The present studies sought to determine whether the lethality of the drug combination [sorafenib + sildenafil] could be enhanced by the anti-inflammatory agent celecoxib, using ovarian cancer and other tumor cell lines as models. Also, in a dose dependent fashion celecoxib enhanced [sorafenib + sildenafil] lethality in multiple ovarian cancer cell lines. In a dose dependent fashion celecoxib enhanced the ability of [sorafenib + sildenafil] to reduce expression of multiple chaperone proteins in parallel with lower levels of the drug efflux pumps ABCB1 and ABCG2. Over-expression of GRP78 and HSP27 maintained pump expression in the presence of drugs. Cell killing by the 3 drug combination was mediated by mitochondrial / caspase 9 -dependent apoptotic signaling and by RIP-1 / caspases 2 and 4 / AIF -dependent necroptotic signaling. Pre-treatment of intrinsically resistant primary ovarian cancer cells with [celecoxib + sorafenib + sildenafil] significantly enhanced tumor cell killing by a subsequent cisplatin exposure. Similar data were obtained in some cancer cell lines, but not all, using the related platinum containing drugs, oxaliplatin and carboplatin. As our prior publications have also validated in vivo the combinations of [celecoxib + sildenafil] and [sorafenib + sildenafil] as cytotoxic to multiple tumor cell types, combined with the present findings, we would argue that the combination of celecoxib/sorafenib/sildenafil should be explored in a new phase I trial in ovarian cancer.

abstract: The effects of specific histone deacetylase inhibitors (HDACi) on transgene expression in combination with a novel polymer as a delivery vehicle are investigated in this research. Polymer vectors, although safer than viruses, are notorious for low levels of gene expression. In this investigation, the use of an emerging chemotherapeutic anti-cancer drug molecule, HDACi, was used to enhance the polymer-mediated gene expression. HDACi are capable of inhibiting deacetylation activities of histones and other non-histone proteins in the cytoplasm and nucleus, as well as increase transcriptional activities necessary for gene expression. In a prior study, a parallel synthesis and screening of polymers yielded a lead cationic polymer with high DNA-binding properties, and even more attractive, high transgene expressions. Previous studies showed the use of this polymer in conjunction with cytoplasmic HDACi significantly enhanced gene expression in PC3-PSMA prostate cancer cells. This led to the basis for the investigation presented in this thesis, but to use nuclear HDACi to potentially achieve similar results. The HDACi, HDACi_A, was a previously discovered lead drug that had potential to significantly enhance luciferase expression in PC3-PSMA cells. The results of this study found that the 20:1 polymer:plasmid DNA weight ratio was effective with 1 uM and 2 uM HDACI_A concentrations, showing up to a 9-fold enhancement. This enhancement suggested that HDACi_A was effectively aiding transfection. While not an astounding enhancement, it is still interesting enough to investigate further. Cell viabilities need to be determined to supplement the results. / Dissertation/Thesis / M.S. Bioengineering 2012

Traumatic brain injury (TBI) represents a significant health crisis. To date there are no FDA-approved pharmacotherapies available to prevent the neurologic deficits caused by TBI. Following TBI, dysfunctional mitochondria generate reactive oxygen and nitrogen species, initiating lipid peroxidation (LP) and the formation of LP-derived neurotoxic aldehydes, which bind mitochondrial proteins, exacerbating dysfunction and opening of the mitochondrial permeability pore (mPTP), resulting in extrusion of mitochondrial sequestered calcium into the cytosol, and initiating a downstream cascade of calpain activation, spectrin degradation, neurodegeneration and neurologic impairment. As central mediators of the TBI secondary injury cascade, mitochondria and LP-derived neurotoxic aldehydes make promising therapeutic targets. In fact, Cyclosporine A (CsA), an FDA-approved immunosuppressant capable of inhibiting mPTP has been shown to be neuroprotective in experimental TBI. Additionally, phenelzine (PZ), an FDA-approved non-selective irreversible monoamine oxidase inhibitor (MAOI) class antidepressant has also been shown to be neuroprotective in experimental TBI due to the presence of a hydrazine (-NH-NH2) moiety allowing for the scavenging of LP-derived neurotoxic aldehydes. The overall goal of this dissertation is to further examine the neuroprotective effects of the mPTP inhibitor, CsA, and the LP-derived neurotoxic aldehyde scavenger, PZ, using a severe controlled cortical impact injury (CCI) model in 3-month old male Sprague-Dawley rats. First, the effects of CsA on cortical synaptic and non-synaptic mitochondria, two heterogeneous populations, are examined. Our results indicate that compared to non-synaptic mitochondria, synaptic mitochondria sustain greater damage 24h following CCI and are protected to a greater degree by CsA. Second, the neuroprotective effects of a novel 72h continuous subcutaneous infusion of CsA combined with PZ are compared to monotherapy. Following CCI, our results indicate that individually both CsA and PZ attenuate modification of mitochondrial proteins by LP-derived neurotoxic aldehydes, PZ is able to maintain mitochondrial respiratory control ratio and cytoskeletal integrity, but together, PZ and CsA, are unable to improve and in some cases negate monotherapy neuroprotective effects. Finally, the effects of PZ (MAOI, aldehyde scavenger), pargyline (PG, MAOI, non-aldehyde scavenger) and hydralazine (HZ, non-MAOI, aldehyde scavenger) are compared. Our results indicate that PZ, PG, and HZ are unable to improve CCI-induced deficits to learning and memory as measured by Morris water maze (post-CCI D3-7). Of concern, PZ animals lost a significant amount of weight compared to all other group, possibly due to MAOI effects. In fact, in uninjured cortical tissue, PZ administration leads to a significant increase in norepinephrine and serotonin. Additionally, although PZ, PG, and HZ did not lead to a statistically significant improvement in cortical tissue sparing 8 days following CCI, the HZ group saw a 10% improvement over vehicle. Overall, these results indicate that pharmacotherapies which improve mitochondrial function and decrease lipid peroxidation should continue to be pursued as neuroprotective approaches to TBI. However, further pursuit of LP-derived aldehyde scavengers for clinical use in TBI may require the development of hydrazine (-NH-NH2)-compounds which lack additional confounding mechanisms of action.