Antagonism of Sorafenib and Regorafenib actions by platelet factors in hepatocellular carcinoma cell lines
© D’Alessandro et al.; licensee BioMed Central Ltd. 2014
Received: 30 December 2013
Accepted: 19 May 2014
Published: 21 May 2014
Platelets are frequently altered in hepatocellular carcinoma (HCC) patients. Platelet lysates (hPL) can enhance HCC cell growth and decrease apoptosis. The aims were to evaluate whether hPL can modulate the actions of Sorafenib or Regorafenib, two clinical HCC multikinase antagonists.
Several human HCC cell lines were grown in the presence and absence of Sorafenib or Regorafenib, with or without hPL. Growth was measured by MTT assay, apoptosis was assessed by Annexin V and by western blot, and autophagy and MAPK growth signaling were also measured by western blot, and migration and invasion were measured by standard in vitro assays.
Both Sorafenib and Regorafenib-mediated inhibition of cell growth, migration and invasion were all antagonized by hPL. Drug-mediated apoptosis and decrease in phospho-ERK levels were both blocked by hPL, which also increased anti-apoptotic phospho-STAT, Bax and Bcl-xL levels. Preliminary data, obtained with epidermal growth factor (EGF) and insulin-like growth factor-I (IGF-I), included in hPL, revealed that these factors were able to antagonized Sorafenib in a proliferation assay, in particular when used in combination.
Platelet factors can antagonize Sorafenib or Regorafenib-mediated growth inhibition and apoptosis in HCC cells. The modulation of platelet activity or numbers has the potential to enhance multikinase drug actions.
KeywordsRegorefenib Platelets HCC Apoptosis Growth Invasion
Platelet activity has been known for a long time to be altered in the presence of cancer, with venous thrombosis being recognized in association with occult malignancy [1, 2]. In addition to the effects of cancer on platelet actions in blood clotting, platelets have been recognized to be involved in cancer development, progression and metastasis [3–8]. Platelet levels have been shown to impact prognosis in several cancers, including those of the ovary, kidney, colon, lung and pancreas [9–14]. Furthermore, whereas hepatocellular carcinoma (HCC) most typically arises on the basis of cirrhosis, with its frequently associated splenomegaly and thrombocytopenia, normal or elevated platelet levels are frequently seen in large size HCCs [15–17]. We recently found that platelet extracts can stimulate HCC cell line growth in vitro, which was associated with a decrease in apoptosis . We now extend those observations, by examining the effects of platelet extracts on the effects of apoptosis-inducing HCC treatment agents and report that platelet extracts can antagonize growth inhibition mediated by Sorafenib or Regorafenib.
Cells and materials
PLC/PRF/5, Hep3B and HepG2 cells were obtained from the ATCC and were cultured as previously described .
Recombinant human EGF was purchased from PeproTech (Rocky Hill, NJ, USA), mouse recombinant IGF-I from Calbiochem (San Diego, CA, USA) and serotonin from Sigma-Aldrich (Saint Louis, MO, USA), all the growth factors were dissolved in water.
The platelet samples were collected from healthy volunteers. The study protocol was approved by the institutional review boards of the University of Bari and “Saverio de Bellis” Institute of Castellana G. (BA), Italy. Additionally, written informed consent was obtained from participants for the use of their blood in this study.
The platelet-rich plasma was obtained using an automated hemapheresis procedure in a local blood transfusion center. The platelets obtained from different volunteers were mixed and then divided into aliquots. Each aliquot was subjected to several freeze-thaw cycles to disrupt their membranes and release the growth factors stored in the granules (human Platelet Lysate, hPL).
Proliferation assay was performed as recently described [19, 20]. The cells were cultured in 1% FBS medium containing different hPL concentrations (2.5 - 3.75 × 107) or equivalent percentage of FBS in presence of 1 μM (HepG2 cell line) or 2.5 μM (Hep3B and PLC/RFP/5) of Sorafenib or Regorafenib. In the same growth condition HCC cell lines were cultured in presence of EGF 10, 25 mg/ml, IGF-I 50, 100 mg/ml and serotonin 1, 10 μM with or without Sorafenib 1 μM.
Medium AFP levels were measured using an automated system (UniCel Integrated Workstations DxC 660i, Beckman Coulter, Fullerton, CA, USA) by a chemoluminescent immunometric method. Sample measurements over the calibration range were automatically re-analyzed according to manufacture’s instructions.
A scratch assay was performed as previously described [19, 20]. Briefly, a wound was generated with a pipette tip, after rinsing, medium containing different concentrations of hPL or equivalent percentage of FBS and 2.5 μM Sorafenib or Regorafenib was added. Photographs were taken of each well immediately and after 24 h and 48 h. The values were expressed as percentage of migration, with 100% being when the wound was completely closed. The results were representative of three independent experiments.
Cell invasion assays were performed using Matrigel (BD Transduction, San Jose, CA, USA)-coated Transwells (8 μm pore PET membrane, Millipore, Billerica, MA, USA) as previously described (19). Briefly, 2.5 μM Sorafenib or Regorafenib treated cells were suspended in low serum medium. Medium containing different hPL or FBS concentrations was added to the bottom wells. After incubation of 24 h, the invading cells were fixed and stained. The images were acquired and analyzed counting the cells with Image J Software (National Institute of Health, USA). Values obtained were expressed as fold increase of invading cells, setting the cell counts of control cells as one. Results were representative of three independent experiments.
The Muse Annexin V/Dead Cell Assay Kit (Millipore, Darmstadt, Germany) for quantitative analysis of live, early/late apoptotic and dead cells was used with a Muse Cell Analyzer (Millipore). Briefly, the assay utilizes Annexin V to detect PS on the external membrane of apoptotic cells. A dead cell marker (7-AAD) is also used. PLC/PRF/5 cell line, including positive and negative controls, were cultured in 1% FBS medium supplemented with a volume of hPL corresponding to 3.75 × 107 platelets/ml or with an equivalent percentage of serum (control cells) for 48 h. The cells were then processed as described in the user’s guide.
Caspase-3/7 quantitative measurements
The Muse Caspase-3/7 kit (Millipore) permits simultaneous evaluation of apoptotic status based on Caspase-3 and -7 activation and cellular plasma membrane permeabilization (cell death). The assay provides relative percentage of cells that are live, early/late apoptotic or dead. Cells were cultured as described above and processed according to the user’s guide.
We analyzed the MAPK signaling and anti-apoptosis markers in Hep3B cells treated with 2.5 μM Sorafenib or Regorafenib and hPL by Western blot, as previously described [19, 20]. In brief, cells were washed twice with cold PBS and then lysed in RIPA buffer (Sigma-Aldrich, Milan; Italy). After quantization of protein concentration, equal amount of protein (50 μg) were resolved on SDS–PAGE and transferred to polyvinyldifluoride (PVDF) filters. The blots were blocked with 5% (w/v) nonfat dry milk for 2 h at room temperature and then probed with primary antibody overnight at 4°C.
The primary antibodies were directed against the following proteins: ERK and phospho-ERK (P-ERK), JNK and phospho-JNK (P-JNK), p38 and phospho-p38 (P-p38), STAT3 and phospho-STAT3 (Tyr705, Ser727) (P-STAT3), AKT and phospho-AKT (P-AKT), survivin, Bcl-xL, Bax, Bim and β-actin (Cell Signaling, Beverly, MA, USA). After three washes, incubation was followed by the reaction with horseradish peroxidase-conjugated secondary antibody for 1 h at room temperature. The immunoreactive bands were visualized and analyzed using the enhanced chemiluminescence detection reagents (Cell Signaling), according to the manufacturer’s instructions, and chemiluminescence detection system (ChemiDoc XRS apparatus and software, Bio-Rad).
GraphPad Prism 5.0 software (La Jolla, CA, USA) was used for all statistical analysis. Mann–Whitney nonparametric test was employed to assess the statistical significance of differences between two groups. For multiple comparisons was used one-way Anova test followed by appropriate post-test. P-values of <0.05 were considered statistically significant. All experiments were done in triplicate and data are presented as mean ± standard deviation (SD).
Platelet factors antagonize drug-mediated inhibition of HCC cell growth
hPL were previously examined for the ability to stimulate human HCC cell line growth . Hep3B, PLC/PRF/5 and HepG2 human HCC cell lines were treated in log phase growth with 1 μM (HepG2) - 2.5 μM (Hep3B and PLC/PRF/5) Regorafenib or Sorafenib, concentrations which are known to decrease in HCC cell proliferation .
Comparing the growth in these different conditions by MTT assay, it was clear that increasing the serum concentration more than 1% had not significant influence on PLTs antagonism (Figure 1G). Identical results were obtained with Sorafenib treatments (data not shown).
The concentrations of medium alpha-fetoprotein (AFP), an HCC cell growth marker, were also measured. We found that Sorafenib-mediated inhibition of AFP levels was also antagonized by the presence of hPL (Figure 1H).
Effects of platelet factors on cell signaling
Platelet factor antagonism of drug-mediated inhibition of migration and invasion
Both Sorafenib and Regorafenib can inhibit both HCC cell migration and invasion through Matrigel membranes. Furthermore, hPL has been shown to stimulate cell motility . We therefore added hPL to 2.5 μM concentrations of Sorafenib or Regorafenib that could inhibit both migration and invasion in Hep3B cells.
Platelet factor antagonism of drug-mediated induction of apoptosis
To evaluate the possible platelet factor mechanisms, we examined their effects on Sorafenib or Regorafenib–mediated apoptosis, since that is one major aspect of their growth-inhibitory actions.
EGF and IGF antagonize drug-mediated inhibition of HCC cell growth
We report here for the first time, the antagonizing effects of platelet extracts on growth inhibition in several HCC cell lines, that was mediated by Sorafenib or Regorafenib. Both agents were similarly antagonized by hPL. Furthermore, the previously demonstrated inhibition of AFP secretion by these drugs, was also antagonized. A main consequence of each drug is a decrease in phospho-ERK levels, secondary to Raf inhibition. hPL antagonized this early consequence of the drug action, without change in ERK levels. There was also an early and strong antagonism of the previously noted inhibitory effects of drug on phospho-p38 levels , and similarly for the p38 downstream target, phospho-STAT3 (Tyr and Ser). These are important molecules in mediating cell proliferation and play a role in the induction of anti-apoptosis mediators. Both Sorafenib and Regorafenib are known to increase apoptosis in treated cells. We found that this apoptosis-induction was antagonized by addition of hPL to cells that were treated with each of these two agents, as measured by both annexin V and caspase 3/7 activation.
Consistent with our findings of increased phospho-STAT3 levels, we also found an increase in the levels of anti-apoptotic Bcl-xL and survivin and a decrease in the levels of pro-apoptotic Bim and Bax, consequent to hPL action.
Due to the important role of platelets in the metastasis mechanisms of many tumors [8, 21], we evaluated hPL for a possible role in stimulating cell migration or invasion. We founds that the extracts also antagonized drug-mediated inhibition of HCC cell migration and invasion on Matrigel-treated membranes. In other systems, the targeting of platelets or experimental decrease in their numbers has been shown to enhance cancer chemotherapy [22, 23].
Platelets are the source of multiple growth factors, cytokines and inflammatory mediators .
Included among them are EGF, IGF-I, fibroblast growth factor (FGF), platelet derived growth factor (PDGF) and serotonin, the modulation of each having been shown to alter cancer chemotherapy sensitivity or resistance [24–30]. Preliminary data, obtained with several growth factors included in hPL, revealed interesting results using EGF and IGF-I. Both these factors were able to antagonized Sorafenib in a proliferation assay, in particular when used in combination. This growth induction was more evident than that observed in absence of drug, suggesting a specific interference of these growth factors with the inhibitory action of Sorafenib.
Interestingly, the clinical insulin modulator and diabetes drug, metformin  and the serotonin modulator Fluoxetine/Prozac that is used in depression treatment [29, 30], each alter chemotherapy sensitivity in cancer cells. Multiple pathways have been found to be involved in Sorafenib-mediated growth inhibition, especially apoptosis and autophagy [19, 31, 32] as well as others [33–37] and several cytokines, or cytokine modulators that are produced by platelets can modulate Sorafenib activity . Since Sorafenib effects have been clinically modest, several approaches are under way to enhance its actions, either on its downstream targets, or by adding inhibitors of parallel pathways in combination therapies . Given the large number of candidate factors in platelets, the identification of those responsible for drug resistance is just beginning. However, FGF, IGF1 and serotonin would seem to be promising possibilities.
The recent finding that platelet inhibitors reduce hepatitis B associated experimental HCC  has led to new interest in the use of aspirin and other platelet inhibitors in HCC prevention, as in colon cancer prevention . Thrombocytosis has been shown to be a negative prognostic factor for renal, breast, ovary, pancreas and colon cancers. Therefore, the results from this paper might be applicable to those tumor types, especially to renal cancer, since Sorafenib is also FDA-approved for treatment of renal cancer.
The current results give support to the idea that platelet inhibitors might also be useful in the drug therapy of patients with unresectable HCC, provided their platelet levels and coagulation systems are normal.
Human Platelets lysate
Portal vein thrombosis
Extracellular signal-regulated kinase
c-Jun NH2-terminal kinase
Signal transducer and activator of transcription-3
Epidermal growth factor
Insulin-like growth factor-I.
This research was supported in part by NIH grant CA82723 (BIC) and by Ministry of Health of Italy. The content is solely the responsibility of the authors and does not necessarily represent the official views of either NIH or Italian Ministry of Health.
- Trousseau A: Phlegmana alba dolens. Clinic Med de l’Hotel-Dieu. 1985, 3: 654-712.Google Scholar
- Levin J, Conley CL: Thrombocytosis associated with malignant disease. Arch Intern Med. 1964, 114: 497-500. 10.1001/archinte.1964.03860100079008.View ArticlePubMedGoogle Scholar
- Leslie M: Cell biology: beyond clotting: the powers of platelets. Science. 2010, 328: 562-564. 10.1126/science.328.5978.562.View ArticlePubMedGoogle Scholar
- Borsig L: The role of platelet activation in tumor metastasis. Expert Rev Anticancer Ther. 2008, 8: 1247-1255. 10.1586/14737188.8.131.527.View ArticlePubMedGoogle Scholar
- Labelle M, Begum S, Hynes RO: Direct signaling between platelets and cancer cells induces an epithelial-mesenchymal-like transition and promotes metastasis. Cancer Cell. 2011, 20: 576-590. 10.1016/j.ccr.2011.09.009.View ArticlePubMedPubMed CentralGoogle Scholar
- Buergy D, Wenz F, Groden C, Brockmann MA: Tumor-platelet interaction in solid tumors. Int J Cancer. 2012, 130: 2747-2760. 10.1002/ijc.27441.View ArticlePubMedGoogle Scholar
- Bambace NM, Holmes CE: The platelet contribution to cancer progression. J Thromb Haemost. 2011, 9: 237-249. 10.1111/j.1538-7836.2010.04131.x.View ArticlePubMedGoogle Scholar
- Goubran HA, Burnouf T, Radosevic M, El-Ekiaby M: The platelet-cancer loop. Eur J Intern Med. 2013, 24: 393-400. 10.1016/j.ejim.2013.01.017.View ArticlePubMedGoogle Scholar
- Stone RL, Nick AM, McNeish IA, Balkwill F, Han HD, Bottsford-Miller J, Rupairmoole R, Armaiz-Pena GN, Pecot CV, Coward J, Deavers MT, Vasquez HG, Urbauer D, Landen CN, Hu W, Gershenson H, Matsuo K, Shahzad MM, King ER, Tekedereli I, Ozpolat B, Ahn EH, Bond VK, Wang R, Drew AF, Gushiken F, Lamkin D, Collins K, DeGeest K, Lutgendorf SK, et al: Paraneoplastic thrombocytosis in ovarian cancer. N Engl J Med. 2012, 366: 610-618. 10.1056/NEJMoa1110352.View ArticlePubMedPubMed CentralGoogle Scholar
- Heng DY, Xie W, Regan MM, Warren MA, Golshayan AR, Sahi C, Eigl BJ, Ruether JD, Cheng T, North S, Venner P, Knox JJ, Chi KN, Kollmannsberger C, McDermott DF, Oh WK, Atkins MB, Bukowski RM, Rini BI, Choueiri TK: Prognostic factors for overall survival in patients with metastatic renal cell carcinoma treated with vascular endothelial growth factor-targeted agents: results from a large, multicenter study. J Clin Oncol. 2009, 27: 5794-5799. 10.1200/JCO.2008.21.4809.View ArticlePubMedGoogle Scholar
- Sasaki K, Kawai K, Tsuno NH, Sunami E, Kitayama J: Impact of preoperative thrombocytosis on the survival of patients with primary colorectal cancer. World J Surg. 2012, 36: 192-200. 10.1007/s00268-011-1329-7.View ArticlePubMedGoogle Scholar
- Wan S, Lai Y, Myers RE, Li B, Hyslop T, London J, Chatterjee D, Palazzo JP, Burkart AL, Zhang K, Xing J, Yang H: Preoperative platelet count associates with survival and distant metastasis in surgically resected colorectal cancer patients. J Gastrointest Cancer. 2013, 44: 293-304. 10.1007/s12029-013-9491-9.View ArticlePubMedPubMed CentralGoogle Scholar
- Yu D, Liu B, Zhang L: Platelet count predicts prognosis in operable non-small cell lung cancer. Exp Ther Med. 2013, 5: 1351-1354.PubMedPubMed CentralGoogle Scholar
- Wang H, Gao J, Bai M, Liu R, Li H, Deng T, Zhou L, Han R, Ge S, Huang D, Ba Y: The pretreatment platelet and plasma fibrinogen level correlate with tumor progression and metastasis in patients with pancreatic cancer. Platelets. 2013, doi:10.3109/09537104.2013.827782Google Scholar
- Hwang SJ, Luo JC, Li CP, Chu CW, Wu JC, Lai CR, Chiang JH, Chau GY, Lui WY, Lee CC, Chang FY, Lee SD: Thrombocytosis: a paraneoplastic syndrome in patients with hepatocellular carcinoma. World J Gastroenterol. 2004, 10: 2472-2477.View ArticlePubMedPubMed CentralGoogle Scholar
- Carr BI, Guerra V: Features of massive hepatocellular carcinomas. Eur J Gastroenterol Hepatol. 2013, doi:10.1097/MEG.0b013e3283644c49Google Scholar
- Carr BI, Guerra V: Thrombocytosis and hepatocellular carcinoma. Dig Dis Sci. 2013, 58: 1790-1796. 10.1007/s10620-012-2527-3.View ArticlePubMedPubMed CentralGoogle Scholar
- Carr BI, D’Alessandro R, Refolo MG, Lippolis C, Cavallini A, Messa C: Effects of Pheripheral Blood Platelets on HCC Cell Growth, Migration and Invasion. The Liver Meeting Proc. 2013, Boston, MA, USA: AASLDGoogle Scholar
- Carr BI, Cavallini A, Lippolis C, D’Alessandro R, Messa C, Refolo MG, Tafaro A: Fluoro- Sorafenib (Regorafenib) effects on hepatoma cells: growth inhibition, quiescence, and recovery. J Cell Physiol. 2013, 228: 292-297. 10.1002/jcp.24148.View ArticlePubMedPubMed CentralGoogle Scholar
- D’Alessandro R, Refolo MG, Lippolis C, Messa C, Cavallini A, Rossi R, Resta L, Di Carlo A, Carr BI: Reversibility of regorafenib effects in hepatocellular carcinoma cells. Cancer Chemother Pharmacol. 2013, 72: 869-877. 10.1007/s00280-013-2269-8.View ArticlePubMedGoogle Scholar
- Dashevsky O, Varon D, Brill A: Platelet-derived microparticles promote invasiveness of prostate cancer cells via upregulation of MMP-2 production. Int J Cancer. 2009, 124: 1773-1777. 10.1002/ijc.24016.View ArticlePubMedGoogle Scholar
- Demers M, Wagner DD: Targeting platelet function to improve drug delivery. Oncoimmunology. 2012, 1: 100-102. 10.4161/onci.1.1.17962.View ArticlePubMedPubMed CentralGoogle Scholar
- Demers M, Ho-Tin-Noé B, Schatzberg D, Yang JJ, Wagner DD: Increased efficacy of breast cancer chemotherapy in thrombocytopenic mice. Cancer Res. 2011, 71: 1540-1549. 10.1158/0008-5472.CAN-10-2038.View ArticlePubMedPubMed CentralGoogle Scholar
- Song S, Wientjes MG, Gan Y, Au JL: Fibroblast growth factors: an epigenetic mechanism of broad spectrum resistance to anticancer drugs. Proc Natl Acad Sci U S A. 2000, 97: 8658-8663. 10.1073/pnas.140210697.View ArticlePubMedPubMed CentralGoogle Scholar
- Shen K, Cui D, Sun L, Lu Y, Han M: Inhibition of IGF-IR increases chemosensitivity in human colorectal cancer cells through MRP-2 promoter suppression. J Cell Biochem. 2012, 113: 2086-2089. 10.1002/jcb.24080.View ArticlePubMedGoogle Scholar
- Jalving M, Gietema JA, Lefrandt JD, de Jong S, Reyners AK, Gans RO, de Vries EG: Metformin: taking away the candy for cancer?. Eur J Cancer. 2010, 46: 2369-2380. 10.1016/j.ejca.2010.06.012.View ArticlePubMedGoogle Scholar
- Atzori F, Traina TA, Ionta MT, Massidda B: Targeting insulin-like growth factor type 1 receptor in cancer therapy. Target Oncol. 2009, 4: 255-266. 10.1007/s11523-009-0123-z.View ArticlePubMedGoogle Scholar
- Kina S, Phonaphonh T, Liang F, Kuang H, Arasaki A, Arakaki K, Nakasone T, Sunakawa H: PDGFα receptor is a mediator for cisplatin-induced Met expression. Eur J Pharmacol. 2013, 699: 227-232. 10.1016/j.ejphar.2012.11.014.View ArticlePubMedGoogle Scholar
- Zhou T, Duan J, Wang Y, Chen X, Zhou G, Wang R, Fu L, Xu F: Fluoxetine synergys with anticancer drugs to overcome multidrug resistance in breast cancer cells. Tumour Biol. 2012, 33: 1299-1306. 10.1007/s13277-012-0377-4.View ArticlePubMedGoogle Scholar
- Peer D, Dekel Y, Melikhov D, Margalit R: Fluoxetine inhibits multidrug resistance extrusion pumps and enhances responses to chemotherapy in syngeneic and in human xenograft mouse tumor models. Cancer Res. 2004, 64: 7562-7569. 10.1158/0008-5472.CAN-03-4046.View ArticlePubMedGoogle Scholar
- Wilhelm SM, Carter C, Tang L, Wilkie D, McNabola A, Rong H, Chen C, Zhang X, Vincent P, McHugh M, Cao Y, Shujath J, Gawlak S, Eveleigh D, Rowley B, Liu L, Adnane L, Lynch M, Auclair D, Taylor I, Gedrich R, Voznesensky A, Riedl B, Post LE, Bollag G, Trail PA: BAY 43–9006 exhibits broad spectrum oral antitumor activity and targets the RAF/MEK/ERK pathway and receptor tyrosine kinases involved in tumor progression and angiogenesis. Cancer Res. 2004, 64: 7099-7109. 10.1158/0008-5472.CAN-04-1443.View ArticlePubMedGoogle Scholar
- Liu L, Cao Y, Chen C, Zhang X, McNabola A, Wilkie D, Wilhelm S, Lynch M, Carter C: Sorafenib blocks the RAF/MEK/ERK pathway, inhibits tumor angiogenesis, and induces tumor cell apoptosis in hepatocellular carcinoma model PLC/PRF/5. Cancer Res. 2006, 66: 11851-11858. 10.1158/0008-5472.CAN-06-1377.View ArticlePubMedGoogle Scholar
- Hikita H, Takehara T, Shimizu S, Kodama T, Shigekawa M, Iwase K, Hosui A, Miyagi T, Tatsumi T, Ishida H, Li W, Kanto T, Hiramatsu N, Hayashi N: The Bcl-xL inhibitor, ABT- 737, efficiently induces apoptosis and suppresses growth of hepatoma cells in combination with sorafenib. Hepatology. 2010, 52: 1310-1321. 10.1002/hep.23836.View ArticlePubMedGoogle Scholar
- Galmiche A, Ezzoukhry Z, François C, Louandre C, Sabbagh C, Nguyen-Khac E, Descamps V, Trouillet N, Godin C, Regimbeau JM, Joly JP, Barbare JC, Duverlie G, Mazière JC, Chatelain D: BAD, a proapoptotic member of the BCL2 family, is a potential therapeutic target in hepatocellular carcinoma. Mol Cancer Res. 2010, 8: 1116-1125. 10.1158/1541-7786.MCR-10-0029.View ArticlePubMedGoogle Scholar
- Shimizu S, Takehara T, Hikita H, Kodama T, Tsunematsu H, Miyagi T, Hosui A, Ishida H, Tatsumi T, Kanto T, Hiramatsu N, Fujita N, Yoshimori T, Hayashi N: Inhibition of autophagy potentiates the antitumor effect of the multikinase inhibitor sorafenib in hepatocellular carcinoma. Int J Cancer. 2012, 131: 548-557. 10.1002/ijc.26374.View ArticlePubMedGoogle Scholar
- Ezzoukhry Z, Louandre C, Trécherel E, Godin C, Chauffert B, Dupont S, Diouf M, Barbare JC, Mazière JC, Galmiche A: EGFR activation is a potential determinant of primary resistance of hepatocellular carcinoma cells to sorafenib. Int J Cancer. 2012, 131: 2961-2969. 10.1002/ijc.27604.View ArticlePubMedGoogle Scholar
- Louandre C, Ezzoukhry Z, Godin C, Barbare JC, Mazière JC, Chauffert B, Galmiche A: Iron-dependent cell death of hepatocellular carcinoma cells exposed to sorafenib. Int J Cancer. 2013, 133: 1732-4172. 10.1002/ijc.28159.View ArticlePubMedGoogle Scholar
- Liu Y, Liu A, Li H, Li C, Lin J: Celecoxib inhibits interleukin-6/interleukin-6 receptor- induced JAK2/STAT3 phosphorylation in human hepatocellular carcinoma cells. Cancer Prev Res (Phila). 2011, 4: 1296-1305. 10.1158/1940-6207.CAPR-10-0317.View ArticleGoogle Scholar
- Dai Y, Grant S: Targeting multiple arms of the apoptotic regulatory machinery. Cancer Res. 2007, 67: 2908-2911. 10.1158/0008-5472.CAN-07-0082.View ArticlePubMedGoogle Scholar
- Sitia G, Iannacone M, Guidotti LG: Anti-platelet therapy in the prevention of hepatitis B virus-associated hepatocellular carcinoma. J Hepatol. 2013, 4: doi:S0168- 8278(13)00373-5Google Scholar
- Nishihara R, Lochhead P, Kuchiba A, Jung S, Yamauchi M, Liao X, Imamura Y, Qian ZR, Morikawa T, Wang M, Spiegelman D, Cho E, Giovannucci E, Fuchs CS, Chan AT, Ogino S: Aspirin use and risk of colorectal cancer according to BRAF mutation status. JAMA. 2013, 309: 2563-2571. 10.1001/jama.2013.6599.View ArticlePubMedPubMed CentralGoogle Scholar
- The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2407/14/351/prepub
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.