New analogs of SYA013 as sigma-2 ligands with anticancer activity.

Our previous study has revealed 4-(4-(4-chlorophenyl)-1,4-diazepan-1-yl)-1-(4-fluorophenyl)butan-1-one·2HCl (SYA013) 1 as a sigma ligand with moderate selectivity for the sigma-2 receptor. Given the overexpression of sigma receptors in solid tumors and reports of sigma ligands with anticancer activities, we selected 1 for evaluation in several solid tumor cell lines. In addition, we have synthesized new analogs of 1 and now report that several of them bind preferentially at the sigma-2 receptor and have shown inhibition of several cancer cell lines including MDA-MB-231, MDA-MB-486, A549, PC-3, MIA PaCa-2 and Panc-1 cells. In particular, compounds 1 and 12 have demonstrated sub-micromolar activity against the Panc-1 cell line. It has also been observed that several of these compounds demonstrate selective toxicity toward cancer cells, when compared to normal cells.

SYA 013 analogs as moderately selective sigma-2 (σ2) ligands: Structure-affinity relationship studies.

Several lines of evidence suggest that selective sigma-2 (σ2) ligands might be useful for the treatment of solid tumors. However, very few selective σ2 ligands have been identified. This study was aimed at identifying new selective σ2 receptor ligands using a previously identified agent, SYA 013 as a lead. Four groups, homopiperazine, piperazine, tropane and selected oxime analogs of the homopiperazines were identified, synthesized and subsequently screened at the σ1 and σ2 receptors. The results demonstrate that these scaffolds can be modified to obtain selective σ2 receptor ligands. 1-(5-Chloropyridin-2-yl)-4-(3-((4-fluorophenyl)thio)propyl)-1,4-diazepane, 7 and 3-(4-chlorophenyl)-8-(3-((2-fluorophenyl)thio)propyl)-8-azabicyclo[3.2.1]octan-3-ol, 21 were identified as the highest binding affinity ligands (σ2Ki = 2.2 nM) and (4-(4-(5-chloropyridin-2-yl)-1,4-diazepan-1-yl)-1-(4-fluorophenyl)-butan-1-one oxime, 22 as a high affinity and the most selective ligand for the σ2 receptor (σ1Ki/σ2Ki = 41.8).

Synthesis and evaluation of the structural elements in alkylated tetrahydroisoquinolines for binding to CNS receptors.

Diseases of the CNS are often complex and involve multiple receptor systems and thus, the treatment options for these diseases must focus on targeting the multiple receptors implicated in the various disorders. Schizophrenia and depression are examples of such diseases and their pharmacotherapy thus depends on agents which target multiple receptors including the dopamine, serotonin and even cholinergic receptors at the same time. In our previous campaign to find multi-receptor ligands, we have identified the benzothiazole 1a as an initial lead molecule. In the current work, we have expanded the structure affinity relationship (SAFIR) of 1a resulting in the identification of a partially restrained butyrophenone 3j as a potent and selective dual 5-HT1A and 5-HT7 receptor ligand. It is expected that compound 3j may serve as a new lead for further development in our search for newer and novel ligands with the potential to treat diseases of CNS origin.

Development of CNS multi-receptor ligands: Modification of known D2 pharmacophores.

Several known D2 pharmacophores have been explored as templates for identifying ligands with multiple binding affinities at dopamine and serotonin receptors considered as clinically relevant receptors in the treatment of neuropsychiatric diseases. This approach has resulted in the identification of ligands that target multiple CNS receptors while avoiding others associated with deleterious effects. In particular, compounds 11, 15 and 22 may have potential for further development as antipsychotic agents as they favorably interact with the clinically relevant receptors including D2R, 5-HT1AR, and 5-HT7R. We have also identified the pair of compounds 11 and 10 as high affinity D2R ligands with and without SERT binding affinities, respectively. These differential binding profiles endow the pair with the potential for evaluating SERT contributions to antipsychotic drug activity in animal behavioral models. In addition, compound 11 has no significant affinity for 5-HT2CR and binds only moderately to the H1R, suggesting it may not induce weight gain or sedation when used clinically. Taken together, compound 11 displays an interesting pharmacological profile that necessitates the evaluation of its functional and in vivo effects in animal models which are currently ongoing.

Design and synthesis of dual 5-HT1A and 5-HT7 receptor ligands.

5-HT1A and 5-HT7 receptors have been at the center of discussions recently due in part to their major role in the etiology of major central nervous system diseases such as depression, sleep disorders, and schizophrenia. As part of our search to identify dual targeting ligands for these receptors, we have carried out a systematic modification of a selective 5HT7 receptor ligand culminating in the identification of several dual 5-HT1A and 5-HT7 receptor ligands. Compound 16, a butyrophenone derivative of tetrahydroisoquinoline (THIQ), was identified as the most potent agent with low nanomolar binding affinities to both receptors. Interestingly, compound 16 also displayed moderate affinity to other clinically relevant dopamine receptors. Thus, it is anticipated that compound 16 may serve as a lead for further exploitation in our quest to identify new ligands with the potential to treat diseases of CNS origin.

Further evaluation of the tropane analogs of haloperidol.

Previous work from our labs has indicated that a tropane analog of haloperidol with potent D2 binding but designed to avoid the formation of MPP(+)-like metabolites, such as 4-(4-chlorophenyl)-1-(4-(4-fluorophenyl)-4-oxobutyl)pyridin-1-ium (BCPP(+)) still produced catalepsy, suggesting a strong role for the D2 receptor in the production of catalepsy in rats, and hence EPS in humans. This study tested the hypothesis that further modifications of the tropane analog to produce compounds with less potent binding to the D2 receptor than haloperidol, would produce less catalepsy. These tests have now revealed that while haloperidol produced maximum catalepsy, these compounds produced moderate to low levels of catalepsy. Compound 9, with the least binding affinity to the D2R, produced the least catalepsy and highest Minimum Adverse Effective Dose (MAED) of the analogs tested regardless of their affinities at other receptors including the 5-HT1AR. These observations support the hypothesis that moderation of the D2 binding of the tropane analogs could reduce catalepsy potential in rats and consequently EPS in man.

Identification of a new selective dopamine D4 receptor ligand.

The dopamine D4 receptor has been shown to play key roles in certain CNS pathologies including addiction to cigarette smoking. Thus, selective D4 ligands may be useful in treating some of these conditions. Previous studies in our laboratory have indicated that the piperazine analog of haloperidol exhibits selective and increased affinity to the DAD4 receptor subtype, in comparison to its piperidine analog. This led to further exploration of the piperazine moiety to identify new agents that are selective at the D4 receptor. Compound 27 (KiD4=0.84 nM) was the most potent of the compounds tested. However, it only had moderate selectivity for the D4 receptor. Compound 28 (KiD4=3.9 nM) while not as potent, was more discriminatory for the D4 receptor subtype. In fact, compound 28 has little or no binding affinity to any of the other four DA receptor subtypes. In addition, of the 23 CNS receptors evaluated, only two, 5HT1AR and 5HT2BR, have binding affinity constants better than 100 nM (Ki <100 nM). Compound 28 is a potentially useful D4-selective ligand for probing disease treatments involving the D4 receptor, such as assisting smoking cessation, reversing cognitive deficits in schizophrenia and treating erectile dysfunction. Thus, further optimization, functional characterization and evaluation in animal models may be warranted.

CoMFA studies and in vitro evaluation of some 3-substituted benzylthio quinolinium salts as anticryptococcal agents.

The 3-dimensional quantitative structure-activity relationship (3D-QSAR) molecular modeling technique or comparative molecular field analysis (CoMFA) has been used to design analogs of the natural product cryptolepine (1). Twenty-three compounds with their in vitro biological activities (IC50 values) against Crytococcus neoformans were used to generate the training set database of compounds for the CoMFA studies. The cross-validated q(2), noncross-validated r(2), and partial least squares (PLS) analysis results were used to predict the biological activity of 11 newly designed test set compounds. The best CoMFA model produced a q(2) of 0.815 and an r(2) of 0.976 indicating high statistical significance as a predictive model. The steric and electrostatic contributions from the contour map were interpreted from the color-coded contour plots generated from the PLS model and the active structural components for potency against C. neoformans were determined and validated in the test set compounds. The 3-substituted benzylthio quinolinium salts (4) that make up the test set were synthesized and evaluated based on the predicted activity from the CoMFA model and the results produced a good correlation between the predicted and experimental activity (R=0.82). Thus, CoMFA has served as an effective tool to aid the design of new analogs and in this case, it has aided the identification of compounds equipotent with amphotericin B, the gold standard in antifungal drug design.

Structure-activity relationship (SAR) and preliminary mode of action studies of 3-substituted benzylthioquinolinium iodide as anti-opportunistic infection agents.

Opportunistic infections are devastating to immunocompromised patients. And in especially sub-Saharan Africa where the AIDS epidemic is still raging, the mortality rate was recently as high as 70%. The paucity of anti-opportunistic drugs, the decreasing efficacy and the development of resistance against the azoles and even amphotericin B have stimulated the search for new drugs with new mechanisms of action. In a previous work, we showed that a new chemotype derived from the natural product cryptolepine displayed selective toxicity against opportunistic pathogens with minimal cytotoxicity to normal cells. In this manuscript, we report the design and synthesis of substituted benzylthioquinolinium iodides, evaluated their anti-infective properties and formulated some initial structure-activity relationships around phenyl ring A from the original natural product. The sensitivity of the most potent analog 10l, to selected strains of C. cerevisiae was also evaluated leading to the observation that this scaffold may have a different mode of action from its predecessor, cryptolepine.

Evaluation of the behavioral and pharmacokinetic profile of SYA013, a homopiperazine analog of haloperidol in rats.

SYA013, a homopiperazine analog of haloperidol, was further evaluated for antipsychotic potential using additional animal models. Previously, SYA013 was tested in mice with an antipsychotic screening model in which it inhibited apomorphine induced climbing behavior, indicating antagonism of the dopaminergic system and the potential for use in the treatment of schizophrenia. In this study, SYA013 was shown to inhibit both d-amphetamine-induced locomotor activity in rats and conditioned avoidance response (CAR) in rats in a dose dependent manner and in the case of CAR, without producing any escape failure responses (EFRs), two tests predictive of antipsychotic action. The selective 5HT(1A) antagonist WAY100,635 was used to determine if binding of SYA013 to the 5HT(1A) receptor contributed to suppression of CAR. The results indicated that 0.63mg/kg WAY100,635 did not have a significant effect on the inhibition of CAR by SYA013. Pharmacokinetic parameters in brain and plasma were determined for SYA013. A log brain/plasma concentration ratio at a t(max) of 1.48 suggests that SYA013 readily crosses the blood brain barrier (BBB). The hypothesis that binding of SYA013 to the 5HT(1A) receptor contributed to the lack of significant catalepsy was investigated using the 5HT(1A) antagonist WAY100,635. The results of acute and semi-chronic tests suggest that binding to the 5HT(1A) receptor alone did not significantly account for the lack of catalepsy. Lack of catalepsy was preserved after the semi-chronic challenge with SYA013. These tests further indicate that SYA013 has a pharmacological profile with the potential for use in the treatment of neuropsychiatric diseases. In addition, the 5HT(1A) receptor does not appear to play a significant role in the pharmacological profile of SYA013.

Benzothiazoles as probes for the 5HT1A receptor and the serotonin transporter (SERT): a search for new dual-acting agents as potential antidepressants.

The synthesis and evaluation of several benzothiazole-based compounds are described in an attempt to identify novel dual-acting 5HT(1A) receptor and SERT inhibitors as new antidepressants. Binding affinities at the 5HT(1A) receptor and the serotonin transporter do not appear to be congruent and other areas of the binding sites would need to be explored in order to improve binding simultaneously at both sites. Compounds 20 and 23 show moderate binding affinity at the 5HT(1A) receptor and the SERT site and thus, have the potential to be further explored as dual-acting agents. In addition, compound 20 binds with low affinity to the dopamine transporter (DAT), the norepinephrine transporter (NET) and 5HT(2C) receptor, which are desirable properties as selectivity for SERT (and not DAT or NET) is associated with an absence of cardiovascular side effects.

Structure-activity relationship studies of SYA 013, a homopiperazine analog of haloperidol.

Structure-activity relationship studies on 4-(4-(4-chlorophenyl)-1,4-diazepan-1-yl)-1-(4-fluorophenyl)butan-1-one (SYA 013), a homopiperazine analog of haloperidol has resulted in an understanding of the effect of structural modifications on binding affinity at dopamine and serotonin receptor subtypes. Further exploration, using bioisosteric replacement strategies has led to the identification of several new agents including compounds 7, 8, 11 and 12 which satisfy the initial criteria for further exploration as new antipsychotic agents. In addition, compound 18, a D(3) selective tropanol, has been identified as having the potential for further optimization into a useful drug which may combat neuropsychiatric diseases.

Multi-receptor drug design: Haloperidol as a scaffold for the design and synthesis of atypical antipsychotic agents.

Using haloperidol as a scaffold, new agents were designed to investigate the structural contributions of various groups to binding at CNS receptors associated with atypical antipsychotic pharmacology. It is clear that each pharmacophoric group, the butyrophenone, the piperidine and the 4-chlorophenyl moieties contributes to changes in binding to the receptors of interest. This strategy has resulted in the identification of several new agents, compounds 16, 18, 19, 23, 24 and 25, with binding profiles which satisfy our stated criteria for agents to act as potential atypical antipsychotics. This research demonstrates that haloperidol can serve as a useful lead in the identification and design of new agents that target multiple receptors associated with antipsychotic pharmacology.

Optimization of 3-(phenylthio)quinolinium compounds against opportunistic fungal pathogens.

Ring-opened benzothieno[3,2-b]quinolinium salts (3) were designed and synthesized with substitution on the thiophene moiety. In vitro screenings were carried out against fungal pathogens including Cryptococcus neoformans, Candida albicans, Candida glabrata, Candida krusei and Aspergillus fumigatus. In all, by replacing the N-methyl group (2) with N-ω-phenylpentyl or ω-cyclohexylpentyl group to form substituted 3-(phenylthio)quinolinium compounds produced remarkable potencies, as high as 300-fold (cf, cryptolepine (1)=250 µg/mL vs 11p=0.8 µg/mL for C. albicans) over the starting tetracyclic parent. In addition, all the N-ω-cyclohexylpentyl analogs produced superior activity against all the microorganisms tested than the N-ω-phenylpentyl substituted compounds. The potential of these compounds to induce toxicity in Vero cells was also investigated and the majority of them showed lower or no cytotoxicity at 10 µg/mL than amphotericin B, the gold standard in antifungal drug development. For instance, the trifluoromethyl substituted analogs (11n-p) have selectivity indices over 2-fold better than those of amphotericin B in C. neoformans. Overall, this ring-opened scafford of benzothienoquinolines, with substitution on the thiophenyl moiety, serves as a new lead for further development.

Benzothieno[3,2-b]quinolinium and 3-(phenylthio)quinolinium compounds: Synthesis and evaluation against opportunistic fungal pathogens.

Substitution around 5-methyl benzothieno[3,2-b]quinolinium (2) ring system was explored in order to identify positions of substitution that could improve its antifungal profile. The 3-methoxy (10b) was active against C. albicans, C. neoformans, and A. fumigatus and the 4-chloro (10f) analog showed moderate increases in anti-cryptococcal and anti-aspergillus activities. The effectiveness of 10b and 10f were validated in murine models of candidiasis and cryptococcosis, respectively. The efficacy of 10f in reducing brain cryptococcal infection and its observation in the brain of mice injected with this quaternary compound confirm the capacity of these compounds to cross the blood-brain barrier of mice. Overall, several of the chloro and methoxy substituted compounds showed significant improvements in activity against A. fumigatus, the fungal pathogen prevalent in patients receiving organ transplant. Opening the benzothiophene ring of 2 to form 1-(5-cyclohexylpentyl)-3-(phenylthio)quinolinium compound (3) resulted in the identification of several novel compounds with over 50-fold increases in potency (cf. 2) while retaining low cytotoxicities. Thus, compound 3 constitutes a new scaffold for development of drugs against opportunistic infections.

Identification of 3-phenylaminoquinolinium and 3-phenylaminopyridinium salts as new agents against opportunistic fungal pathogens.

Previous studies on the indoloquinoline alkaloid, cryptolepine (2), revealed that it has antii-nfective properties among other activities. Using Structure-activity relationship (SAR) techniques, several ring-opened analogs of cryptolepine (3-phenylaminopyridinium and 3-phenylaminoquinolinium derivatives) were designed to improve the potency and lower the cytotoxicity shown by several of the precursor agents. Results indicate that these ring-opened analogs constitute new anti-infective agents with over a 100-fold potency and several fold lower cytotoxicity than cryptolepine from which they are derived.

Synthesis and evaluation of ligands for D2-like receptors: the role of common pharmacophoric groups.

Arylcycloalkylamines, such as phenyl piperidines and piperazines and their arylalkyl substituents, constitute pharmacophoric groups exemplified in several antipsychotic agents. A review of previous reports indicates that arylalkyl substituents can improve the potency and selectivity of the binding affinity at D(2)-like receptors. In this paper, we explored the contributions of two key pharmacophoric groups, that is, 4'-fluorobutyrophenones and 3-methyl-7-azaindoles, to the potency and selectivity of synthesized agents at D(2)-like receptors. Preliminary observation of binding affinities indicates that there is little predictability of specific effects of the arylalkyl moieties but the composite structure is responsible for selectivity and potency at these receptors.

Identification of a butyrophenone analog as a potential atypical antipsychotic agent: 4-[4-(4-chlorophenyl)-1,4-diazepan-1-yl]-1-(4-fluorophenyl)butan-1-one.

The synthesis and exploration of novel butyrophenones have led to the identification of a diazepane analogue of haloperidol, 4-[4-(4-chlorophenyl)-1,4-diazepan-1-yl]-1-(4-fluorophenyl)butan-1-one (compound 13) with an interesting multireceptor binding profile. Compound 13 was evaluated for its binding affinities at DA subtype receptors, 5HT subtype receptors, H-1, M-1 receptors and at NET, DAT, and SERT transporters. At each of these receptors, compound 13 was equipotent or better than several of the standards currently in use. In in vivo mouse and rat models to evaluate its efficacy and propensity to elicit catalepsy and hence EPS in humans, compound 13 showed similar efficacy as clozapine and did not produce catalepsy at five times its ED(50) value.

Synthesis and evaluation of isosteres of N-methyl indolo[3,2-b]-quinoline (cryptolepine) as new antiinfective agents.

Isosteres of cryptolepine (1) were synthesized and evaluated for their antiinfective activities. Overall, the sulfur isostere, 5-methyl benzothieno[3,2-b]quinolinium salt (5b), was equipotent to 1 and has shown no cytotoxicity at 23.8 microg/mL. Compound 5b was also found to have a broad spectrum of activity. Both the carbon and oxygen isosteres were less potent than cryptolepine. A limited library of 2-substituted analogs of 5b has been synthesized and evaluated in antifungal screens but did not show increase in potency compared to the unsubstituted 5b. Similarly, evaluation of tricyclic benzothieno[3,2-b]pyridines while showing promise in individual screens did not produce an overall increase in potency. Overall, the evaluation of the activities of 5b compared with standard antifungal/anti-protozoal agents suggests that the benzothienoquinoline scaffold could serve as a lead for optimization.

Identification of bis-quindolines as new antiinfective agents.

Several N-substituted quindolines were made to further evaluate the role of N-alkylation on the activity of indoloquinolines as antifungal agents. While N-5 substitution is required for these activities, N-10 alkylation alone leads to inactive products but is tolerated in the presence of N-5 alkyl groups. It was also discovered that bis-quindolines appear to have a more expanded antimicrobial spectrum and lower cytotoxicity than their monomeric counterparts.