DISCOVERY OF NOVEL Se-ASPIRIN MOLECULES AS POTENTIAL THERAPEUTICS FOR GASTROINTESTINAL CANCERS

Open Access
Author:
Karelia, Deepkamal Nilkamal
Graduate Program:
Molecular Toxicology
Degree:
Doctor of Philosophy
Document Type:
Dissertation
Date of Defense:
June 14, 2016
Committee Members:
  • Arun Kumar Sharma, Dissertation Advisor
  • Arun Kumar Sharma, Committee Chair
  • Shantu G Amin, Committee Member
  • Jong Kak Yun, Committee Member
  • Victor J Ruiz-Velasco, Outside Member
  • Rosalyn Bryson Irby, Committee Member
Keywords:
  • Pancreatic cancer
  • colorectal cancer
  • small molecule
  • NSAIDs
  • apoptotsis
  • inflammatory pathways
  • selenium
  • aspirin
  • cell cycle
  • MTT
  • EMSA
  • Panc-1
Abstract:
In today’s era there have been a lot of advances in cancer screening, surgery, and chemotherapy, however, gastrointestinal (GI) cancer remains the second leading cause of cancer related deaths. Among all the GI cancers, colorectal cancer (CRC) (New cases: 134,490 deaths: 49,190) and pancreatic cancer (PC) (New cases: 53,070 deaths: 41,780) have high mortality rates. Current therapies result in minimal survival advantage and are linked to multiple adverse events and drug resistance. Further, inflammation has been linked towards initiation and metastasis of both PC and CRC. Literature reports show that the inflammatory pathways like NF-ĸB and STAT3 are constitutively active and have shown to play an important role in resistance towards chemotherapeutic agents. Therefore, there is an unmet need for novel chemotherapeutic agents, that target such inflammatory pathways, with enhanced potency and reduced toxicity to enhance survival and improve patient’s quality of life. Non-steroidal anti-inflammatory drugs (NSIADs), specifically aspirin (ASA), have been highlighted for their chemopreventive properties and anti-inflammatory activities. Recent studies suggest that long term use of ASA reduces the risk of both CRC and PC. However, long term use, as is required in a prevention setting, and the high dose, lead to GI toxicity; e.g. GI bleeding and ulcer formation. Therefore, many efforts are being made to generate novel ASA analogs with reduced GI toxicity, so as not only to achieve a preventive effect at lower doses but increased efficacy towards killing cancer cells to extend their use as therapeutics. Further, selenium containing molecules have gained a lot of attention for their chemopreventive potential and anticancer activities. Selenium containing molecules have also demonstrated to have gastro-protective effects like accelerating ulcer healing as well as reducing NSAIDs induced ulcer formation. Hence, we hypothesize that combining both ASA and selenium will lead to discovery of novel agents with potent anti-cancer activities while having minimal associated GI toxicity. Through extensive structure-activity relationship studies of novel compounds designed by incorporating selenium into the NSAIDs, we recently identified two novel ASA-selenium hybrid molecules (AS-10 and Comp 8), that showed potent anti-cancer activities against both CRC and PC. Comp 8, was designed by incorporating ethyl selenocyanate moiety into carboxylic acid moiety of ASA. AS-10 on the other hand, was generated via novel synthesis pattern to contain a cyclic selenazolidine ring flanked by two acetyl salicylate moieties similar to ASA. Our mechanistic studies show that both agents exhibit similar mechanism of action in PC and CRC. They both inhibited the activation of pro-inflammatory and pro-survival NF-ĸB pathway, when PC cells were stimulated with inflammatory stimuli (TNF-α), at a similar potency. They also reduced the expression of NF-ĸB down-stream anti-apoptotic targets like Bcl-xL, Mcl-1 and survivin. Further, they induced intrinsic form of apoptosis via caspase 9 and caspase 3 activation. Notably, AS-10 was more effective at inhibiting PC growth, and hence we explored its mechanism of action further. AS-10 increased the expression of p21 and p27 at early time points (compared to Comp 8) which arrested cell cycle at G1/G0 and G2/M phase. Cell cycle arrest was later translated to apoptosis. AS-10 also increased reactive oxygen species (ROS) levels, and quenching of ROS via N-acetylcysteine, led to reduction of AS-10’s anti-cancer activity. Moreover, similar mechanism of action was seen in CRC, where both agents were able to induce cell cycle arrest and activation of apoptosis. Our findings show that both agents are more potent at inhibiting cancer cell growth compared to current first line therapies for both PC and CRC. Notably, AS-10 also potentiated the effectiveness of gemcitabine, first line therapy for PC, in gemcitabine resistant PC cell line. Additionally, both agents are more selective towards cancer cells than normal cells, making them clinically more significant. Moreover, we also tested the broad spectrum activity of AS-10 and Comp 8 in different cancer types. Effectiveness of AS-10 was further tested in an even broader range of cancers via National Cancer institute’s (NCI’s) drug screening program in 60 human cancer cell lines which contains cell lines from nine different types of cancer; AS-10 was effective in different cancer types, out of which renal cancer cells were most sensitive. Taken together, our in vitro results suggest both Comp 8 and AS-10 as promising novel chemotherapeutic agents for treatment of CRC and PC. However, based on our preliminary MTD studies in nude mice, AS-10 was found ~5 times more tolerable, and hence could prove to be a potent anti-cancer agent with high therapeutic index.