Racial Health Disparity and COVID-19.

The infection by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and resultant coronavirus diseases-19 (COVID-19) disproportionally affects minorities, especially African Americans (AA) compared to the Caucasian population. The AA population is disproportionally affected by COVID-19, in part, because they have high prevalence of underlying conditions such as obesity, diabetes, and hypertension, which are known to exacerbate not only kidney diseases, but also COVID-19. Further, a decreased adherence to COVID-19 guidelines among tobacco smokers could result in increased infection, inflammation, reduced immune response, and lungs damage, leading to more severe form of COVID-19. As a result of high prevalence of underlying conditions that cause kidney diseases in the AA population coupled with tobacco smoking make the AA population vulnerable to severe form of both COVID-19 and kidney diseases. In this review, we describe how tobacco smoking interact with SARS-CoV-2 and exacerbates SARS-CoV-2-induced kidney diseases including renal failure, especially in the AA population. We also explore the role of extracellular vesicles (EVs) in COVID-19 patients who smoke tobacco. EVs, which play important role in tobacco-mediated pathogenesis in infectious diseases, have also shown to be important in COVID-19 pathogenesis and organ injuries including kidney. Further, we explore the potential role of EVs in biomarker discovery and therapeutics, which may help to develop early diagnosis and treatment of tobacco-induced renal injury in COVID-19 patients, respectively.

PLGA Nanoparticle-Based Formulations to Cross the Blood-Brain Barrier for Drug Delivery: From R&D to cGMP.

The blood-brain barrier (BBB) is a natural obstacle for drug delivery into the human brain, hindering treatment of central nervous system (CNS) disorders such as acute ischemic stroke, brain tumors, and human immunodeficiency virus (HIV)-1-associated neurocognitive disorders. Poly(lactic-co-glycolic acid) (PLGA) is a biocompatible polymer that is used in Food and Drug Administration (FDA)-approved pharmaceutical products and medical devices. PLGA nanoparticles (NPs) have been reported to improve drug penetration across the BBB both in vitro and in vivo. Poly(ethylene glycol) (PEG), poly(vinyl alcohol) (PVA), and poloxamer (Pluronic) are widely used as excipients to further improve the stability and effectiveness of PLGA formulations. Peptides and other linkers can be attached on the surface of PLGA to provide targeting delivery. With the newly published guidance from the FDA and the progress of current Good Manufacturing Practice (cGMP) technologies, manufacturing PLGA NP-based drug products can be achieved with higher efficiency, larger quantity, and better quality. The translation from bench to bed is feasible with proper research, concurrent development, quality control, and regulatory assurance.

Drug Resistance in Metastatic Breast Cancer: Tumor Targeted Nanomedicine to the Rescue.

Breast cancer, specifically metastatic breast, is a leading cause of morbidity and mortality in women. This is mainly due to relapse and reoccurrence of tumor. The primary reason for cancer relapse is the development of multidrug resistance (MDR) hampering the treatment and prognosis. MDR can occur due to a multitude of molecular events, including increased expression of efflux transporters such as P-gp, BCRP, or MRP1; epithelial to mesenchymal transition; and resistance development in breast cancer stem cells. Excessive dose dumping in chemotherapy can cause intrinsic anti-cancer MDR to appear prior to chemotherapy and after the treatment. Hence, novel targeted nanomedicines encapsulating chemotherapeutics and gene therapy products may assist to overcome cancer drug resistance. Targeted nanomedicines offer innovative strategies to overcome the limitations of conventional chemotherapy while permitting enhanced selectivity to cancer cells. Targeted nanotheranostics permit targeted drug release, precise breast cancer diagnosis, and importantly, the ability to overcome MDR. The article discusses various nanomedicines designed to selectively target breast cancer, triple negative breast cancer, and breast cancer stem cells. In addition, the review discusses recent approaches, including combination nanoparticles (NPs), theranostic NPs, and stimuli sensitive or "smart" NPs. Recent innovations in microRNA NPs and personalized medicine NPs are also discussed. Future perspective research for complex targeted and multi-stage responsive nanomedicines for metastatic breast cancer is discussed.

Challenges in Biomaterial-Based Drug Delivery Approach for the Treatment of Neurodegenerative Diseases: Opportunities for Extracellular Vesicles.

Biomaterials have been the subject of numerous studies to pursue potential therapeutic interventions for a wide variety of disorders and diseases. The physical and chemical properties of various materials have been explored to develop natural, synthetic, or semi-synthetic materials with distinct advantages for use as drug delivery systems for the central nervous system (CNS) and non-CNS diseases. In this review, an overview of popular biomaterials as drug delivery systems for neurogenerative diseases is provided, balancing the potential and challenges associated with the CNS drug delivery. As an effective drug delivery system, desired properties of biomaterials are discussed, addressing the persistent challenges such as targeted drug delivery, stimuli responsiveness, and controlled drug release in vivo. Finally, we discuss the prospects and limitations of incorporating extracellular vesicles (EVs) as a drug delivery system and their use for biocompatible, stable, and targeted delivery with limited immunogenicity, as well as their ability to be delivered via a non-invasive approach for the treatment of neurodegenerative diseases.

Methamphetamine augment HIV-1 Tat mediated memory deficits by altering the expression of synaptic proteins and neurotrophic factors.

Methamphetamine (METH) abuse is common among individuals infected with HIV-1 and has been shown to affect HIV replication and pathogenesis. These HIV-1 infected individuals also exhibit greater neuronal injury and higher cognitive decline. HIV-1 proteins, specifically gp120 and HIV-1 Tat, have been earlier shown to affect neurocognition. HIV-1 Tat, a viral protein released early during HIV-1 replication, contributes to HIV-associated neurotoxicity through various mechanisms including production of pro-inflammatory cytokines, reactive oxygen species and dysregulation of neuroplasticity. However, the combined effect of METH and HIV-1 Tat on neurocognition and its potential effect on neuroplasticity mechanisms remains largely unknown. Therefore, the present study was undertaken to investigate the combined effect of METH and HIV-1 Tat on behavior and on the expression of neuroplasticity markers by utilizing Doxycycline (DOX)-inducible HIV-1 Tat (1-86) transgenic mice. Expression of Tat in various brain regions of these mice was confirmed by RT-PCR. The mice were administered with an escalating dose of METH (0.1 mg/kg to 6 mg/kg, i.p) over a 7-day period, followed by 6 mg/kg, i.p METH twice a day for four weeks. After three weeks of METH administration, Y maze and Morris water maze assays were performed to determine the effect of Tat and METH on working and spatial memory, respectively. Compared with controls, working memory was significantly decreased in Tat mice that were administered METH. Moreover, significant deficits in spatial memory were also observed in Tat-Tg mice that were administered METH. A significant reduction in the protein expressions of synapsin 1, synaptophysin, Arg3.1, PSD-95, and BDNF in different brain regions were also observed. Expression levels of Calmodulin kinase II (CaMKII), a marker of synaptodendritic integrity, were also significantly decreased in HIV-1 Tat mice that were treated with METH. Together, this data suggests that METH enhances HIV-1 Tat-induced memory deficits by reducing the expression of pre- and postsynaptic proteins and neuroplasticity markers, thus providing novel insights into the molecular mechanisms behind neurocognitive impairments in HIV-infected amphetamine users.

An Overview of Human Immunodeficiency Virus Type 1-Associated Common Neurological Complications: Does Aging Pose a Challenge?

With increasing survival of patients infected with human immunodeficiency virus type 1 (HIV-1), the manifestation of heterogeneous neurological complications is also increasing alarmingly in these patients. Currently, more than 30% of about 40 million HIV-1 infected people worldwide develop central nervous system (CNS)-associated dysfunction, including dementia, sensory, and motor neuropathy. Furthermore, the highly effective antiretroviral therapy has been shown to increase the prevalence of mild cognitive functions while reducing other HIV-1-associated neurological complications. On the contrary, the presence of neurological disorder frequently affects the outcome of conventional HIV-1 therapy. Although, both the children and adults suffer from the post-HIV treatment-associated cognitive impairment, adults, especially depending on the age of disease onset, are more prone to CNS dysfunction. Thus, addressing neurological complications in an HIV-1-infected patient is a delicate balance of several factors and requires characterization of the molecular signature of associated CNS disorders involving intricate cross-talk with HIV-1-derived neurotoxins and other cellular factors. In this review, we summarize some of the current data supporting both the direct and indirect mechanisms, including neuro-inflammation and genome instability in association with aging, leading to CNS dysfunction after HIV-1 infection, and discuss the potential strategies addressing the treatment or prevention of HIV-1-mediated neurotoxicity.

4-(E)-{(p-tolylimino)-methylbenzene-1,2-diol} (TIMBD) suppresses HIV1-gp120 mediated production of IL6 and IL8 but not CCL5.

Human immunodeficiency virus (HIV) has been associated with inflammatory effects that may potentially result in neurodegenerative changes and a number of newer chemotherapeutic agents are being tested to ameliorate these effects. In this study, we investigated the anti-neuroinflammatory activity of a novel resveratrol analog 4-(E)-{(p-tolylimino)-methylbenzene-1,2-diol} (TIMBD) against HIV1-gp120 induced neuroinflammation in SVG astrocytes. SVG astrocytic cells were pretreated with TIMBD or resveratrol (RES) and then transfected with a plasmid encoding HIV1-gp120. The mRNA and protein expression levels of proinflammatory cytokines IL6, IL8 and CCL5 were determined. Protein expression levels of NF-κB, AP1, p-STAT3, p-AKT, p-IKKs and p-p38 MAPK were also determined. TIMBD inhibited gp120-induced RNA and protein expression levels of IL6 and IL8, but not that of CCL5 in SVG astrocytes. Moreover, TIMBD attenuated gp120-induced phosphorylation of cJUN, cFOS, STAT3, p38-MAPK, AKT and IKKs, and the nuclear translocation of NF-κB p-65 subunit whereas RES mostly affected NF-κB protein expression levels. Our results suggest that TIMBD exerts anti-inflammatory effects better than that of RES in SVG astrocytes in vitro. These effects seem to be regulated by AP1, STAT-3 and NF-κB signaling pathways. TIMBD may thus have a potential of being a novel agent for treating HIV1-gp120-mediated neuroinflammatory diseases.

Effect of Methamphetamine on Spectral Binding, Ligand Docking and Metabolism of Anti-HIV Drugs with CYP3A4.

Cytochrome P450 3A4 (CYP3A4) is the major drug metabolic enzyme, and is involved in the metabolism of antiretroviral drugs, especially protease inhibitors (PIs). This study was undertaken to examine the effect of methamphetamine on the binding and metabolism of PIs with CYP3A4. We showed that methamphetamine exhibits a type I spectral change upon binding to CYP3A4 with δAmax and KD of 0.016±0.001 and 204±18 µM, respectively. Methamphetamine-CYP3A4 docking showed that methamphetamine binds to the heme of CYP3A4 in two modes, both leading to N-demethylation. We then studied the effect of methamphetamine binding on PIs with CYP3A4. Our results showed that methamphetamine alters spectral binding of nelfinavir but not the other type I PIs (lopinavir, atazanavir, tipranavir). The change in spectral binding for nelfinavir was observed at both δAmax (0.004±0.0003 vs. 0.0068±0.0001) and KD (1.42±0.36 vs.2.93±0.08 µM) levels. We further tested effect of methamphetamine on binding of 2 type II PIs; ritonavir and indinavir. Our results showed that methamphetamine alters the ritonavir binding to CYP3A4 by decreasing both the δAmax (0.0038±0.0003 vs. 0.0055±0.0003) and KD (0.043±0.0001 vs. 0.065±0.001 nM), while indinavir showed only reduced KD in presence of methamphetamine (0.086±0.01 vs. 0.174±0.03 nM). Furthermore, LC-MS/MS studies in high CYP3A4 human liver microsomes showed a decrease in the formation of hydroxy ritonavir in the presence of methamphetamine. Finally, CYP3A4 docking with lopinavir and ritonavir in the absence and presence of methamphetamine showed that methamphetamine alters the docking of ritonavir, which is consistent with the results obtained from spectral binding and metabolism studies. Overall, our results demonstrated differential effects of methamphetamine on the binding and metabolism of PIs with CYP3A4. These findings have clinical implication in terms of drug dose adjustment of antiretroviral medication, especially with ritonavir-boosted antiretroviral therapy, in HIV-1-infected individuals who abuse methamphetamine.

Effect of mild-to-moderate smoking on viral load, cytokines, oxidative stress, and cytochrome P450 enzymes in HIV-infected individuals.

Mild-to-moderate tobacco smoking is highly prevalent in HIV-infected individuals, and is known to exacerbate HIV pathogenesis. The objective of this study was to determine the specific effects of mild-to-moderate smoking on viral load, cytokine production, and oxidative stress and cytochrome P450 (CYP) pathways in HIV-infected individuals who have not yet received antiretroviral therapy (ART). Thirty-two human subjects were recruited and assigned to four different cohorts as follows: a) HIV negative non-smokers, b) HIV positive non-smokers, c) HIV negative mild-to-moderate smokers, and d) HIV positive mild-to-moderate smokers. Patients were recruited in Cameroon, Africa using strict selection criteria to exclude patients not yet eligible for ART and not receiving conventional or traditional medications. Those with active tuberculosis, hepatitis B or with a history of substance abuse were also excluded. Our results showed an increase in the viral load in the plasma of HIV positive patients who were mild-to-moderate smokers compared to individuals who did not smoke. Furthermore, although we did not observe significant changes in the levels of most pro-inflammatory cytokines, the cytokine IL-8 and MCP-1 showed a significant decrease in the plasma of HIV-infected patients and smokers compared with HIV negative non-smokers. Importantly, HIV-infected individuals and smokers showed a significant increase in oxidative stress compared with HIV negative non-smoker subjects in both plasma and monocytes. To examine the possible pathways involved in increased oxidative stress and viral load, we determined the mRNA levels of several antioxidant and cytochrome P450 enzymes in monocytes. The results showed that the levels of most antioxidants are unaltered, suggesting their inability to counter oxidative stress. While CYP2A6 was induced in smokers, CYP3A4 was induced in HIV and HIV positive smokers compared with HIV negative non-smokers. Overall, the findings suggest a possible association of oxidative stress and perhaps CYP pathway with smoking-mediated increased viral load in HIV positive individuals.

Molecular mechanisms involved in HIV-1 Tat-mediated induction of IL-6 and IL-8 in astrocytes.

BACKGROUND: HIV-associated neurocognitive disorders (HAND) exist in approximately 50% of infected individuals even after the introduction of highly active antiretroviral therapy. HIV-1 Tat has been implicated in HIV-associated neurotoxicity mediated through production of pro-inflammatory cytokines like IL-6 and IL-8 by astrocytes among others as well as oxidative stress. However, the underlying mechanism(s) in the up-regulation of IL-6 and IL-8 are not clearly understood. The present study was designed to determine the mechanism(s) responsible for IL-6 and IL-8 up-regulation by HIV-1 Tat. METHODS: SVG astrocytes were transiently transfected with a plasmid encoding HIV-1 Tat. The HIV-1 Tat-mediated mRNA and protein expression levels of both IL-6 and IL-8 in SVG astrocytes were quantified using real time RT-PCR and multiplex cytokine assay respectively. We also employed immunocytochemistry for staining of IL-6 and IL-8. The underlying signaling mechanism(s) were identified using pharmacological inhibitors and siRNA for different intermediate steps involved in PI3K/Akt, p38 MAPK and JNK MAPK pathways. Appropriate controls were used in the experiments and the effect of pharmacological antagonists and siRNA were observed on both mRNA expression and protein levels. RESULTS: Both IL-6/IL-8 mRNA and protein showed peak expressions at 6 hours and 96 hours post-transfection, respectively. Elevated levels of IL-6/IL-8 were also confirmed by immunocytochemistry. Our studies indicated that both NF-kB and AP-1 transcription factors were involved in IL-6 and IL-8 expression mediated by HIV-1 Tat; however, AP-1 was differentially activated for either cytokine. In the case of IL-6, p38δ activated AP-1 whereas JNK but not p38 MAPK was involved in AP-1 activation for IL-8 production. On the other hand both PI3K/Akt and p38 MAPK (ß subunit) were found to be involved in activation of NF-κB that led to IL-6 and IL-8 production. CONCLUSION: Our results demonstrate HIV-1 Tat-mediated induction of both IL-6 and IL-8 in a time-dependent manner in SVG astrocytes. Furthermore, we also showed the involvement of NF-κB and AP-1 transcription factors regulated by PI3/Akt, p38 MAPK and JNK MAPK upstream signaling molecules. These results present new therapeutic targets that could be used in management of HAND.

Enhanced methamphetamine metabolism in rhesus macaque as compared with human: an analysis using a novel method of liquid chromatography with tandem mass spectrometry, kinetic study, and substrate docking.

Methamphetamine (MA), which remains one of the widely used drugs of abuse, is metabolized by the cytochrome P450 (P450) family of enzymes in humans. However, metabolism of methamphetamine in macaques is poorly understood. Therefore, we first developed and validated a very sensitive liquid chromatography with tandem mass spectrometry (LC-MS/MS) method using solid phase extraction of rhesus plasma with a lower limit of quantitation at 1.09 ng/ml for MA and its metabolites, 4-hydroxy methamphetamine (4-OH MA), amphetamine (AM), 4-OH amphetamine (4-OH AM), and norephedrine. We then analyzed plasma samples of MA-treated rhesus, which showed >10-fold higher concentrations of AM (∼29 ng/ml) and 4-OH AM (∼28 ng/ml) than MA (∼2 ng/ml). Because the plasma levels of MA metabolites in rhesus were much higher than in human samples, we examined MA metabolism in human and rhesus microsomes. Interestingly, the results showed that AM and 4-OH AM were formed more rapidly and that the catalytic efficiency (Vmax/Km) for the formation of AM was ∼8-fold higher in rhesus than in human microsomes. We further examined the differences in these kinetic characteristics using three selective inhibitors of each human CYP2D6 and CYP3A4 enzymes. The results showed that each of these inhibitors inhibited both d- and l-MA metabolism by 20%-60% in human microsomes but not in rhesus microsomes. The differences between human and rhesus CYP2D6 and CYP3A4 enzymes were further assessed by docking studies for both d and l-MA. In conclusion, our results demonstrated an enhanced MA metabolism in rhesus compared with humans, which is likely to be caused by differences in MA-metabolizing P450 enzymes between these species.

HIV-1 Tat-mediated induction of CCL5 in astrocytes involves NF-κB, AP-1, C/EBPα and C/EBPγ transcription factors and JAK, PI3K/Akt and p38 MAPK signaling pathways.

The incidence of HIV-associated neurological disorders (HAND) has increased during recent years even though the highly active antiretroviral therapy (HAART) has significantly curtailed the virus replication and increased the life expectancy among HIV-1 infected individuals. These neurological deficits have been attributed to HIV proteins including HIV-1 Tat. HIV-1 Tat is known to up-regulate CCL5 expression in mouse astrocytes, but the mechanism of up-regulation is not known. The present study was undertaken with the objective of determining the mechanism(s) underlying HIV-1 Tat-mediated expression of CCL5 in astrocytes. SVGA astrocytes were transiently transfected with a plasmid encoding Tat, and expression of CCL5 was studied at the mRNA and protein levels using real time RT-PCR and multiplex cytokine bead array, respectively. HIV-1 Tat showed a time-dependent increase in the CCL5 expression with peak mRNA and protein levels, observed at 1 h and 48 h post-transfection, respectively. In order to explore the mechanism(s), pharmacological inhibitors and siRNA against different pathway(s) were used. Pre-treatment with SC514 (NF-κB inhibitor), LY294002 (PI3K inhibitor), AG490 (JAK2 inhibitor) and Janex-1 (JAK3 inhibitor) showed partial reduction of the Tat-mediated induction of CCL5 suggesting involvement of JAK, PI3K/Akt and NF-κB in CCL5 expression. These results were further confirmed by knockdown of the respective genes using siRNA. Furthermore, p38 MAPK was found to be involved since the knockdown of p38δ but not other isoforms showed partial reduction in CCL5 induction. This was further confirmed at transcriptional level that AP-1, C/EBPα and C/EBPγ were involved in CCL5 up-regulation.