Unexpectedly stable homopurine parallel triplex of SNA:RNA*SNA and L-aTNA:RNA*L-aTNA.

Homopurine strands are known to form antiparallel triplexes stabilized by G*G and A*A Hoogsteen pairs, which have two hydrogen bonds. But there has been no report on the parallel triplex formation of homopurine involving both adenosine and guanosine to the duplex. In this paper, we first report parallel triplex formation between a homopurine serinol nucleic acid (SNA) strand and an RNA/SNA duplex. Melting profiles revealed that the parallel SNA:RNA*SNA triplex was remarkably stable, even though the A*A pair has a single hydrogen bond. An L-acyclic threoninol nucleic acid (L-aTNA) homopurine strand also formed a stable parallel triplex with an L-aTNA/RNA duplex.

A H2O2-specific fluorescent probe for evaluating oxidative stress in pesticides-treated cells, rice roots and zebrafish.

Hydrogen peroxide (H2O2) plays an irreplaceable role in the evaluation of the redox status in versatile circumstances. The levels of H2O2 can be affected by both internal and external stimuli, including environmental hazards. Abnormal production of H2O2 is a common characteristic of pesticide-caused damage. Therefore, H2O2 levels can intuitively and conveniently reflect the oxidative stress caused by various pesticides in cells and organisms. However, reliable and convenient monitoring of H2O2 in living cells is still limited by the lack of specific imaging probes. In this study, a fluorescent probe (HBTM-HP) was developed for in situ observation of H2O2 fluctuations caused by pesticide treatment over time in mammalian cells, rice roots and zebrafish. HBTM-HP showed high sensitivity and selectivity for H2O2. Fluorescence imaging results confirmed that HBTM-HP could be applied to reveal H2O2 production induced by multiple pesticides. This study revealed that HBTM-HP could serves as a versatile tool to monitor the redox status related to H2O2 both in vitro and in vivo upon exposure to pesticides, and also provides a basis for clarifying the mechanisms of pesticides in physiological and pathological processes.

Stereoselective Synthesis and Antiproliferative Activity of Steviol-Based Diterpene 1,3-Aminoalcohol Regioisomers.

A series of novel diterpene-type 1,3-aminoalcohols and their regioisomers have been synthesised from natural stevioside in a stereoselective manner. The key intermediate ß-keto alcohol was prepared using Wagner-Meerwein rearrangement of the epoxide derived from steviol methyl ester. The primary aminoalcohol was formed via Raney-nickel-catalysed hydrogenation of an oxime, and a versatile library of aminoalcohols was synthesised using a Schiff base with the primary amines. The aminoalcohol regioisomers were prepared from the mesylate of the ß-keto alcohols. The corresponding primary aminoalcohol was formed via the palladium-catalysed hydrogenation of hydroxyl-azide, and click reactions of the latter were also carried out. The new compounds were characterised using 1D- and 2D-NMR techniques and HRMS measurements. The in vitro investigations showed high inhibition of cell growth in human cancer cell lines (HeLa, SiHa, A2780, MCF-7 and MDA-MB-231) in the case of naphthalic N-substituted derivatives. The antiproliferative effects were assayed using the MTT method.

Preparation of Chiral γ-Secondary Amino Alcohols via Ni-Catalyzed Asymmetric Reductive Coupling of 2-Aza-butadiene with Aldehydes.

The first Ni-catalyzed asymmetric reductive coupling of 2-aza-butadiene with aldehydes was achieved to synthesize chiral γ-secondary amino alcohols. This transformation features good enantioselectivity and tolerance to various functional groups, which may serve as a complementary method to previously reported noble-metal-catalyzed protocols. Through competition reaction, 2-aza-butadiene was proved to be a more reactive coupling component than its full-carbon analogue, 1,3-butadiene. Notably, this reaction delivers ß-siloxyl imine, an aza-aldol-type product which is difficult to access by conventional methods.

The Interplay of GPER1 with 17ß-Aminoestrogens in the Regulation of the Proliferation of Cervical and Breast Cancer Cells: A Pharmacological Approach.

The G-protein-coupled receptor for estrogen (GPER1) is a transmembrane receptor involved in the progression and development of various neoplasms whose ligand is estradiol (E2). 17ß-aminoestrogens (17ß-AEs) compounds, analogs to E2, are possible candidates for use in hormone replacement therapy (HRT), but our knowledge of their pharmacological profile is limited. Thus, we explored the molecular recognition of GPER1 with different synthetic 17ß-AEs: prolame, butolame, and pentolame. We compared the structure and ligand recognition sites previously reported for a specific agonist (G1), antagonists (G15 and G36), and the natural ligand (E2). Then, the biological effects of 17ß-AEs were analyzed through cell viability and cell-cycle assays in two types of female cancer. In addition, the effect of 17ß-AEs on the phosphorylation of the oncoprotein c-fos was evaluated, because this molecule is modulated by GPER1. Molecular docking analysis showed that 17ß-AEs interacted with GPER1, suggesting that prolame joins GPER1 in a hydrophobic cavity, similarly to G1, G15, and E2. Prolame induced cell proliferation in breast (MCF-7) and cervical cancer (SIHA) cells; meanwhile, butolame and pentolame did not affect cell proliferation. Neither 17ß-AEs nor E2 changed the activation of c-fos in MCF-7 cells. Meanwhile, in SIHA cells, E2 and 17ß-AEs reduced c-fos phosphorylation. Thus, our data suggest that butolame and pentolame, but not prolame, could be used for HRT without presenting a potential risk of inducing breast- or cervical-cancer-cell proliferation. The novelty of this work lies in its study of compound analogs to E2 that may represent important therapeutic strategies for women in menopause, with non-significant effects on the cell viability of cancer cells. The research focused on the interactions of GPER1, a molecule recently associated with promoting and maintaining various neoplasms.

Effective Perturbations by Small-Molecule Modulators on Voltage-Dependent Hysteresis of Transmembrane Ionic Currents.

The non-linear voltage-dependent hysteresis (Hys(V)) of voltage-gated ionic currents can be robustly activated by the isosceles-triangular ramp voltage (Vramp) through digital-to-analog conversion. Perturbations on this Hys(V) behavior play a role in regulating membrane excitability in different excitable cells. A variety of small molecules may influence the strength of Hys(V) in different types of ionic currents elicited by long-lasting triangular Vramp. Pirfenidone, an anti-fibrotic drug, decreased the magnitude of Ih's Hys(V) activated by triangular Vramp, while dexmedetomidine, an agonist of α2-adrenoceptors, effectively suppressed Ih as well as diminished the Hys(V) strength of Ih. Oxaliplatin, a platinum-based anti-neoplastic drug, was noted to enhance the Ih's Hys(V) strength, which is thought to be linked to the occurrence of neuropathic pain, while honokiol, a hydroxylated biphenyl compound, decreased Ih's Hys(V). Cell exposure to lutein, a xanthophyll carotenoid, resulted in a reduction of Ih's Hys(V) magnitude. Moreover, with cell exposure to UCL-2077, SM-102, isoplumbagin, or plumbagin, the Hys(V) strength of erg-mediated K+ current activated by triangular Vramp was effectively diminished, whereas the presence of either remdesivir or QO-58 respectively decreased or increased Hys(V) magnitude of M-type K+ current. Zingerone, a methoxyphenol, was found to attenuate Hys(V) (with low- and high-threshold loops) of L-type Ca2+ current induced by long-lasting triangular Vramp. The Hys(V) properties of persistent Na+ current (INa(P)) evoked by triangular Vramp were characterized by a figure-of-eight (i.e., ∞) configuration with two distinct loops (i.e., low- and high-threshold loops). The presence of either tefluthrin, a pyrethroid insecticide, or t-butyl hydroperoxide, an oxidant, enhanced the Hys(V) strength of INa(P). However, further addition of dapagliflozin can reverse their augmenting effects in the Hys(V) magnitude of the current. Furthermore, the addition of esaxerenone, mirogabalin, or dapagliflozin was effective in inhibiting the strength of INa(P). Taken together, the observed perturbations by these small-molecule modulators on Hys(V) strength in different types of ionic currents evoked during triangular Vramp are expected to influence the functional activities (e.g., electrical behaviors) of different excitable cells in vitro or in vivo.

Lipid nanoparticle-mediated lymph node-targeting delivery of mRNA cancer vaccine elicits robust CD8+ T cell response.

The targeted delivery of messenger RNA (mRNA) to desired organs remains a great challenge for in vivo applications of mRNA technology. For mRNA vaccines, the targeted delivery to the lymph node (LN) is predicted to reduce side effects and increase the immune response. In this study, we explored an endogenously LN-targeting lipid nanoparticle (LNP) without the modification of any active targeting ligands for developing an mRNA cancer vaccine. The LNP named 113-O12B showed increased and specific expression in the LN compared with LNP formulated with ALC-0315, a synthetic lipid used in the COVID-19 vaccine Comirnaty. The targeted delivery of mRNA to the LN increased the CD8+ T cell response to the encoded full-length ovalbumin (OVA) model antigen. As a result, the protective and therapeutic effect of the OVA-encoding mRNA vaccine on the OVA-antigen-bearing B16F10 melanoma model was also improved. Moreover, 113-O12B encapsulated with TRP-2 peptide (TRP2180-188)-encoding mRNA also exhibited excellent tumor inhibition, with the complete response of 40% in the regular B16F10 tumor model when combined with anti-programmed death-1 (PD-1) therapy, revealing broad application of 113-O12B from protein to peptide antigens. All the treated mice showed long-term immune memory, hindering the occurrence of tumor metastatic nodules in the lung in the rechallenging experiments that followed. The enhanced antitumor efficacy of the LN-targeting LNP system shows great potential as a universal platform for the next generation of mRNA vaccines.

Eugenol ß-Amino/ß-Alkoxy Alcohols with Selective Anticancer Activity.

Eugenol, 4-allyl-2-methoxyphenol, is the main constituent of clove essential oil and has demonstrated relevant biological activity, namely anticancer activity. Aiming to increase this activity, we synthesized a series of eugenol ß-amino alcohol and ß-alkoxy alcohol derivatives, which were then tested against two human cancer cell lines, namely gastric adenocarcinoma cells (AGS) and lung adenocarcinoma cells (A549). An initial screening was performed to identify the most cytotoxic compounds. The results demonstrated that three ß-amino alcohol derivatives had anticancer activity that justified subsequent studies, having been shown to trigger apoptosis. Importantly, the most potent molecules displayed no appreciable toxicity towards human noncancer cells. Structure-activity relationships show that changes in eugenol structure led to enhanced cytotoxic activity and can contribute to the future design of more potent and selective drugs.

Functionalized Acyclic (l)-Threoninol Nucleic Acid Four-Way Junction with High Stability In Vitro and In Vivo.

Oligonucleotides are increasingly being used as a programmable connection material to assemble molecules and proteins in well-defined structures. For the application of such assemblies for in vivo diagnostics or therapeutics it is crucial that the oligonucleotides form highly stable, non-toxic, and non-immunogenic structures. Only few oligonucleotide derivatives fulfil all of these requirements. Here we report on the application of acyclic l-threoninol nucleic acid (aTNA) to form a four-way junction (4WJ) that is highly stable and enables facile assembly of components for in vivo treatment and imaging. The aTNA 4WJ is serum-stable, shows no non-targeted uptake or cytotoxicity, and invokes no innate immune response. As a proof of concept, we modify the 4WJ with a cancer-targeting and a serum half-life extension moiety and show the effect of these functionalized 4WJs in vitro and in vivo, respectively.

Thiol-based chemical probes exhibit antiviral activity against SARS-CoV-2 via allosteric disulfide disruption in the spike glycoprotein.

The development of small-molecules targeting different components of SARS-CoV-2 is a key strategy to complement antibody-based treatments and vaccination campaigns in managing the COVID-19 pandemic. Here, we show that two thiol-based chemical probes that act as reducing agents, P2119 and P2165, inhibit infection by human coronaviruses, including SARS-CoV-2, and decrease the binding of spike glycoprotein to its receptor, the angiotensin-converting enzyme 2 (ACE2). Proteomics and reactive cysteine profiling link the antiviral activity to the reduction of key disulfides, specifically by disruption of the Cys379-Cys432 and Cys391-Cys525 pairs distal to the receptor binding motif in the receptor binding domain (RBD) of the spike glycoprotein. Computational analyses provide insight into conformation changes that occur when these disulfides break or form, consistent with an allosteric role, and indicate that P2119/P2165 target a conserved hydrophobic binding pocket in the RBD with the benzyl thiol-reducing moiety pointed directly toward Cys432. These collective findings establish the vulnerability of human coronaviruses to thiol-based chemical probes and lay the groundwork for developing compounds of this class, as a strategy to inhibit the SARS-CoV-2 infection by shifting the spike glycoprotein redox scaffold.

Structure, DFT studies and evaluation of catechol oxidase (CO) mimic activity of mononuclear Co(II) complexes derived from aminoalcohols: an experimental and theoretical approach.

The impetus to modeling of enzyme mimics comes from their potential to provide insight to the alternate mechanistic pathways of the native enzymes. The present study demonstrates the syntheses and characterization of two different cobalt(II) complexes, [Co(pdm)(Phen)Cl]Cl·H2O (1) and [Co(pmmH)2(SCN)2] (2) with the aminoalcohol ligands such as pyridine-2,6-dimethanol (pdmH2) or 2-pyridinemonomethanol (pmmH) and their assessment as catechol oxidase (CO) enzyme mimic. Single Crystal X-ray diffraction and powder X-ray diffraction data suggest the octahedral environment around the Co(II) ion and the complexes form extensive 1D or 2D propagating network as a result of non-covalent interactions (O···H and C-H···π). TD-DFT calculations were used to explain the spectral bands obtained during the UV-Vis absorption studies and it is ascertained that the transitions were mainly of the intra-ligand charge transfer (ILCT) type. The catecholase biomimetic catalytic activity of the synthesized complexes has been investigated in detail and the kinetics is also performed. The results obtained show that both the complexes catalyze the aerobic oxidation of catechol to the corresponding o-quinone. The Kcat value for 1 is 106.99 h‒1 and for 2 is 90.32 h‒1 in methanol. It may be mentioned here that 1 and 2 are effective catalysts, with the order of activity being 1 > 2. The order of enzymatic activity is well justified by CV and DFT studies.Communicated by Ramaswamy H. Sarma.

Pre-Clinical Activity of Amino-Alcohol Dimeric Naphthoquinones as Potential Therapeutics for Acute Myeloid Leukemia.

BACKGROUND: The clinical outcomes of patients with Acute Myeloid Leukemia (AML) remain unsatisfactory. Therefore the development of more efficacious and better-tolerated therapy for AML is critical. We have previously reported anti-leukemic activity of synthetic halohydroxyl dimeric naphthoquinones (BiQ) and aziridinyl BiQ. OBJECTIVE: This study aimed to improve the potency and bioavailability of BiQ compounds and investigate antileukemic activity of the lead compound in vitro and a human AML xenograft mouse model. METHODS: We designed, synthesized, and performed structure-activity relationships of several rationally designed BiQ analogues with amino alcohol functional groups on the naphthoquinone core rings. The compounds were screened for anti-leukemic activity and the mechanism as well as in vivo tolerability and efficacy of our lead compound was investigated. RESULTS: We report that a dimeric naphthoquinone (designated BaltBiQ) demonstrated potent nanomolar anti-leukemic activity in AML cell lines. BaltBiQ treatment resulted in the generation of reactive oxygen species, induction of DNA damage, and inhibition of indoleamine dioxygenase 1. Although BaltBiQ was tolerated well in vivo, it did not significantly improve survival as a single agent, but in combination with the specific Bcl-2 inhibitor, Venetoclax, tumor growth was significantly inhibited compared to untreated mice. CONCLUSION: We synthesized a novel amino alcohol dimeric naphthoquinone, investigated its main mechanisms of action, reported its in vitro anti-AML cytotoxic activity, and showed its in vivo promising activity combined with a clinically available Bcl-2 inhibitor in a patient-derived xenograft model of AML.

Design, synthesis, and characterization of novel aminoalcohol quinolines with strong in vitro antimalarial activity.

Malaria is the fifth most lethal parasitic infections in the world. Herein, five new series of aminoalcohol quinolines including fifty-two compounds were designed, synthesized and evaluated in vitro against Pf3D7 and PfW2 strains. Among them, fourteen displayed IC50 values below or near of 50.0 nM whatever the strain with selectivity index often superior to 100.17b was found as a promising antimalarial candidate with IC50 values of 14.9 nM and 11.0 nM against respectively Pf3D7 and PfW2 and a selectivity index higher than 770 whatever the cell line is. Further experiments were achieved to confirm the safety and to establish the preliminary ADMET profile of compound 17b before the in vivo study performed on a mouse model of P. berghei ANKA infection. The overall data of this study allowed to establish new structure-activity relationships and the development of novel agents with improved pharmacokinetic properties.

Kinetic Studies of Newly Patented Aminoalkanol Derivatives with Potential Anticancer Activity as Competitive Inhibitors of Prostate Acid Phosphatase.

BACKGROUND: Acid phosphatase and its regulation are important objects of biological and clinical research and play an important role in the development and treatment of prostate and bone diseases. The newly patented aminoalkanol (4-[2-hydroxy-3-(propan-2-ylamino)propyl]-1,7-dimethyl-8,9-diphenyl-4-azatricyclo[5.2.1.02,6]dec-8-ene-3,5,10-trione hydrochloride) (I) and (4-[3-(dimethylamino)-2-hydroxypropyl]-1,7-dimethyl-8,9-diphenyl-4-azatricyclo[5.2.1.02,6]dec-8-ene-3,5,10-trione hydrochloride) (II) derivatives have potential anticancer activity, and their influence on enzymatic activity can significantly impact the therapeutic effects of acid phosphatase against many diseases. Therefore, in this study, we investigated the action of compounds (I) and (II) on acid phosphatase. METHODS: Capillary electrophoresis was used to evaluate the inhibition of acid phosphatase. Lineweaver-Burk plots were constructed to compare the Km of this enzyme in the presence of inhibitors (I) or (II) with the Km in solutions without these inhibitors. RESULTS: Compound (I) showed a stronger competitive inhibition against acid phosphatase, whereas derivative (II) showed a weaker competitive type of inhibition. The detailed kinetic studies of these compounds showed that their type and strength of inhibition as well as affinity depend on the kind of substituent occurring in the main chemical molecule. CONCLUSIONS: This study is of great importance because the disclosed inhibition of acid phosphatase by compounds (I) and (II) raises the question of whether these compounds could have any effect on the treatment possibilities of prostate diseases.

Synthesis and Biological Application of Isosteviol-Based 1,3-Aminoalcohols.

Starting from isosteviol, a series of diterpenoid 1,3-aminoalcohol derivatives were stereoselectively synthesised. The acid-catalysed hydrolysis and rearrangement of natural stevioside gave isosteviol, which was transformed to the key intermediate methyl ester. In the next step, Mannich condensation of diterpenoid ketone, paraformaldehyde, and secondary amines resulted in the formation of 1,3-aminoketones with different stereoselectivities. During the Mannich condensation with dibenzylamine, an interesting N-benzyl → N-methyl substituent exchange was observed. Reduction of 1,3-aminoketones produced diastereoisomeric 1,3-aminoalcohols. Alternatively, aminoalcohols were obtained via stereoselective hydroxy-formylation, followed by oxime preparation, reduction, and finally, reductive alkylation of the obtained primary aminoalcohols. An alternative 1,3-aminoalcohol library was prepared by reductive amination of the intermediate 3-hydroxyaldehyde obtained from isosteviol in two-step synthesis. Cytotoxic activity of compounds against human tumour cell lines (A2780, SiHa, HeLa, MCF-7 and MDA-MB-231) was investigated. In our preliminary study, the 1,3-aminoalcohol function and N-benzyl substitution seemed to be essential for the reliable antiproliferative activity. To extend their application, a diterpenoid condensed with 2-phenylimino-1,3-thiazine and -1,3-oxazine was also attempted to prepare, but only formation of thioether intermediate was observed.

Carboxyboranylamino ethanol: unprecedented discovery of boron agents for neutron capture therapy in cancer treatment.

Carboxyboranylamino ethanol (Me2N(BH2CO2H)CH2CH2OH, 1) was prepared in 75.0% yield by an amine-exchange reaction. Compound 1 shows lower cytotoxicity and higher anti-tumor efficacy in vitro towards the SCCVII cell line in comparison with 4-borono-L-phenylalanine (BPA) and methyl 2-hydroxyl-5-(1'-ortho-carbonylmethyl-1',2',3'-triazol-4'-yl)-benzonate (2). The bio-enhancement is interpreted using molecular docking calculations.

In Silico Prediction of the Toxic Potential of Neuroprotective Bifunctional Molecules Based on Chiral N-Propargyl-1,2-amino Alcohol Derivatives.

N-Propargylamines are useful synthetic scaffolds for the synthesis of bioactive molecules, and in addition, they possess important pharmacological activities. We obtained several neuroprotective molecules, chiral 1,2-amino alcohols and 1,2-diamines, able to reduce by almost 70% the rotenone and oligomycin A-induced damage in SH-SY5Y cells. Furthermore, some molecules assessed also counteracted the toxicity evoked by the Ser/Thr phosphatase inhibitor okadaic acid. Before extrapolating these data to preclinical studies, we analyze the molecules through an in silico prediction system to detect carcinogenicity risk or other toxic effects. In light of these promising results, these molecules may be considered as a lead family of neuroprotective and relatively safe compounds.

A new lipophilic amino alcohol, chemically similar to compound FTY720, attenuates the pathogenesis of experimental autoimmune encephalomyelitis by PI3K/Akt pathway inhibition.

Experimental autoimmune encephalomyelitis (EAE) is one of the main animal models used for the study of Multiple Sclerosis (MS). Long-chain lipophilic amino alcohols with immunoregulatory activities have already been studied in some models of inflammatory diseases, but the action of these compounds in EAE and MS is still unknown. In this study, we investigated whether the lipophilic amino alcohol 4b would act to improve the clinical signs of EAE and reduce the demyelination process and the neuroinflammatory parameters in the spinal cord, as well as the inflammatory process in the inguinal lymph nodes, of C57Bl/6 mice induced with EAE after stimulation with MOG35-55 and pertussis toxin. The 4b treatment (1.0 mg/kg/day) was orally administered, starting on the day of onset of clinical signs of the disease (10th) and ending on the 20th day after immunization. This treatment was able to reduce the cell count on the inguinal lymph nodes, the migration of inflammatory cells into the central nervous system (CNS), as well as the processes of microgliosis, astrogliosis, and the production of chemokines and pro-inflammatory cytokines, thus increasing the IL-10 anti-inflammatory cytokine levels in EAE mice. The inhibition of Akt phosphorylation in the CNS of EAE mice after treatment with 4b indicates that the immunoregulatory action of 4b is related to the PI3K/Akt signaling pathway. Our results indicate the immunoregulatory efficacy of the new compound 4b in the control of some inflammatory parameters and in the glial proliferation. In addition, 4b was able to reduce the demyelination of neurons and the worsening of clinical signs of EAE as effectively as the compound FTY720, the first oral drug approved by the FDA for the treatment of MS.

Secondary Amino Alcohols: Traceless Cleavable Linkers for Use in Affinity Capture and Release.

Capture and release of peptides is often a critical operation in the pathway to discovering materials with novel functions. However, the best methods for efficient capture impede facile release. To overcome this challenge, we report linkers based on secondary amino alcohols for the release of peptides after capture. These amino alcohols are based on serine (seramox) or isoserine (isoseramox) and can be incorporated into peptides during solid-phase peptide synthesis through reductive amination. Both linkers are quantitatively cleaved within minutes under NaIO4 treatment. Cleavage of isoseramox produced a native peptide N-terminus. This linker also showed broad substrate compatibility; incorporation into a synthetic peptide library resulted in the identification of all sequences by nanoLC-MS/MS. The linkers are cell compatible; a cell-penetrating peptide that contained this linker was efficiently captured and identified after uptake into cells. These findings suggest that such secondary amino alcohol based linkers might be suitable tools for peptide-discovery platforms.

Amino Alcohol Acrylonitriles as Activators of the Aryl Hydrocarbon Receptor Pathway: An Unexpected MTT Phenotypic Screening Outcome.

Lead (Z)-N-(4-(2-cyano-2-(3,4-dichlorophenyl)vinyl)phenyl)acetamide, 1 showed MCF-7 GI50 =30 nM and 400-fold selective c.f. MCF10A (normal breast tissue). Acetamide moiety modification (13 a-g) to introduce additional hydrophobicity was favoured with MCF-7 breast cancer cell activity enhanced at 1.3 nM. Other analogues were potent against the HT29 colon cancer cell line at 23 nM. Textbook SAR data was observed in the MCF-7 cell line, in an MTT assay, via the ortho (17 a), meta (17 b) and para (13 f). The amino alcohol -OH moiety was pivotal, but no stereochemical preference noted. But, these data did not fit our homology modelling expectations. Aberrant MTT ((3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) screening results and metabolic interference confirmed by sulforhodamine B (SRB) screening. Interfering analogues resulted in 120 and 80-fold CYP1A1 and CYP1A2 amplification, with no upregulation of SULT1A1. This is consistent with activation of the AhR pathway. Piperidine per-deuteration reduced metabolic inactivation. 3-OH / 4-OH piperidine analogues showed differential MTT and SRB activity supporting MTT assay metabolic inactivation. Data supports piperidine 3-OH, but not the 4-OH, as a CYP substrate. This family of ß-amino alcohol substituted 3,4-dichlorophenylacetonitriles show broad activity modulated via the AhR pathway. By SRB analysis the most potent analogue was 23 b, (Z)-3-(4-(3-(4-phenylpiperidin-1-yl)-2-hydroxypropoxy)phenyl)-2-(3,4-dichlorophenyl)-acrylonitrile.