Subramaniam Ananthan

14-Alkoxy- and 14-acyloxypyridomorphinans: μ agonist/δ antagonist opioid analgesics with diminished tolerance and dependence side effects.

In the search for opioid ligands with mixed functional activity, a series of 5-(4-chlorophenyl)-4,5α-epoxypyridomorphinans possessing alkoxy or acyloxy groups at C-14 was synthesized and evaluated. In this series, the affinity and functional activity of the ligands were found to be influenced by the nature of the substituent at C-14 as well as by the substituent at N-17. Whereas the incorporation of a 3-phenylpropoxy group at C-14 on N-methylpyridomorhinan gave a dual MOR agonist/DOR agonist 17h, its incorporation on N-cyclopropylmethylpyridomorphinan gave a MOR agonist/DOR antagonist 17d. Interestingly, 17d, in contrast to 17h, did not produce tolerance or dependence effects upon prolonged treatment in cells expressing MOR and DOR. Moreover, 17d displayed greatly diminished analgesic tolerance as compared to morphine upon repeated administration, thus supporting the hypothesis that ligands with MOR agonist/DOR antagonist functional activity could emerge as novel analgesics devoid of tolerance, dependence, and related side effects.

Design, Synthesis, and Structure–Activity Relationship Studies of a Series of [4-(4-Carboxamidobutyl)]-1-arylpiperazines: Insights into Structural Features Contributing to Dopamine D3 versus D2 Receptor Subtype Selectivity

Antagonist and partial agonist modulators of the dopamine D3 receptor (D3R) have emerged as promising therapeutics for the treatment of substance abuse and neuropsychiatric disorders. However, development of druglike lead compounds with selectivity for the D3 receptor has been challenging because of the high sequence homology between the D3R and the dopamine D2 receptor (D2R). In this effort, we synthesized a series of acylaminobutylpiperazines incorporating aza-aromatic units and evaluated their binding and functional activities at the D3 and D2 receptors. Docking studies and results from evaluations against a set of chimeric and mutant receptors suggest that interactions at the extracellular end of TM7 contribute to the D3R versus D2R selectivity of these ligands. Molecular insights from this study could potentially enable rational design of potent and selective D3R ligands.

Studies of the Biogenic Amine Transporters. 13. Identification of “Agonist” and “Antagonist” Allosteric Modulators of Amphetamine-Induced Dopamine Release

Recent studies identified novel allosteric modulators of the dopamine (DA) transporter (DAT). N-(Diphenylmethyl)-2-phenyl-4-quinazolinamine (SoRI-9804), N-(2,2-diphenylethyl)-2-phenyl-4-quinazolinamine (SoRI-20040), and N-(3,3-diphenylpropyl)-2-phenyl-4-quinazolinamine (SoRI-20041) partially inhibited [125I]3β-(4′-iodophenyl)tropan-2β-carboxylic acid methyl ester (RTI-55) binding, slowed the dissociation rate of [125I]RTI-55 from the DAT, and partially inhibited [3H]dopamine uptake. In the present study, we report that SoRI-9804 and SoRI-20040, at doses that do not alter release, partially inhibited d-amphetamine-induced DAT-mediated release of [3H]1-methyl-4-phenylpyridinium (MPP+)or[3H]dopamine from striatal synaptosomes (“DAT-mediated DA release”) in a dose-dependent manner. SoRI-20041, which does not alter DAT-mediated DA release measured with [3H]DA, reversed the effect of SoRI-20040. SoRI-20040 and SoRI-9804 also partially inhibited DAT-mediated DA release induced by DA or (±)-3,4-methylenedioxyamphetamine, demonstrating that the observed partial inhibition is not specific for a particular DAT substrate. SoRI-9804 and SoRI-20040 did not attenuate d-amphetamine-induced release of [3H]5-hydroxytryptamine from serotonergic, or [3H]MPP+ from noradrenergic, nerve terminals. Kinetic experiments demonstrated that SoRI-9804, in contrast to cocaine, slowed d-amphetamine-induced release of [3H]MPP+ from dopaminergic nerve terminals without altering the apparent rate constants. The two major findings of this study are 1) the identification of both “agonist” (SoRI-9804 and SoRI-20040) and “antagonist” (SoRI-20041) allosteric modulators of d-amphetamine-induced DAT-mediated DA release and 2) [3H]DA uptake and d-amphetamine-induced DAT-mediated efflux can be separately modulated. Such agents may have therapeutic potential for the treatment of stimulant addiction, Parkinsons disease, and other psychiatric disorders.

Studies of the Biogenic Amine Transporters. 12. Identification of Novel Partial Inhibitors of Amphetamine-Induced Dopamine Release

Previous studies identified partial inhibitors and allosteric modulators of 5-hydroxytryptamine ([5-amino-3-(3,4-dichlorophenyl)-1,2-dihydropyrido[3,4-b]pyrazin-7-yl]carbamic acid ethyl ester [SoRI-6238], 4-(2-[bis(4-fluorophenyl)methoxy]ethyl)-1-(2-trifluoromethyl-benzyl)-piperidine [TB-1-099]) and dopamine transporters N-(diphenylmethyl)-2-phenyl-4-quinazolinamine, [SoRI-9804]). We report here the identification of three novel allosteric modulators of the dopamine transporter [N-(2,2-diphenylethyl)-2-phenyl-4-quinazolinamine [SoRI-20040], N-(3,3-diphenylpropyl)-2-phenyl-4-quinazolinamine [SoRI-20041], and [4-amino-6-[(diphenylmethyl)amino]-5-nitro-2-pyridinyl]carbamic acid ethyl ester [SoRI-2827]]. Membranes were prepared from human embryonic kidney cells expressing the cloned human dopamine transporter (hDAT). [125I]3β-(4′-Iodophenyl)tropan-2β-carboxylic acid methyl ester ([125I]RTI-55) binding and other assays followed published procedures. SoRI-20040, SoRI-20041, and SoRI-2827 partially inhibited [125I]RTI-55 binding, with EC50 values ranging from ∼1.4 to 3 μM and Emax values decreasing as the [125I]RTI-55 concentrations increased. All three compounds decreased the [125I]RTI-55 Bmax value and increased the apparent Kd value in a manner well described by a sigmoid dose-response curve. In dissociation rate experiments, SoRI-20040 (10 μM) and SoRI-20041 (10 μM), but not SoRI-2827 (10 μM), slowed the dissociation of [125I]RTI-55 from hDAT by ∼30%. Using rat brain synaptosomes, all three agents partially inhibited [3H]dopamine uptake, with EC50 values ranging from 1.8 to 3.1 μM and decreased the Vmax value in a dose-dependent manner. SoRI-9804 and SoRI-20040 partially inhibited amphetamine-induced dopamine transporter-mediated release of [3H]1-methyl-4-phenylpyridinium ion from rat caudate synaptosomes in a dose-dependent manner. Viewed collectively, we report several compounds that allosterically modulate hDAT binding and function, and we identify novel partial inhibitors of amphetamine-induced dopamine release.

Identification of quinazoline compounds as novel potent inhibitors of Wnt/β-catenin signaling in colorectal cancer cells.

The Wnt/β-catenin signaling pathway is critical for the initiation and progression of most colon cancers, and has emerged as one of the most promising targets for colorectal cancer chemoprevention and treatment. In this study, we have discovered a structurally related series of quinazolines as potent inhibitors of Wnt/β-catenin signaling in colorectal cancer cells harboring mutations in CTNNB1 or APC. We showed that the quinazoline leads suppressed Wnt/β-catenin signaling without altering the level of β-catenin protein in colorectal cancer cells, suggesting that they act on the downstream elements of the pathway. Moreover, the quinazoline leads displayed potent anticancer activities with IC50 values between 4.9 and 17.4 μM in colorectal cancer cells. Importantly, we also found that a structurally related quinazoline lacking inhibitory effect on Wnt/β-catenin signaling was unable to suppress colorectal cancer cell proliferation. Together, these results suggest that the quinazoline lead compounds identified in this study have therapeutic potential for the prevention and treatment of colorectal cancer.

Discovery of novel frizzled-7 inhibitors by targeting the receptor’s transmembrane domain

Frizzled (Fzd) proteins are seven transmembrane receptors that belong to a novel and separated family of G-protein-coupled receptors (GPCRs). The Fzd receptors can respond to Wnt proteins to activate the canonical β-catenin pathway which is important for both initiation and progression of cancers. Disruption of the Wnt/β-catenin signal thus represents an opportunity for rational cancer prevention and therapy. Of the 10 members of the Fzd family, Fzd7 is the most important member involved in cancer development and progression. In the present studies, we applied structure-based virtual screening targeting the transmembrane domain (TMD) of Fzd7 to select compounds that could potentially bind to the Fzd7-TMD and block the Wnt/Fzd7 signaling and further evaluated them in biological assays. Six small molecule compounds were confirmed as Fzd7 inhibitors. The best hit, SRI37892, significantly blocked the Wnt/Fzd7 signaling with IC50 values in the sub-micromolar range and inhibited cancer cell proliferation with IC50 values around 2 μM. Our results provide the first proof of concept of targeting Fzd-TMD for the development of Wnt/Fzd modulators. The identified small molecular Fzd7 inhibitors can serve as a useful tool for studying the regulation mechanism(s) of Wnt/Fzd7 signaling as well as a starting point for the development of cancer therapeutic agents.

Interactions Between Cocaine and the Putative Allosteric Dopamine Transporter Ligand SRI-31142

Drugs that inhibit the dopamine (DA) transporter (DAT) include both therapeutic agents and abused drugs. Recent studies identified a novel series of putative allosteric DAT inhibitors, but the in vivo effects of these compounds are unknown. This study examined the abuse-related behavioral and neurochemical effects produced in rats by SRI-31142 [2-(7-methylimidazo[1,2-a]pyridin-6-yl)-N-(2-phenyl-2-(pyridin-4-yl)ethyl)quinazolin-4-amine], one compound from this series. In behavioral studies, intracranial self-stimulation (ICSS) was used to compare the effects produced by SRI-31142, the abused and nonselective DAT inhibitor cocaine, and the selective DAT inhibitor GBR-12935 [1-[2-(diphenylmethoxy)ethyl]-4-(3-phenylpropyl)piperazine]. In neurochemical studies, in vivo microdialysis was used to compare the effects of SRI-31142 and cocaine on levels of DA and serotonin in nucleus accumbens (NAc). The effects of SRI-31142 in combination with cocaine were also examined in both procedures. In contrast to cocaine and GBR-12935, SRI-31142 failed to produce abuse-related increases in ICSS or NAc DA; instead, SRI-31142 only decreased ICSS and NAc DA at a dose that was also sufficient to block cocaine-induced increases in ICSS and NAc DA. Pharmacokinetic studies suggested low but adequate brain penetration of SRI-31142, in vitro binding studies failed to identify likely non-DAT targets, and in vitro functional assays failed to confirm DA uptake inhibition in an assay of DAT-mediated fluorescent signals in live cells. These results indicate that SRI-31142 does not produce cocaine-like abuse-related effects in rats. SRI-31142 may have utility to block cocaine effects and may warrant further study as a candidate pharmacotherapy; however, the role of DAT in mediating these effects is unclear, and side effects may be a limiting factor.