Strategy For Pancreatic Cancer Therapy Research Articles

Dual Starvations Induce Pyroptosis for Orthotopic Pancreatic Cancer Therapy through Simultaneous Deprivation of Glucose and Glutamine.

Pancreatic cancer is a highly fatal disease, and existing treatment methods are ineffective, so it is urgent to develop new effective treatment strategies. The high dependence of pancreatic cancer cells on glucose and glutamine suggests that disrupting this dependency could serve as an alternative strategy for pancreatic cancer therapy. We identified the vital genes glucose transporter 1 (GLUT1) and alanine-serine-cysteine transporter 2 (ASCT2) through bioinformatics analysis, which regulate glucose and glutamine metabolism in pancreatic cancer, respectively. Human serum albumin nanoparticles (HSA NPs) for delivery of GLUT1 and ASCT2 inhibitors, BAY-876/V-9302@HSA NPs, were prepared by a self-assembly process. This nanodrug inhibits glucose and glutamine uptake of pancreatic cancer cells through the released BAY-876 and V-9302, leading to nutrition deprivation and oxidative stress. The inhibition of glutamine leads to the inhibition of the synthesis of the glutathione, which further aggravates oxidative stress. Both of them lead to a significant increase in reactive oxygen species, activating caspase 1 and GSDMD and finally inducing pyroptosis. This study provides a new effective strategy for orthotopic pancreatic cancer treatment by dual starvation-induced pyroptosis. The study for screening metabolic targets using bioinformatics analysis followed by constructing nanodrugs loaded with inhibitors will inspire future targeted metabolic therapy for pancreatic cancer.

Abstract B035: Targeting ribosome biogenesis addition is a novel strategy for pancreatic cancer therapy

Abstract Pancreatic cancer (PanCa) is the third leading cause of cancer-related deaths in the United States with limited therapeutic options available. Gemcitabine (GEM), a deoxycytidine nucleoside analog is currently considered the most effective therapy for PanCa; however, it shows only a marginal survival benefit of 6 months. PanCa cells are addicted to ribosome biogenesis (RiBi) which supports their high rate of growth and proliferation. Therefore, strategically targeting the process of RiBi could be one of the ideal strategies for the prevention and treatment of PanCa. In this study, we for the first-time report that RPA194, a catalytic subunit of RNA Pol I is differentially expressed in normal pancreatic ductal epithelial (NPDE) and cancer cells. We also observed differential expression of RPA194 in various grades of pancreatic tumor tissues and its level was increased in high-grade pancreatic tumor tissues. We tested our hypothesis that targeting RPA194 with a non-toxic pharmacological inhibitor (BMH-21) will inhibit the growth of advanced PanCa. We observed that BMH-21 inhibits the growth and induced apoptosis of PanCa cells in a dose-dependent manner and this inhibition was correlated with the expression of RPA194. We observed proteasomal mediated degradation of RPA194 in PanCa cells. BMH-21 treatment inhibited RPA194 occupancy on rDNA but did not affect other nucleolar proteins (UBF, fibrillarin, nucleolin, and nucleophosmin). Two times the effective concentration of BMH-21 (5 µM) did not show any cytotoxic effect in NPDE cells. However, BMH-21 degraded RPA 194 protein in these cells in a similar manner as in PanCa cells. Therefore, we speculate that other proteins are involved in the selective toxicity of BMH-21 in PanCa cells. These findings were translated to in vivo model systems. We observed that BMH-21 significantly (P<0.01) inhibited the growth of pancreatic tumors in an orthotopic xenograft model. This pancreatic tumor growth inhibition was correlated with degradation of RPA194 protein in BMH-21 treated mice tumor tissues followed by inhibition of cell proliferation markers (ki67 and PCNA). Taken together, our results strongly suggest that targeting the RiBi process is a novel approach and BMH-21 is a non-toxic potential targeting agent for the prevention and treatment of advanced PaCa. Citation Format: Mudassier Ahmad, Carlos Perez, Andrew Massey, Vivek Kashyap, Neeraj Chauhan, Haider Ahsan, Jasmine Jasmine, Manish Tripathi, Subhash Chauhan, Bilal Hafeez. Targeting ribosome biogenesis addition is a novel strategy for pancreatic cancer therapy [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer; 2022 Sep 13-16; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2022;82(22 Suppl):Abstract nr B035.

Neutrophil membrane-coated nanoparticles for enhanced nanosecond pulsed electric field treatment of pancreatic cancer

Objective Pancreatic cancer is one of the leading causes of cancer-related deaths worldwide. Poor prognosis and low survival rates have driven the development of novel therapeutic strategies. Nanosecond pulsed electric field has emerged as a novel, minimal invasive and non-thermal treatment for solid tumors. It is of great significance to study the combination therapy of nsPEF and other treatment strategies for pancreatic cancer. Methods We developed neutrophil membrane-wrapped liposomal nanoparticles loaded with gemcitabine (NE/Lip-GEM) and investigated their use as a complementary agent for nsPEF treatment. Results Our results showed that neutrophil-mediated delivery of liposomal-gemcitabine (NE/Lip-GEM) efficiently inhibited the growth of pancreatic tumors in mice whose has been treated with incomplete nsPEF ablation. Conclusions The combination of nsPEF and NE/Lip-GEM may be a promising synergistic strategy for pancreatic cancer therapy.

Development of a Tumor-Responsive Nanopolyplex Targeting Pancreatic Cancer Cells and Stroma.

Desmoplasia plays a pivotal role in promoting pancreatic cancer progression and is associated with poor clinical outcome. Targeting the desmoplastic tumor microenvironment in combination with chemotherapy is therefore a promising strategy for pancreatic cancer therapy. Here, we report a novel biodegradable copolymer to codeliver LY2109761 (a TGF-β receptor I/II inhibitor) and CPI-613 (a novel chemotherapy agent) to desmoplastic stroma and tumor cells, respectively, in the tumor microenvironment. Hydrophobic CPI-613 is conjugated to the hydrophilic copolymer via a newly designed MMP-2-responsive linker to form a trigger-responsive nanopolyplex. LY2109761 is hydrophobic and encapsulated into the hydrophobic core of the nanopolyplex. The resulting nanopolyplex is modified with a plectin-1-targeting peptide to enhance the accumulation of the nanopolyplex in pancreatic tumors. The nanopolyplex aims to normalize the stroma by blocking the interaction between tumor cells and pancreatic stellate cells to inhibit the activation of pancreatic stellate cells and subsequently reduce the dense extracellular matrix. Normalized stroma increases the penetration of the nanopolyplex into the tumor. The nanopolyplex shows enhanced accumulation in xenograft pancreatic tumors in a biodistribution study. Moreover, the targeted nanopolyplex markedly inhibits tumor growth in an orthotopic pancreatic cancer mouse model by dual-targeting tumor cells and stroma. Overall, the multifunctional nanopolyplex is a promising platform for pancreatic cancer therapy.

CUDC-907 displays potent antitumor activity against human pancreatic adenocarcinoma in vitro and in vivo through inhibition of HDAC6 to downregulate c-Myc expression.

Pancreatic adenocarcinoma is a highly malignant cancer that often involves a deregulation of c-Myc. It has been shown that c-Myc plays a pivotal role in the regulation of a variety of physiological processes and is involved in early neoplastic development, resulting in poor progression. Hence, suppression of c-Myc overexpression is a potential strategy for pancreatic cancer therapy. CUDC-907 is a novel dual-acting inhibitor of phosphoinositide 3-kinase (PI3K) and histone deacetylase (HDAC). It has shown potential efficiency in patients with lymphoma, multiple myeloma, or thyroid cancer, as well as in solid tumors with c-Myc alterations, but the evidence is lacking for how CUDC-907 regulates c-Myc. In this study, we investigated the effect of CUDC-907 on human pancreatic cancer cells in vitro and in vivo. Our results showed that CUDC-907 potently inhibited the proliferation of 9 pancreatic cancer cell lines in vitro with IC50 values ranging from 6.7 to 54.5 nM. Furthermore, we revealed the antitumor mechanism of CUDC-907 in Aspc-1, PANC-1, and Capan-1 pancreatic cancer cells: it suppressed the HDAC6 subunit, thus downregulating c-Myc protein levels, which was a mode of action distinct from the existing mechanisms. Consistently, the extraordinary antitumor activity of CUDC-907 accompanied by downregulation of c-Myc and Ki67 expression in tumor tissue was observed in a human pancreatic cancer Aspc-1 xenograft nude mouse model in vivo. Our results suggest that CUDC-907 can be a valuable therapeutic option for treating pancreatic adenocarcinoma.

Targeting glypican-4 overcomes 5-FU resistance and attenuates stem cell-like properties via suppression of Wnt/β-catenin pathway in pancreatic cancer cells.

The existences of cancer stem cells in patients with pancreatic cancer are considered as pivotal factors contributing to chemoresistance and disease relapse. Glypican-4 (GPC4) is one of the members of the glypicans family, which underlies human congenital malformations and multiple diseases. However, its potential biological function in pancreatic cancer still remains elusive. In this study, we are the first to demonstrate that GPC4 was involved in 5-fluorouracil (5-FU) resistance and pancreatic cancer stemness through comprehensive bioinformatical analysis. Functional experiments showed that knockdown of GPC4 sensitized pancreatic cancer cells to 5-FU and attenuated stem cell-like properties. In terms of mechanism research, knockdown of GPC4 suppressed the activation of Wnt/β-catenin pathway and its downstream targets. Furthermore, the expression of GPC4 was significantly upregulated in pancreatic cancer tissues compared with normal tissues and remarkably correlated with patients' overall survival according to the data derived from the Cancer Genome Atlas database. Taken together, our results suggest that GPC4 is a key regulator in chemoresistance and pancreatic cancer stemness. Thus, targeting GPC4 may serve as a promising strategy for pancreatic cancer therapy.

Effects of obatoclax combined with gemcitabine on the biological activity of pancreatic cancer cells under hypoxic conditions.

The present study aimed to investigate the effects of obatoclax (OBX) combined with gemcitabine (GEM) treatment on the proliferation, migration, invasion and epithelial‑mesenchymal transition (EMT) related proteins of pancreatic cancer cell line BxPC‑3 under hypoxic conditions. Protein expression levels of hypoxia‑inducible factor 1α (HIF‑1α) in BxPC‑3 pancreatic cancer cells under normoxic and hypoxic conditions were detected by western blotting. Cells were divided into four groups: Normoxia group, hypoxia group, OBX group and OBX + GEM group. The proliferation activity of BxPC‑3 cells was detected by Cell Counting kit‑8. The migratory and invasive abilities of BxPC‑3 cells were detected by the scratch test and Matrigel assay, respectively. The protein expression levels of vimentin, E‑cadherin and p53 in BxPC‑3 cells were also detected by western blotting. HIF‑1α expression under hypoxic conditions was significantly increased compared with expression under normoxic conditions. Under hypoxic conditions, OBX treatment reduced cell activity, decreased cell migration and invasion, promoted the expression of E‑cadherin and p53. In the OBX + GEM group, BxPC‑3 cell activity decreased significantly, cell migration and invasion decreased significantly, the expression of vimentin was significantly reduced and the expression of E‑cadherin and p53 further increased. In conclusion, the present results demonstrated that under hypoxic conditions, OBX combined with a small dose of GEM may be able to inhibit the growth, migration and invasion of pancreatic cancer cells, possibly via inhibition of EMT process. These results may provide a promising strategy for pancreatic cancer therapy.

Doublecortin-Like Kinase 1 (DCLK1) Regulates B Cell-Specific Moloney Murine Leukemia Virus Insertion Site 1 (Bmi-1) and is Associated with Metastasis and Prognosis in Pancreatic Cancer

Background/Aims: Cancer stem cells (CSCs) are largely responsible for tumor relapse and metastatic behavior. Doublecortin-like kinase 1 (DCLK1) was recently reported to be a biomarker for gastrointestinal CSCs and involved in the epithelial-mesenchymal transition (EMT) and tumor progression. B cell-specific Moloney murine leukemia virus insertion site 1 (Bmi-1) is a crucial regulator of CSC self-renewal, malignant transformation and EMT, and a previous study from our group showed that Bmi-1 is upregulated in pancreatic cancer progression and participates in EMT. However, it remains unclear whether DCLK1 is involved in pancreatic cancer or whether DCLK1 is associated with the altered level of Bmi-1 expression. Methods: The correlation of DCLK1 expression and clinical features of pancreatic cancer was analyzed in 210 paraffin-embedded archived pancreatic cancer specimens by immunohistochemical analysis. The biological effects of DCLK1 siRNA on cells were investigated by examining cell proliferation using a cell counting kit and cell colony assays, cell migration by wound healing assay and cell invasion by Transwell invasion assay. We further investigated the effect of therapeutic siRNA targeting DCLK1 on pancreatic cancer cell growth in vivo. Moreover, the molecular mechanism by which DCLK1 upregulates Bmi-1 expression was explored using real-time PCR, western blotting and Co-immunoprecipitation assay. Results: DCLK1 is overexpressed in pancreatic cancer and is related to metastasis and prognosis. Knockdown of DCLK1 markedly suppressed cell growth in vitro and in vivo and also inhibited the migration and invasion of pancreatic cancer cells. Furthermore, we found that DCLK1 silencing could inhibit EMT in cancer cells via downregulation of Bmi-1 and the mesenchymal markers Snail and Vimentin and upregulation of the epithelial marker E-cadherin. Moreover, high DCLK1 expression in human pancreatic cancer samples was associated with a mesenchymal phenotype and increased cell proliferation. Further co-immunoprecipitation indicated that DCLK1 did not interact with Bmi-1 directly. Conclusion: Our data suggest that upregulation of DCLK1 may contribute to pancreatic cancer metastasis and poor prognosis by increasing Bmi-1 expression indirectly. The findings indicate that inhibiting DCLK1 expression might be a novel strategy for pancreatic cancer therapy.

Chaetospirolactone reverses the apoptotic resistance towards TRAIL in pancreatic cancer

The pancreatic cancer is one of the most aggressive tumors. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) can trigger apoptosis by interaction with death receptors. However, in TRAIL-resistant pancreatic cancer, responsiveness to TRAIL treatment is terribly poor. In current work, we have demonstrated that a natural product chaetospirolactone (CSL) isolated from an endophytic fungus Chaetomium sp. NF00754 can enhance the susceptibility of TRAIL-resistant pancreatic cancer cells to apoptosis. CSL can induce apoptosis in TRAIL-treated pancreatic cancer cells. Furthermore, combined CSL and TRAIL treatment significantly inhibits viability and migration of pancreatic cancer cells. Combinatorial TRAIL and CSL treatment repressed xenograft tumor growth without substantially toxic side effects. CSL can specifically upregulate expression of death receptor 4 (DR4). Further study revealed that CSL represses the activities of an epigenetic regulator enhancer of zeste homolog 2 (EZH2) and consistently reduces histone H3 lysine 27 trimethylation (H3K27me3) to allow DR4 transcription. Taken together, CSL treatment may reverse TRAIL resistance in pancreatic cancer cells via epigenetic regulation of DR4 implying that administration of CSL might represent a putative strategy for pancreatic cancer therapy.

Co-delivery of microRNA-21 antisense oligonucleotides and gemcitabine using nanomedicine for pancreatic cancer therapy.

Tumor metastasis occurs naturally in pancreatic cancer, and the efficacy of chemotherapy is usually poor. Precision medicine, combining downregulation of target genes with chemotherapy drugs, is expected to improve therapeutic effects. Therefore, we developed a combined therapy of microRNA‐21 antisense oligonucleotides (ASO‐miR‐21) and gemcitabine (Gem) using a targeted co‐delivery nanoparticle (NP) carrier and investigated the synergistic inhibitory effects on pancreatic cancer cells metastasis and growth. Polyethylene glycol–polyethylenimine–magnetic iron oxide NPs were used to co‐deliver ASO‐miR‐21 and Gem. An anti‐CD44v6 single‐chain variable fragment (scFvCD 44v6) was used to coat the particles to obtain active and targeted delivery. Our results showed that the downregulation of the oncogenic miR‐21 by ASO resulted in upregulation of the tumor‐suppressor genes PDCD4 and PTEN and the suppression of epithelial–mesenchymal transition, which inhibited the proliferation and induced the clonal formation, migration, and invasion of pancreatic cancer cells in vitro. The co‐delivery of ASO‐miR‐21 and Gem induced more cell apoptosis and inhibited the growth of pancreatic cancer cells to a greater extent than single ASO‐miR‐21 or Gem treatment in vitro. In animal tests, more scFvCD 44v6‐PEG‐polyethylenimine/ASO‐magnetic iron oxide NP/Gem accumulated at the tumor site than non‐targeted NPs and induced a potent inhibition of tumor proliferation and metastasis. Magnetic resonance imaging was used to observed tumor homing of NPs. These results imply that the combination of miR‐21 gene silencing and Gem therapy using an scFv‐functionalized NP carrier exerted synergistic antitumor effects on pancreatic cancer cells, which is a promising strategy for pancreatic cancer therapy.

Overcoming chemo/radio-resistance of pancreatic cancer by inhibiting STAT3 signaling.

Chemo/radio-therapy resistance to the deadly pancreatic cancer is mainly due to the failure to kill pancreatic cancer stem cells (CSCs). Signal transducer and activator of transcription 3 (STAT3) is activated in pancreatic CSCs and, therefore, may be a valid target for overcoming therapeutic resistance. Here we investigated the potential of STAT3 inhibition in sensitizing pancreatic cancer to chemo/radio-therapy. We found that the levels of nuclear pSTAT3 in pancreatic cancer correlated with advanced tumor grade and poor patient outcome. Liposomal delivery of a STAT3 inhibitor FLLL32 (Lip-FLLL32) inhibited STAT3 phosphorylation and STAT3 target genes in pancreatic cancer cells and tumors. Consequently, Lip-FLLL32 suppressed pancreatic cancer cell growth, and exhibited synergetic effects with gemcitabine and radiation treatment in vitro and in vivo. Furthermore, Lip-FLLL32 reduced ALDH1-positive CSC population and modulated several potential stem cell markers. These results demonstrate that Lip-FLLL32 suppresses pancreatic tumor growth and sensitizes pancreatic cancer cells to radiotherapy through inhibition of CSCs in a STAT3-dependent manner. By targeting pancreatic CSCs, Lip-FLLL32 provides a novel strategy for pancreatic cancer therapy via overcoming radioresistance.

Combination of Anti-Diabetic Drug Metformin and Boswellic Acid Nanoparticles: A Novel Strategy for Pancreatic Cancer Therapy

Pancreatic cancer has an infaust prognosis and is the fourth common cause of cancer related death in India. It is highly resistant to conventional treatment modalities such as chemotherapy, radiation therapy and surgery. The association of pancreatic cancer and diabetes mellitus is explored in our study. Pancreatic cancer is more likely to occur in people who have diabetes than people devoid of it, which is supported by the observation that hyperglycaemia occurs at an early stage of pancreatic cancer and is indeed a risk factor. In the present study, we have demonstrated a synergistic relationship between metformin and boswellic acid nanoparticles with varying doses of boswellic acid nanoparticles and constant metformin (20 mM). The effect revealed increased synergism between metformin and boswellic acid nanoparticles through the inhibition of cell proliferation with an effect of 80% for the combination with 0.3 mg/mL and 0.4 mg/mL and a constant concentration of metformin. We examined the effect of combination on cell migration which revealed time dependent inhibitory effect on pancreatic cell line (MiaPaCa-2). Also, we found that the combinatorial approach significantly decreased colony formation and exhibited high rate of induction of apoptosis through DNA fragmentation in pancreatic cancer cells. In-vitro hemolysis confirmed the hemocompatibility of the combination therapy with metformin and boswellic acid nanoparticles. Flow cytometry based apoptosis assay and Caspase mediated apoptosis proved apoptosis mediated cell death. Further, the cells were analysed with mitochondrial membrane potential kit which revealed depolarization of mitochondrial membrane potential due to apoptosis after treatment with drug combination. Hence, the combination approach proved to be a promising therapy towards pancreatic cancer.

Embryonic stem cell factors and pancreatic cancer

Pancreatic ductal adenocarcinoma (PDAC), the most common type of pancreatic tumor, is a highly aggressive human cancer with the lowest five-year survival rate of any human maligancy primarily due to its early- metastasis and lack of response to chemotherapy and radiation. Recent research suggests that PDAC cells comprise a hierarchy of tumor cells that develop around a population of cancer stem cells (CSCs), a small and distinct population of cancer cells that mediates tumoregenesis, metastasis and resistance to standard treatments. Thus, CSCs could be a target for more effective treatment options. Interestingly, pancreatic CSCs are subject to regulation by some of key embryonic stem cell (ESC) transctiption factors abberently expressed in PDAC, such as SOX2, OCT4 and NANOG. ESC transcription factors are important DNA-binding proteins present in both embryonic and adult somatic cells. The critical role of these factors in reprogramming processes makes them essential not only for embryonic development but also tumorigenesis. Here we provide an overview of stem cell transcription factors, particularly SOX2, OCT4, and NANOG, on their expression and function in pancreatic cancer. In contrast to embryonic stem cells, in which OCT4 and SOX2 are tightly regulated and physically interact to regulate a wide spectrum of target genes, de novo SOX2 expression alone in pancreatic cancer cells is sufficient to promote self-renewal, de-differentiation and imparting stemness characteristics via impacting specific cell cycle regulatory genes and epithelial-mesnechymal transtion driver genes. Thus, targeting ESC factors, particularly SOX2, could be a worthy strategy for pancreatic cancer therapy.

Reduction of Decoy Receptor 3 Enhances TRAIL-Mediated Apoptosis in Pancreatic Cancer

Most human pancreatic cancer cells are resistant to tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL)-mediated apoptosis. However, the mechanisms by which pancreatic cancer cells utilize their extracellular molecules to counteract the proapoptotic signaling mediated by the TNF family are largely unknown. In this study, we demonstrate for the first time that DcR3, a secreted decoy receptor that malignant pancreatic cancer cells express at a high level, acts as an extracellular antiapoptotic molecule by binding to TRAIL and counteracting its death-promoting function. The reduction of DcR3 with siRNA unmasked TRAIL and greatly enhanced TRAIL-induced apoptosis. Gemcitabine, a first-line drug for pancreatic cancer, also reduced the level of DcR3. The addition of DcR3 siRNA further enhanced gemcitabine-induced apoptosis. Notably, our in vivo study demonstrated that the therapeutic effect of gemcitabine could be enhanced via further reduction of DcR3, suggesting that downregulation of DcR3 in tumor cells could tip the balance of pancreatic cells towards apoptosis and potentially serve as a new strategy for pancreatic cancer therapy.

Stem cell factor/c-kit signaling enhances invasion of pancreatic cancer cells via HIF-1α under normoxic condition

The SCF/c-kit signaling plays an important role in invasion of c-kit-expressing tumor cells, however, the molecular mechanisms have not been studied yet. Using a pancreatic cancer model, we demonstrate that SCF/c-kit binding up-regulates the expression of invasion-related genes through the accumulation of HIF-1α. Furthermore, the expression of HIF-1α induced by SCF is not dependent on the oxygen level, but rather on both the PI3K/Akt and Ras/MEK/ERK signaling pathways. In conclusion, under normoxic conditions, SCF/c-kit binding increases expression of HIF-1α through the PI3K/Akt and Ras/MEK/ERK pathways, and the accumulation of HIF-1α up-regulates expression of invasion-related genes that augment the invasiveness of pancreatic cancer, a fatal cancer. Therefore, our results suggest that the inhibition of both c-kit and HIF-1α may be an effective strategy for pancreatic cancer therapy.

Abstract 5378: Targeting CXCL12/CXCR4 signaling axis for pancreatic cancer therapy

Abstract Pancreatic cancer is a lethal malignancy with an extremely poor prognosis. Furthermore, it is also highly resistant to currently available therapeutic modalities. Hence, it is very important to better understand the drug-resistance mechanisms and to develop novel and effective therapeutic strategies. Chemokine signaling has been shown to play an important role in cancer progression and metastasis. Importantly, a chemokine receptor, CXCR4, has been shown to be expressed in pancreatic cancer tissues, precancerous lesions, and cancer stem cells. CXCR4 signaling induced by binding to its ligand (CXCL12) potentiates pancreatic tumor cell growth, migration and invasion, and distant metastasis in an autocrine and/or paracrine fashion. CXCL12/CXCR4 signaling has also been shown to facilitate the tumor-microenvironment interaction and induce desmoplastic alterations in the stroma, which is a common feature of pancreatic adenocarcinoma. Therefore, targeting CXCR4, alone or in combination with cytotoxic drug, might be an effective strategy for pancreatic cancer therapy. Here, we examined the expression of CXCR4 in a panel of twelve pancreatic cancer cell lines and pancreatic cancer tissues. Our data document that CXCR4 is expressed by all pancreatic cancer cell lines and overexpressed in pancreatic cancer tissues as compared to corresponding normal pancreas. Furthermore, treatment with CXCL12 induced the growth of pancreatic cancer cells, indicating the functionality of the CXCL12/CXCR4 signaling axis and suggesting its potential suitability as a therapeutic target. Interestingly, pancreatic cancer cells treated with CXCL12 showed reduced cytotoxicity to the cytotoxic drug 5-fluoro-uracil (5-FU) as compared to cells treated with drug alone. Treatment with CXCR4 antagonist, AMD3100, arrested the CXCL12-induced pancreatic cancer cell growth, and resulted in a cooperative growth inhibitory effect with 5-FU treatment. Taken together, our findings suggest that targeting of CXCL12/CXCR4 signaling axis can be exploited as a novel and effective approach for pancreatic cancer therapy, alone or in combination with cytotoxic chemotherapeutic agents. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5378.

Nonpeptidic Small-Molecule Inhibitor of Bcl-2 and Bcl-XL, (???)-Gossypol, Enhances Biological Effect of Genistein Against BxPC-3 Human Pancreatic Cancer Cell Line

In pancreatic cancer, several important survival molecules such as EGFR, NF-kappaB, and Bcl-2 or Bcl-XL are highly activated. Thus, agents that inhibit NF-kappaB activation, together with agents that directly inhibit Bcl-2 or Bcl-XL protein function, may lead to enhanced cell killing. (-)-Gossypol, a natural polyphenolic compound isolated from cottonseeds, is a dual and potent small-molecule inhibitor of Bcl-2 and Bcl-XL proteins, with a Ki value in the 300-600 nM range for both proteins. : The BxPC-3 human pancreatic cell line was used in this study. (-)-Gossypol was dissolved in DMSO at 20 mmol/L as stock solution, and genistein was dissolved in 0.1 M Na2CO3 to make a 10 mM stock solution. For cell viability, apoptosis, and NF-kappaB studies, MTT assay, histone/DNA ELISA, and Electrophoretic Mobility Shift Assay (EMSA) were used, respectively. Coimmunoprecipitation experiments were designed to study Bcl-XL/Bim heterodimerization, and Western blots to study cytochrome c release. (-)-Gossypol showed a concentration-dependent growth inhibition effect against BxPC-3 pancreatic cancer cell line and induced apoptosis with no effect on normal peripheral blood lymphocytes. Results from coimmunoprecipitation experiments indicate that the effect of (-)-gossypol is mediated, at least, in part via disrupting the heterodimerization of Bcl-XL with Bim in BxPC-3 pancreatic cancer cells. (-)-Gossypol completely disrupts Bcl-XL/Bim heterodimerization with no change in the total Bcl-XL or Bim protein, indicating that (-)-gossypol treatment does not affect the levels of Bcl-XL and Bim proteins. We have previously shown that genistein, a prominent soy isoflavone, transcriptionally down-regulates Bcl-2, Bcl-XL, VEGF, MMP-9, and uPAR via inhibiting NF-kappaB activity. In this study, genistein down-regulated NF-kappaB DNA binding activity and inhibited the growth of BxPC-3 pancreatic cancer cells. In addition, the combination of (-)-gossypol with genistein showed significantly greater growth inhibition compared with either agent alone. From these results, we conclude that inhibition of NF-kappaB activity and direct inhibition of Bcl-2 or Bcl-XL function should serve as a novel strategy for pancreatic cancer therapy.