene]hydrazides 2a and 2c. Compounds 2a and 2c evaluated in luciferase reporter assays exhibited high antiestrogenic potency which was superior to that of tamoxifen. These hit compounds were characterized by high activity against MCF-7 cells that retained towards multidrug-resistant NCI/ADR-RES cells.

Synthesis and Biological Evaluation of Chalconesulfonamides: En Route to Proapoptotic Agents with Antiestrogenic Potency.

Breast and other estrogen receptor α-positive cancers tend to develop resistance to existing drugs. Chalcone derivatives possess anticancer activity based on their ability to form covalent bonds with targets acting as Michael acceptors. This study aimed to evaluate the anticancer properties of a series of chalcones (7a-l) with a sulfonamide group attached to the vinyl ketone moiety. Chalconesulfonamides showed a potent antiproliferative effect at low micromolar concentrations against several cancer cell lines, including ERα-positive 4-hydroxytamoxifen-resistant MCF7/HT2. Immunoblotting of samples treated with the lead compound 7e revealed its potent antiestrogenic activity (ERα/GREB1 axis) and induction of PARP cleavage (an apoptosis marker) in breast cancer cells. The obtained compounds represent a promising basis for further development of targeted drugs blocking hormone pathways in cancer cells.

Indoline-5-Sulfonamides: A Role of the Core in Inhibition of Cancer-Related Carbonic Anhydrases, Antiproliferative Activity and Circumventing of Multidrug Resistance.

The overexpression and activity of carbonic anhydrase (CA, EC 4.2.1.1) isoforms CA IX and CA XII promote the accumulation of exceeding protons and acidosis in the extracellular tumor environment. Sulfonamides are effective inhibitors of most families of CAs. In this study, using scaffold-hopping, indoline-5-sulfonamide analogs 4a-u of the CA IX-selective inhibitor 3 were designed and synthesized to evaluate their biological properties. 1-Acylated indoline-5-sulfonamides demonstrated inhibitory activity against tumor-associated CA IX and XII with KI values up to 132.8 nM and 41.3 nM. Compound 4f, as one of the most potent inhibitors of CA IX and XII, exhibits hypoxic selectivity, suppressing the growth of MCF7 cells at 12.9 µM, and causes partial inhibition of hypoxia-induced CA IX expression in A431 skin cancer cells. 4e and 4f reverse chemoresistance to doxorubicin of K562/4 with overexpression of P-gp.

Design, Synthesis and In Vitro Investigation of Cabozantinib-Based PROTACs to Target c-Met Kinase.

(1) Background: This investigation aimed at developing a series of c-Met-targeting cabozantinib-based PROTACs. (2) Methods: Purification of intermediate and target compounds was performed using column chromatography, in vitro antiproliferation activity was measured using a standard MTT assay and a c-Met degradation assay was performed via the immunoblotting technique. (3) Results: Several compounds exhibited antiproliferative activity towards different cell lines of breast cancer (T47D, MDA-MB-231, SKBR3, HCC1954 and MCF7) at the same level as parent cabozantinib and 7-demethyl cabozantinib. Two target conjugates, bearing a VHL-ligand as an E3-ligase binding moiety and glycol-based linkers, exhibited the effective inhibition of c-Met phosphorylation and an ability to decrease the level of c-Met in HCC1954 cells at micromolar concentrations. (4) Conclusions: Two compounds exhibit c-Met inhibition activity in the nanomolar range and can be considered as PROTAC molecules due to their ability to decrease the total level of c-Met in HCC1954 cells. The structures of the offered compounds can be used as starting points for further evaluation of cabozantinib-based PROTACs.

Synthesis and biological activity of 21,22-cyclosteroids and their derivatives.

Synthesis of 21,22-cyclosteroids has been achieved starting from pregnenolone acetate. The key transformation was the Kulinkovich reaction of 17-vinyl steroids with esters. The resulting cyclopropanols were further subjected to three-membered ring-opening under various conditions including to base-, palladium or visible light-promoted isomerization and cross-coupling reaction. A number of steroidal Δ2-6-ketones and 3ß-hydroxy-Δ5-enes with functional groups at C-21 - C-23 have been synthesized via the 21,22-cyclosteroids. The antiproliferative and antihormonal activity of the obtained compounds on the cell lines of prostate (22Rv1) and breast (MCF-7) cancer was studied. The androgen receptor activity was assessed by reporter assay when the expression of signalling proteins was evaluated by immunoblotting. (20S,22R)-22-Acetoxy-21,22-cyclo-5α-cholest-5-ene with the moderate antiandrogenic potency revealed IC50 values of 18.4 ± 1.2 and 14.6 ± 1.4 µM against MCF-7 and 22Rv1 cells, respectively, and its effects on the expression of AR-V7, cyclin D1 and BCL2 were explored.

Synthesis, Molecular Docking, In Vitro and In Vivo Studies of Novel Dimorpholinoquinazoline-Based Potential Inhibitors of PI3K/Akt/mTOR Pathway.

A (series) range of potential dimorpholinoquinazoline-based inhibitors of the PI3K/Akt/mTOR cascade was synthesized. Several compounds exhibited cytotoxicity towards a panel of cancer cell lines in the low and sub-micromolar range. Compound 7c with the highest activity and moderate selectivity towards MCF7 cells which express the mutant type of PI3K was also tested for the ability to inhibit PI3K-(signaling pathway) downstream effectors and associated proteins. Compound 7c inhibited the phosphorylation of Akt, mTOR, and S6K at 125-250 nM. It also triggered PARP1 cleavage, ROS production, and cell death via several mechanisms. Inhibition of PI3Kα was observed at a concentration of 7b 50 µM and of 7c 500 µM and higher, that can indicate minority PI3Kα as a target among other kinases in the titled cascade for 7c. In vivo studies demonstrated an inhibition of tumor growth in the colorectal tumor model. According to the docking studies, the replacement of the triazine core in gedatolisib (8) by a quinazoline fragment, and incorporation of a (hetero)aromatic unit connected with the carbamide group via a flexible spacer, can result in more selective inhibition of the PI3Kα isoform.

Photostability and Antiproliferative Activity of Furan Analogues of Combretastatin A-4.

Cancer is one of the most serious health problems that usually require heavy medical treatment. It is important to ensure that no additional burden is placed on patients due to the modes of administration and/or poor quality of pharmaceuticals. In this regard, understanding, quantifying, and improving the photostability (resistance to UV light or sunlight) of drugs is among the important elements that can improve the patient's quality of life. In this work, the photochemical properties of a wide range of furanone analogues of combretastatin A-4 and their antiproliferative activity against A-431 epidermoid carcinoma cells were studied in a search for compounds with improved photostability and antiproliferative activity. It was found that the incorporation of an arylidene moiety led to a significant improvement in photostability, while the antiproliferative activity strongly depends on the nature of the aryl residue in the arylidene moiety. The high photostability of arylidenes was achieved due to the delocalization of the central double bond of the 1,3,5-hexatriene system, which limited the 6π-electrocyclization. The best results in terms of antiproliferative activity were obtained for thiophene arylidene (IC50 = 0.6 µM) and 3,4-diarylfuran (IC50 = 0.047 µM). The obtained results address the lack of data available now in scientific literature on the photodegradation of combretastatin A-4 analogues and should be taken into account in studies of the side effects of pharmaceuticals based on them.

Synthesis, biological evaluation, and in silico studies of potential activators of apoptosis and carbonic anhydrase inhibitors on isatin-5-sulfonamide scaffold.

Carbonic anhydrase IX is a promising target for the search for new antitumor compounds with improved properties. Using the molecular hybridization approach, on the basis of structures of a selective carbonic anhydrase IX inhibitor 3 and an activator of apoptosis 2 (1), a series of 1-substituted isatin-5-sulfonamides 5a-5u were designed and synthesized. The study of the inhibitory activity of isatin-5-sulfonamides showed the ability to inhibit I, II, IX, XII isoforms at nano- and micromolar concentrations. Docking of compounds 5e and 5k into the active site of II and IX carbonic anhydrase isoforms showed the coordination of sulfonamidate anions with zinc cations, as well as a number of additional hydrophobic interactions. The trifluoromethylthio derivative 5r suppressed the growth of tumor cells at low micromolar concentrations, maintaining activity on resistant lines and under hypoxic conditions. Immunoblotting of MCF7 cells treated with the 5r revealed its antiestrogenic activity and ability to activate apoptosis in tumor cells.

Secosteroidal hydrazides: Promising scaffolds for anti-breast cancer agents.

A convenient and selective approach to 13,17-secoestra-1,3,5(10)-trien-17-oic acid hydrazides and their N'-(het)arylmethylene derivatives was disclosed and these novel types of secosteroids were screened for cytotoxicity against hormone-dependent human breast cancer cell line MCF-7. A number of 13,17-secoestra-1,3,5(10)-trien-17-oic acid [N'-(het)arylmethylene]hydrazides show significant cytotoxic effect comparable or superior to that for reference drug cisplatin. Compound 3l exhibits the highest activity with the IC50 value of about 2 µM and is 2.8 times more active than cisplatin. Hit 13,17-secoestra-1,3,5(10)-trien-17-oic acid [N'-(het)arylmethylene]hydrazides 3d, 3l and 3q are characterized by high cytotoxicity and good selectivity towards MCF-7 breast cancer cells. The synthesized secosteroids may be considered as new promising antitumor agents.

Antiestrogenic and antiproliferative potency of secoisolariciresinol diglucoside derivatives on MCF-7 breast cancer cells.

Secoisolariciresinol diglucoside (SDG) is isolated from Linum usitatissimum seeds. The antiproliferative effects of SDG (1) and its derivatives secoisolariciresinol (2) and secoisolariciresinol-4', 4″-diacetate (3) have been evaluated on MCF-7 breast cancer cells and normal breast epithelial line MCF-10A. Lignan 1 has not shown cytotoxic effects on MCF-7 cells, while derivatives 2 and 3 have inhibited cell growth with IC50 values of 25 and 11 µM, respectively. Estrogen receptor alpha is a key growth driver in MCF-7 cells. Compound 1 did not affect the activity of ERα, while derivatives 2 and 3 showed significant antiestrogenic effects. Compounds 2 and 3 caused apoptosis in the MCF-7 line, determined by the cleavage of PARP. SDG derivative 3 enhanced the effect of doxorubicin. SDG derivatives can be considered as promising agents that exhibit a combined antiestrogen and proapoptotic effect in hormone-dependent breast cancer cells.

Fe(iii)-Catalyzed synthesis of steroidal imidazoheterocycles as potent antiproliferative agents.

An efficient and practical method has been developed for the synthesis of steroidal imidazoheterocycles via cost-effective and environmentally benign FeCl3-catalyzed oxidative amination. A library of steroidal imidazo[1,2-a]pyridines was directly synthesized from readily available 2-aminopyridines and steroidal ketones in aerobic conditions. The synthesized compounds were screened for activity on human microsomal cytochrome P450s CYP7, CYP17 and CYP21. Antiproliferative activity of two lead compounds 3ia and 3la was additionally evaluated against the human MCF-7 (breast cancer), SKOV3 (ovarian cancer), and 22Rv1 (prostate cancer) cell lines. Steroidal imidazo[1,2-a]pyridine 3la which is a substrate molecule for CYP17A1 with IC50 = 1.7 µM (MCF-7), 3.0 (SKOV3), and 6.0 µM (22Rv1) has proved to be more active than reference drug cisplatin.