Incorporation of aza-β3-amino acids into endogenous neuropeptide from mollusks (ALSGDAFLRF-NH2) with weak antimicrobial activities allows us to design new AMPs sequences. We find that, depending on the nature of the substitution, these could result either in inactive pseudopeptides or in a drastic enhancement of the antimicrobial activity without high cytotoxicity resulted. Structural studies perform by NMR and circular dichroism on the pseudopeptides show the impact of aza-β3-amino acids on the peptide structures. We obtain the first three-dimensional structures of pseudopeptides containing aza-β3-amino acids in aqueous micellar SDS and demonstrate that hydrazino turn can be formed in aqueous solution. Overall, these results demonstrate the ability to modulate AMPs activities through structural modifications induced by the nature and the position of these amino acid analogs in the peptide sequences.
Amino Acids; chemical synthesis; chemistry; pharmacology; Animals; Anti-Bacterial Agents; chemical synthesis; chemistry; pharmacology; Antimicrobial Cationic Peptides; chemistry; Aza Compounds; chemical synthesis; chemistry; pharmacology; CHO Cells; Circular Dichroism; Cricetinae; Cricetulus; Gram-Negative Bacteria; drug effects; Gram-Positive Bacteria; drug effects; Hemolysis; Magnetic Resonance Spectroscopy; Microbial Sensitivity Tests; Models, Molecular; Molecular Conformation; Neuropeptides; chemical synthesis; chemistry; pharmacology; Oligopeptides; chemical synthesis; chemistry; pharmacology; Rabbits; Stereoisomerism; Structure-Activity Relationship
This G-rich region of the c-MYC promoter has been shown to form a G-quadruplex structure, acting as a silencer element for c-MYC transcriptional control. In the present work, we have synthesized a series of 11-substituted quindoline analogs as c-MYC G-quadruplex–stabilizing compounds, and the cell-free and in vitro activity of these compounds were evaluated. Two lead compounds (4 and 12) demonstrated good cell-free profiles, and compound 4 (2-(4-(10H-indolo[3,2-b]quinolin-11-yl)piperazin-1-yl)-N,N-dimethylethanamine) significantly downregulated c-MYC expression. However, despite the good cell-free activity and the effect of these compounds on c-MYC gene expression, we have demonstrated, using a cellular assay in a Burkitt’s lymphoma cell line (CA46-specific), that these effects were not mediated through targeting the c-MYC G-quadruplex. Thus, caution should be used in assigning the effects of G-quadruplex-interactive compounds that lower c-MYC to direct targeting of these promoter elements unless this assay, or similar ones, demonstrates direct targeting of the G-quadruplex in cells.
Compounds bactericidal against both replicating and non-replicating Mtb may shorten the length of TB treatment regimens by eliminating infections more rapidly. Screening of a panel of antimicrobial and anticancer drug classes that are bioreduced into cytotoxic species revealed that 1,2,4-benzotriazine di-N-oxides (BTOs) are potently bactericidal against replicating and non-replicating Mtb. Medicinal chemistry optimization, guided by semi-empirical molecular orbital calculations, identified a new lead compound (20q) from this series with an MIC of 0.31 μg/mL against H37Rv and a cytotoxicity (CC50) against Vero cells of 25 μg/mL. 20q also had equivalent potency against a panel of single-drug resistant strains of Mtb and remarkably selective activity for Mtb over a panel of other pathogenic bacterial strains. 20q was also negative in a L5178Y MOLY assay, indicating low potential for genetic toxicity. These data along with measurements of the physiochemical properties and pharmacokinetic profile demonstrate that BTOs have the potential to be developed into a new class of antitubercular drugs.
Benzotriazine oxides; drug resistance; bioreductive drugs; antitubercular compounds; antibiotics; nonreplicating persistence; tirapazamine
We recently described the discovery of oxygenated N-alkyl deoxynojirimycin (DNJ) derivative 7 (CM-10-18) with antiviral activity against dengue virus (DENV) infection both in vitro and in vivo. This imino sugar was promising, but had an EC50 against DENV in BHK cells of 6.5 μM, which limited its use in in vivo. Compound 7 presented structural opportunities for activity relationship analysis, which we exploited and report here. These structure-activity relationship studies led to analogs 2h, 2l, 3j, 3l, 3v and 4b–4c with nanomolar antiviral activity (EC50 = 0.3–0.5 μM) against DENV infection, while maintaining low cytotoxicity (CC50 > 500 μM, SI > 1000). In male Sprague-Dawley rats, compound 3l was well tolerated at a dose up to 200 mg/kg and displayed desirable PK profiles, with significantly improved bioavailability (F = 92 ± 4%).
The 6-aminoglucosamine ring of the aminoglycoside antibiotic neomycin B (ring II) was conjugated to a 16 mer peptide nucleic acid (PNA) targeting HIV-1 TAR RNA. For this purpose we prepared the aminoglucosamine monomer 15 and attached it to the protected PNA prior to its cleavage from the solid support. We found that the resulting PNA-aminoglucosamine conjugate is stable under acidic condition, efficiently taken up by the human cells and fairly distributed in both cytosol and nucleus without endosomal entrapment since co-treatment with endosome-disrupting agent had no effect on its cellular distribution. The conjugate displayed very high target specificity in vitro and strongly inhibited Tat mediated transactivation of HIV-1 LTR transcription in cell culture system. The unique properties of this new class of PNA conjugate suggest it to be a potential candidate for therapeutic application.
Peptide nucleic acids; polyamide nucleic acids; aminosugar; aminoglycoside; 6-aminoglucosamine; neosamine; neamine; antiviral activity; HIV; cellular penetration; TAR; Tat
The binding pocket of both NPFF receptors was investigated, focusing on subtype-selective behavior. By using four non-peptidic compounds and the peptide mimetics RF9 and BIBP3226 agonistic and antagonistic properties were characterized. A set of Ala receptor mutants was generated, the binding pocket was narrowed down to the upper part of transmembrane helices V, VI, VII, and the extracellular loop 2. Positions 5.27 and 6.59 have been shown to have a strong impact on receptor activation and were suggested to form an acidic, negatively charged binding pocket in both NPFF receptor subtypes. Additionally, position 7.35 was identified to play an important role in functional selectivity. According to docking experiments, the aryl group of AC-216 interacts with position 7.35 in the NPFF1 but not in the NPFF2 receptor. These results provide distinct insights into the receptor specific binding pockets, which is necessary for the development of drugs to address the NPFF system.
functional selectivity; neuropeptide FF; NPFF receptors; RFamide; small non-peptidic compounds; subtype-selectivity
A unique heterocyclic carbamate prodrug of seco-CBI-indole2 that releases no residual byproduct is reported as a new member of a class of hydrolyzable prodrugs of the duocarmycin and CC-1065 family of natural products. The prodrug was designed to be activated by hydrolysis of a carbamate releasing the free drug without the cleavage release of a traceable extraneous group. Unlike prior carbamate prodrugs examined that are rapidly cleaved in vivo, the cyclic carbamate was found to be exceptionally stable to hydrolysis under both chemical and biological conditions providing a slow, sustained release of the exceptionally potent free drug. An in vivo evaluation of the prodrug found that its efficacy exceeded that of the parent drug, that its therapeutic window of efficacy versus toxicity is much larger than the parent drug, and that its slow free drug release permitted the safe and efficacious use of doses 150-fold higher than the parent compound.
In our effort to develop multifunctional drugs against Parkinson’s disease, a structure-activity-relationship study was carried out based on our hybrid molecular template targeting D2/D3 receptors. Competitive binding with [3H]spiroperidol was used to evaluate affinity (Ki) of test compounds. Functional activity of selected compounds in stimulating [35S]GTPγS binding was assessed in CHO-cells expressing either human D2 or D3 receptors. Our results demonstrated development of highly selective compounds for D3 receptor ((−)-40; Ki D3 = 1.84 nM, D2/D3 = 583.2, (−)-45; Ki D3 = 1.09 nM, D2/D3 = 827.5). Functional data identified (−)-40 (EC50 D2 = 114 nM and D3 = 0.26 nM, D2/D3 = 438) as one of the highest D3 selective agonists known to date. In addition, high affinity, non-selective, D3 agonist, (−)-19 (EC50 D2 = 2.96 nM and D3 = 1.26 nM), was also developed. Lead compounds with antioxidant activity were evaluated using an in vivo PD animal model.
Due to structural and mechanistic differences between eukaryotic and prokaryotic fatty acid synthesis enzymes, the bacterial pathway, FAS-II, is an attractive target for the design of antimicrobial agents. We have previously reported the identification of a novel series of benzimidazole compounds with particularly good antibacterial effect against Francisella tularensis, a Category A biowarfare pathogen. Herein we report the crystal structure of the F. tularensis FabI enzyme in complex with our most active benzimidazole compound bound with NADH. The structure reveals that the benzimidazole compounds bind to the substrate site in a unique conformation that is distinct from the binding motif of other known FabI inhibitors. Detailed inhibition kinetics have confirmed that the compounds possess a novel inhibitory mechanism that is unique among known FabI inhibitors. These studies could have a strong impact on future antimicrobial design efforts and may reveal new avenues for the design of FAS-II active antibacterial compounds.
Compound 2 (KU-32) is a first-generation novologue (a novobiocin-based, C-terminal, heat shock protein 90 (Hsp90) inhibitor), that decreases glucose-induced death of primary sensory neurons and reverses numerous clinical indices of diabetic peripheral neuropathy in mice. The current study sought to exploit the C-terminal binding site of Hsp90 to determine whether the optimization of hydrogen bonding and hydrophobic interactions of second generation novologues could enhance neuroprotective activity. Using a series of substituted phenylboronic acids to replace the coumarin lactone of 2, we identified electronegative atoms placed at the meta-position of the B-ring exhibit improved cytoprotective activity, which is believed to result from favorable interactions with Lys539 in the Hsp90 C-terminal binding pocket. Consistent with these results, a meta-3-fluorophenyl substituted novologue (13b) exhibited a 14-fold lower ED50 compared to 2 for protection against glucose-induced toxicity of primary sensory neurons.
Mithramycin is an antitumor compound produced by Streptomyces argillaceus that has been used for the treatment of several types of tumors and hypercalcaemia processes. However, its use in humans has been limited because its side effects. Using combinatorial biosynthesis approaches, we have generated seven new mithramycin derivatives, which either differ from the parental compound in the sugar profile or in both the sugar profile and the 3-side chain. From these studies three novel derivatives were identified, demycarosyl-3D-β-d-digitoxosyl-mithramycin SK, demycarosyl-mithramycin SDK and demycarosyl-3D-β-d-digitoxosyl-mithramycin SDK, which show high antitumor activity. The first one, which combines two structural features previously found to improve pharmacological behavior, was generated following two different strategies, and it showed less toxicity than mithramycin. Preliminary in vivo evaluation of its antitumor activity through hollow fiber assays, and in subcutaneous colon and melanoma cancers xenografts models, suggests that demycarosyl-3D-β-d-digitoxosyl-mithramycin SK could be a promising antitumor agent, worthy of further investigation.
Streptomyces; antitumor; mithramycin; mithramycin SK; combinatorial biosynthesis; digitoxosyl-mithramycin SK
Activation of the retinoid X receptor (RXR), which is involved in cell proliferation, differentiation and apoptosis, is a strategy for cancer chemotherapy and chemoprevention, and 3-amino-6-(3'-aminopropyl)-5H-indeno[1,2-c]isoquinoline-5,11-(6H)dione (AM6-36) (3) is among the few RXR ligands known. The presently reported studies of 3 include its binding to human plasma proteins, metabolic stability using human liver microsomes, metabolism by human liver microsomes and hepatocytes, and in vivo disposition in rat serum, liver and mammary tissue. Compound 3 was 75% bound to human plasma proteins, and its metabolic stability was much greater than propranolol. One phase I metabolite was formed by human liver microsomes, 7 phase I and II metabolites were formed by human hepatocytes, and 5 metabolites were detected in rat serum and liver after oral administration. The putative metabolites predicted using LC-MS-MS were synthesized to confirm their structures and to provide sufficient material for investigation of induction of RXRE transcriptional activity and inhibition of NFκB.
A major challenge in the field of Gaucher disease has been the development of new therapeutic strategies including molecular chaperones. All previously described chaperones of glucocerebrosidase are enzyme inhibitors, which complicates their clinical development, because their chaperone activity must be balanced against the functional inhibition of the enzyme. Using a novel high throughput screening methodology, we identified a chemical series that does not inhibit the enzyme, but can still facilitate its translocation to the lysosome as measured by immunostaining of glucocerebrosidase in patient fibroblasts. These compounds provide the basis for the development of a novel approach towards small molecule treatment for patients with Gaucher disease.
Gaucher disease; glucocerebrosidase; chaperone; pyrazolopyrimidine; activator
We have devised a chemocentric informatics methodology for drug discovery integrating independent approaches to mining biomolecular databases. As a proof of concept, we have searched for novel putative cognition enhancers. First, we generated Quantitative Structure- Activity Relationship (QSAR) models of compounds binding to 5-hydroxytryptamine-6 receptor (5HT6R), a known target for cognition enhancers, and employed these models for virtual screening to identify putative 5-HT6R actives. Second, we queried chemogenomics data from the Connectivity Map (http://www.broad.mit.edu/cmap/) with the gene expression profile signatures of Alzheimer’s disease patients to identify compounds putatively linked to the disease. Thirteen common hits were tested in 5-HT6R radioligand binding assays and ten were confirmed as actives. Four of them were known selective estrogen receptor modulators that were never reported as 5-HT6R ligands. Furthermore, nine of the confirmed actives were reported elsewhere to have memory-enhancing effects. The approaches discussed herein can be used broadly to identify novel drug-target-disease associations.
Of great interest in recent years has been computationally predicting the novel polypharmacology of drug molecules. Here, we applied an “induced-fit” protocol to improve the homology models of 5-HT2A receptor, and we assessed the quality of these models in retrospective virtual screening. Subsequently, we computationally screened the FDA approved drug molecules against the best induced-fit 5-HT2A models, and chose six top scoring hits for experimental assays. Surprisingly, one well-known kinase inhibitor, sorafenib has shown unexpected promiscuous 5-HTRs binding affinities, Ki = 1959, 56 and 417 nM against 5-HT2A, 5-HT2B and 5-HT2C, respectively. Our preliminary SAR exploration supports the predicted binding mode, and further suggests sorafenib to be a novel lead compound for 5HTR ligand discovery. Although it has been well known that sorafenib produces anticancer effects through targeting multiple kinases, carefully designed experimental studies are desirable to fully understand whether its “off-target” 5-HTR binding activities contribute to its therapeutic efficacy or otherwise undesirable side effects.
GPCR; 5-HTR; induced-fit; molecular docking; molecular dynamics; sorafenib
Novel dimethyl-4,4′-dimethoxy-5,6,5′,6′-dimethylenedioxybiphenyl-2,2′-dicarboxylate (DDB) analogs were designed and synthesized to improve their chemosensitizing action on KBvin (vincristine resistant nasopharyngeal carcinoma) cells, a multi-drug resistant cell line over-expressing P-glycoprotein (P-gp). Structure-activity relationship analysis showed that aromatic and bulky aliphatic side chains at the 2,2′-positions effectively and significantly sensitized P-gp overexpressing multidrug resistant (MDR) cells to anticancer drugs, such as paclitaxel (TAX), vincristine (VCR), and doxorubicin (DOX). DDB derivatives 16 and 23 showed 5–10 times more effective reversal ability than verapamil (VRP) for TAX and VCR. Analog 6 also exhibited five times greater chemosensitizing effect against DOX than VRP. Importantly, no cytotoxicity was observed by the active DDB analogs against both non-MDR and MDR cells, suggesting that DDB analogs serve as the novel lead compounds for the development of chemosensitizers to overcome MDR phenotype. The mechanism of action studies demonstrated that effective inhibition of P-glycoprotein by DDB analogs dramatically elevated cellular concentration of anticancer drugs.
Five cationic platinum(II) complexes of general formula, [Pt(NH3)2(β-diketonate)]X are reported, where X is a non-coordinating anion and β-diketonate = acetylacetonate (acac), 1,1,1,-trifluoroacetylacetonate (tfac), benzoylacetonate (bzac), 4,4,4-trifluorobenzoylacetonate (tfbz), or dibenzoylmethide (dbm), corresponding respectively to complexes 1–5. The log P values and the stabilities of 1–5 in aqueous solution were evaluated. The phenyl ring substituents of 3–5 increase the lipophilicity of the resulting complexes, whereas the trifluoromethyl groups of 2 and 4 decrease the stability of the complexes in aqueous solution. The uptake of 1–5 in HeLa cells increase as the lipophilicity of the investigated complex increases. Cancer cell cytotoxicity studies indicate that 1 and 3 are the least active complexes whereas 2, 4, and 5 are comparable to cisplatin.
A series of novel 3-nitro-1H-1,2,4-triazole-(and in some cases 2-nitro-1H-imidazole)-based amides and sulfonamides were characterized for their in vitro anti-trypanosomal and antileishmanial activities as well as mammalian toxicity. Out of 36 compounds tested, 29 (mostly 3-nitro-1H-1,2,4-triazoles) displayed significant activity against T. cruzi intracellular amastigotes (IC50 ranging from 28 nM to 3.72 μM) without concomitant toxicity to L6 host cells (selectivity 66 to 2782). Twenty three of these active compounds were more potent (up to 58 fold) than the reference drug benznidazole, tested in parallel. In addition, 9 nitrotriazoles which were moderately active (0.5 μM ≤ IC50 < 6.0 μM) against T. b. rhodesiense trypomastigotes, were 5 to 31 fold more active against bloodstream-form T. b. brucei trypomastigotes engineered to overexpress NADH-dependent nitroreductase (TbNTR). Finally, 3 nitrotriazoles displayed a moderate activity against the axenic form of Leishmania donovani. Therefore, 3-nitro-1H-1,2,4-triazole-based amides and sulfonamides are potent anti-trypanosomal agents.
nitrotriazoles; amides; sulfonamides; T. cruzi; Chagas disease; anti-trypanosomal agents
Tubulin, the major structural component of microtubules, is a target for the development of anticancer agents. A series of (Z)-1-Aryl-3-arylamino-2-propen-1-one (10) were synthesized and evaluated for anti-proliferative activity in cell based assay. The most active compound (Z)-1-(2- bromo-3,4,5-trimethoxyphenyl)-3-(3-hydroxy-4-methoxyphenylamino)-prop-2-en-1-one (10ae) was tested in 20 tumor cell lines including multidrug resistant phenotype and was found to induce apoptosis in all these cell lines with similar GI50 values. Flow cytometry studies showed that 10ae arrested the cells in G2/M phase of cell cycle. In addition to G2/M block, these compounds caused microtubule stabilization like paclitaxel and induced apoptosis via activation of the caspase family. The observations made in this investigation demonstrate that (Z)-1-Aryl-3- arylamino-2-propen-1-one (10) represents a new class of microtubule – stabilizing agents.
The BRAF oncoprotein is mutated in about half of malignant melanomas and other cancers, and a kinase activating single valine to glutamate substitution at residue 600 (BRAFV600E) accounts for over 90% of BRAF-mediated cancers. Several BRAFV600E inhibitors have been developed, although they harbor some liabilities, thus motivating the development of other BRAFV600E inhibitor options. We report here the use of an ELISA based high-throughput screen to identify a family of related quinolol/naphthol compounds that preferentially inhibit BRAFV600E over BRAFWT and other kinases. We also report the X-ray crystal structure of a BRAF/quinolol complex revealing the mode of inhibition, employ structure-based medicinal chemistry efforts to prepare naphthol analogs that inhibit BRAFV600E
in vitro with IC50 values in the 80–200 nM under saturating ATP concentrations, and demonstrate that these compounds inhibit MAPK signaling in melanoma cells. Prospects for improving the potency and selectivity of these inhibitors are discussed.
A new series of tubulin polymerization inhibitors based on the 2-aryl/heteroaryl-4-amino-5-(3′,4′,5′-trimethoxybenzoyl)thiazole scaffold was synthesized and evaluated for growth inhibition activity on a panel of cancer cell lines, cell cycle effects, and in vivo potency. Structure–activity relationships were elucidated with various substitutions at the 2-position of the thiazole skeleton. Hydrophobic moieties, such as phenyl and 3-thienyl, were well tolerated at this position, and variation of the phenyl substituents had remarkable effects on potency. The most active compound (3b) induced apoptosis through the mitochondrial pathway with activation of caspase-3. We also showed that it has potential antivascular activity since it reduced in vitro endothelial cell migration and disrupted capillary-like tube formation at noncytotoxic concentrations. Furthermore, compound 3b significantly reduced the growth of the HT-29 xenograft in a nude mouse model, suggesting that 3b is a promising new antimitotic agent with clinical potential.
The vitamin D receptor (VDR) is a nuclear hormone receptor that regulates cell proliferation, cell differentiation, and calcium homeostasis. The receptor is activated by vitamin D analogs that induce the disruption of VDR-corepressor binding and promote VDR-coactivator interactions. The interactions between VDR and coregulators are essential for VDR-mediated transcription. Small molecule inhibition of VDR–coregulator binding represents an alternative method to the traditional ligand-based approach in order to modulate the expression of VDR target genes. A high throughput fluorescence polarization screen that quantifies the inhibition of binding between VDR and a fluorescently labeled steroid receptor coactivator 2 peptide was applied to discover the new small molecule VDR–coactivator inhibitors, 3-indolyl-methanamines. Structure-activity relationship studies with 3-indolyl-methanamine analogs were used to determine their mode of VDR-binding and to produce the first VDR-selective and irreversible VDR–coactivator inhibitors with the ability to regulate the transcription of the human VDR target gene, TRPV6.
Vitamin D receptor; steroid receptor coactivator; fluorescence polarization; high throughput screening; 3-indolyl-methanamines; TRPV6
(N)-Methanocarba adenosine 5′-methyluronamides containing known A3 AR (adenosine receptor)-enhancing modifications, i.e. 2-(arylethynyl)adenine and N6-methyl or N6-(3-substituted-benzyl), were nanomolar full agonists of human (h) A3AR and highly selective (Ki ~0.6 nM, N6-methyl 2-(halophenylethynyl) analogues 13, 14). Combined 2-arylethynyl-N6-3-chlorobenzyl substitutions preserved A3AR affinity/selectivity in the (N)-methanocarba series (e.g. 3,4-difluoro full agonist MRS5698 31, Ki 3 nM, human and mouse A3) better than for ribosides. Polyaromatic 2-ethynyl N6-3-chlorobenzyl analogues, such as potent linearly extended 2-p-biphenylethynyl MRS5679 34 (Ki hA3 3.1 nM; A1, A2A: inactive) and fluorescent 1-pyrene adduct MRS5704 35 (Ki hA3 68.3 nM) were conformationally rigid; receptor docking identified a large, mainly hydrophobic binding region. The vicinity of receptor-bound C2 groups was probed by homology modeling based on recent X-ray structure of an agonist-bound A2AAR, with a predicted helical rearrangement requiring an agonist-specific outward displacement of TM2 resembling opsin. Thus, X-ray structure of related A2AAR is useful in guiding design of new A3AR agonists.
G protein-coupled receptor; purines; molecular modeling; structure activity relationship; radioligand binding; adenylate cyclase
The CD4 binding site on HIV-1 gp120 has been validated as a drug target to prevent HIV-1 entry to cells. Previously, we identified two small molecule inhibitors consisting of a 2,2,6,6-tetramethylpiperidine ring linked by an oxalamide to a p-halide-substituted phenyl group, which target this site, specifically, a cavity termed “Phe43 cavity”. Here we use synthetic chemistry, functional assessment and structure-based analysis to explore variants of each region of these inhibitors for improved antiviral properties. Alterations of the phenyl group and of the oxalamide linker indicated that these regions were close to optimal in the original lead compounds. Design of a series of compounds, where the tetramethylpiperidine ring was replaced with new scaffolds, lead to improved antiviral activity. These new scaffolds provide insight into the surface chemistry at the entrance of the cavity and offer additional opportunities by which to optimize further these potential-next-generation therapeutics and microbicides against HIV-1.
Employing a structure-based strategy, we have designed a new class of potent small-molecule inhibitors of the anti-apoptotic proteins Bcl-2 and Bcl-xL. An initial lead compound with a new scaffold was designed based upon the crystal structure of Bcl-xL and FDA-approved drugs and was found to have an affinity of 100 μM to both Bcl-2 and Bcl-xL. Linking this weak lead to another weak-affinity fragment derived from Abbott's ABT-737 led to an improvement of the binding affinity by a factor of >10,000. Further optimization ultimately yielded compounds with subnanomolar binding affinities to both Bcl-2 and Bcl-xL and potent cellular activity. The best compound (21) binds to Bcl-xL and Bcl-2 with Ki < 1 nM, inhibits cell growth in the H146 and H1417 small-cell lung cancer cell lines with IC50 values of 60–90 nM and induces robust cell death in the H146 cancer cell line at 30–100 nM.