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Item Combination Effects of Aminolevulinic Acid and Mycophenolic Acid on Hacat Cell Proliferation and Inhibition of Inosine Monophosphate Dehydrogenase.(Molecules, 2025-03-18) Venkatesh, Manisha*; Capriglione, Noelle*; Rehberg, Kaitlyn*; Voigt, Jeffrey; Hass, Martha ADerivatives of mycophenolic acid (MPA) and 5-aminolevulinic acid photodynamic therapy (ALA-PDT) have been used separately to treat psoriasis, a chronic, inflammatory skin disease that is characterized by the unregulated hyperproliferation of epidermal keratinocytes and a T-cell-mediated immune response. However, the combination of these two therapies has not previously been explored. This study investigated the in vitro effects of combining MPA with ALA-PDT to suppress keratinocytes and the in vitro inhibition of inosine monophosphate dehydrogenase, a key enzyme. The effects of ALA, MPA, and their combination on protoporphyrin IX (PpIX) generation and cell viability in HaCaT cells, as well as the inhibition of IMPDH, were evaluated. Treatment of HaCaT cells with ALA, MPA, and their 1:1 molar combination showed that ALA alone induced PpIX production, with concentrations increasing from 5.25 ng/mL at 10 μM to 157.5 ng/mL at 1 mM. MPA did not increase PpIX on its own but had a modest synergistic effect with ALA at low concentrations (10 μM and 50 μM). The impact of blue light irradiation (465 nm) on cell viability was also assessed, revealing that ALA and ALA + MPA treatment led to significant reductions in HaCaT cell viability at higher concentrations (500 μM-1 mM), while MPA alone with blue light irradiation showed no cytotoxicity. The reduction in skin cell viability was enhanced when ALA was combined with MPA. Additionally, MPA effectively inhibited IMPDH activity in a dose-dependent manner, with 94-96% inhibition at concentrations of 100 μM and above. Interestingly, ALA weakly inhibited IMPDH, with a peak inhibition of 46% at 5 μM. At higher ALA concentrations, its inhibitory effect diminished, and it interfered with the potency of MPA's IMPDH2 inhibition, suggesting that ALA could modulate MPA's therapeutic action. These findings suggest that the combination of MPA with ALA-PDT may be a viable new treatment for psoriasis.Item Stipulations of cell and gene therapy and the ties to biomanufacturing.(Biotechnology Progress, 2025-01-23) Allisha, Justin*; Das, Juthika*; Dunnigan, Thomas*; Sharfstein, Susan T; Datta, PayelCell and gene therapy (CGT) products are emerging and innovative biopharmaceuticals that hold promise for treating diseases that are otherwise beyond the scope of conventional medicines. The evolution of CGT from a research idea to a promising therapeutic product is due to the complementary advancements across various scientific disciplines. First, the innovations and advancements in gene editing and delivery technology have provided fundamental tools to manipulate genes and cells for therapeutic pursuits. Second, advancements in applied and translational research, including how clinical trials are designed, performed, evaluated, and analyzed, have transformed the technology into a potential therapeutic product. Third, advancements in scaling up the production of CGT products have been critical in delivering the product for preclinical studies, clinical trials, and approved treatments. In parallel, regulatory requirements have continuously evolved, with lessons learned from translational studies and biomanufacturing. These combined efforts have transformed CGT products from a promising concept into a reality with the potential to treat a wide range of diseases. However, continued R&D and regulatory oversight are crucial to further improve the safety, efficacy, and accessibility of CGT products.Item Structural and biophysical analysis of cytochrome P450 2C9*14 and *27 variants in complex with losartan.(Journal of Inorganic Biochemistry, 2024-09) Parikh, Sonia J*; Edara, Sreeja*; Deodhar, Shruti*; Singh, Ajit K; Maekawa, Keiko; Zhang, Qinghai; Glass, Karen C; Shah, Manish BThe human cytochrome P450 (CYP) 1, 2 and 3 families of enzymes are responsible for the biotransformation of a majority of the currently available pharmaceutical drugs. The highly polymorphic CYP2C9 predominantly metabolizes many drugs including anticoagulant S-warfarin, anti-hypertensive losartan, anti-diabetic tolbutamide, analgesic ibuprofen, etc. There are >80 single nucleotide changes identified in CYP2C9, many of which significantly alter the clearance of important drugs. Here we report the structural and biophysical analysis of two polymorphic variants, CYP2C9*14 (Arg125His) and CYP2C9*27 (Arg150Leu) complexed with losartan. The X-ray crystal structures of the CYP2C9*14 and *27 illustrate the binding of two losartan molecules, one in the active site near heme and another on the periphery. Both losartan molecules are bound in an identical conformation to that observed in the previously solved CYP2C9 wild-type complex, however, the number of losartan differs from the wild-type structure, which showed binding of three molecules. Additionally, isothermal titration calorimetry experiments reveal a lower binding affinity of losartan with *14 and *27 variants when compared to the wild-type. Overall, the results provide new insights into the effects of these genetic polymorphisms and suggests a possible mechanism contributing to reduced metabolic activity in patients carrying these alleles.Item HIV-Associated Neurocognitive Disorder: A Look into Cellular and Molecular Pathology.(International Journal of Molecular Sciences, 2024-04-25) Thompson, Landon John-Patrick*; Genovese, Jessica*; Hong, Zhenzi*; Singh, Meera Vir; Singh, Vir BahadurDespite combined antiretroviral therapy (cART) limiting HIV replication to undetectable levels in the blood, people living with HIV continue to experience HIV-associated neurocognitive disorder (HAND). HAND is associated with neurocognitive impairment, including motor impairment, and memory loss. HIV has been detected in the brain within 8 days of estimated exposure and the mechanisms for this early entry are being actively studied. Once having entered into the central nervous system (CNS), HIV degrades the blood-brain barrier through the production of its gp120 and Tat proteins. These proteins are directly toxic to endothelial cells and neurons, and propagate inflammatory cytokines by the activation of immune cells and dysregulation of tight junction proteins. The BBB breakdown is associated with the progression of neurocognitive disease. One of the main hurdles for treatment for HAND is the latent pool of cells, which are insensitive to cART and prolong inflammation by harboring the provirus in long-lived cells that can reactivate, causing damage. Multiple strategies are being studied to combat the latent pool and HAND; however, clinically, these approaches have been insufficient and require further revisions. The goal of this paper is to aggregate the known mechanisms and challenges associated with HAND.Item The role of CXCL1 in crosstalk between endocrine resistant breast cancer and fibroblast.(Molecular Biology Reports, 2024-02-23) Pandithar, Sneha*; Galke, Daniel*; Akume, Ahone*; Belyakov, Artem*; Lomonaco, Dominick*; Guerra, Amirah A*; Park, Jay*; Reff, Olivia*; Jin, KideokBackground: ER positive breast cancer is currently targeted using various endocrine therapies. Despite the proven therapeutic efficacy, resistance to the drug and reoccurrence of tumor appears to be a complication that many patients deal with. Molecular pathways underlying the development of resistance are being widely studied. Methods and results: In this study, using four established endocrine resistant breast cancer (ERBC) cell lines, we characterized CXCL1 as a secreted factor in crosstalk between ERBC cells and fibroblasts. Protein array revealed upregulation of CXCL1 and we confirmed the CXCL1 expression by real-time qRT-PCR and U-Plex assay. Co-culturing ERBC cells with fibroblasts enhanced the cell growth and migration compared to monoculture. The crosstalk of ERBC cells with fibroblasts significantly activates ERK/MAPK signaling pathway while reparixin, CXCR1/2 receptor inhibitor, attenuates the activity. Reparixin displayed the ERBC cell growth inhibition and the combination treatment with reparixin and CDK4/6 inhibitor (palbociclib and ribociclib) increased these inhibitory effect. Conclusions: Taken together, our study implicates CXCL1 as a critical role in ERBC growth and metastasis via crosstalk with fibroblast and cotargeting CXCR1/2 and CDK4/6 could potentially overcome endocrine resistant breast cancer.