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Review
Recent advances in pancreatic cancer: biology, treatment, and prevention
Divya Singh a, Ghanshyam Upadhyay a,⁎, Rakesh K. Srivastava b,⁎, Sharmila Shankar b,c,⁎⁎a Department of Biology, City College of New York, 160 Convent Avenue, New York, NY 10031, USAb Kansas City VA Medical Center, 4801 Linwood Boulevard, Kansas City, MO 64128, USAc Department of Pathology, School of Medicine, University of Missouri-Kansas City, Kansas City, MO 64108, USA
Abbreviations:Oct4,octamer-bindingtranscriptionfactFrizzled-9, frizzled class receptor 9; Glut1, glucose tranmyelocytomatosis viral oncogene homolog; FGF, fibroblafamilyBmember1;MDR1,multidrugresistanceprotein1;Dlocus 4; STAT3, signal transducer and activator of transcripgrowth factor receptor; PDGF, platelet-derived growthtransforminggrowth factorbeta; Chk2, checkpoint kinase2oxiainduciblefactor1α;MMP,matrixmetalloproteinase,Tlarge,Bad,Bcl-2-associateddeathpromoter;Bak,Bcl-2homcreatic and duodenal homeobox 1, uPA, urokinase-type pforkhead box O1; FoxO3, forkhead box O3; PI3K, phosphalpha-type platelet-derived growth factor receptor; IGF2forkhead box O1; AFX, forkhead box O4; TP53, tumor prooncogene serine/threonine-protein kinase;Her-2, humanhesionmolecule;vWF,vonWillebrandfactor;PCNA,prolifePdk1, phosphoinositide-dependent kinase-1; mTOR, mdeacetylases; p38, P38mitogen-activated protein kinases;⁎ Corresponding author.⁎⁎ Correspondence to: S. Shankar, Kansas City VA Medic
Pancreatic cancer (PC) is the fourth leading cause of cancer-related death in United States. Efforts have beenmade towards the development of the viable solution for its treatment with constrained accomplishmentbecause of its complex biology. It is well established that pancreatic cancer stem cells (CSCs), albeit present ina little count, contribute incredibly to PC initiation, progression, and metastasis. Customary chemo andradiotherapeutic alternatives, however, expands general survival, the related side effects are the significantconcern. Amid the most recent decade, our insight about molecular and cellular pathways involved in PC androle of CSCs in its progression has increased enormously. Presently the focus is to target CSCs. The herbal productshave gained much consideration recently as they, usually, sensitize CSCs to chemotherapy and target molecularsignaling involved in various tumors including PC. Some planned studies have indicated promising resultsproposing that examinations in this course have a lot to offer for the treatment of PC. Although preclinical studiesuncovered the importance of herbal products in attenuating pancreatic carcinoma, limited studies have beenconducted to evaluate their role in clinics. The present review provides a new insight to recent advances inpancreatic cancer biology, treatment and current status of herbal products in its anticipation.
The burden of pancreatic cancer (PC) has continuously increasedworldwide. It is a serious health concern and fourth leading causeof cancer-related death in United States of America [1,2]. PC is describedas a type of gastrointestinal tumor with a poor anticipation and a highlevel of danger and death rate [3]. More than 90% of pancreatic tumorshave inception from the ductal epithelium of pancreas consequentlytermed as pancreatic ductal adenocarcinoma (PDAC). It is disturbingto see that frequency rate of the pancreatic tumor is relentlesslyexpanding in the western world [4]. The danger components for pan-creatic cancer incorporate smoking, obesity and high utilization ofprocessed meat. Age is positively correlated with pancreatic cancerincidences, and the larger part of cases are diagnosed over the ageof 60 [5]. The introductory indications of patients with PDAC areback agony and dyspepsia with additional disturbing manifestationslike the new onset of diabetes, jaundice, unconstrained profoundvein thrombosis and weight reduction. When one begins perceiving,the tumor typically, spreads to the encompassing tissues or distantorgans. For the tumors spotted in the head region of pancreas, thedetermination is actually productive and they are diagnosed rela-tively early because of biliary impediment. Nonetheless, the tumorsin the body and tail of pancreas regularly stay asymptomatic untillate in disease stage. Most of the patients (~80%) are identifiedwith unresectable locally advanced or metastatic stage and themajor cause is the delayed diagnosis and lack of specific bloodor urine biomarkers to identify patients with increased risk of devel-oping pancreatic cancer [6–9]. The routine diagnostics incorporatetransabdominal ultrasound in the introductory assessment of thejaundiced patient alongside computed tomography (CT) scan ormagnet-ic resonance imaging (MRI).
Despite the fact that the survival rate for most cancers has beenincreased lately in a couple of decades, little change is seen in the caseof pancreatic cancer. The usual survival rate for pancreatic cancerpatients is under six months, and just 3% patients survive over 5-years[6–9]. The reason is attributed to various factors including silent naturein early stages, aggressive tumor biology, the low scope of surgicalmanagement, and lack of effective systemic therapies. Although, thecurrent procedures including surgery, chemotherapy, radiation, and im-munosuppressants, have made great advances in diminishing tumorfrequencies and death rates, pancreatic cancer remains a continuingchallenge to the researchers. The treatment strategies at present utilizedare not very encouraging [10]. There are exceptionally poor post-surgery survival rates even when the pancreatic tumor is surgicallyresected. Safety concerns related with these medications/techniquesare likewise a significant issue for their accomplishment in the treatment
of the disease [6–9]. The prevalent chemotherapeutic choices for thecancer treatment prolong the life of pancreatic cancer patientsminimally,and the survival span in a large portion of the cases is not over one year.Since limited treatment choices are accessible, and it additionally showsresistance against chemo- and radiotherapies, it is important to findnovel and viable methodologies for the treatment of pancreatic cancer[10].
Although the potential use of herbal components for the protectionagainst various cancers began several decades ago, studies to under-stand the mechanism of their action at biochemical, genomic, and pro-teomic levels started very recently. Many plant products, such astriterpenes, flavonoids or polyphenols, are now established potent che-mopreventive agents [11–15]. The phenolic substances are isolatedfrom the wide range of vascular plants and have the ability to reduceand scavenge free radicals [16,17]. Epidemiological studies haveshown the reduced risk of pancreatic cancer by increased consumptionof fruits and vegetables [18]. In the recent past, a number of preclinicalstudies have demonstrated various degrees of the efficacy of herbalproducts both in vitro and in vivo [18]. Certain dietary agents, for exam-ple, resveratrol and curcumin, have been demonstrated to potentiatethe standard chemotherapy [18]. It has been observed that herbal prod-ucts target different pathways simultaneously therefore any solution in-cluding these products may be a smart thought for better results. Manygroups are working in this direction, and the outcomes are promisingtowards the improvement of new helpful cure. In this review, we willexamine the biology of the pancreatic tumor, diagnosis, treatment tech-niques and clinical trials. We will likewise concentrate on the plausiblerole of herbal products, alone or in combination with systemic chemo-preventive medications, in the treatment of the pancreatic tumor.
2. Biology of pancreatic cancer
The biology of pancreatic cancer is perplexing and inadequatelycaught on. Pancreas has both exocrine and endocrine cells that canstructure tumors; however, the likelihood is more for exocrine cells.The vast majority of the exocrine tumors are adenocarcinomas thatbegin in organ cells in the ductal epithelium and advances from prema-lignant injuries to the entirely invasive tumor. Tumors of the endocrinepancreas, commonly termed as islet cell tumors or neuroendocrinetumors, are less common and can be characterized into gastrinomas,insulinomas, glucagonomas, somatostatinomas, VIPomas, PPomas andso forth.
Themicroenvironment of the pancreatic tumor is made out of a fewcomponents, for example, pancreatic cancer cells, pancreatic cancerstem cells (pancreatic CSCs), and the thick, ineffectively vascularizedstroma. The studies suggest that the stroma likewise regulate the
Fig. 1.Generation of cancer cells by various cell types. Somatic cells can be converted to progenitor cells which give rise to various types of differentiated cells. Cancer stem cells, progenitorcells, and cells with oncogenic mutations may lead to formation of tumor mass.
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pancreatic tumor growth and development, intrusion and metastasisseparated from its movement as the mechanical boundary. These stro-mal cells consistently interface with cancer cells by different autocrineand paracrine secretion of the pancreas, for example, platelet-derivedgrowth factor (PDGF), transforming growth factor β (TGF-β) andcytokines [19]. These development variables fortify a critical segmentof stroma called pancreatic stellate cells, which in turn express α–smooth-muscle actin and produce rich collagen fibers. These fibersadd to tumor hypoxia (a primary stimulator of tumor movement andmetastasis by influencing angiogenesis), cell survival, and apoptoticpathways [19]. Hypoxia is likewise proposed to be a major cause fordrug/therapy resistance in different tumors [20,21].
Pancreatic tumor cells apparently grow around a population of can-cer stem cells (CSCs) that have the capability of self-renewal andmulti-lineage differentiation [22] (Fig. 1). Pancreatic CSCs can be isolated byflow cytometry utilizing CD44, CD24, and ESA as surface markers [23].Although CD44+/CD24+/ESA+ cells constitute only 0.2-0.8% of thetotal cell population, they are capable of forming tumor spheres [24].Other markers for pancreatic CSCs are CD133, CXCR4, c-Met andALDH1 [25–28]. CXCR4 assumes an essential part in the tumor invasionand metastasis and the cells positive for both CD133 and CXCR4, showhigher metastatic potential than other populations [26,29]. In a recentstudy, it has been demonstrated that human PDACs contain CSCs withhigh levels of CXCR4 and ABCB1, and such patients had reduced survivalrate [30]. Recently Shankar et al. effectively demonstrated the tumori-genic potential of pancreatic CSCs isolated from human pancreatictumors in NOD/SCID mice utilizing surface markers CD44, ESA, CD133,and CD24 [10]. These cells were highly tumorigenic and were likewiseexpressing ALDH and pluripotency maintaining factor, Oct-4 [10].They further indicated high expression of CD133, CD24, CD44, ESA,Nanog, Notch1, MDR1 and ABCG2 in these CSCs (CD133+CD44+CD24+
ESA+ cell population) contrasted with CD133−CD44−CD24−ESA− cellpopulation [10].
Various elements regulate the properties and conduct of pancreaticCSCs, for example, nestin can balance attack or metastasis of pancreatic
CSCs, Oct-4 andNanog can direct pancreatic CSC's conduct andmetastasisto different organs, DCLK1 can segregate between normal and tumoralstem cells and Sox2 controls cell proliferation and differentiation [31].Furthermore, c-Kit and Kras likewise tweak the movement of pancreaticadenocarcinoma [32]. Epithelial tomesenchymal transition (EMT), a pro-cess of change of epithelial attributes into mesenchymal properties, is anurgent process for tumor progression and is proposed to be in charge forthe appearance of cells with stem cell-like properties [33].
CSCs are thought to be themajor contributory element to the absenceof effective treatment for pancreatic malignancy and are in charge of tu-morigenesis, metastasis, and development of chemo and radioresistance[24]. A recent study exhibited that PANC-1 cancer cell line, steadily over-expressing Oct4 and Nanog, show chemoresistance, multiplication, relo-cation, intrusion, and tumorigenesis in vitro and in vivo [34]. Moreover,the ALDH+CD44+CD24+cell population iswell reported to be impervi-ous to treatment with gemcitabine [35]. Removal of CSCs is essentialfor robust tumor treatment, as CSCs stay untouched. Hence, drugs thatcan specifically target CSCs could be a superior alternative for pancreaticcancer prevention.
Pancreatic cancer is hereditarily complex and heterogeneous innature. Different malady conditions, for example, pancreatitis, cysticfibrosis, and inflammation have their effect on the initiation of pancre-atic cancer and its malignant progression [36,37]. Mutations in fourcritical genes namely Kras2, Cdkn2a, TP53 and Dpc4 (or Smad4) are fre-quently seen in pancreatic malignancy patients. The vastmajority of thecancer patients carry one ormore of thesemutations [38]. The observedfrequency of Kras mutation is more than 90%, however, the rate of inac-tivation mutation of Cdkn2a, TP53, and Dpc4 are 95%, 75% and 50%respectively [39,40]. The mutation in Kras2 brings about the consistentexpression of irregular Ras protein that causes aberrant activation of cellproliferation and survival pathways [19]. With increasing age, the like-lihood of acquiring activating mutations in Kras2 gene increases in themajor organs such as lung, pancreas, colon, and other tissues. On theother hand Cdkn2a, TP53 and Dpc4 are the tumor suppressors and mu-tations in these genes result in their inactivation, which facilitate the
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proliferation and survival signaling [19,41]. Moreover, a recent studycalled attention to the loss of functionmutation in SWI/SNF nucleosomeremodeling complex in 23% pancreatic adenocarcinomas [42].
3. Signaling pathways in pancreatic cancer
Genetic mutations serve as the basis for abbrent signaling pathways.A comprehensive study with more than 24 pancreatic cancer cases, anaverage of 63 relevent genetic abnormalities (mainly point mutations)per tumor were classified as likely to be relevent in its pathogenesis[43] (reviewed in [19]). Thesemutations can be clubbed in 12 notewor-thy signaling pathways including STAT3, Smad/TGF-β, Wnt, Notch,PI3K/Akt, sonic hedgehog and so forth [43] (reviewed in [19]). Aberrantsignaling in these cellular events has been implicated in the develop-ment and progression of pancreatic tumors by permitting increasedproliferation, angiogenesis, survival, and metastasis (Fig. 2).
4. STAT3 pathway
Signal transducer and activator of transcription 3 (STAT3), encodedby STAT3 gene, is activated in a wide variety of signaling pathways.It mediates diverse responses, including transmission of the signals ofcytokines and growth factors from the cell membrane to the nucleusto regulate gene expression for cell development, differentiation, prolif-eration, survival, and angiogenesis [44–46] (Fig. 3). A range of cytokinesincluding interleukin (IL)-6, IL-9, IL-10, IL-27, tumor necrosis factor α(TNF-α), and monocyte chemotactic protein-1 (MCP-1) activate theSTAT3 pathway [47–49]. Various growth factors, for example, epidermalgrowth factor (EGF), platelet-derived growth factor (PDGF), granulocytecolony-stimulating factor (G-CSF), and granulocyte-macrophage colony-stimulating factor (GM-CSF) also activate this pathway [44–49]. Primaryfunctions of activated STAT3pathway incorporate cell proliferationbyup-regulating cyclin D1 and cyclin B1 and inhibition of apoptosis by up-regulating Bcl-2, Bcl-xL, Mcl-1. Initiated STAT3 assumes a discriminatingpart in tumorigenesis by controlling angiogenesis (VEGF, FGF, andHIF1α), invasion and metastasis (MMP2, MMP9, TWIST, and ICAM1)[44–49].
STAT3 activation has been detected in diverse type of malignancies,and its inhibition by use of inhibitors or short interfering RNA has
Fig. 2.Molecular targets of cancer preventive agents. Cancer preventive agents regulates varioumodulating the expression of various genes.
prompted to reverse the malignant phenotype. STAT3 activation hasbeen portrayed in almost 70% of solid and hematological malignancies[50]. Studies in conditional knockout mice have demonstrated thatSTAT3 pathway is latent in typical pancreas and is not needed for anyvital process related to pancreatic development and homeostasis [51].Nevertheless, it is constitutively activated in PDAC by phosphorylationof Tyr705 in human tumor specimens and also in various PDAC celllines [52–55]. Further, STAT3 is necessary for the development of ADMprocess (acinar-to-ductal metaplasia), which is an early event in PDACpathogenesis mediated by ectopic expression of the Pdx1 [54]. It is re-markable that Pdx1 is a transcription factior and a key regulator ofearly pancreatic development [54]. Another study showed that withmalignant transformation, activated STAT3 promotes proliferation ofcells by regulating G1/S-phase progression and supports the malignantphenotype of human pancreatic cancer [52]. IL-6 signaling dependentactivation of STAT3 plays an important role in promoting PanIN pro-gression and thePDACdevelopment, in addition to oncogenic KrasG12Dtransformation [56]. Themyeloid cells in the pancreas induce STAT3 ac-tivation by releasing IL-6, which promote PanIN progression and thePDAC development.
5. Smad/TGF-β pathway
The transforming growth factor beta (TGF-β) signalingpathway reg-ulates various cellular processes like cell growth, cell differentiation, ap-optosis and cellular homeostasis in both the adult organism and thedeveloping embryo [57]. TGF-β signaling occurs from the membraneto the nucleus via Smad proteins [58]. Smads can be classified into 3major groups; receptor-regulated Smads (R-Smad; Smad1, Smad2,Smad3, Smad5 and Smad8/9), common-mediator Smad (co-Smad;Smad4) and agonistic or inhibitory Smads (I-Smad; Smad6 andSmad7) [58]. Cascade is triggered by the binding of TGF-β superfamilyligand to a type II receptor which catalyzes the recruitment and phos-phorylation of a type I receptor. Subsequently R-Smads are phosphory-lated, linkedwith the coSmad andother factors, andfinally accumulatedin the nucleus to regulate the target gene expression. TGF-β receptor ac-tivation results in Smad2 and Smad3 phosphorylation, which then formheteromeric complexes with Smad4. Furthermore, Smad6 and Smad7,
s cell signaling pathways, cell cycle, cell viability, apoptosis, angiogenesis andmetastasis by
Fig. 3. STAT3 signaling pathway. Activation of STAT3 signaling pathway regulates invasion and metastasis, angiogenesis, cell metabolism, inflammation, cell survival and proliferationthrough modulation of several genes.
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can prevent TGF-β signaling by interacting either with the receptor orwith Smad2 and Smad3 [58].
TGF-β signaling pathway impairment because of inactivated Smad4(DPC4) is often recognized in pancreatic carcinomas [59,60]. Jonsonet al., investigated a series of pancreatic carcinoma cell lines with re-spect to alterations of five Smad genes involved in TGF-β signaling,and demonstrated the structural rearrangement of Smad4 in 42% ofthese tumor cells [59]. Since, this pathway could likewise be influencedby other factors that regulate the activation of TGF-β and its receptorgenes, they further assessed expression of uPA, uPAR, IGF2R, TGF-βR1-3,ENG, ALK1, TGF-β1-3, mutations of TGF-βR1-2, cell surface localizationof TGF-βR2 and ENG, and TGF-β1 response in 14 pancreatic carcinomacell lines [59]. The study suggested ALK5- Smad4 as a major target for in-activation in pancreatic carcinomas and that the expression of TGF-βR2,TGF-βR3, and receptors involved in TGF-β activation aremaintained [59].
6. Wnt pathway
Wnt signaling pathway is a complex process and plays an importantrole in tumor development apart from its involvement in other physio-logical and pathological processes [61]. In canonical pathway, the ligandbinding to its receptor (Frizzled/LRP receptor complexes) triggers a cas-cade of events that prevents β-catenin degradation inside the cytoplasmand permits its stabilization and translocation in nucleus where it bindsto transcriptional factors of the Tcf/Lef family forming an activator com-plex. AbnormalWnt/β-catenin signaling is reported in pancreatic cancer[62]. Activated Wnt signal leads to the accumulation of β-catenin in thenucleus where it activates specific target genes [63]. The accumulationof β-catenin is observed both in nucleus and cytoplasm in pancreaticcancer [61,64–66]. The functional evidences are also accumulating thatimplicate a supporting role for β-catenin in PDACmaintenance and pro-gression [61]. It has been also found that β-catenin accumulation and
signaling could be increased through paracrine signaling taking placein the PDAC micro-environment [61]. In a recent investigation, it hasbeen found that Wnt/β-catenin signaling inhibition by wnt-c59 re-sults in reversal of TSA sensitivity, migration ability, and the EMTphenotype in trichostatin A-resistant Panc-1 cells (Panc-1/TSA) [67].
In spite of the fact that our understanding of the role of this pathwayhas increased during the last decade, the general mechanism by whichβ-catenin accumulation occurs in PDAC is poorly understood and needsfurther elucidation.
7. Notch pathway
Notch signaling is shown to regulate proliferation and apoptosisevents in various cell types. The alterations in Notch signaling have vari-ous consequences including tumorigenesis [68,69]. In mammals, fourNotch receptors (Notch1-4) and five ligands (Jagged1, Jagged2, Delta-like 1(Dll-1), Dll-3, and Dll-4) have been accounted for to date [68,70].Binding of Notch ligand to an adjacent Notch receptor activates Notch sig-naling prompting the cleavage of Notch through a cascade of proteolyticcleavages by themetalloprotease, tumor necrosis factor-α-converting en-zyme (TACE) and γ-secretase complexes [68,71]. The cleavage by TACEgenerates Notch extracellular truncation (NEXT) which is subsequentlycleaved by the γ-secretase complex releasing the active fragment Notchintracellular domain (NICD) from the plasma membrane. NICD translo-cates into the nucleus where it binds tomembers of the CSL transcriptionfactor family and activates Notch target genes [68,72].
Impaired Notch signaling is well reported in pancreatic cancers.Recently, it has been reported that inhibition of Notch signalingpathway by Notch1 siRNA or gamma-secretase inhibitors, such as,MRK-003, MK-0752 etc., enhances chemosensitivity to gemcitabinein pancreatic cancer cells through activating apoptosis [73]. Abelet al. reported the significance of Notch pathway in maintaining
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cancer stem cell population in pancreatic cancer and investigated theconnection of Notch pathway and percentage of the CSCs population.The inhibition of this pathway resulted in a reduced percentage ofCSCs and tumor sphere formation; notwithstanding, activation dem-onstrated the inverse impact [74]. Further, Lee et al. suggested thatthe activation of the Notch pathway and the increase in CSCs mightcontribute to the failure of treatment in pancreatic cancer [75].Notch has also been shown to be associated with the EMT in pancre-atic cancer [72,76]. It is remarkable that during the EMT process,epithelial cells gain the mesenchymal over endothelial characteristicthereby increasing in migratory and invasive capacity, leading to in-vasion and metastasis [77,78].
8. PI3K/Akt pathway
PI3K pathway acts via phosphorylation of FoxO proteins via Akt,which in turn impairs the DNA-binding ability and increases its affinityfor 14-3-3 proteins [79]. These complexes are exported from the nucle-us to cytoplasm leading to the inhibition of FoxO-mediated survivalpathways. Some other downstream effectors that regulate cell cyclearrest and apoptosis, such as, active FKHRL1, FKHR, and AFX are alsotranslocated to the cytoplasm by similar mechanism [79]. Downstreameffectors of the PI3K-Akt pathway are actively involved in a variety ofvital and specialized functions such as differentiation and proliferationin diversified cells including adipocytes, hepatocytes, myoblasts, thy-mocytes and cancer cells [79].
PI3K/Akt pathway is activated in a variety of cells including fibro-blastic, epithelial, and neuronal cells as survival signal. An elevatedlevel of Akt has been reported in many types of tumors. Studies haveshown the requirement of activated PI3K-Akt/FoxO signaling for thegrowth and survival of the pancreatic tumor. It has been found thatthe cells with elevated Akt levels are less sensitive to apoptosis stimuli.Akt regulates apoptosis directly by regulating its primary targets, Bad,and Caspase 9 and indirectly by controlling human telomerase reversetranscriptase subunit, FoxOs and IkappaB kinases and so forth [79].
9. Sonic Hedgehog pathway
Sonic hedgehog (Shh) is a member of the Hedgehog (Hh) family ofsecreted signaling proteins. Shh signaling is triggered by binding of thesecreted Shh peptide to Patched (Ptch), which leads to inhibition ofPtch activity. Consequently, Smoothened (Smo) gets phosphorylatedresulting in the activation of the Gli family of zinc-finger transcription fac-tors and therefore target gene expression [80]. Shh signaling has diversefunctions during vertebrate development and post-embryonically in tis-sue homeostasis [81,82]. Alteration in this pathway have been linked tovarious tumor types including pancreatic cancer [81,82]. Activation ofShh signaling pathway has been reported to be involved in the regulationof the pancreatic CSC's expansion,whereas its inhibition (by impairing Glibinding to its promoters) has been demonstrated to upregulate DRs andFas expression, curb Bcl-2 and PDGFRα expressions, and encourage apo-ptotic cell death in pancreatic CSCs [24]. Further, its inhibition has beenshown to reduce tumor-associated stromal tissues, enhance gemcitabineuptake in tumor cells and prolong the average survival rate in pancreaticcancer mouse model [83]. Recently, Rodova et al. showed that inhibitionof Shh pathway components, Gli transcriptional activity and its down-stream targets by sulforaphane inhibited human pancreatic CSCs derivedspheres and induced apoptosis by inhibition of Bcl-2 and activation ofcaspases in vitro [84]. Further, Li et al. showed that inhibition of Shh path-way by sulforaphane results in a marked reduction in EMT, metastatic,angiogenic markers with significant inhibition of tumor growth in mice.Since aberrant Shh signaling is frequently observed in pancreatic cancers,therapeutics that target Shh pathway and therefore CSCs, may improvethe outcomes of patients with this devastating disease [85].
10. Treatment of pancreatic cancer: chemotherapy and radiotherapy
The essential choice for pancreatic cancer treatment is its surgicalremoval. However, in advance metastatic stages, the treatment mainlyaims to increase the survival by optimal control of metastases. Systemicchemotherapy may be utilized at any phase of pancreatic malignancywith the objective to minimize the patient's disease-related symptomsand to prolong survival. Presently, a limited number of drugs are acces-sible for the treatment of the pancreatic tumor. Gemcitabine has beenthe reference regimen since 1997 when it was indicated better than 5-Fluorouracil (5-FU) in a phase III clinical trial [86]. A significant concernwith 5-FU was the associated toxicity with its treatment, in particular,gastrointestinal toxicity. A combination of 5-FU and Fluoropyrimidine(S-1) could likewise be a decent choice as S-1 potentiates the antitumoractivity of 5-FU and decreases gastrointestinal toxicity in pancreatictumor mouse models [87]. In recent years different combinations, forexample, the systemic treatment with Abraxane and Gemcitabineand the multidrug combination, FOLFIRINOX, have been attempted[88–109]. Despite the fact that FOLFIRINOX has been promising, it canhave a remarkable reaction profile, constraining its utility in patientswith poor baseline performance status [110].
Radiation therapy, combined with chemotherapy, may be used inpatients whose cancers have grown beyond the pancreas and cannotbe removed by surgery. Uses of radiation treatment alongside high-energy x-beams to kill cancer cells is extremely regular for the treat-ment of advanced stages of tumors. Pancreatic neuroendocrine tumors(NETs) usually do not respond to radiation, and therefore it is rarelyused to treat these tumors. However, it can be used in case of pancreaticNETs that have spread to the bone and other tissues. Furthermore, thetypical radiation treatment utilized for the treatment of the exocrinepancreatic malignancies is External Shaft Radiation Treatment that fo-cuses the radiation on the cancer from a machine outside the body. Ra-diation treatment is typically associated with different side effects, likeskin changes in areas getting radiation, nausea and vomiting, diarrhea,fatigue, poor appetite, weight loss, lower blood counts and last but notthe least increased risk of severe infection.
11. Clinical trials in pancreatic cancer
The chemotherapy drug, Gemcitabine, has been a standard initialtreatment for patients with metastatic pancreatic cancer for over 15years [86]. Various clinical trials have tried new medications, eitheralone or in combination with Gemcitabine (Table 1); however, the ad-vancement is moderate amid the most recent decade [88–109,111].Gemcitabine alone or in combination with Capecitabine or Erlotinibremained the favored systemic treatment alternatives until 2010[88–109]. Since 2010, use of FOLFIRINOX has increased both inmetasta-tic and locally advanced cancer [110,111]. Various drugs and combina-tions have been evaluated in the last couple of years, and some ofthem have shown promising results. Sunitinib and Everolimus haveshown significant improvement in survival. Sunitinib treatmentshowed median progression-free survival of 11.4 months as comparedwith 5.5 months for patients who received the placebo. On the otherhand, the patients receiving Everolimus showed median progression-free survival of 11 months as compared with 4.6 months for patientswho received the placebo. Although these drugs have shown prom-ising results, severe side effects as anemia and neutropenia werealso observed.
Combination of Nab-Paclitaxel (Abraxane®), a form of the che-motherapy drug Paclitaxel bound to the human protein albuminand contained in nanoparticles, and Gemcitabine (Gemzar®) wasalso investigated in an international randomized phase III trial show-ing improved survival [112]. Patients who received the drug combi-nation had a median overall survival of 8.5 months, compared with6.7 months for patients treated with gemcitabine alone [112]. FDAapproved therapy to treat patients with metastatic pancreatic cancer
Table 1List of some of the completed clinical trials on pancreatic cancer (Source: www. ClinicalTrials.gov).
Rank Interventions NCT Number Enrollment Conditions
Cancer|Stage IV Pancreatic Cancer8 Drug: bortezomib|Drug: carboplatin|Other: laboratory biomarker
analysisNCT00416793 9 Acinar Cell Adenocarcinoma of the Pancreas|Duct Cell
Adenocarcinoma of the Pancreas|Stage IV Pancreatic Cancer9 Drug: bevacizumab [Avastin] NCT01214720 607 Pancreatic Cancer10 Drug: Erlotinib, escalating dose|Drug: Erlotinib, standard
dose|Drug: GemcitabineNCT00652366 467 Pancreatic Cancer
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based on the results of the MPACT trial. Some other combinations,such as, gemcitabine with gamma-secretase inhibitor (MK-0752)or FG-3019 (a human monoclonal antibody that suppresses connec-tive tissue growth factor), have also shown promising results [113].
12. Herbal products, cancer prevention, and pancreatic cancer
The relation between the consumption of certain dietary herbals anda reduced risk of cancer is becoming evident as many epidemiologicaland pre-clinical studies have shown the effect of herbals on health[114]. In the past few years, the cancer chemoprevention approach is
directed towards polyphenols and their health-related properties anda wide range of dietary constituents show potential biological activities[114]. Studies in this line on cell/animal models and human epidemio-logical trials have shown the potential of dietary polyphenols as anti-carcinogenic agents. The reports have shown that phenolic compoundshave the capability to inhibit the molecular events in the cancer initia-tion, promotion, and progression stages. They may increase the expres-sion of pro-apoptotic components in initiated proliferating cells andthereby prevent or delay tumor development. Although it seems thatphenolic compounds induce apoptosis in a precise manner in cancercells but in some human studies no promising results were obtained.
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A Cohort Study of Diet and Cancer in Netherlands suggested no effect ofthe consumption of black tea on the risk for colorectal, stomach, lungand breast cancers [115]. A similar studywasperformed in Japan involv-ing more than 25,000 stomach cancer patients with a similarobservation i.e. no association of consumption of green tea with gastriccancer risk [116]. Someother studies ondifferent types of cancer also in-dicated the same conclusion [117–119]. On the contrary, a decreasedrisk for the different types of cancer has been reported after the con-sumption of flavonoids or certain foods or drinks rich in these phenoliccompounds [118–122].
The use of herbals in the treatment of pancreatic cancer is a novelapproach and is continuously gaining the attention of investigators.Previous research in this area was focused on inducing apoptosis butrecently these herbals have beenused in the targeting of other key path-ways of cell survival, angiogenesis, metastasis, and differentiation. Easyavailability and less or no toxicity even at higher doses have made themthe preferable choice over other cancer chemopreventive options (Fig. 2).
Table 3Summary of recent findings showing chemopreventive potential of curcumin againstpancreatic cancer.
Cells/animals Effect Reference
MIA PaCa-2 and mouse (tumorxenograft model)
Inhibits the proliferation andenhances apoptosis in MIA PaCa-2cells and inhibits tumor growthand the expression of thetranscription nuclear factor NF-κBand NF-κB-regulated geneproducts in xenograft mousemodel
[220]
MiaPaCa-2 and Panc-1 cells Down-regulates the expression ofmiR-221 resulting in upregulationof PTEN, p27(kip1), p57(kip2),and PUMA leading to the
[221]
13. Resveratrol
Resveratrol, a phytoalexin, is commonly found as an ingredient in redwine, skins of grapes, peanuts and so forth. It possess anti-tumorigenic,anti-inflammatory, and anti-oxidant properties [123]. Its preventive roleagainst cancers, cardiovascular diseases, and various neurological disor-ders has been widely reported [124–126]. Hydroxylation of resveratrolby CYP1B1 generates two major metabolites namely piceatannol and3,4,5,4′-tetrahydroxystilbene that substantially contribute to its chemo-preventive activities by inhibiting tyrosine kinase and inducing apoptosis[127–129].
Chemopreventive effects of resveratrol against various cancers havebeen extensively investigated in both in vitro and in vivo. It has beenshown to have anti-tumor activity by inhibiting angiogenesis, endothe-lial cell migration, tumor formation and by blockage of oxygen free rad-ical formation [130–132] (Table 2). Due to its lipophilic nature, it readilycrosses the plasmamembrane and establishes dynamic homeostasis byinhibiting the phase I (mainly CYP450s) and inducing phase II enzymes(UDP-glucuronosyl transferase, NAD(P)H quinone oxidoreductase,and glutathione-s-transferases) during stress conditions [133–138]. Incancerous cells, it inhibits the expression of inducible nitric oxide (NO)synthase and NO production [139]. It also inhibits the formation of apreneoplastic lesion inmousemammary glands and proliferation of a va-riety of cancer cells in culture including, human colon, breast, andprostatecancer cells [140–143].
Resveratrol sensitizes a broad spectrum of tumors including lungcarcinoma, acute myeloid leukemia, promyelocytic leukemia, multiple
Table 2Summary of recent findings showing chemopreventive potential of resveratrol againstpancreatic cancer.
Cells Effect Reference
MIA PaCa-2 cells Inhibits proliferation and induces apoptosis [213]Panc-28 and Hs766Tcells
Increases calcium levels and preventsmigration of TG2-expressing cells
myeloma, prostate cancer, oral epidermoid carcinoma, and pancreaticcancer. Zhou et al. showed that it enhances caspase-3 activationand p53 and p21 expression in capan-2 and colo357 pancreatic cancercell lines [144]. A recent approach, using human pancreatic CSCs(CD133+CD44+CD24+ESA+), showed that resveratrol sensitizes andinhibits the growth and development of pancreatic cancer lesion inKrasG12D mice [10]. This study further showed that the resveratrolinhibits pluripotency maintaining factors (Nanog, Sox-2, c-Myc and Oct-4), drug resistance gene ABCG2, CSC's migration, invasion, self-renewal,and components of EMT (Zeb-1, Slug, and Snail) [10]. Resveratrol inhibitscell growth, proliferation and expression of the anti-apoptotic proteinsBcl-2, Bcl-xL, and XIAP and induces apoptosis, cell cycle arrests, caspasesand pro-apoptotic gene Bax in pancreatic cancer cell lines [145]. Addition-ally, resveratrol has been found to suppress proliferation and anchorage-independent growth of pancreatic cancer by inhibiting leukotriene B4(LTB4) production and expression of the LTB4 receptor 1 (LTB4R1)[146]. It is remarkable that LTB4 is a hydrolysis product of the leukotrieneA4 (LTA4), and the process is catalyzed by LTA4hydrolase, a known targetfor prevention and therapy of cancers including PC [147]. Resveratrol candirectly bind to leukotriene A4 hydrolase (LTA-4H) and inhibit its activityand, therefore, LTB4 production [146].
14. Curcumin
Curcumin is one of themost commonly used and highly investigatedphytochemical. During the last decade, our understanding of its thera-peutic potential and themultiplemechanismsbywhich it offers chemo-prevention against various cancers has been increased [148] (Table 3).The various pharmacological effects of curcumin include apoptotic,anti-proliferative, anti-oxidant, and anti-angiogenic properties. The
inhibition of cell proliferation andmigration
MIA PaCa-2 and mouse (tumorxenograft model)
Inhibits cell proliferation, reducestumor growth and angiogenesis asdetermined by a reduced numberof blood vessels and decreasedexpression of vascular endothelialgrowth factor and annexin A2proteins
[222]
BxPC-3 and MiaPaCa-2 Activates TNFR, CASP 8, CASP3,BID, BAX, and down-regulatesNFκB, NDRG 1, and BCL2L10 gene
[223]
TGF-β1-stimulated PANC-1 cells Inhibits proliferation, inducesapoptosis and reverses the EMT
[224]
AsPC-1 and MiaPaCa-2 Decreases pancreatic cancer cellsurvival, clonogenicity, formationof pancreatospheres, invasive cellmigration, and CSC function
[225]
AsPC-1, MiaPaCa-2, Panc-1,BxPC-3 human and Pan02mouse pancreatic cancer cells
Inhibits tumor growth throughmitotic catastrophe by increasingthe expression of RNA bindingprotein CUGBP2, therebyinhibiting the translation of COX-2and VEGF expression
[151]
21D. Singh et al. / Biochimica et Biophysica Acta 1856 (2015) 13–27
previous studies have highlighted that curcumin targets multiple signaltransduction pathways and that it suppresses a number of essentialelements in cellular signaling pathways, for example, phosphorylationcatalyzed by protein kinases, c-Jun-activated protein 1 (AP-1) activationand prostaglandin biosynthesis.
It has also been found that curcumin potentiates radiotherapy in PCcure probably by involving selective regulation of radiotherapy-inducedNF-κB [149]. Studies have shown that curcumin inhibits cell prolifera-tion and induces apoptotic cell death mediated by PARP cleavage andCaspase-3 in MIAPaCa-2, Panc-1 and BxPC-3 pancreatic cancer cells[150]. Subramaniam et al. showed a significant reduction in tumor vol-ume and angiogenesis in curcumin treated tumor xenografts [151].They further showed that curcumin inhibits cell proliferation, inducesof G2-M arrest and apoptosis, enhances phosphorylation of checkpointkinase 2 (Chk2) coupled with higher levels of nuclear cyclin B1 andCdc-2, and increases expression of cyclooxygenase-2 (COX-2) [151].Curcumin also inhibits ERK activity and suppresses EGFR andNotch-1 sig-naling leading to increased apoptosis in pancreatic cancer. Glienke et al.showed that incubation with curcumin results in down-regulation ofWilms' tumor gene 1 (WT1; a gene frequently expressed in pancreaticcancer) in a dose-dependent manner [152]. Additionally, curcumin hasbeen shown to restrain STAT3 and induce apoptosis by inhibiting theexpression of the anti-apoptotic gene Survivin/BIRC4 in pancreatic cancercells [153].
Recently Bar-Sela et al. reviewed the accomplished and continuingclinical trials with curcumin as an anticancer agent [154]. In one trial,17 patients were treated with the oral dose of 8 gm/day of curcuminin combination with Gemcitabine. Although the results showed thatthis combined treatment is tolerable in patients; nevertheless, it hasbeen suggested to reduce the dose of curcumin [155]. Dhillon et al.used only curcumin as the 1st line treatment for the 25 patients. Theyfound that curcumin down regulates the expression of NFκB, COX-2,and the phosphorylation of STAT3 in peripheral blood [156]. In spite ofthese encouraging results, extensive clinical trials are needed beforedrawing any conclusion.
15. Epigallocatechin gallate (EGCG)
Epigallocatechin gallate (EGCG) is a most extensively studied cate-chin and the major polyphenol present in green tea. Various studieshave shown that EGCG offers protection against pancreatic canceramong the other tumors (Table 4); however, the exactmolecularmech-anismbywhich EGCG suppresses humanpancreatic cancer cell prolifer-ation is unclear. Kürbitz et al. showed anticancer properties of EGCG onhuman pancreatic ductal adenocarcinoma (PDAC) cells PancTu-I, Panc1,Panc89 and BxPC3 in vitro [157]. They found that EGCG inhibits prolifer-ation of PDAC cells in a dose- and time-dependent manner. The proteinexpression analysis performed with PancTu-I cells evidently showedEGCG-mediated modulation of cell cycle regulatory proteins (cyclins,
Table 4Summary of recent findings showing chemopreventive potential of EGCG against pancreatic ca
Cells/animals Effect
PANC-1 Suppresses proliferation and induces apopAsPC-1 cells Regulates RKIP/ERK/NF-κB and/or RKIP/NBalb c nude mice (tumor xenograft model) Inhibits pancreatic cancer orthotopic tumo
pathways and activation of FKHRL1/FoxO3Colo357 human pancreatic adenocarcinomacells
With PGHS-2-specific inhibitor celecoxib,down-regulates release of pro-angiogenicmatrix metalloproteinase (MMP)-2
PANC-1 Inhibits HIF-1α protein expression, P-gp mAsPC-1 and PANC-1 Suppresses the growth, invasion, and migr
and enhances the therapeutic potential ofHuman pancreatic cancer stem cells(CD133+/CD44+/CD24+/ESA+)
Inhibits Nanog, c-Myc and Oct-4 expressio(smoothened, patched, Gli1 and Gli2) andXIAP and activating caspase-3
cyclin-dependent kinases, and inhibitors). The study further statedthat EGCG inhibits TNFα-induced activation of NF-κB and consequentlysecretion of pro-inflammatory and invasion-promoting proteins likeIL-8 and uPA [157]. Moreover, previous studies have demonstratedthat EGCG decreases cell adhesion ability on micro-pattern dots, ac-companied by dephosphorylation of both focal adhesion kinase andinsulin-like growth factor-1 receptor (IGF-1R) in AsPC-1 and BxPC-3 cells [158,159,64]. EGCG has been also found to aid retained activa-tion of MAPK signaling, suppressed growth, reduced cell viability,and increased apoptosis in these cells in a dose-dependent manner[158].
Effects of EGCG on heat shock proteins were also investigated. Astudy by Li Y et al. showed that the binding of EGCG to Hsp90 impairsthe association of Hsp90 with its co-chaperones, thereby inducingdegradation of Hsp90 client proteins (Akt, Cdk4, Raf-1, Her-2, andpERK) consequently anti-proliferating effects in pancreatic cancer cells[160]. Basu and Haldar showed that EGCG causes the disappearance ofintact 21 kDa Bid protein and induces activation of caspase-8 leadingto cell death in MIA PaCa-2 cells [161]. Further, involvement of trans-membrane extrinsic signaling in this polyphenol triggered pancreaticcarcinoma cell death was confirmed by RNase protection assay thatclearly showed up-regulation of the members of death receptor family[161,160].
Shankar et al. examined the role of EGCG in inhibiting growth, inva-sion, metastasis and angiogenesis of human pancreatic cancer cells in axenograft model system [162]. They found that EGCG inhibits viability,capillary tube formation, and migration of HUVEC. Additionally, theyobserved EGCG-mediated inhibition of proliferation (Ki-67 and PCNAstaining), angiogenesis (vWF, VEGF and CD31) and metastasis (MMP-2, MMP-7, MMP-9 and MMP-12), and induction of apoptosis (TUNEL),caspase-3 activity and growth arrest (p21/WAF1) in vivo [162]. Theyalso found a significant reduction in the circulating vascular endothelialgrowth factor receptor 2 (VEGF-R2) positive endothelial cells, ERK ac-tivity, and induction of p38 and JNK activities in vivo following EGCGtreatment [162].
16. Genistein
Genistein is found in a number of plants including lupin, fava beans,soybeans, kudzu, and psoralea, in themedicinal plant, Flemingia vestita,and coffee [159,160,163–165]. It has multiple effects in living cells, suchas activation of PPARs, estrogen receptor-β, Nrf2 anti-oxidative re-sponse, stimulation of autophagy and inhibition of several tyrosinekinases, topoisomerase, and mammalian hexose transporter GLUT-1[73,166–174]. Genistein also affects tumor formation, cellmultiplicationand differentiation, angiogenesis, and signaling triggered by growthfactors [175–184]. The most critical activity that contributes to the che-mopreventive potential of genistein is tyrosine kinase inhibition,mostlyof epidermal growth factor receptor EGFR. Additionally, the inhibitory
ncer.
Reference
tosis, modulates the PI3K/Akt/mTOR signaling pathway [226]F-κB/Snail and inhibits invasive metastasis [227]r growth, angiogenesis, and metastasis, inhibits PI3K/Akt and ERKa
[162]
synergistically diminishes metabolic activity via apoptosis induction andvascular endothelial growth factor (VEGF) and invasiveness-promoting
[228]
RNA and protein levels and cell proliferation [229]ation, induces apoptosis by interfering with the STAT3 signaling pathwaygemcitabine and CP690550
[230]
n, self-renewal, proliferation, EMT, components of Shh pathwayGli transcriptional activity, and induces apoptosis by inhibiting Bcl-2 and
[231]
Table 5Summary of recent findings showing chemopreventive potential of sulforaphane againstpancreatic cancer.
Cells/animals Effect Reference
Established BxPc-3 and AsPC-1 PDAcell lines and immortalizedCRL-4023 hTERT-HPNE humanPDA cells
Increases Cx43 and E-cadherinlevels, inhibits c-Met andCD133, improved thefunctional morphology andcommunication of gapjunctions.
[232]
MIA PaCa-2 and Panc-1 Inhibits cell viability and NF-κBDNA binding activity, inducescell apoptosis by activation ofcaspase-3 and PARP cleavage,increases pERK1/2, c-Jun, p38MAPK, p53 protein expressionwhen used in combinationwith aspirin and curcumin
[233]
NOD/SCID/IL2Rgamma mice (tumorxenograft model)
Inhibits growth of tumors,expression of Shh pathwaycomponents, EMT,pluripotency maintainingtranscription factors,angiogenic markers andinduces apoptosis
[85]
Human normal pancreatic stem cells(HPSC) and human pancreaticcancer stem cells(CD133+/CD44+/CD24+/ESA+)
Inhibits CSC's derived spheres,components of Shh pathwayand Gli transcriptional activity,expression of pluripotencymaintaining factors (Nanogand Oct-4) as well as PDGFRαand Cyclin D1
[84]
Pancreatic cancer xenograft mousemodel
Disrupts protein-proteininteraction in Hsp90 complexfor its chemopreventiveactivity
[234]
MIA-PaCa2 Potentiates the inhibitoryeffects of gemcitabine and5-flurouracil on clonogenicity,spheroid formation, ALDH1activity, Notch-1 and c-Relexpression
[235]
PANC-1, MIA PaCa-2 and AsPC-1 Inhibits cell proliferation,colony formation,phosphorylation of Akt andERK, activates FoxOtranscription factors andinduces apoptosis
[79]
22 D. Singh et al. / Biochimica et Biophysica Acta 1856 (2015) 13–27
effect of genistein onDNA topoisomerase II is also amajor contributor toits cytotoxic activity [170,172].
Genistein inhibits cell growth, clonogenicity, cell migration and in-vasion, EMT phenotype, and formation of pancreatospheres consistentwith reduced expression of CD44 and EpCAM [185].Wang et al., showedthat genistein restricts pancreatic cancer cell invasion by inhibiting cellgrowth and inducing apoptosis alongwith attenuation of FoxM1 and itsdownstream genes (survivin, Cdc25a, MMP-9, and VEGF) [186].
17. Sulforaphane
Sulforaphane has been reported to inhibit the growth of establishedtumors and prevent chemically induced cancers in animal models[187–189] (Table 5). It has been shown to inhibit Akt pathway in ovar-ian, prostate and colorectal cancers [189–191] and down-regulateβ-catenin in HeLa and HepG2 cells [192]. Additionally, it also targetsbreast cancer stem/progenitor cells effectively in both in vitro andin vivo conditions [193]. The studies have shown that recipient NOD/SCID mice inoculated with tumor cells derived from sulforaphane-treated primary xenograft failed to develop tumor growth, whereascontrol tumor cells quickly generate large tumors [194].
Furthermore, sulforaphane has also been shown to inhibit theself-renewal capacity of pancreatic CSCs. Srivastava et al., showed thatinhibition of Nanog enhances the inhibitory effects of sulforaphane on
the self-renewal capacity of CSCs [195]. Sulforaphane induces apoptosisby activating caspase-3 and inhibiting the expression of Bcl-2 and XIAP,as well as phosphorylation of FKHR. Additionally, sulforaphane issuggested to block signaling involved in early metastasis by inhibitingthe expression of proteins involved in the epithelial-mesenchymal tran-sition (beta-catenin, vimentin, TWIST1, and ZEB1) [195]. Additionally,sulforaphane, in combination of with TRAIL, has been suggested to bea promising strategy for targeting pancreatic tumor initiating cells(TICs). It has been found that it could abrogate the resistance of pancre-atic TICs to TRAIL (tumor necrosis factor-related apoptosis-inducing li-gand) by interfering with TRAIL-activated NF-κB signaling [196].
18. Garlic
Garlic has been traditionally used for varied human ailments aroundthe world. Epidemiological observations and preclinical studies, both incell and animal models, suggest the anti-carcinogenic potential of garlicand its constituents [197]. Chemical analysis revealed that the protec-tive effects of garlic are due to the presence of organosulfur compoundsmainly allyl derivatives [197]. Additionally, it modulates the activity ofseveralmetabolizing enzymes involved in the activation and detoxifica-tion of carcinogens and inhibits DNA adduct formation. It possessanti-oxidative and free radicals scavenging properties and regulatescell proliferation, apoptosis, and immune responses. Recent data sug-gest that garlic also modulates cell-signaling pathways to avoid prolifera-tion of unwanted cells thereby imparting strong cancer chemopreventive,as well as cancer therapeutic effects [197].
19. Benzyl isothiocyanate (BITC)
Due to their capability to induce apoptosis, modulate signalingpathways and inhibit angiogenesis, isothiocyanates (ITCs) have showna great promise as chemopreventive agents against various tumors inrecent years [198]. Benzyl isothiocyanate (BITC) is a major ITC com-pound present in cruciferous vegetables. BITC suppresses the initiationand progression of a variety of cancers including lung, esophageal,forestomach, urinary bladder, mammary, liver, colon, and pancreatictumors [198–203]. Various preclinical and mechanistic studies havesupported the anticancer efficacy of BITC, and it has been found to sup-press the growth of human pancreatic cancer cells both in vitro andin vivo. BITC is reported to induceG(2)/Mphase cell cycle arrest, and ap-optotic cell death in pancreatic cancer cell/animal models [204–207].The apoptotic potential of BITC is attributed to its capability to activateMAPK family members i.e. ERK, JNK and P38 by catalyzing their phos-phorylation at Thr202/Tyr204, Thr183/Tyr185, and Thr180/Tyr182respectively in a dose-dependent manner [206]. Additionally, the po-tential to inhibit the phosphorylation and expression of NF-kB mostlikely via inhibition of HDAC1/HDAC3, is suggested to be another con-tributory factor to the apoptotic potential of BITC [204].
Furthermore, it has been found to inhibit angiogenesis andmetasta-sis by suppressing VEGF and MMP-2 expression in pancreatic cancercells [208]. Recently Boreddy et al., showed that BITC offers protectionagainst pancreatic tumor growth by effectively containing STAT-3 andHIF-1α and VEGF expression in BxPC-3 and PanC-1 pancreatic cancercells [208]. It reduces the phosphorylation of PI3K, Akt, Pdk1, mTOR,FoxO1, and FoxO3a and increases apoptosis in tumor xenograft mousemodel. BITC treatment also decreases the binding of FoxO1 with 14-3-3protein suggesting it's nuclear retention and subsequent elevationof FoxO-responsive proteins involved in apoptosis (Bim) and cell cyclearrest (p27 and p21) [209].
BITC has also been shown to sensitize pancreatic tumors forradiotherapy. BITC treatment in a combination of X-rays or gamma-irradiation reduces cell survival as compared with individual (X-raysor gamma-irradiation) exposure in pancreatic cancer cells. This effectis suggested to be due to the inhibition of cell proliferation and anti-apoptotic genes like XIAP/IAP, and augmentation of apoptosis protease
23D. Singh et al. / Biochimica et Biophysica Acta 1856 (2015) 13–27
activating factor-1 (Apaf-1) triggered by BITC [208]. It is remarkablethat Apaf-1 is essential for activation of caspase-9 in stress-inducedapoptosis [208].
20. Piperlongumine
Piperlongumine (PL) is an alkaloid found in the fruits of long pepperplants that displays potent growth-inhibitory properties in a varietyof cancer cell lines and various animal models. It has been identifiedto target cancer cells selectively over normal cells through an ROS-dependent mechanism in a cell-based small-molecule screening andquantitative proteomics approach [210]. It increases ROS levels andcancer-selective cell death by directly binding and inhibiting the antiox-idant enzyme glutathione S-transferase pi 1 (GSTP1) [210,211]. Rajet al., have shown that PL selectively targets pancreatic cancer cells,PANC-1, and MIA PaCa-2- both of which harbor mutated K-ras [210].A recent study by Dhillon et al. further supported this finding and sug-gested that PL also targets BxPC-3 pancreatic cancer cells that containwild-type K-ras [212]. They further showed the anti-cancer effects ofPL in vivo. PL reduces tumor volume, increases oxidative DNA damage(8-OHdGhigh), and reduces proliferation (Ki-67low) in nude mice xeno-grafts for PANC-1 [212].
21. Conclusion and future perspectives
Pancreatic cancer is continuously posing a challenge to the cliniciansand researchers. We are still relying on the old traditional therapies.The major drawback of current therapy is their unilateral actions onone or two pathways whereas the approach should be to target severalevents simultaneously. The combination therapy is a right approach inthis direction, but associated side effects are a major concern. Sincethe small population of pancreatic CSCs is mostly responsible for thepathogenesis of pancreatic cancer, an efficient therapy targeted to pan-creatic CSCs is also an excellent approach. However, the problem is theresistance of pancreatic CSCs against conventional treatment but stilldevelopmental pathways such as the hedgehog, Wnt, Notch, etc. canbe targeted.
Flavonoids have emerged as potential chemopreventive candidatesfor cancer treatment, especially by their ability to induce apoptosis.These can interfere with the initiation, development and progressionof cancer by the modulation of cellular proliferation, differentiation,apoptosis, angiogenesis, and metastasis. Flavonoids have been shownto target cancer cells specifically with no or insignificant effects onhealthy cells in vitro. Nevertheless, some studies suggest to includeexperimental conditions (dose, cell type, culture conditions and treat-ment length) while interpreting the results of in vitro studies becausethe biological outcome can be affected. Since the apparent phenomenonis a result of complex interaction of different cellular events, the mech-anisms for inducing the apoptosis of these polyphenols may overlapwith other signaling cascades. Therefore, using polyphenols to promoteprogrammed cell death through the modulation of different proteins inother pathways that can contribute to apoptosis could be an alternativeapproach.
In summary, the characteristic effects of flavonoids, such as, inductionof apoptosis, activation of caspases, down-regulation or up-regulation ofBcl-2 family members, induction of cell cycle arrest and inhibition of sur-vival/proliferation make them ideal therapeutic candidate for pancreaticcancer prevention. Nevertheless, although the results from in vitroexperiments constitute a valuable tool for elucidating the pathwaysinvolved in the overall carcinogenesis process, these cannot be directlyextrapolated to clinical effects. Despite the fact that the herbal productstarget multiple signaling events simultaneously, more studies will beneeded to understand clearly the mechanisms of their action and thera-peutic potential.
Disclosure of potential conflicts of interest
The authors have declared that no Conflicts of Interest exist.
Acknowledgments
We acknowledge our lab members for critical reading of the manu-script, insightful discussions, and valuable advice. The project was fundedby the National Institutes of Health (RKS) and The VA Merit Award (SS).
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