Deoxynyboquinones as NQO1-targeted anticancer compounds and deoxynybomycins as potent and selective antibiotics

NAD(P)H:quinone oxidoreductase-1 (NQO1);reduction-oxidation cycling;deoxynyboquinone;anticancer;fluoroquinolones;antibiotic resistance;deoxynybomycin;DNA gyrase

Parkinson, Elizabeth Ivy

关键词: NAD(P)H:quinone oxidoreductase-1 (NQO1);    reduction-oxidation cycling;    deoxynyboquinone;    anticancer;    fluoroquinolones;    antibiotic resistance;    deoxynybomycin;    DNA gyrase;   

Others  :  https://www.ideals.illinois.edu/bitstream/handle/2142/89176/PARKINSON-DISSERTATION-2015.pdf?sequence=1&isAllowed=y

美国|英语

来源: The Illinois Digital Environment for Access to Learning and Scholarship

【 摘 要 】

Cancer and antibiotic-resistant bacterial infections are currently two of the major health concerns facing the United States. Novel therapeutics capable of specifically targeting either cancer or resistant bacteria are greatly needed. Described herein are three separate efforts to address these needs.Described in Chapter 2 is the development of a targeted anticancer agent deoxynyboquinone (DNQ) which is specifically activated by the enzyme NAD(P)H:quinone oxidoreductase-1 (NQO1). NQO1 is a 2-electron reductase that is known to be overexpressed in many solid tumors. Development of an anticancer quinone that is bioactivated by NQO1 has long been a goal of cancer therapy. Previously, several putative NQO1 substrates have been developed including mitomycin C, RH1, streptonigrin, and β-Lapachone (β-Lap). Recently the Hergenrother laboratory discovered the small molecule DNQ which has potent anticancer activity. Due to its quinone moiety and the fact that it causes reactive oxygen species (ROS) dependent cell death, we hypothesized that its activity was due to activation by NQO1. Described herein is a set of assays that was developed to determine the NQO1-dependence of anticancer compounds. Of the putative NQO1 substrates, only β-Lap and DNQ were found to be selectively activated by NQO1. Due to its excellent potency and pharmacokinetic profile, DNQ was explored further. Mechanistic evaluation of DNQ revealed that after reduction by NQO1, DNQ undergoes reduction-oxidation cycling which concurrently results in the formation of ROS. ROS causes extensive DNA damage that then activates poly(ADP-ribose) polymerase-1 dependent cell death. DNQ was found to be efficacious a murine model of lung cancer. Utilizing a modified version of the DNQ synthesis previously developed by the Hergenrother laboratory, derivatives were synthesized and evaluated. Several were found that have potent activity against a panel of breast and lung cancers along with improved solubility and toxicity profiles compared to DNQ. These derivatives are currently under investigation for in vivo activity.Described in Chapter 3 is the development of deoxynybomycin (DNM) as an antibiotic with potent activity against fluoroquinolone-resistant (FQR) bacteria. DNM is a natural product that has been shown previously to have antibiotic activity. Recently DNM was found to have potent activity against FQR Methicillin-resistant S. aureus (MRSA).This activity is due to the ability of DNM to inhibit the mutant DNA gyrase (specifically S84L gyrA) responsible for FQR. At the start of the studies described here, two main challenges to the further development of DNM existed: 1) Difficulty in attaining significant quantities of pure DNM for biological evaluation and 2) The poor solubility of DNM. The first issue was addressed by the development of a synthesis of DNM. A single reaction from a late stage intermediate of the DNQ synthesis allowed for the generation of DNM. The modular nature of the synthesis also allowed for the synthesis of a variety of derivatives some of which showed similar potency against FQR MRSA and greatly improved solubility. DNM and its derivative DNM-2 were tested against a variety of bacterial species to determine the activity profile for this class of compounds. The best activity was observed for FQR MRSA with S84L mutant of DNA gyrase and FQR VRE with S84I mutation. Less potent activity was observed for bacteria that commonly have other mutations such as S84F or S84Y. In vitro inhibition assays suggest that DNM is less potent against DNA gyrase with these mutations, but further studies need to be performed to confirm this. Additionally, DNM is inactive against Gram-negative bacteria likely due to its inability to traverse the outer membrane. Further studies to identify compounds active against Gram-negative bacteria are ongoing. Resistance to DNM was found to occur via regeneration of the WT DNA gyrase, thus re-sensitizing bacteria to FQs. This resistance cycling suggests that bacteria which develop resistance to DNM would be treatable.After determining that DNM and DNM-2 have good potency against FQR MRSA, studies evaluating their in vivo activity were performed. Initial pharmacokinetic analysis revealed that oral administration of DNM is not a useful administration route likely due to its poor solubility. However, DNM-2 has excellent oral absorption with area under the curve values which predict good in vivo efficacy. DNM-2 was used in further studies. Toxicity studies revealed no significant effects of DNM-2 on mice when treated at 50 mg/kg for ten consecutive days. Excitingly, DNM-2 was the first compound in the deoxynybomycin class to show in vivo activity, saving mice with FQR MRSA sepsis.Described in Chapter 4 is the analysis of the anticancer compound ersindole, an actiniophyllic acid analogue synthesized by the Martin laboratory. The anticancer activity of ersindole was discovered by the Hergenrother laboratory via a high throughput screen for compounds which induce breast cancer cell death. One of the most striking features of ersindole-induced cancer cell death is the shape of the dose response curve. Specifically, it has a very steep Hill slope and high Emax. These attributes reflect consistent and efficient induction of cancer cell death and suggest that ersindole is a very promising anticancer drug. Analysis of multiple cell lines and timepoints reveal that the steep Hill slope and high Emax of the ersindole dose response curve are general attributes of the compound. Previous mechanistic studies with ersindole suggested that it induced cancer cell death via induction of endoplasmic reticulum stress. This was further confirmed here via Western blot analysis and siRNA knockdown studies. Future efforts should focus on determining the molecular target of ersindole.Unfortunately, ersindole was found to induce hemolysis of red blood cells. A set of derivatives was investigated in an effort to find compounds that do not lyse red blood cells. Ersindole-9 was found to be nearly as potent as ersindole against a panel of cancer cell lines and to have a similarly shaped dose-response curve. Gratifyingly, ersindole-9 does not induce significant hemolysis. For this reason, ersindole-9 was studied in a murine model of breast cancer where it was found to have good efficacy.Evaluation of a second set of derivatives was then performed in order to find additional derivatives that are potent and do not induce hemolysis. Several leads were discovered. Further analysis of these compounds is needed to determine the best compound for future evaluation.