Are the metal identity and stoichiometry of metal complexes important for colchicine site binding and inhibition of tubulin polymerization?

Quite recently we discovered that copper(II) complexes with isomeric morpholine-thiosemicarbazone hybrid ligands show good cytotoxicity in cancer cells and that the molecular target responsible for this activity might be tubulin. In order to obtain better lead drug candidates, we opted to exploit the power of coordination chemistry to (i) assemble structures with globular shape to better fit the colchicine pocket and (ii) vary the metal ion. We report the synthesis and full characterization of bis-ligand cobalt(III) and iron(III) complexes with 6-morpholinomethyl-2-formylpyridine 4N-(4-hydroxy-3,5-dimethylphenyl)-3-thiosemicarbazone (HL1), 6-morpholinomethyl-2-acetylpyridine 4N-(4-hydroxy-3,5-dimethylphenyl)-3-thiosemicarbazone (HL2), and 6-morpholinomethyl-2-formylpyridine 4N-phenyl-3-thiosemicarbazone (HL3), and mono-ligand nickel(II), zinc(II) and palladium(II) complexes with HL1, namely [CoIII(HL1)(L1)](NO3)2 (1), [CoIII(HL2)(L2)](NO3)2 (2), [CoIII(HL3)(L3)](NO3)2 (3), [FeIII(L2)2]NO3 (4), [FeIII(HL3)(L3)](NO3)2 (5), [NiII(L1)]Cl (6), [Zn(L1)Cl] (7) and [PdII(HL1)Cl]Cl (8). We discuss the effect of the metal identity and metal complex stoichiometry on in vitro cytotoxicity and antitubulin activity. The high antiproliferative activity of complex 4 correlated well with inhibition of tubulin polymerization. Insights into the mechanism of antiproliferative activity were supported by experimental results and molecular docking calculations.

Copper(II) Complexes with Isomeric Morpholine-Substituted 2-Formylpyridine Thiosemicarbazone Hybrids as Potential Anticancer Drugs Inhibiting Both Ribonucleotide Reductase and Tubulin Polymerization: The Morpholine Position Matters.

The development of copper(II) thiosemicarbazone complexes as potential anticancer agents, possessing dual functionality as inhibitors of R2 ribonucleotide reductase (RNR) and tubulin polymerization by binding at the colchicine site, presents a promising avenue for enhancing therapeutic effectiveness. Herein, we describe the syntheses and physicochemical characterization of four isomeric proligands H2L3-H2L6, with the methylmorpholine substituent at pertinent positions of the pyridine ring, along with their corresponding Cu(II) complexes 3-6. Evidently, the position of the morpholine moiety and the copper(II) complex formation have marked effects on the in vitro antiproliferative activity in human uterine sarcoma MES-SA cells and the multidrug-resistant derivative MES-SA/Dx5 cells. Activity correlated strongly with quenching of the tyrosyl radical (Y•) of mouse R2 RNR protein, inhibition of RNR activity in the cancer cells, and inhibition of tubulin polymerization. Insights into the mechanism of antiproliferative activity, supported by experimental results and molecular modeling calculations, are presented.

Synthesis and biological evaluation of structurally diverse 6-aryl-3-aroyl-indole analogues as inhibitors of tubulin polymerization.

The synthesis and evaluation of small-molecule inhibitors of tubulin polymerization remains a promising approach for the development of new therapeutic agents for cancer treatment. The natural products colchicine and combretastatin A-4 (CA4) inspired significant drug discovery campaigns targeting the colchicine site located on the beta-subunit of the tubulin heterodimer, but so far these efforts have not yielded an approved drug for cancer treatment in human patients. Interest in the colchicine site was enhanced by the discovery that a subset of colchicine site agents demonstrated dual functionality as both potent antiproliferative agents and effective vascular disrupting agents (VDAs). Our previous studies led to the discovery and development of a 2-aryl-3-aroyl-indole analogue (OXi8006) that inhibited tubulin polymerization and demonstrated low nM IC50 values against a variety of human cancer cell lines. A water-soluble phosphate prodrug salt (OXi8007), synthesized from OXi8006, displayed promising vascular disrupting activity in mouse models of cancer. To further extend structure-activity relationship correlations, a series of 6-aryl-3-aroyl-indole analogues was synthesized and evaluated for their inhibition of tubulin polymerization and cytotoxicity against human cancer cell lines. Several structurally diverse molecules in this small library were strong inhibitors of tubulin polymerization and of MCF-7 and MDA-MB-231 human breast cancer cells. One of the most promising analogues (KGP591) caused significant G2/M arrest of MDA-MB-231 cells, disrupted microtubule structure and cell morphology in MDA-MB-231 cells, and demonstrated significant inhibition of MDA-MB-231 cell migration in a wound healing (scratch) assay. A phosphate prodrug salt, KGP618, synthesized from its parent phenolic precursor, KGP591, demonstrated significant reduction in bioluminescence signal when evaluated in vivo against an orthotopic model of kidney cancer (RENCA-luc) in BALB/c mice, indicative of VDA efficacy. The most active compounds from this series offer promise as anticancer therapeutic agents.

4-(3-Phenyl-4-(3,4,5-trimethoxybenzoyl)-1H-pyrrol-1-yl)benzenesulfonamide, a Novel Carbonic Anhydrase and Wnt/ß-Catenin Signaling Pathway Dual-Targeting Inhibitor with Potent Activity against Multidrug Resistant Cancer Cells.

We synthesized new pyrrole and indole derivatives as human carbonic anhydrase (hCA) inhibitors with the potential to inhibit the Wnt/ß-catenin signaling pathway. The presence of both N1-(4-sulfonamidophenyl) and 3-(3,4,5-trimethoxyphenyl) substituents was essential for strong hCA inhibitors. The most potent hCA XII inhibitor 15 (Ki = 6.8 nM) suppressed the Wnt/ß-catenin signaling pathway and its target genes MYC, Fgf20, and Sall4 and exhibited the typical markers of apoptosis, cleaved poly(ADP-ribose)polymerase, and cleaved caspase-3. Compound 15 showed strong inhibition of viability in a panel of cancer cells, including colorectal cancer and triple-negative breast cancer cells, was effective against the NCI/ADR-RES DOX-resistant cell line, and restored the sensitivity to doxorubicin (DOX) in HT29/DX and MDCK/P-gp cells. Compound 15 is a novel dual-targeting compound with activity against hCA and Wnt/ß-catenin. It thus has a broad targeting spectrum and is an anticancer agent with specific potential in P-glycoprotein overexpressing cell lines.

Structure Guided Design, Synthesis, and Biological Evaluation of Oxetane-Containing Indole Analogues.

The oxetane functional group offers a variety of potential advantages when incorporated within appropriate therapeutic agents as a ketone surrogate. OXi8006, a 2-aryl-3-aroyl-indole analogue, functions as a small-molecule inhibitor of tubulin polymerization that has a dual mechanism of action as both an antiproliferative agent and a tumor-selective vascular disrupting agent. Replacement of the bridging ketone moiety in OXi8006 with an oxetane functional group has expanded structure activity relationship (SAR) knowledge and provided insights regarding oxetane incorporation within this class of molecules. A new synthetic method using an oxetane-containing tertiary alcohol subjected to Lewis acid catalyzed conditions led to successful Friedel-Crafts alkylation and yielded fourteen new oxetane-containing indole-based molecules. This synthetic approach represents the first method to successfully install an oxetane ring at the 3-position of a 2-aryl-indole system. Several analogues showed potent cytotoxicity (micromolar GI50 values) against human breast cancer cell lines (MCF-7 and MDA-MB-231) and a pancreatic cancer cell line (PANC-1), although they proved to be ineffective as inhibitors of tubulin polymerization. Molecular docking studies comparing colchicine with the OXi8006-oxetane analogue 5m provided a rationale for the differential interaction of these molecules with the colchicine site on the tubulin heterodimer.

Indolo[2,3-e]benzazocines and indolo[2,3-f]benzazonines and their copper(II) complexes as microtubule destabilizing agents.

A series of four indolo[2,3-e]benzazocines HL1-HL4 and two indolo[2,3-f]benzazonines HL5 and HL6, as well as their respective copper(II) complexes 1-6, were synthesized and characterized by 1H and 13C NMR spectroscopy, ESI mass spectrometry, single crystal X-ray diffraction (SC-XRD) and combustion analysis (C, H, N). SC-XRD studies of precursors Vd, VIa·0.5MeOH, of ligands HL4 and HL6·DCM, and complexes 2·2DMF, 5·2DMF, 5'·iPrOH·MeOH provided insights into the energetically favored conformations of eight- and nine-membered heterocycles in the four-ring systems. In addition, proton dissociation constants (pKa) of HL1, HL2 and HL5, complexes 1, 2 and 5, overall stability constants (log ß) of 1, 2 and 5 in 30% (v/v) DMSO/H2O at 298 K, as well as thermodynamic solubility of HL1-HL6 and 1-6 in aqueous solution at pH 7.4 were determined by UV-vis spectroscopy. All compounds were tested for antiproliferative activity against Colo320, Colo205 and MCF-7 cell lines and showed IC50 values in the low micromolar to sub-micromolar concentration range, while some of them (HL1, HL5 and HL6, 1, 2 and 6) showed remarkable selectivity towards malignant cell lines. Ethidium bromide displacement studies provided evidence that DNA is not the primary target for these drugs. Rather, inhibition of tubulin assembly is likely the underlying mechanism responsible for their antiproliferative activity. Tubulin disassembly experiments showed that HL1 and 1 are effective microtubule destabilizing agents binding to the colchicine site. This was also confirmed by molecular modelling investigations. To the best of our knowledge, complex 1 is the first reported transition metal complex to effectively bind to the tubulin-colchicine pocket.

Identification of pyrrolo[3',4':3,4]cyclohepta[1,2-d][1,2]oxazoles as promising new candidates for the treatment of lymphomas.

Unsatisfactory outcomes for relapsed/refractory lymphoma patients prompt continuing efforts to develop new therapeutic strategies. Our previous studies on pyrrole-based anti-lymphoma agents led us to synthesize a new series of twenty-six pyrrolo[3',4':3,4]cyclohepta[1,2-d] [1,2]oxazole derivatives and study their antiproliferative effects against a panel of four non-Hodgkin lymphoma cell lines. Several candidates showed significant anti-proliferative effects, with IC50's reaching the sub-micromolar range in at least one cell line, with compound 3z demonstrating sub-micromolar growth inhibitory effects towards the entire panel. The VL51 cell line was the most sensitive, with an IC50 value of 0.10 µM for 3z. Our earlier studies had shown that tubulin was a prominent target of many of our oxazole derivatives. We therefore examined their effects on tubulin assembly and colchicine binding. While 3u and 3z did not appear to target tubulin, good activity was observed with 3d and 3p. Molecular docking and molecular dynamics simulations allowed us to rationalize the binding mode of the synthesized compounds toward tubulin. All ligands exhibited a better affinity for the colchicine site, confirming their specificity for this binding pocket. In particular, a better affinity and free energy of binding was observed for 3d and 3p. This result was confirmed by experimental data, indicating that, although both 3d and 3p significantly affected tubulin assembly, only 3d showed activity comparable to that of combretastatin A-4, while 3p was about 4-fold less active. Cell cycle analysis showed that compounds 3u and especially 3z induced a block in G2/M, a strong decrease in S phase even at low compound concentrations and apoptosis through the mitochondrial pathway. Thus, the mechanism of action of 3u and 3z remains to be elucidated. Very high selectivity toward cancer cells and low toxicity in human peripheral blood lymphocytes were observed, highlighting the good potential of these agents in cancer therapy and encouraging further exploration of this compound class to obtain new small molecules as effective lymphoma treatments.

RS6077 induces mitotic arrest and selectively activates cell death in human cancer cell lines and in a lymphoma tumor in vivo.

We synthesized a new inhibitor of tubulin polymerization, the pyrrole (1-(7H-pyrrolo[2,3- d]pyrimidin-4-yl)-1H-pyrrol-3-yl)(3,4,5-trimethoxy-phenyl)methanone 6 (RS6077). Compound 6 inhibited the growth of multiple cancer cell lines, with IC50 values in the nM range, without affecting the growth of non-transformed cells. The novel agent arrested cells in the G2/M phase of the cell cycle in both transformed and non-transformed cell lines, but single cell analysis by time-lapse video recording revealed a remarkable selectivity in cell death induction by compound 6: in RPE-1 non-transformed cells mitotic arrest induced was not necessarily followed by cell death; in contrast, in HeLa transformed and in lymphoid-derived transformed AHH1 cell lines, cell death was effectively induced during mitotic arrest in cells that fail to complete mitosis. Importantly, the agent also inhibited the growth of the lymphoma TMD8 xenograft model. Together these findings suggest that derivative 6 has a selective efficacy in transformed vs non-transformed cells and indicate that the same compound has potential as novel therapeutic agent to treat lymphomas. Compound 6 showed good metabolic stability upon incubation with human liver microsomes.

Induction of Ferroptosis in Glioblastoma and Ovarian Cancers by a New Pyrrole Tubulin Assembly Inhibitor.

We synthesized new aroyl diheterocyclic pyrrole (ARDHEP) 15 that exhibited the hallmarks of ferroptosis. Compound 15 strongly inhibited U-87 MG, OVCAR-3, and MCF-7 cancer cells, induced an increase of cleaved PARP, but was not toxic for normal human primary T lymphocytes at 0.1 µM. Analysis of the levels of lactoperoxidase, malondialdehyde, lactic acid, total glutathione, and ATP suggested that the in vivo inhibition of cancer cell proliferation by 15 went through stimulation of oxidative stress injury and Fe2+ accumulation. Quantitative polymerase chain reaction analysis of the mRNA expression in U-87 MG and SKOV-3 tumor tissues from 15-treated mice showed the presence of Ptgs2/Nfe2l2/Sat1/Akr1c1/Gpx4 genes correlated with ferroptosis in both groups. Immunofluorescence staining revealed significantly lower expressions of proteins Ki67, CD31, and ferroptosis negative regulation proteins glutathione peroxidase 4 (GPX4) and FTH1. Compound 15 was found to be metabolically stable when incubated with human liver microsomes.

Development of [1,2]oxazoloisoindoles tubulin polymerization inhibitors: Further chemical modifications and potential therapeutic effects against lymphomas.

Lymphomas are among the ten most common cancers, and, although progress has been achieved in increasing survival, there is still an unmet need for more effective therapeutic approaches, including better options for patients with refractory tumors that initially respond but then relapse. The lack of effective alternative treatment options highlights the need to develop new therapeutic strategies capable of improving survival prospects for lymphoma patients. Herein, we describe the identification and exploration of the SAR of a series of [1,2]oxazolo[5,4-e]isoindoles as potent small molecules that bind to the colchicine site of tubulin and that have promise for the treatment of refractory lymphomas. Exploration of the chemical space of this class of compounds at the pyrrole moiety and at the [1,2]oxazole ring highlighted two compounds bearing a 3,5-dimethoxybenzyl and a 3,4,5-trimethoxybenzyl group as potent candidates and showed that structural modifications at the isoxazole moiety are generally not favorable for activity. The two best candidates showed efficacy against different lymphoma histotypes and displayed 88 and 80% inhibition of colchicine binding fitting well into the colchicine pocket, as demonstrated by X-ray crystallography T2R-TTL-complexes, docking and thermodynamic analysis of the tubulin-colchicine complex structure. These results were confirmed by transcriptome data, thus indicating [1,2]oxazolo[5,4-e]isoindoles are promising candidates as antitubulin agents for the treatment of refractory lymphomas.

2-Difluoromethoxy-Substituted Estratriene Sulfamates: Synthesis, Antiproliferative SAR, Antitubulin Activity, and Steroid Sulfatase Inhibition.

2-Difluoromethoxyestratriene derivatives were designed to improve potency and in vivo stability of the drug candidate 2-methoxyestradiol (2ME2). Compound evaluation in vitro against the proliferation of MCF-7 and MDA MB-231 breast cancer cells, as inhibitors of tubulin polymerisation and also steroid sulfatase (STS) both in cell lysates and in whole cells, showed promising activities. In antiproliferative assays 2-difluoromethoxyestradiol was less potent than 2ME2, but its sulfamates were often more potent than their corresponding non-fluorinated analogues. The fluorinated bis-sulfamate is a promising antiproliferative agent in MCF-7 cells (GI50 0.28 µM) vs the known 2-methoxyestradiol-3,17-O,O-bissulfamate (STX140, GI50 0.52 µM), confirming the utility of our approach. Compounds were also evaluated in the NCI 60-cell line panel and the fluorinated bis-sulfamate derivative displayed very good overall activities with a sub-micromolar average GI50 . It was a very potent STS inhibitor in whole JEG-3 cells (IC50 3.7 nM) similar to STX140 (4.2 nM) and additionally interferes with tubulin assembly in vitro and colchicine binding to tubulin. An X-ray study of 2-difluoromethoxy-3-benzyloxyestra-1,3,5(10)-trien-17-one examined conformational aspects of the fluorinated substituent. The known related derivative 2-difluoromethyl-3-sulfamoyloxyestrone was evaluated for STS inhibition in whole JEG-3 cells and showed an excellent IC50 of 55 pM.

The interaction of spongistatin 1 with tubulin.

A tritiated derivative of the sponge-derived natural product spongistatin 1 was prepared, and its interactions with tubulin were examined. [3H]Spongistatin 1 was found to bind rapidly to tubulin at a single site (the low specific activity of the [3H]spongistatin 1, 0.75 Ci/mmol, prevented our defining an association rate), and the inability of spongistatin 1 to cause an aberrant assembly reaction was confirmed. Spongistatin 1 bound to tubulin very tightly, and we could detect no significant dissociation reaction from tubulin. The tubulin-[3H]spongistatin 1 complex did dissociate in 8 M urea, so there was no evidence for covalent bond formation. Apparent KD values were obtained by Scatchard analysis of binding data and by Hummel-Dreyer chromatography (3.5 and 1.1 µM, respectively). The effects of a large cohort of vinca domain drugs on the binding of [3H]spongistatin 1 to tubulin were evaluated. Compounds that did not cause aberrant assembly reactions (halichondrin B, eribulin, maytansine, and rhizoxin) caused little inhibition of [3H]spongistatin 1 binding. Little inhibition also occurred with the peptides dolastatin 15, its active pentapeptide derivative, vitilevuamide, or diazonamide A, nor with the vinca alkaloid vinblastine. Strong inhibition was observed with dolastatin 10, hemiasterlin, and cryptophycin 1, all of which cause aberrant assembly reactions that might actually mask the spongistatin 1 binding site. Spongistatin 5 was found to be a competitive inhibitor of [3H]spongistatin 1 binding, with an apparent Ki of 2.2 µM. We propose that the strong picomolar cytotoxicity of spongistatin 1 probably derives from its extremely tight binding to tubulin.

Discovery of pyrrole derivatives for the treatment of glioblastoma and chronic myeloid leukemia.

Long-term survivors of glioblastoma multiforme (GBM) are at high risk of developing second primary neoplasms, including leukemia. For these patients, the use of classic tyrosine kinase inhibitors (TKIs), such as imatinib mesylate, is strongly discouraged, since this treatment causes a tremendous increase of tumor and stem cell migration and invasion. We aimed to develop agents useful for the treatment of patients with GBM and chronic myeloid leukemia (CML) using an alternative mechanism of action from the TKIs, specifically based on the inhibition of tubulin polymerization. Compounds 7 and 25, as planned, not only inhibited tubulin polymerization, but also inhibited the proliferation of both GMB and CML cells, including those expressing the T315I mutation, at nanomolar concentrations. In in vivo experiments in BALB/cnu/nu mice injected subcutaneously with U87MG cells, in vivo, 7 significantly inhibited GBM cancer cell proliferation, in vivo tumorigenesis, and tumor growth, tumorigenesis and angiogenesis. Compound 7 was found to block human topoisomerase II (hTopoII) selectively and completely, at a concentration of 100 µM.

Deciphering the key heterocyclic scaffolds in targeting microtubules, kinases and carbonic anhydrases for cancer drug development.

Heterocyclic scaffolds are widely utilized for drug design by taking into account the molecular structure of therapeutic targets that are related to a broad spectrum of ailments, including tumors. Such compounds display various covalent and non-covalent interactions with the specific residues of the target proteins while causing their inhibition. There is a substantial number of heterocyclic compounds approved for cancer treatment, and these compounds function by interacting with different therapeutic targets involved in tumorogenesis. In this review, we trace and emphasize the privileged heterocyclic pharmacophores that have immense potency against several essential chemotherapeutic tumor targets: microtubules, kinases and carbonic anhydrases. Potent compounds currently undergoing pre-clinical and clinical studies have also been assessed for ascertaining the effective class of chemical scaffolds that have significant therapeutic potential against multiple malignancies. In addition, we also describe briefly the role of heterocyclic compounds in various chemotherapy regimens. The optimized molecular hybridization of delineated motifs may result in the discovery of more active anticancer therapeutics and circumvent the development of resistance by specific targets in the future.

Novel pyrazolo[4,3-d]pyrimidine microtubule targeting agents (MTAs): Synthesis, structure-activity relationship, in vitro and in vivo evaluation as antitumor agents.

The design, synthesis, and biological evaluation of a series novel N1­methyl pyrazolo[4,3-d]pyrimidines as inhibitors of tubulin polymerization and colchicine binding were described here. Synthesis of target compounds involved alkylation of the pyrazolo scaffold, which afforded two regioisomers. These were separated, characterized and identified with 1H NMR and NOESY spectroscopy. All compounds, except 10, inhibited [3H]colchicine binding to tubulin, and the potent inhibition was similar to that obtained with CA-4. Compounds 9 and 11-13 strongly inhibited the polymerization of tubulin, with IC50 values of 0.45, 0.42, 0.49 and 0.42 µM, respectively. Compounds 14-16 inhibited the polymerization of tubulin with IC50s near ∼1 µM. Compounds 9, 12, 13 and 16 inhibited MCF-7 breast cancer cell lines and circumvented ßIII-tubulin mediated cancer cell resistance to taxanes and other MTAs, and compounds 9-17 circumvented Pgp-mediated drug resistance. In the standard NCI testing protocol, compound 9 exhibited excellent potency with low to sub nanomolar GI50 values (≤10 nM) against most tumor cell lines, including several multidrug resistant phenotypes. Compound 9 was significantly (P < 0.0001) better than paclitaxel at reducing MCF-7 TUBB3 (ßIII-tubulin overexpressing) tumors in a mouse xenograft model. Collectively, these studies support the further preclinical development of the pyrazolo[4,3-d]pyrimidine scaffold as a new generation of tubulin inhibitors and 9 as an anticancer agent with advantages over paclitaxel.

Insight on [1,3]thiazolo[4,5-e]isoindoles as tubulin polymerization inhibitors.

A series of [1,3]thiazolo[4,5-e]isoindoles has been synthesized through a versatile and high yielding multistep sequence. Evaluation of the antiproliferative activity of the new compounds on the full NCI human tumor cell line panel highlighted several compounds that are able to inhibit tumor cell proliferation at micromolar-submicromolar concentrations. The most active derivative 11g was found to cause cell cycle arrest at the G2/M phase and induce apoptosis in HeLa cells, following the mitochondrial pathway, making it a lead compound for the discovery of new antimitotic drugs.

Pyrrolo[2',3':3,4]cyclohepta[1,2-d][1,2]oxazoles, a New Class of Antimitotic Agents Active against Multiple Malignant Cell Types.

A new class of pyrrolo[2',3':3,4]cyclohepta[1,2-d][1,2]oxazoles was synthesized for the treatment of hyperproliferative pathologies, including neoplasms. The new compounds were screened in the 60 human cancer cell lines of the NCI drug screen and showed potent activity with GI50 values reaching the nanomolar level, with mean graph midpoints of 0.08-0.41 µM. All compounds were further tested on six lymphoma cell lines, and eight showed potent growth inhibitory effects with IC50 values lower than 500 nM. Mechanism of action studies showed the ability of the new [1,2]oxazoles to arrest cells in the G2/M phase in a concentration dependent manner and to induce apoptosis through the mitochondrial pathway. The most active compounds inhibited tubulin polymerization, with IC50 values of 1.9-8.2 µM, and appeared to bind to the colchicine site. The G2/M arrest was accompanied by apoptosis, mitochondrial depolarization, generation of reactive oxygen species, and PARP cleavage.

Diaryl disulfides and thiosulfonates as combretastatin A-4 analogues: Synthesis, cytotoxicity and antitubulin activity.

Diaryl disulfides and diaryl thiosulfonates were synthesized with the two phenyl rings of all compounds bearing identical halide substituents. Because of structural similarity to the potent antimitotic natural product combretastatin A-4 (CA-4), the compounds were examined for inhibition of tubulin polymerization, and the thiosulfonates were more active than the disulfides. The nine thiosulfonates had IC50 values ranging from 1.2 to 9.1 µM, as compared with 1.3 µM obtained with CA-4. The compounds thus ranged from equipotent with CA-4 to 7-fold less active. The nine disulfides had IC50 values ranging from 1.2 to 5.1 µM, as compared with 0.54 µM obtained with CA-4. The compounds thus ranged from less than half as active as CA-4 to over 9-fold less active. The most active members of each group, 2 g and 3c, in the assembly assay were modeled into the colchicine site. Compound 3c had significant hydrophobic interactions with ß-tubulin residues CYS 241 and ALA 250, and its thiosulfonate bridge made a hydrogen bond with ß-tubulin residue ASN 258. Compound 2 g had hydrophobic interactions with ß-tubulin residues ALA 250, CYS 241 and ALA 254, but there was no significant interaction of the disulfide bridge with tubulin.

Structure-activity relationship studies and in vitro and in vivo anticancer activity of novel 3-aroyl-1,4-diarylpyrroles against solid tumors and hematological malignancies.

Novel 3-aroyl-1,4-diarylpyrrole derivatives were synthesized to explore structure-activity relationships at the phenyls at positions 1 and 4 of the pyrrole. The presence of amino phenyl rings at positions 1 and 4 of the pyrrole ring were found to be a crucial requirement for potent antitumor activity. Several compounds strongly inhibited tubulin assembly through binding to the colchicine site. Compounds 42, 44, 48, 62 and 69 showed antitumor activity with low nanomolar IC50 values in several cancer cell lines. Compound 48 was generally more effective as an inhibitor of glioblastoma, colorectal and urinary bladder cancer cell lines; 69 consistently inhibited CML cell lines and demonstrated superiority in nilotinib and imatinib resistant LAMA84-R and KBM5-T315I cells. In animal models, compound 48 exhibited significant inhibition of the growth of T24 bladder carcinoma and ES-2 ovarian clear cell carcinoma tumors. Compounds 48 and 69 represent robust lead compounds for the design of new broad-spectrum anticancer agents active in different types of solid and hematological tumors.

S-(4-Methoxyphenyl)-4-methoxybenzenesulfonothioate as a Promising Lead Compound for the Development of a Renal Carcinoma Agent.

Organosulfur compounds show cytotoxic potential towards many tumor cell lines. Disulfides and thiosulfonates act through apoptotic processes, inducing proteins associated with apoptosis, endoplasmic reticulum stress, and the unfolded protein response. Three p-substituted symmetric diaryl disulfides and three diaryl thiosulfonates were synthesized and analyzed for inhibition of tubulin polymerization and for human cancer cell cytotoxic activity against seven tumor cell lines and a non-tumor cell line. S-(4-methoxyphenyl)-4-methoxybenzenesulfonothioate (6) exhibited inhibition of tubulin polymerization and showed the best antiproliferative potential, especially against the 786-0 cell line, being six times more selective as compared with the non-tumor cell line. In addition, compound 6 was able to activate caspase-3 after 24 and 48 h treatments of the 786-0 cell line and induced cell-cycle arrest in the G2/M stage at the highest concentration evaluated at 24 and 48 h. Compound 6 was able to cause complete inhibition of proliferation, inducing the death of 786-0 cells, by increasing the number of cells at G2/M and greater activation of caspase-3.