Agent For Treatment Of Pancreatic Cancer Research Articles

Abstract 7170: Pharmacokinetic profiling of Re-Phenformin in vivo: A novel approach in pancreatic cancer therapy

Abstract Phenformin, a biguanide medication, has long been utilized in the treatment of diabetes. Recent research indicates that phenformin may also serve as an effective anticancer agent. This study introduces the synthesis and evaluation of Re tricarbonyl phenformin complex (Re-(CO)3-Phenformin, Re-Phen), as a derivative of Phenformin and novel anticancer agent for pancreatic cancer treatment. Re-Phen was synthesized, purified, and characterized through preparative HPLC, NMR, mass spectrometry, and FT-IR. Subsequent in vitro studies, utilizing the MTS assay, assessed its cytotoxicity against pancreatic cancer cell lines, including Kras* murine PDAC and MiaPaca-2 human PDAC. Notably, Re-Phen exhibited significantly higher cytotoxic effects in MiaPaca-2 cells (IC50= 0.755 mM) and Kras* cells (IC50= 0.410 mM) compared to Phenformin treatment (IC50= 4.02 mM and 2.36 mM, respectively). These results highlight Re-Phen's enhanced anticancer activity, outperforming Phenformin in potency. Further investigation into the pharmacokinetics of Re-Phen involved an in vivo study with rats intravenously administered 2 mg/kg of Re-Phenformin. Blood samples were collected at intervals (10 min to 24 h post-dose) and were analyzed by validated LC-MS/MS. The pharmacokinetic profile of Re-Phen was best described by a 2-compartment model, using Phoenix® WinNonlin (version 8.0), with a particular focus on comparing the profiles of Phenformin and Re-Phen. The study revealed that Re-Phen has a significantly extended half-life compared to Phenformin (9.13 hr vs 2.11 hr), indicating prolonged systemic retention. Additionally, Re-Phen demonstrated an increased area under the curve (AUC), suggesting greater systemic exposure at equivalent dosing (90.55 hr*ng/ml vs 7.13 hr*ng/ml). This could imply enhanced bioavailability or altered metabolic clearance of Re-Phen compared to its parent compound. The two-Compartment of Re-Phe, provide a deeper understanding of its pharmacokinetic characteristics. In conclusion, Re-Phen not only demonstrates superior anticancer efficacy in vitro but also exhibits a more favorable pharmacokinetic profile in vivo. The extended half-life and increased AUC of Re-Phen suggest the potential of offering improved therapeutic efficacy in pancreatic cancer treatment. These promising findings warrant further investigation and could position Re-Phen as a promising therapeutic agent for the pharmacological management of pancreatic cancer. Citation Format: Amir Mohammad Gholizadeh, Lu Dai, Fatima Dagher, Chiyi Xiong, Chun Li, Diana S-L Chow. Pharmacokinetic profiling of Re-Phenformin in vivo: A novel approach in pancreatic cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7170.

Repositioning of Montelukast to inhibit proliferation of mutated KRAS pancreatic cancer through a novel mechanism that interfere the binding between KRAS and GTP/GDP

Pancreatic cancer is one of the most lethal cancer types with 5-year survival rate of ∼10.8%. Various KRAS mutations exist in ∼85% pancreatic cancer cell lines. Mutated KRAS is a major cause that leads cancer cell proliferation. Chemotherapy is still the major treatment for pancreatic cancer. Alternatively, repositioning old drug to inhibit mutated KRAS may be a cost-effective way for pancreatic cancer treatment. In this study, we choose mutated KRAS (G12D) as a target. Based on mutated KRAS GTP binding domain (hydrolyze GTP to GDP), we perform virtual screening on FDA-approved drugs. Montelukast shows strong binding affinity to mutated KRAS as well as interfering both GTP and GDP binding to mutated KRAS. Furthermore, Montelukast shows very strong anti-proliferation effect on mutated KRAS pancreatic cancer cells both in vitro and in vivo. Our results support repositioning of Montelukast as single agent for pancreatic cancer treatment.

Gemcitabine‑fucoxanthin combination in human pancreatic cancer cells.

Gemcitabine is a chemotherapeutic agent for pancreatic cancer treatment. It has also been demonstrated to inhibit human pancreatic cancer cell lines, MIA PaCa-2 and PANC-1. The aim of the present study was to investigate the suppressive effect of fucoxanthin, a marine carotenoid, in combination with gemcitabine on pancreatic cancer cells. MTT assays and cell cycle analysis using flow cytometry were performed to study the mechanism of action. The results revealed that combining a low dose of fucoxanthin with gemcitabine enhanced the cell viability of human embryonic kidney cells, 293, while a high dose of fucoxanthin enhanced the inhibitory effect of gemcitabine on the cell viability of this cell line. In addition, the enhanced effect of fucoxanthin on the inhibitory effect of gemcitabine on PANC-1 cells was significant (P<0.01). Fucoxanthin combined with gemcitabine also exerted significant enhancement of the anti-proliferation effect in MIA PaCa-2 cells in a concentration dependent manner (P<0.05), compared with gemcitabine treatment alone. In conclusion, fucoxanthin improved the cytotoxicity of gemcitabine on human pancreatic cancer cells at concentrations that were not cytotoxic to non-cancer cells. Thus, fucoxanthin has the potential to be used as an adjunct in pancreatic cancer treatment.

Corrole Nanoparticles for Chemotherapy of Castration-Resistant Prostate Cancer and as Sonodynamic Agents for Pancreatic Cancer Treatment.

A nanoparticle-based system, composed of the gallium(III) complex of a minimally substituted corrole that is coated by transferrin as a targeting vehicle (3-Ga NPs), has been used for pre-clinical evaluation of its efficacy against human metastatic castration-resistant prostate cancer (mCRPC) tumor xenografts. All mice (N = 9) responded to a dose of 10 mg/kg, with a remarkable tumor growth inhibition of 400% following 2 weeks of treatment; Ames and hERG tests excluded potential concerns regarding mutagenicity and cardiotoxicity, respectively. Also demonstrated is the potential application of these 3-Ga NPs as sonodynamic agents for the preclinical treatment of pancreatic cancer. 10 mg/kg 3-Ga NPs combined with exposure to ultrasound waves (2 min of 1 MHz 0.1 w/cm2 twice a week) induced up to 77% tumor shrinkage. Consistently, tumor/tissue distribution and serum levels of 3-Ga NPs in mice revealed high tumor specificity, favorable pharmacokinetics, fast absorption, slower redistribution, and very slow drug clearance.

Novel compound C150 inhibits pancreatic cancer through induction of ER stress and proteosome assembly.

Pancreatic cancer is a devastating disease with a dismal prognosis and poor treatment outcomes. Searching for new agents for pancreatic cancer treatment is of great significance. We previously identified a novel activity of compound C150 to inhibit pancreatic cancer epithelial-to-mesenchymal transition (EMT). Here, we further revealed its mechanism of action. C150 induced ER stress in pancreatic cancer cells and subsequently increased proteasome activity by enhancing proteasome assembly, which subsequently enhanced the degradation of critical EMT transcription factors (EMT-TFs). In addition, as cellular responses to ER stress, autophagy was elevated, and general protein synthesis was inhibited in pancreatic cancer cells. Besides EMT inhibition, the C150-induced ER stress resulted in G2/M cell cycle arrest, which halted cell proliferation and led to cellular senescence. In an orthotopic syngeneic mouse model, an oral dose of C150 at 150 mg/kg 3× weekly significantly increased survival of mice bearing pancreatic tumors, and reduced tumor growth and ascites occurrence. These results suggested that compound C150 holds promises in comprehensively inhibiting pancreatic cancer progression.

Selection of Optimal Adjuvant Chemotherapeutic Agents for Pancreatic Cancer Treatment

Pancreatic cancer generally has a dismal prognosis, with a 5-year overall survival rate of approximately 10%. At present, surgical resection is the only treatment with a curative aim; however, it is applicable to only 15–20% patients with pancreatic cancer at the time of diagnosis and the median survival following treatment with surgery alone is only 11–20 months. Many trials have shown that adjuvant chemotherapy offers improved outcomes over observation following surgical resection. Recently, modified folinic acid–fluorouracil–irinotecan–oxaliplatin (FOLFIRINOX) treatment following surgical resection in patients with good performance status has shown an impressive 9-month increase in disease-free survival. However, patients included in the study were selected based on a narrow set of criteria, and adverse events during adjuvant chemotherapy remain a limitation. In this article, we review some key trials of adjuvant chemotherapy treatment for pancreatic cancer and discuss future perspectives including neoadjuvant treatment approaches.

Abstract 6216: Therapeutic potential of IAP inhibitor APG-1387 in combination with PARP- or MEK-targeted therapy, or chemotherapy in pancreatic cancer

Abstract APG-1387 is a small molecule antagonist of the inhibitor of apoptosis proteins (IAPs) which blocks the activity of IAPs family proteins (XIAP, cIAP-1, cIAP-2, and ML-IAP) and consequently induces apoptosis. It is currently in phase I clinical development in patients with solid tumors, including pancreatic cancer. Targeting IAPs has been reported to effectively inhibit tumor growth in the preclinical models of pancreatic cancer. Currently, PARP and MEK inhibitors are also in clinical development for pancreatic cancer with BRCA1/2 or KRAS mutations. To further explore the therapeutic potential of APG-1387 in combination with PARP or MEK inhibitors, a genomic biomarker guided avatar mouse trial (n=2) using pancreatic cancer patient-derived xenografts (PDX) was conducted to evaluate antitumor activity of APG-1387 in combination with (i) the PARP inhibitor, olaparib, in PDXs carrying BRCA1/2 mutations, and (ii) the MEK inhibitor, trametinib, in PDXs carrying KRAS mutations. The results revealed that APG-1387 improved the antitumor activity of olaparib in 4 out of 6 (67%) BRCA1/2 mutant PDXs with a 44% reduction of T/C (%) value (T, treatment group, C, vehicle control), including one partial tumor regression (PR). In combination with trametinib, the antitumor activity was improved in 4 out of 9 (44%) KRAS mutant PDXs, showing a 34% reduction in the T/C value. Furthermore, synergy of the combinations of APG-1387 with PARP or MEK inhibitor was confirmed in a follow-up efficacy study (n=5) using a PDX model carrying both BRCA2 and KRAS mutations. The molecular mechanisms underlying the synergy are currently being investigated. In summary, our results suggest that APG-1387 is a potential agent for pancreatic cancer treatment, ascribing to its potential synergistic antitumor effect by combining with either PARP inhibitors in BRCA1/2 mutant or MEK inhibitors in KRAS mutant pancreatic cancers. Overall, our data provide the scientific rationale for the future clinical development of these combinations in pancreatic cancer patients with distinct genomic alterations. Citation Format: Qiang Li, Douglas D. Fang, Shoulai Gu, Dajun Yang, Yifan Zhai. Therapeutic potential of IAP inhibitor APG-1387 in combination with PARP- or MEK-targeted therapy, or chemotherapy in pancreatic cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6216.

PO-515 Novel water-solube [60]fullerene nanotherapeutic agent for pancreatic cancer treatment

IntroductionThe rapid development of nanotechnology is of great interest to researchers focused on translational medicine and novel targeted cancer treatment. This novel ‘nano’ approach has been used by medicinal chemist to design potential drugs which could easily broke almost any biological barriers. It has been published earlier, that engineered nanoparticles, including carbon nanomaterials, could enter tumours via their leaky vessels system and retain due to a weak drainage in the tumour microenvironment. Here we prestent the synthesis of water-soluble d-glucosamine derivatives used as drug delivery tools and selective FYN and LCK kinase inhibitor for pancreatic cancer treatment.Material and methodsHuman PDAC line PANC-1, ASCP-1 and PAN-O2 was obtained from American Type Culture Collection (ATCC, USA) or from National Cancer Institute. Nuclear magnetic resonance spectra were measured on a Bruker 400 MHz NMR Spectrometer with TMS as an internal standard. MS datasets were collected using Autoflex II MALDI-TOF mass spectrometer, and MS electrospray ionisation time-of-flight (ESI-microTOF) mass spectrometer, both instruments from Bruker Daltonics Inc (Fremont, CA). High-resolution spectra were performed using Shimadzu IT-ToF LC-MS System and flash chromatography was performed using Agilent 971-FP Flash Purification System.Results and discussionsTo date we have designed and synthesised three glucosamine-based, water-soluble [60]fullerene derivatives with high wate solubility up to 400 mg/ml. It was observed that all fullerenes form two aggregate fraction 20–30 nm and 400–500 nm. Initial dark cytotoxicity studies on pancreatic cancer cell line PANC1 have been carried out using flow cytometry and propidium iodide (PI) apoptosis staining. It has been shown that all 3 glycofullerenes are non-toxic even in high concentrations (up to 1 mg/ml, incubation 3 and 24 hours). Moreover, synthesised [60]fullerene derivative localises preferentially in the nucleus of PSC cells, with some localization in the cell cytoplasm.ConclusionOur results show predominant cellular nuclear internalisation of our novel hexakis glucosamine C60 derivative. In addition, synthesised nanotherapeutic is inherently non-toxic up to concentrations of 1 mg/ml, and displays strong photodynamic cytotoxic behaviour, when illuminated with both blue and green light. Moreover designed nanotherapeutic is showing selective inhibition to two kinases FYN A and LCK.MS thanks National Science Centre (Poland) for the support (grant UMO- 2016/23/D/NZ7/00912)

A new goniothalamin N-acylated aza-derivative strongly downregulates mediators of signaling transduction associated with pancreatic cancer aggressiveness

In this study, a novel concise series of molecules based on the structure of goniothalamin (1) was synthesized and evaluated against a highly metastatic human pancreatic cancer cell line (Panc-1). Among them, derivative 8 displayed a low IC50 value (2.7 μM) and its concentration for decreasing colony formation was 20-fold lower than goniothalamin (1). Both compounds reduced the levels of the receptor tyrosine kinase (AXL) and cyclin D1 which are known to be overexpressed in pancreatic cancer cells. Importantly, despite the fact that goniothalamin (1) and derivative 8 caused pancreatic cancer cell cycle arrest and cell death, only derivative 8 was able to downregulate pro-survival and proliferation pathways mediated by mitogen activated protein kinase ERK1/2. Another interesting finding was that Panc-1 cells treated with derivative 8 displayed a strong decrease in the transcription factor (c-Myc), hypoxia-inducible factor-1α (HIF-1α) and vascular endothelial growth factor (VEGF) protein levels. Notably, the molecular effects caused by derivative 8 might not be related to ROS generation, since no significant production of ROS was observed in low concentrations of this compound (from 1.5 up to 3 μM). Therefore, the downregulation of important mediators of pancreatic cancer aggressiveness by derivative 8 reveals its great potential for the development of new chemotherapeutic agents for pancreatic cancer treatment.

Abstract 1685: HER2 upregulation induced by gemcitabine treatment augments antitumor effect of trastuzumab emtansine against pancreatic cancer

Abstract [Background] Although human pancreatic cancer (PC) frequently expresses human epidermal growth factor receptor type 2 (HER2), Trastuzumab has not provided sufficient therapeutic effect on PC probably due to low HER2 expression level. Recently we discovered HER2 expression of human PC cell lines was enhanced by gemcitabine (GEM) treatment. Furthermore, Trastuzumab Emtansine (T-DM1), Trastuzumab conjugated with a cytotoxic agent, showed promising results in breast cancer therapy. We investigated the possibility of new molecular targeting therapy against PC by combined treatment with GEM and T-DM1. [Materials and Methods] HER2 expression was analyzed by quantitative Real Time-PCR, western blotting and flowcytometry. The cell proliferation was examined by spectrophotometry using the counting reagents. T-DM1 was provided by Genentech Inc. T-DM1 bindinig to HER2 on PC cell surface was analyzed by flowcytometry. [Results] GEM treatment enhanced HER2 expression at around two fold in three out of five human PC cell lines. GEM was the most potent agent to enhance HER2 expression of PC cells among the standard chemotherapeutic agents for PC treatment. T-DM1 binding to HER2 on GEM-treated MiaPaca-2 cells was increased in accordance with the up-regulation of HER2 expression. Combined treatment with GEM and T-DM1 synergistically inhibited the cell growth of MiaPaca-2, and this cytotoxic effect was inhibited by adding to Trastuzumab which did not show direct cytotoxic activity to PC cells. [Conclusions] GEM treatment enhances HER2 expression of PC cells which promotes T-DM1 binding to HER2 on PC cells. Combined treatment with GEM and T-DM1 might become a promising modality against HER2 positive PC. Citation Format: Shin Kan, Shigeo Koido, Masato Okamoto, Kazumi Hayashi, Masaki Ito, Yuko Kamata, Hideo Komita, Eijiro Nagasaki, Sadamu Homma. HER2 upregulation induced by gemcitabine treatment augments antitumor effect of trastuzumab emtansine against pancreatic cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1685. doi:10.1158/1538-7445.AM2014-1685

Perifosine inhibits S6K1–Gli1 signaling and enhances gemcitabine-induced anti-pancreatic cancer efficiency

The pancreatic cancer has extremely low overall 5-year survival, and gemcitabine is the only approved single agent for pancreatic cancer treatment. In the present study, we investigated the potential effect of perifosine, a novel Akt inhibitor on gemcitabine-induced anti-pancreatic cancer effect both in vivo and in vitro. We showed that sub-cytotoxic low concentration of perifosine dramatically enhanced gemcitabine-induced cytotoxicity in cultured pancreatic cancer cells. Perifosine inhibited Akt-mammalian target of rapamycin and Erk-mitogen-activated protein kinase activation in pancreatic cancer cells. Meanwhile, perifosine suppressed the hedgehog signaling, as it inhibited glioma-associated oncogenes (Gli) 1 activation and decreased its target protein patched 1 (PTCH1) expression. Our data demonstrated that perifosine blocked p70S6K1 (S6K1) activation, thus disrupting S6K1-Gli1 association and subsequent Gli1 activation. The reduction of S6K1 or Gli1 expression by target siRNAs inhibited PTCH1 expression and enhanced gemcitabine-induced cytotoxicity in pancreatic cancer cells. Significantly, perifosine dramatically enhanced gemcitabine-mediated antitumor effect in a PANC-1 xenograft severe combined immunodeficiency mice model. In summary, we conclude that perifosine sensitizes gemcitabine-mediated anti-pancreatic cancer efficiency through regulating multiple signaling pathways.

Bufalin exerts antitumor effects by inducing cell cycle arrest and triggering apoptosis in pancreatic cancer cells

As one of the most aggressive human malignancies, pancreatic cancer is a leading cause of cancer-related deaths worldwide and only about 4% of patients will live 5 years after diagnosis. Eighty to approximately eighty-five percent of patients are diagnosed with an unresectable or metastatic disease, which is correlated with poor prognosis and low survival rate. Therefore, it is tremendously significant to exploit novel chemicals to prevent and treat pancreatic cancer. Previous research and clinical studies have demonstrated that many natural products derived from traditional Chinese medicine (TCM) such as camptothecin derivatives and vinca alkaloids could be effective antitumor compounds, hinting that TCM is a promising source for developing new antitumor drugs. In this report, we investigated the effects of bufalin, a primary active ingredient of the traditional Chinese medicine Chan-Su, on pancreatic cancer cell lines PANC-1 and CFPAC-1 and studied the underlying molecular mechanism. We found that exposure to bufalin could suppress the proliferation of pancreatic cancer cells time and dose dependently. We used flow cytometry to study the effects of bufalin on apoptosis and cell cycle distribution in PANC-1 and CFPAC-1 cells. The results indicated that bufalin could significantly induce both apoptosis and G2/M cell cycle arrest in pancreatic cancer cells. With western blotting, we found that the expression level of an antiapoptotic protein heat shock protein 27 (Hsp27) and its partner molecule p-Akt was decreased upon the treatment with bufalin. Besides, bufalin activated pro-caspase-3 and pro-caspase-9 and modulated the expression level of Bcl-2 and Bax. These data suggested that bufalin may trigger apoptosis by targeting Hsp27, which could inhibit apoptosis by interfering with key apoptotic proteins. The influence on the level of cylinB1, CDK1, and p21 was also observed after bufalin treatment, and the relationship between Hsp27 and the cell cycle-related proteins mentioned above deserves much more research. In addition, our data showed that bufalin could enhance the growth inhibition effect of gemcitabine in above pancreatic cancer cells. Taken together, bufalin might be worthy of further study for its potential as a therapeutic agent for pancreatic cancer treatment.

Therapy of Smac mimetic SM-164 in combination with gemcitabine for pancreatic cancer

Pancreatic cancer (PC) is a lethal solid malignancy with resistance to traditional chemotherapy. We investigated therapy of PC with SM-164 and gemcitabine alone and in combination. Survival of PC cells was reduced as the dose of SM-164 increased. SM-164 and/or gemcitabine increased the number of apoptotic and dead PC cells, and expression of cleavage fragments of caspase-3 and PARP1, and inhibited tumor xenograft growth in nude mice. The inhibitory effect of combination treatment was greater and of longer duration than monotherapy. Neither combination nor monotherapy showed any significant toxicity in vivo. Apoptosis and necrosis, decreased expression of Ki67, and increased expression of cleaved caspase-3 were observed in xenograft tumor tissues in SM164/gemcitabine-treated mice. SM-164 could be a promising new agent for treatment of PC in combination with gemcitabine.

The Role of EMT in Pancreatic Cancer Progression

The Role of EMT in Pancreatic Cancer Progression

Novel Thiosemicarbazone Iron Chelators Induce Up-Regulation and Phosphorylation of the Metastasis Suppressor N-myc Down-Stream Regulated Gene 1: A New Strategy for the Treatment of Pancreatic Cancer

Pancreatic cancer is an aggressive neoplasm, with a mortality rate close to 100%. The most successful agent for pancreatic cancer treatment is gemcitabine, although the overall effect in terms of patient survival remains very poor. This study was initiated to evaluate a novel class of anticancer agents against pancreatic cancer. This group of compounds belongs to the dipyridyl thiosemicarbazone class that have been shown to have potent and selective activity against a range of different neoplasms in vitro and in vivo. We demonstrate for the first time in pancreatic cancer that these agents increase the expression of the growth and metastasis suppressor N-myc downstream-regulated gene 1 and its phosphorylation at Ser330 and Thr346 that is important for its activity against this tumor. In addition, these agents increased expression of the cyclin-dependent kinase inhibitor p21(CIP1/WAF1), whereas decreasing cyclin D1 in pancreatic cancer cells. Together, these molecular alterations account, in part, for the pronounced antitumor activity observed. Indeed, these agents had significantly higher antiproliferative activity in vitro than the established treatments for pancreatic cancer, namely gemcitabine and 5-fluorouracil. Studies in vivo demonstrated that a novel thiosemicarbazone, namely di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone hydrochloride, completely inhibited the growth of pancreatic cancer xenografts with no evidence of marked alterations in normal tissue histology. Together, our studies have identified molecular effectors of a novel and potent antitumor agent that could be useful for pancreatic cancer treatment.

Abstract 4234: Ursolic acid induces apoptosis in pancreatic cancer cells associated with inhibition of STAT3 and down-regulation of Bcl-XL

Abstract Pancreatic cancer is the fourth leading cause of cancer death and without effective medical therapy. New agents acting on novel targets need to be developed. Natural products provide a promising source for developing effective anticancer agents. Oleanolic acid (OA) and ursolic acid (UA) are two natural triterpenoids with reported antitumor activities in several types of cancer. The antitumor effects of OA, oleanolic acid methyl ester (OAME) and UA were determined in pancreatic cancer Aspc-1 cells. Cell growth inhibition of the three compounds was determined using the MTT assay. The IG50s of OA, OAME and UA after treatment with UA for 4 days are &amp;gt;30, 8.9, 12.5 μM, respectively. OA is a weak cell growth inhibitor and methyl esterification improves its antitumor activity. UA is more effective than OA to inhibit cell growth in Aspc-1 cells and was used to study the mechanism of action. Aspc-1 cells treated with UA at a concentration higher than 25 uM induced apoptosis as determined by PARP cleavage detection. Western blot analysis revealed that the levels of procaspase-3, -9 and Bcl-XL were markedly decreased while the levels of Mcl-1, XIAP and survivin were weakly or not decreased. Correlated with the decrease in the level of Bcl-XL, the level of phosphorylated STAT3 was significantly inhibited. STAT3 is activated in pancreatic cancer cells and contributes to pancreatic tumorigenesis. Activated STAT3 increases the levels of antiapoptotic protein such as Bcl-XL, Mcl-1 and survivin, are being considered as a target for developing agents for pancreatic cancer treatment. Our data suggest that UA is a lead candidate for targeting STAT3 and structural modification of UA may result in a new group of compounds with improved antitumor activity in pancreatic cancer cells. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4234. doi:10.1158/1538-7445.AM2011-4234

Reactivation of p53 by Novel MDM2 Inhibitors: Implications for Pancreatic Cancer Therapy

The present study is the first to show in pancreatic cancer (PC) the growth inhibition and apoptosis by novel MDM2 inhibitors (MI-319 & 219) through reactivation of p53 pathway. Our results highlight two new secondary targets of MDM2 inhibitor 'SIRT1' and Ku70. SIRT1 which has a role in ageing and cancer and is known to regulate p53 signaling through acetylation. Ku70 is a key component of non-homologous end joining machinery in the DNA damage pathway and is known to regulate apoptosis by blocking Bax entry into mitochondria. Growth inhibition and apoptosis by MI-219, MI-319 was accompanied by increase in levels of p53 along with p21(WAF1) and the proapoptotic Puma. SiRNA against p21(WAF1) abrogated the growth inhibition of PC cells confirming p21(WAF1) as a key player downstream of activated p53. Immunoprecipitation-western blot analysis revealed reduced association of MDM2-p53 interaction in drug exposed PC cells. In combination studies, the inhibitors synergistically augmented anti-tumor effects of therapeutic drug gemcitabine both in terms of cell growth inhibition as well as apoptosis. Surface plasmon resonance studies confirmed strong binding between MI-319 and Ku70 (K(D) 170 nM). Western blot revealed suppression of SIRT1 and Ku70 with simultaneous upregulation of acetyl-p53 (Lys379) and Bax. Co-Immunoprecipitation studies confirmed that MI-319 could disrupt Ku70-Bax and SIRT1-Bax interaction. Further, using wt-p53 xenograft of Capan-2, we found that oral administration of MI-319 at 300 mg/kg for 14 days resulted in significant tumor growth inhibition without any observed toxicity to the animals. No tumor inhibition was found in mut-p53 BxPC-3 xenografts. In light of our results, the inhibitors of MDM2 warrant clinical investigation as new agents for PC treatment.

Rottlerin stimulates apoptosis in pancreatic cancer cells through interactions with proteins of the Bcl-2 family

Rottlerin is a polyphenolic compound derived from Mallotus philipinensis. In the present study, we show that rottlerin decreased tumor size and stimulated apoptosis in an orthotopic model of pancreatic cancer with no effect on normal tissues in vivo. Rottlerin also induced apoptosis in pancreatic cancer (PaCa) cell lines by interacting with mitochondria and stimulating cytochrome c release. Immunoprecipitation results indicated that rottlerin disrupts complexes of prosurvival Bcl-xL with Bim and Puma. Furthermore, siRNA knockdown showed that Bim and Puma are necessary for rottlerin to stimulate apoptosis. We also showed that rottlerin and Bcl-2 and Bcl-xL inhibitor BH3I-2' stimulate apoptosis through a common mechanism. They both directly interact with mitochondria, causing increased cytochrome c release and mitochondrial depolarization, and both decrease sequestration of BH3-only proteins by Bcl-xL. However, the effects of rottlerin and BH3I-2' on the complex formation between Bcl-xL and BH3-only proteins are different. BH3I-2' disrupts complexes of Bcl-xL with Bad but not with Bim or Puma, whereas rottlerin had no effect on the Bcl-xL interaction with Bad. Also BH3I-2', but not rottlerin, required Bad to stimulate apoptosis. In conclusion, our results demonstrate that rottlerin has a potent proapoptotic and antitumor activity in pancreatic cancer, which is mediated by disrupting the interaction between prosurvival Bcl-2 proteins and proapoptotic BH3-only proteins. Thus rottlerin represents a promising novel agent for pancreatic cancer treatment.

Effects of recombinant human canstatin protein in the treatment of pancreatic cancer

To examine the effect of canstatin, a newly discovered endogenous inhibitor of angiogenesis, in the treatment of pancreatic cancer in vivo. The canstatin cDNA fragment was synthesized and amplified from the total RNA extracted from human placenta tissues by RT-PCR. The resulting product was firstly cloned into pUCm-T vector, then into plasmid pET-22b (+) and transformed into E. coli BL21. Isopropyl-1-thio-b-Dgalactopyran-oside (IPTG) was used to induce the expression of canstatin protein and affinity chromatography was used to purify the protein. To determine the activity of purified recombinant human canstatin (rhCanstatin), orthotopic xenograft human pancreatic cancer models were established. Human pancreatic cancer cells (SW1990) were injected into the pancreas of BALB/c nude mice. Twenty-four nude mice with orthotopic xenograft tumor were randomly divided into 3 groups 10 d after the inoculation, and were treated with PBS 0.3 mL, or canstatin 5 mg/kg, or 10 mg/kg per day for 3 wk intraperitoneally. When the experiment was over, all tumors were resected and the effects of rhCanstatin on tumor growth, microvessel density (MVD) were analyzed. After IPTG induction, SDS-PAGE showed a new monomeric 24 kDa protein band. This protein was purified through affinity chromatography and refolded through dialysis with a final concentration of 60 mg/L. In orthotopic pancreatic cancer models, the final tumor volume in groups treated with PBS, canstatin 5 mg/kg, 10 mg/kg were 355.21+/-39.54 mm3, 112.73+/-10.47 mm3, and 61.75+/-6.99 mm3 respectively. The immunohistochemical examination showed that the MVD in tumors treated with canstatin was significantly less than that in other group. These findings demonstrate that the rhCanstatin effectively retards the growth of pancreatic cancer in a dose-dependent manner through inhibiting angiogenesis and may be a promising therapeutic agent for pancreatic cancer treatment in the clinic.

Arotinoid mofarotene (RO40-8757) up-regulates p21 and p27 during growth inhibition of pancreatic cancer cell lines.

Effective chemotherapy for pancreatic cancer is urgently needed. The anti-proliferative activity of a new retinoid, mofarotene (RO40-8757), was compared with that of other retinoids, such as all trans-retinoic acid, 13-cis retinoic acid and 9-cis retinoic acid, on 9 pancreatic cancer cell lines in relation to the effects on various cell cycle-regulating factors. After treatment with each retinoid, anti-proliferative effect was determined by the MTT method and expression of cell cycle-regulating factors, such as cyclins (D1, E and A), cyclin-dependent kinases (2 and 4), cyclin-dependent kinase inhibitors (p21 and p27) and retinoblastoma protein, was analyzed by Western blotting. Mofarotene showed half-maximal inhibition of cell proliferation at concentrations between 0.14 x 10(-6) and 3.8 x 10(-6) mol/l with little cytotoxicity. In contrast, the other retinoids did not inhibit the growth of all cell lines by over 50% compared to controls. A marked increase in the fraction of cells in G1 phase of the cell cycle was observed after mofarotene treatment; this was associated with marked up-regulation of p21/p27 and a shift of retinoblastoma protein into the hypophosphorylated form. In conclusion, mofarotene inhibits the growth of pancreatic cancer cells by inducing G1-phase cell cycle-inhibitory factors (p21, p27 and hypophosphorylated form of Rb protein) and is considered to be a useful agent for pancreatic cancer treatment.