Understanding the patient and supporter journey in cocaine use disorder.

Background: There is a paucity of literature describing experiences and journey of individuals with cocaine use disorder (CUD) and supporters who care for them. The aim of this study was to understand and document the journey of individuals with current CUD, those in CUD remission, and supporters. Methods: The online bulletin board (OBB) is a qualitative tool where participants engage in an interactive discussion on a virtual forum. After completing a 15-minute screening questionnaire determining eligibility, individuals in CUD remission and supporters participated in an OBB for 60 minutes, split across 8 days over 2 weeks. Individuals with current CUD participated in a one-time virtual focus group discussion for 90 minutes. Results: Individuals in CUD remission (n=35) were from Brazil, France, Spain, the UK, and the US; those with current CUD (n=5) and supporters (n=6) were from the US. Key insights were that individuals with current CUD were seeking a 'euphoric high' that cocaine provides. Those in CUD remission described a 'euphoric high' when they first tried cocaine, but over time it became harder to re-create this feeling. Individuals in CUD remission expressed a 'rollercoaster' of emotions from when they first started using cocaine to when they stopped. Supporters were sad, isolated, and worried about a potential cocaine overdose for their loved ones with CUD. Conclusion: The study provides valuable insights into the experiences and journey of individuals with CUD and their supporters. Data generated from this study gives insights into this under-served and growing population.

Patient perspectives on current and potential therapies and clinical trial approaches for cocaine use disorder.

Background: Cocaine use disorder (CUD) is characterized by the continued use of cocaine despite serious impacts on life. This study focused on understanding the perspective of individuals with current CUD, individuals in CUD remission, and their supporters regarding current therapies, future therapies, and views on clinical trials for CUD. Methods: The online bulletin board (OBB) is a qualitative tool where participants engage in an interactive discussion on a virtual forum. Following completion of a screening questionnaire to determine eligibility, individuals in CUD remission and their supporters logged in to the OBB and responded to questions posed by the moderator. Individuals with current CUD participated in a one-time virtual focus group. Results: All individuals with current CUD and 94% of those in CUD remission reported a diagnosis consistent with CUD or substance use disorder during screening. Individuals with current CUD and their supporters were recruited from the United States (US). Individuals in CUD remission were recruited from five countries, including the US. Individuals with current CUD reported hesitation about seeking treatment due to stigma, a lack of privacy, and being labeled as a drug seeker; barriers to therapy included time, cost, and a lack of privacy. Participants wanted a safe therapy to stop cravings and withdrawal symptoms. Seven clinical trial outcomes, including long-term abstinence and craving control, were suggested based on collected insights. Conclusion: This study can help inform the design of clinical trials and emphasize the need for effective, safe, and accessible therapies. Recruiting participants will require significant trust building.

Current trends in small molecule discovery targeting key cellular signaling events towards the combined management of diabetes and obesity.

Non-insulin dependent diabetes mellitus, also known as Type 2 diabetes is a polygenic disorder leading to abnormalities in the carbohydrate and lipid metabolism. The major contributors in the pathophysiology of type 2 diabetes (T2D) include resistance to insulin action, ß cell dysfunction, an abnormality in glucose metabolism and storage, visceral obesity and to some extent inflammation and oxidative stress. Insulin resistance, along with a defect in insulin secretion by the pancreatic ß cells is instrumental towards progression to hyperglycemia. Increased incidence of obesity is also a major contributing factor in the escalating rates of type 2 diabetes. Drug discovery efforts are therefore crucially dependent on identifying individual molecular targets and validating their relevance to human disease. The current review discusses bioactive compounds from medicinal plants offering enhanced therapeutic potential for the combined patho-physiology of diabetes and obesity. We have demonstrated that 3ß-taraxerol a pentacyclic triterpenoid (14-taraxeren-3-ol) isolated from the ethyl acetate extract of Mangifera indica, chlorogenic acid isolated from the methanol extract of Cichorium intybus, methyl tetracosanoate from the methanol extract of Costus pictus and vitalboside A derived from methanolic extract of Syzygium cumini exhibited significant effects on insulin stimulated glucose uptake causing insulin sensitizing effects on 3T3L1 adipocytes (an in vitro model mimicking adipocytes). Whereas, (3ß)-stigmast-5-en-3-ol isolated from Adathoda vasica and Aloe emodin isolated from Cassia fistula showed significant insulin mimetic effects favoring glucose uptake in L6 myotubes (an in vitro model mimicking skeletal muscle cells). These extracts and molecules showed glucose uptake through activation of PI3K, an important insulin signaling intermediate. Interestingly, cinnamic acid isolated from the hydro-alcohol extract of Cinnamomum cassia was found to activate glucose transport in L6 myotubes through the involvement of GLUT4 via the PI3K-independent pathway. However, the activation of glucose storage was effective in the presence of 3ß-taraxerol and aloe emodin though inhibition of GSK3ß activity. Therefore, the mechanism of improvement of glucose and lipid metabolism exhibited by the small molecules isolated from our lab is discussed. However, Obesity is a major risk factor for type-2 diabetes leading to destruction of insulin receptors causing insulin resistance. Identification of compounds with dual activity (anti-diabetic and antiadipogenic activity) is of current interest. The protein tyrosine phosphatase 1B (PTP1B) is an important negative regulator of the insulin and leptin-signaling pathway is of significance in target definition and discovery.

Selective Inhibition of PTP1B by Vitalboside A from Syzygium cumini Enhances Insulin Sensitivity and Attenuates Lipid Accumulation Via Partial Agonism to PPARγ: In Vitro and In Silico Investigation.

Although antidiabetic drugs show good insulin-sensitizing property for T2DM, they also exhibit undesirable side-effects. Partial peroxisome proliferator-activated receptor γ agonism with protein tyrosine phosphatase 1B inhibition is considered as an alternative therapeutic approach toward the development of a safe insulin sensitizer. Bioactivity-based fractionation and purification of Syzygium cumini seeds led to the isolation and identification of bifunctional Vitalboside A, which showed antidiabetic and anti-adipogenic activities, as measured by glucose uptake in L6 and 3T3-L1 adipocytes and Nile red assay. A non-competitive allosteric inhibition of protein tyrosine phosphatase 1B by Vitalboside A was observed, which was confirmed by docking studies. Inhibitor studies with wortmannin and genistein showed an IRTK- and PI3K-dependent glucose uptake. A PI3K/AKT-dependent activation of GLUT4 translocation and an inactivation of GSK3ß were observed, confirming its insulin-sensitizing potential. Vitalboside A exhibited partial transactivation of peroxisome proliferator-activated receptor γ with an increase in adiponectin secretion, which was confirmed using docking analysis. Vitalboside A is a bifunctional molecule derived from edible plant showing inhibition of PTP1B and partial agonism to peroxisome proliferator-activated receptor γ which could be a promising therapeutic agent in the management of obesity and diabetes.

In Vitro Investigation of the Immunomodulatory Potential of Probiotic Lactobacillus casei.

The current study investigated the immunomodulatory potential of ethyl acetate soluble supernatant of Lactobacillus casei (LC-EAS) in vitro. The effect of LC-EAS on nitric oxide release was analyzed in RAW 264.7 cells, wherein, an inhibition in nitric oxide production through suppression of inducible nitric oxide synthase mRNA expression was observed. Evaluation of LC-EAS on LPS-induced peripheral blood mononuclear cells showed a down-regulation in TNF-α and IL-6 genes and an upregulation of IL-10. An inhibition in the protein expression of NF-κB, ERK1/2 and STAT3 phosphorylation confirms the immunomodulatory potential of LC-EAS. The effect of LC-EAS on in vitro intestinal epithelial cells was investigated using HT-29 human colon adenocarcinoma cancer cells. LC-EAS exhibited an inhibition of NF-κB and ERK1/2 phosphorylation, whereas STAT3 phosphorylation was unregulated. To evaluate the downstream target of STAT3 upregulation, expression of the intestinal trefoil factor TFF3 which is a NF-κB regulator and STAT3 downstream target was studied. LC-EAS was observed to elevate TFF3 mRNA expression. Overall the study shows that the anti-inflammatory potential of LC-EAS is through inhibition of NF-κB in different cell types.

Molecular dynamics approach to probe the allosteric inhibition of PTP1B by chlorogenic and cichoric acid.

Protein tyrosine phosphatase 1B (PTP1B), a major negative regulator of the insulin and leptin signaling pathway, is a potential target for therapeutic intervention against diabetes and obesity. The recent discovery of an allosteric site in PTP1B has created an alternate strategy in the development of PTP1B targeted therapy. The current study investigates the molecular interactions between the allosteric site of PTP1B with two caffeoyl derivatives, chlorogenic acid (CGA) and cichoric acid (CHA), using computational strategies. Molecular docking analysis with CGA and CHA at the allosteric site of PTP1B were performed and the resulting protein-ligand complexes used for molecular dynamics simulation studies for a time scale of 10 ns. Results show stable binding of CGA and CHA at the allosteric site of PTP1B. The flexibility of the WPD loop was observed to be constrained by CGA and CHA in the open (inactive), providing molecular mechanism of allosteric inhibition. The allosteric inhibition of CGA and CHA of PTP1B was shown to be favorable due to no restriction by the α-7 helix in the binding of CGA and CHA at the allosteric binding site. In conclusion, our results exhibit an inhibitory pattern of CGA and CHA against PTP1B through potent binding at the allosteric site.

3ß-hydroxylup-20(29)-ene-27,28-dioic acid dimethyl ester, a novel natural product from Plumbago zeylanica inhibits the proliferation and migration of MDA-MB-231 cells.

Plumbago zeylanica, a traditional Indian herb is being used for the therapy of rheumatism and has been approved for anti-tumor activity. However, the molecular mechanisms involved in the biological action are not very well understood. In this study, the anti-invasive activities of P. zeylanica methanolic extract (PME) and pure compound 3ß-hydroxylup-20(29)-ene-27,28-dioic acid (PZP) isolated from it are investigated in vitro. PME and PZP were noted to have the ability to induce apoptosis as assessed by flow cytometry. Further, the molecular mechanism of apoptosis induced by PME and PZP was found by the loss of mitochondrial membrane potential with the down regulation of Bcl-2, increased expression of Bad, release of cytochrome c, activation of caspase-3 and cleavage of PARP leading to DNA fragmentation. Importantly, both PME and PZP were observed to suppress MDA-MB-231 cells adhesion to the fibronectin-coated substrate and also inhibited the wound healing migration and invasion of MDA-MB-231 cells through the reconstituted extracellular matrix. Gelatin zymography revealed that PME and PZP decreased the secretion of matrix metalloproteinases-2 (MMP-2) and metalloproteinases-9 (MMP-9). Interestingly both PME and PZP exerted an inhibitory effect on the protein levels of p-PI3K, p-Akt, p-JNK, p-ERK1/2, MMP-2, MMP-9, VEGF and HIF-1α that are consistent with the observed anti-metastatic effect. Collectively, these data provide the molecular basis of the anti-proliferative and anti-metastatic effects of PME and PZP.

3beta-taraxerol of Mangifera indica, a PI3K dependent dual activator of glucose transport and glycogen synthesis in 3T3-L1 adipocytes.

BACKGROUND: The present study focuses on identifying and developing an anti-diabetic molecule from plant sources that would effectively combat insulin resistance through proper channeling of glucose metabolism involving glucose transport and storage. METHODS: Insulin-stimulated glucose uptake formed the basis for isolation of a bioactive molecule through column chromatography followed by its characterization using NMR and mass spectroscopic analysis. Mechanism of glucose transport and storage was evaluated based on the expression profiling of signaling molecules involved in the process. RESULTS: The study reports (i) the isolation of a bioactive compound 3beta-taraxerol from the ethyl acetate extract (EAE) of the leaves of Mangifera indica (ii) the bioactive compound exhibited insulin-stimulated glucose uptake through translocation and activation of the glucose transporter (GLUT4) in an IRTK and PI3K dependent fashion. (iii) the fate of glucose following insulin-stimulated glucose uptake was ascertained through glycogen synthesis assay that involved the activation of PKB and suppression of GSK3beta. GENERAL SIGNIFICANCE: This study demonstrates the dual activity of 3beta-taraxerol and the ethyl acetate extract of Mangifera indica as a glucose transport activator and stimulator of glycogen synthesis. 3beta-taraxerol can be validated as a potent candidate for managing the hyperglycemic state.

Probing key targets in insulin signaling and adipogenesis using a methanolic extract of Costus pictus and its bioactive molecule, methyl tetracosanoate.

A methanolic extract of Costus pictus (CPME) showed optimum anti-diabetic activity at 100 ng/ml. Bioactivity-guided purification of CPME led to the isolation of methyl tetracosanoate (MT) which showed an optimum glucose uptake at 1 ng/ml. CPME at 10 mug/ml inhibited adipogenesis whereas fully differentiated adipocytes exhibited a 3-fold increase in lipid accumulation compared to pre-adipocytes. Gene and protein expression of key targets in insulin signaling and adipogenesis pathway revealed that CPME exhibited anti-diabetic activity along with anti-adipogenic activity whereas MT demonstrated only anti-diabetic activity.