Katrina M Miranda

Katrina M Miranda

Associate Professor, Chemistry and Biochemistry-Sci
Associate Professor, BIO5 Institute
Primary Department
Department Affiliations
Contact
(520) 626-3655

Work Summary

We seek to produce new drugs that harness molecules produced during the natural immune response in order to treat cancer and pain. Such compounds may also provide new treatments for heart failure and alcoholism.

Research Interest

Katrina Miranda, PhD, claims nitric oxide (NO), which is synthesized in the body via enzymatic oxidation of L-arginine, is critical to numerous physiological functions, but also can contribute to the severity of diseases such as cancer or pathophysiological conditions such as stroke. This diversity in the responses to NO biosynthesis is a reflection of the diverse chemistry of NO. For instance, NO can alter the function of enzymes by binding to metal centers. This type of interaction could result in outcomes as disparate as control of blood pressure or death of an invading bacterium. NO can also be readily converted to higher nitrogen oxides such as N2O3 or ONOOH, which have very different chemical and biological properties. The ultimate result will depend upon numerous factors, particularly the location and concentration of NO produced. Therefore, site-specific modulation of NO concentration offers intriguing therapeutic possibilities for an ever expanding list of diseases, including cancer, heart failure and stroke. As a whole, Dr. Miranda is interested in elucidating the fundamental molecular redox chemistry of NO and in developing compounds to deliver or scavenge NO and other nitrogen oxides. These projects are designed to answer questions of potential medical importance through a multi-disciplinary approach, including analytical, synthetic, inorganic and biochemical techniques.The project categories include five major disciplines. First, she will work on the development and utilization of analytical techniques for detection and measurement of NO and other nitrogen oxides as well as the resultant chemistry of these species. Second, she will synthesize potential donors or scavengers of NO and other nitrogen oxides. Third, it’s necessary to describe chemical characterization of these compounds (spectroscopic features, kinetics, mechanisms and profiles of nitrogen oxide release, etc.). Fourth, Dr. Miranda will try to describe the biological characterization of these compounds (assay of effects on biological compounds, mechanisms and pathways, in vitro determination of potential for therapeutic utility, etc.). Fifth, she will identify of potential targets, such as enzymes, for treatment of disease through exposure to nitrogen oxide donors. Keywords: cancer treatment, pain treatment

Publications

Boitano, S., Omsland, A., Miranda, K. M., Friedman, R. L., & Boitano, S. A. (2008). Bordetella bronchiseptica responses to physiological reactive nitrogen and oxygen stresses. FEMS microbiology letters, 284(1).
BIO5 Collaborators
Scott A Boitano, Katrina M Miranda

Bordetella bronchiseptica can establish prolonged airway infection consistent with a highly developed ability to evade mammalian host immune responses. Upon initial interaction with the host upper respiratory tract mucosa, B. bronchiseptica are subjected to antimicrobial reactive nitrogen species (RNS) and reactive oxygen species (ROS), effector molecules of the innate immune system. However, the responses of B. bronchiseptica to redox species at physiologically relevant concentrations (nM-microM) have not been investigated. Using predicted physiological concentrations of nitric oxide (NO), superoxide and hydrogen peroxide (H2O2) on low numbers of CFU of B. bronchiseptica, all redox active species displayed dose-dependent antimicrobial activity. Susceptibility to individual redox active species was significantly increased upon introduction of a second species at subantimicrobial concentrations. An increased bacteriostatic activity of NO was observed relative to H2O2. The understanding of Bordetella responses to physiologically relevant levels of exogenous RNS and ROS will aid in defining the role of endogenous production of these molecules in host innate immunity against Bordetella and other respiratory pathogens.

Hofseth, L., Saito, S., Hussain, S., Espey, M., Miranda, K., Araki, Y., Jhappan, C., Higashimoto, Y., He, P., Linke, S., Quezado, M., Zurer, ., Rotter, ., Wink, D., Appella, E., & Harris, C. (2003). Nitric oxide-induced cellular stress and p53 activation in chronic inflammation. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 100(1), 143-148.

Free radical-induced cellular stress contributes to cancer during chronic inflammation. Here, we investigated mechanisms of p53 activation by the free radical, NO. NO from donor drugs induced both ataxia-telangiectasia mutated (ATM)- and ataxia-telangiectasia mutated and Rad3-related-dependent p53 posttranslational modifications, leading to an increase in p53 transcriptional targets and a G(2)/M cell cycle checkpoint. Such modifications were also identified in cells cocultured with NO-releasing macrophages. In noncancerous colon tissues from patients with ulcerative colitis (a cancer-prone chronic inflammatory disease), inducible NO synthase protein levels were positively correlated with p53 serine 15 phosphorylation levels. Immunostaining of HDM-2 and p21(WAF1) was consistent with transcriptionally active p53. Our study highlights a pivotal role of NO in the induction of cellular stress and the activation of a p53 response pathway during chronic inflammation.

Väänänen, A. J., Salmenperä, P., Hukkanen, M., Miranda, K. M., Harjula, A., Rauhala, P., & Kankuri, E. (2008). Persistent susceptibility of cathepsin B to irreversible inhibition by nitroxyl (HNO) in the presence of endogenous nitric oxide. Free Radical Biology and Medicine, 45(6), 749-755.

PMID: 18572022;Abstract:

Nitrosation of enzyme regulatory cysteines is one of the key posttranslational modification mechanisms of enzyme function. Frequently such modifications are readily reversible; however, cysteine proteases, such as cathepsin B, have been shown to be covalently and permanently inactivated by nitroxyl (HNO), the one-electron reduction product of NO. Owing to the high reactivity of HNO with NO, endogenous NO production could provide direct protection for the less reactive protein cysteines by scavenging HNO. Additionally, endogenous cellular production of NO could rescue enzyme function by protective nitrosation of cysteines prior to exposure to HNO. Thus, we studied the effect of endogenous NO production, induced by LPS or IFN-γ, on inhibition of cysteine protease cathepsin B in RAW macrophages. Both LPS and IFN-γ induce iNOS with generation of nitrate up to 9 μM in the media after a 24-h stimulation, while native RAW 264.7 macrophages neither express iNOS nor generate nitrate. After the 24-h stimulation, the HNO-releasing Angeli's salt (0-316 μM) caused dose-dependent and DTT-irreversible loss of cathepsin B activity, and induction of iNOS activity did not protect the enzyme. The lack of protection was also verified in an in vitro setup, where papain, a close structural analogue of cathepsin B, was inhibited by Angeli's salt (2.7 μM) in the presence of the NO donor DEA/NO (0-316 μM). This clearly showed that a high molar excess of DEA/NO (EC50 406 μM) is needed to protect papain from the DTT-irreversible covalent modification by HNO. Our results provide first evidence on a cellular level for the remarkably high sensitivity of active-site cysteines in cysteine proteases for modification by HNO. © 2008 Elsevier B.V. All rights reserved.

Zarpelon, A. C., Souza, G. R., Cunha, T. M., Schivo, I. R., Marchesi, M., Casagrande, R., Pinge-Filho, P., Cunha, F. Q., Ferreira, S. H., Miranda, K. M., & Verri Jr., W. A. (2013). The nitroxyl donor, Angeli's salt, inhibits inflammatory hyperalgesia in rats. Neuropharmacology, 71, 1-9.

PMID: 23541720;PMCID: PMC3666861;Abstract:

Nitric oxide modulates pain development. However, there is no evidence on the effect of nitroxyl (HNO/NO-) in nociception. Therefore, we addressed whether nitroxyl inhibits inflammatory hyperalgesia and its mechanism using the nitroxyl donor Angeli's salt (AS; Na2N2O 3). Mechanical hyperalgesia was evaluated using a modified Randall and Selitto method in rats, cytokine production by ELISA and nitroxyl was determined by confocal microscopy in DAF (a cell permeable reagent that is converted into a fluorescent molecule by nitrogen oxides)-treated dorsal root ganglia neurons in culture. Local pre-treatment with AS (17-450 μg/paw, 30 min) inhibited the carrageenin-induced mechanical hyperalgesia in a dose- and time-dependent manner with maximum inhibition of 97%. AS also inhibited carrageenin-induced cytokine production. AS inhibited the hyperalgesia induced by other inflammatory stimuli including lipopolysaccharide, tumor necrosis factor-α, interleukin-1β and prostaglandin E2. Furthermore, the analgesic effect of AS was prevented by treatment with ODQ (a soluble guanylate cyclase inhibitor), KT5823 (a protein kinase G [PKG] inhibitor) or glybenclamide (an ATP-sensitive K+ channel blocker), but not with naloxone (an opioid receptor antagonist). AS induced concentration-dependent increase in fluorescence intensity of DAF-treated neurons in a l-cysteine (nitroxyl scavenger) sensitive manner. l-cysteine did not affect the NO+ donor S-Nitroso-N-acetyl-DL- penicillamine (SNAP)-induced anti-hyperalgesia or fluorescence of DAF-treated neurons. This is the first study to demonstrate that nitroxyl inhibits inflammatory hyperalgesia by reducing cytokine production and activating the cGMP/PKG/ATP-sensitive K+ channel signaling pathway in vivo. © 2013 Elsevier Ltd. All rights reserved.

Wink, D. A., Miranda, K. M., & Espey, M. G. (2001). Cytotoxicity related to oxidative and nitrosative stress by nitric oxide. Experimental Biology and Medicine, 226(7), 621-623.