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Journal of Cancer Research and Clinical Oncology logoLink to Journal of Cancer Research and Clinical Oncology
. 2010 May 21;137(3):505–512. doi: 10.1007/s00432-010-0914-8

Bevacizumab enhances chemosensitivity of hepatocellular carcinoma to adriamycin related to inhibition of survivin expression

Yu-Quan Xiong 1,2,3, Hui-Chuan Sun 1,2, Xiao-Dong Zhu 1,2, Wei Zhang 1,2, Peng-Yuan Zhuang 1,2, Ju-Bo Zhang 1,2, Hua-Xiang Xu 1,2, Ling-Qun Kong 1,2, Wei-Zhong Wu 1,2, Lun-Xiu Qin 1,2,3, Zhao-You Tang 1,2,3,✉
PMCID: PMC11828078  PMID: 20490863

Abstract

Purpose

In recent years, anti-angiogenesis drugs have shown promising clinical effects against many tumors, particularly in combination with chemotherapy. Although the combination has become a standard of care for many tumors, the mechanisms of the chemosensitizing activity of anti-angiogenic drugs are not fully understood. Here, we sought to determine if anti-angiogenesis drug bevacizumab could enhance the chemosensitivity of HCC by inhibition of survivin.

Methods

After treatment of human umbilical vein endothelial cells (HUVECs) and hepatocellular carcinoma (HCC) cell line PLC/PRF/5 (PLC) with bevacizumab or/and adriamycin, the direct effects were examined by survival assays, and the expression of Akt, Phospho-Akt and survivin were evaluated by western blot. Tumor growth was observed in a human HCC xenograft nude mouse model treated with different drugs, and the expression of PCNA, CD31 and survivin in tumor tissues were evaluated by means of immunohistochemistry.

Results

Bevacizumab enhanced the chemosensitivity of HCC by inhibiting the VEGF-PI3 K/Akt-survivin signaling cascade in endothelial cells. The combination of bevacizumab with adriamycin therapy resulted in better outcomes compared with monotherapy in hepatocellular carcinoma xenografts; bevacizumab significantly inhibited tumor angiogenesis and growth. In addition, bevacizumab reduced survivin expression in tumor tissues, including tumor vascular endothelial cells in vivo, although it did not inhibit survivin expression in tumor cells in vitro.

Conclusion

These results implicate the bevacizumab-increased efficacy of adriamycin via an inhibition of survivin expression in malignant cells as well as tumor vasculature cells, which provides other insights into the mechanism of enhanced efficacy by combination of VEGF blocker and chemotherapeutic agents.

Keywords: Chemosensitivity, Bevacizumab, Survivin, Adriamycin, Hepatocellular carcinoma

Introduction

Tumor angiogenesis is critical for the progression and metastasis of cancer (Bergers and Benjamin 2003), which is particularly relevant in the highly vascularized tumors. As an important angiogenesis factor, vascular endothelial growth factor (VEGF) is required for growth and differentiation of tumor endothelial cells. Most tumors, including hepatocellular carcinoma (HCC), express elevated VEGF. Inadvertently, elevated VEGF induced by stresses such as radiotherapy or chemotherapy can contribute to enhanced survival of tumor endothelial cells, which render them less sensitive to conventional chemotherapy and radiotherapy treatments (Le Gouill et al. 2004; Riedel et al. 2004). Bevacizumab, a recombinant humanized monoclonal antibody that targets VEGF, has become an important therapeutic agent for tumors, including HCC (Finn and Zhu 2009; Zhu et al. 2006). Most clinical trials to date indicate that bevacizumab is more effective when combined with chemotherapy; however, the mechanism of the chemosensitizing activity of bevacizumab is not fully understood (Kerbel 2006).

Survivin is a member of the family of anti-apoptotic proteins, playing a role in both cell division and apoptosis by controlling the interface between mitotic progression and regulation of caspase activity, which is expressed during fetal development and in cancer tissues, including HCC (Altieri 2008; Fields et al. 2004; Ito et al. 2000). Growing evidence has indicated that survivin expression plays an essential role in drug resistance and that genetic or pharmacological modulation of survivin expression affects drug effectiveness in apoptosis induction (Li and Ling 2006; Moriai et al. 2008). In endothelial cells (ECs), VEGF can up-regulate survivin expression through the PI3 K/Akt signaling pathway to escape from apoptosis (O’Connor et al. 2000), which can be suppressed by survivin targeting treatment (Mesri et al. 2001). When treated with cytotoxin drugs, VEGF-induced survivin expression by the PI3 K/Akt pathway worked as a major chemoresistance mechanism in HUVECs (Tran et al. 2002). In tumor-associated endothelial cells isolated from human glioma, Virrey et al. (2008) reported that increased survivin expression was protective against cytotoxicity drugs. Recently, it was reported that anti-angiogenesis agent sunitinib and pazopanib can decrease survivin expression in renal carcinoma cells and multiple myeloma cells, respectively (Podar et al. 2006; Xin et al. 2009). However, it is not known whether anti-angiogenesis treatment can decrease survivin expression in tumors in vivo.

In this study, we sought to determine if bevacizumab can enhance the chemosensitivity of HCC by inhibition of survivin. Here, we report that bevacizumab, by inhibiting the VEGF-PI3 K/Akt-survivin signaling cascade, increased the cytotoxicity of adriamycin to HUVECs exposed to VEGF. In HCC xenograft models, the combination of bevacizumab with adriamycin therapy significantly suppressed tumor growth compared with monotherapy. Additionally, we first report that bevacizumab can reduce survivin expression in tumor cells and vascular endothelial cells in vivo, although it did not inhibit survivin expression in tumor cells in vitro.

Materials and methods

Cell lines and culture conditions

Human umbilical vein endothelial cells (HUVECs, ScienCell Research Laboratory, San Diego, CA) were grown in EBM-2 medium (Cambrex BioScience, Walkersville, MD) at 5% CO2 and used between passages 2 and 6. The HCC cell line PLC/PRF/5 (PLC, American Type Culture Collection, Manassas, VA) was cultured in DMEM containing supplements (10% FCS, penicillin/streptomycin, and L-glutamine).

Animals

Male athymic BALB/c-nu/nu nude mice, 5–6 weeks of age, were obtained from the Shanghai Institute of Materia Medica, Chinese Academy of Sciences, and housed in laminal-flow cabinets under specific pathogen-free (SPF) conditions. Experimental protocol was approved by the Shanghai Medical Experimental Animal Care Committee.

Survival assays

Cells (2 × 103) were plated in triplicate into 96-well plates in complete EBM-2 medium. After 10 h, cells were washed once with serum-free EBM-2. For detecting cells’ response to treatment with VEGF (R&D Systems, Minneapolis, MN), doxorubicin hydrochloride (Shenzhen Main Luck Pharmaceuticals Inc., Shenzhen, China) and bevacizumab (Roche, Welwyn Garden City, UK) cells were cultured for 48 h in serum-free EBM-2 medium containing different drugs at indicated concentrations with or without the PI3 K inhibitor LY294002. The supernatant was carefully removed and discarded without disturbing the cell pellet, and the cell pellets were frozen at −70°C. Then the viable cells grown in microplates were detected with CyQUANT Cell Proliferation Assay Kit (Molecular Probes–Invitrogen, Carlsbad, CA).

Western blot analysis

Cells were harvested in a lysis buffer (Pierce, Rockford, IL), and an equal amount of protein was subjected to 12% SDS–PAGE. After gel electrophoresis, the proteins were transferred to the polyvinylidene difluoride membranes (Immobilon PVDF, Millipore, Bedford, MA). The membranes were blocked for 1 h at room temperature in 5% nonfat dry milk in Tris-buffered saline containing 0.05% Tween-20 (TBST), followed by an overnight incubation at 4°C with primary antibodies. The membranes were then incubated with horseradish peroxidase (HRP)-labeled anti-rabbit secondary antibody (Chemicon, Temecula, CA) for 1 h at room temperature. Peroxidase activity was detected via chemiluminescence (SuperSignal West Femto luminol substrate and peroxide buffer; Pierce). Primary antibodies include anti-Akt, anti-phosphor-Akt, anti-Survivin, and anti-β-Actin (Cell Signaling Technology, Beverly, MA).

Nude mice xenograft models

PLC/PRF/5 tumors were initially established by s.c. injection of 5 × 106 cells in PBS. For the therapeutic experiments in mice, mice were randomized into four groups (n = 5 in each group), as follows: (a) control animals received i.p. injections of 100 μl of 0.9% sodium chloride solution twice weekly; (b) animals received i.p. injections of 2.5 mg/kg doxorubicin hydrochloride in 100 μl of 0.9% sodium chloride solution twice weekly; (c) animals received i.p. injections of 5 mg/kg bevacizumab in 100 μl of 0.9% sodium chloride solution twice weekly; (d) animals received i.p. injections of 2.5 mg/kg doxorubicin hydrochloride and 5 mg/kg bevacizumab in 100 μl of 0.9% sodium chloride solution twice weekly. Tumor growth was monitored by periodic caliper measurements every 3 days, and tumor volume was calculated according to the formula tumor volume = largest diameter × (perpendicular2/2). After 3 weeks, both untreated and treated nude mice were killed, and tumors were excised for histological study.

Immunohistochemical analysis

Immunohistochemical analysis was done with the following antibodies: rabbit anti-mouse PCNA and rabbit anti-mouse survivin (1:100; Cell Signaling Technology); rat anti-mouse CD31 (1:200; BD PharMingen, San Diego, CA); donkey anti-rabbit antibody conjugated to Alexa Fluor 448; HRP-labeled anti-rabbit secondary antibody; and HRP-labeled anti-rat secondary antibody (Jackson ImmunoResearch Laboratories Inc., West Grove, PA).

For PCNA and survivin staining, paraffin-embedded sections were immunostained with anti-PCNA and anti-Survivin. For evaluation of tumor angiogenesis, frozen sections were immunostained with anti-CD31 antibody. Immunostaining was carried out using DAKO EnVision Plus System, peroxidase (3,3Vdiaminobenzidine; DakoCytomation California Inc., Carpinteria, CA). For the quantification of mean vessel density in sections stained for CD31, five random fields at × 100 magnification were captured for each tumor, and microvessels were quantified. The data were expressed as mean ± standard error (SE). For double staining with CD31-Survivin, the frozen sections were incubated with rabbit anti-survivin antibody for 18 h at 4°C and incubated with anti-rabbit antibody conjugated to Alexa Fluor 488 for 1 h at room temperature, then the sections were incubated with rat anti-CD31 antibody for 18 h at 4°C and incubated with HRP-labeled anti-rat secondary antibody for 1 h at room temperature. Staining results were analyzed with an inverted fluorescence microscope (Olympus IX81, Olympus America Inc., Center Valley, PA) equipped with an Olympus Qcolor 3 digital camera (Olympus America Inc.).

Statistical analysis

Each experiment was repeated two to four times. Continuous data were expressed as mean ± SEM. A standard t test was used. Results were considered statistically significant at P < 0.05.

Results

Bevacizumab enhances chemosensitivity of ECs to adriamycin

Because the majority of chemotherapeutic drugs cause apoptosis, we set out to determine whether VEGF could reduce drug toxicity in ECs induced by chemotherapy. HUVECs were treated with different concentrations of adriamycin for 48 h in the presence or absence of VEGF (20 ng/ml); the addition of VEGF significantly increased survival when compared with ECs grown in growth factor–deprived conditions (Fig. 1a). As the significant rescue result was observed at 0.01 μg/mL of adriamycin when the VEGF concentration was 20 ng/ml, it was used in the following in vitro experiments. To test whether inhibition of VEGF can enhance chemosensitivity of ECs, HUVECs were treated with bevacizumab (1 μg/ml) before being treated with VEGF and adriamycin. Results showed that bevacizumab neutralized the rescue function of VEGF and increased cytotoxicity of HUVECs to adriamycin (Fig. 1b). However, treatments of PLC cells with the same condition as HUVECs did not exhibit a significant effect (Fig. 1c).

Fig. 1.

Fig. 1

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Effect of bevacizumab on HUVECs and PLC cells. a HUVECs were cultured for 48 h in the presence of 20 ng/ml VEGF and various concentrations of adriamycin (Adri). Proliferation assay showed that VEGF protected cells from adriamycin-induced apoptosis. b With or without PI3 K inhibitor LY294002, HUVECs were cultured for 48 h in the presence of different drugs. Bevacizumab (Beva) neutralized the chemoprotection function of VEGF; however, the addition of LY294002 abolished the significant effects of VEGF and Bevacizumab to adriamycin cytotoxicity. c PLC cells were cultured for 48 h in the presence of different drugs. There were no significant effects of VEGF and Bevacizumab on adriamycin cytotoxicity with or without LY294002. Points and columns, mean from three individual experiments with three to four samples per group; bars, SEM

Bevacizumab-driven chemosensitivity of ECs requires PI3 K-dependent induction of survivin

Consistent with other reports, survivin and p-Akt protein levels were up-regulated in HUVECs stimulated with VEGF in the present study, while survivin and p-Akt protein levels were strikingly decreased after the addition of bevacizumab (Fig. 2a). To determine whether bevacizumab-mediated survivin down-regulation requires PI3 K activation, we treated HUVECs with the same condition in the presence or absence of LY294002 (Merck, Nottingham, UK). PI3 K inhibition resulted in down-regulation of VEGF-mediated chemoprotection and VEGF-induced survivin protein expression, which is consistent with the down-regulation of bevacizumab-mediated chemosensitivity and survivin protein induction (Figs. 1b, 2a). Those results demonstrated that PI3 K activity is required for the bevacizumab-mediated reduction of survivin in HUVECs.

Fig. 2.

Fig. 2

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Bevacizumab inhibited survivin expression by the VEGF-PI3 K/Akt-survivin pathway. a Lysates of HUVECs treated with various drugs in the presence or absence of PI3 K inhibitor LY294002 were subjected to Western blot analysis. When cultured in adriamycin (A), the up-regulation of p-Akt and survivin induced by VEGF (V) were eliminated by bevacizumab (B); after dealing with LY294002, the changes in p-Akt and survivin induced by VEGF and bevacizumab disappeared. b There were no significant changes of p-Akt and survivin after treating with VEGF and bevacizumab in PLC cells

We also treated PLC cells with the same conditions as HUVECs. There was no significant difference in the response of PLC cells to the drug treatment with or without LY294002. Moreover, VEGF and bevacizumab cannot change survivin protein expression in PLC cells (Fig. 2b).

Effect of combined therapy with bevacizumab and adriamycin on PLC tumor growth

To assess whether bevacizumab combined with adriamycin would have an enhanced inhibitory effect on tumor growth, mice bearing PLC xenografts were treated with bevacizumab, adriamycin, and bevacizumab/adriamycin. As shown in Fig. 3a, after being treated for 21 days, a significant tumor growth delay can be observed in all groups; the bevacizumab/adriamycin combination group exhibits a more significant anti-tumor activity than groups treated by bevacizumab or adriamycin alone.

Fig. 3.

Fig. 3

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Effect of combined therapy with bevacizumab and adriamycin on PLC xenografts. a tumor volume at a given time for PLC xenografts treated with various treatments of adriamycin (Adri) or/and bevacizumab (Beva). Columns, mean; bars, SE. *P < 0.05; **P < 0.01. b The density of microvessels was significantly lower in bevacizumab/adriamycin groups compared with the other groups. Columns, mean; bars, SE. *P < 0.05; **P < 0.01; versus Beva + Adri groups. c Expression of CD31 (×100), PCNA (×200), and survivin (×200) in PLC xenografts. Adriamycin and/or bevacizumab significantly decreased CD31 and PCNA expression; meanwhile, bevacizumab, but not adriamycin, reduced survivin expression

Effect of combined therapy with bevacizumab and adriamycin on tumor angiogenesis and growth

After 21 days of treatment, a significant decrease in tumor microvessel density (MVD) was observed in all treatment groups, and bevacizumab/adriamycin combination groups showed less MVD compared with the other groups in association with reduced tumor growth (Fig. 3b, c).

Effect of combined therapy with bevacizumab and adriamycin on survivin expression in PLC xenograft

After 21 days of treatment, survivin expression was significantly decreased in bevacizumab- and bevacizumab/adriamycin-treated groups compared with untreated groups, while the adriamycin-treated group showed no significant difference with untreated groups (Fig. 3c). Decreased survivin expression can be found in tumor cells and tumor endothelial cells after being treated with bevacizumab (Fig. 4).

Fig. 4.

Fig. 4

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Survivin expression in tumor vascular endothelial cells of PLC xenografts. Frozen sections of PLC xenografts were stained with anti-survivin antibody (green immunofluorescence), followed by vessel staining with anti-CD31 antibody (brown immunohistochemistry). Tumor vascular endothelial cells expressing survivin are indicated by arrows. Original magnifications, ×400

Discussion

HCC is a major health problem worldwide. More than 70% of patients are not candidates for surgical treatment when diagnosis is established. Conventional cytotoxic chemotherapy is not effective in most cases, mainly for reasons of chemoresistance (Bruix and Llovet 2002). As a novel anti-apoptotic protein, it has been shown that survivin expression is invariably up-regulated in human cancers and is associated with resistance to chemotherapy (Yamamoto et al. 2008), while survivin inhibition can increase the sensitivity of chemotherapeutic drugs in many tumors (Nakagawa et al. 2006; Zaffaroni and Daidone 2002; Zaffaroni et al. 2005). Over the past several years, bevacizumab has been shown to have significant clinical benefit in combination with chemotherapy in advanced colon, breast, and liver cancers (Hurwitz et al. 2004; Miller et al. 2007; Zhu et al. 2006), but the mechanisms of the chemosensitizing activity of bevacizumab are not clear.

Considering that VEGF-induced survivin expression works as a major chemoresistance mechanism in HUVECs (Tran et al. 2002). We hypothesized that bevacizumab can increase chemosensitivity by inhibiting survivin expression.

The present study showed that pretreatment with bevacizumab blocked up-regulation of survivin and p-Akt in HUVECs and the chemoprotective effect induced by VEGF, which is consistent with previous reports (Tran et al. 2002). By Western blot analysis, we confirmed that VEGF-induced chemoprotection of HUVECs is associated with an induction of survivin downstream of PI3 K/Akt activation, as has been reported previously (Tran et al. 2002). However, neither VEGF nor bevacizumab has a marked effect on the survival of PLC cells or the PI3 K/Akt-survivin signaling pathway. Although tumor cells usually represent the main source of VEGF in tumors, including HCC, VEGF receptors (VEGFR-1, its soluble form sVEGFR-1, VEGFR-2, and neuropilin-1) are expressed predominantly by endothelial cells (Ferrara 2005; Sakurai et al. 2006), and this makes HUVECs more sensitive to VEGF and bevacizumab than tumor cells. These data suggest that bevacizumab can increase the chemosensitivity of HUVECs by inhibiting the VEGF-PI3 K/Akt-survivin signaling pathway but has no significant effects on PLC cells in vitro.

The animal experiments showed that bevacizumab/adriamycin was more effective than monotherapy in the treatment of HCC. Bevacizumab/adriamycin-treated tumors have a gross reduction in proliferation and fewer visible blood vessels compared with bevacizumab and adriamycin groups. There were many reports that angiogenic factors that induce the expression of survivin may act to shield endothelial cells from the apoptotic effects (Caldas et al. 2007; O’Connor et al. 2000; Singh et al. 2005; Virrey et al. 2008), and anti-angiogenesis therapy may enhance apoptosis of endothelial cells by inhibiting survivin expression, but there has been no report on the effect of anti-angiogenesis drugs on expression of survivin in tumor cells. Here, we observed that survivin expression was decreased by bevacizumab in PLC cells in vivo, but not in vitro. Compared with endothelial cells, PLC cells express fewer VEGFRs and present more resistance to serum-starved situations, which may result in PLC cells relying less on VEGF; meanwhile, they showed no big response to bevacizumab in the same treatment as in HUVECs in vitro, and survivin expression was not changed. While in vivo, survivin expression is associated with the tumor microenvironment, such as hypoxia (Yang et al. 2004), the hepatitis B virus X protein (HBx) expression (Marusawa et al. 2003), inflammation (Altznauer et al. 2004), and growth factor expression (Cosgrave et al. 2006), which may be changed by bevacizumab treatment, yet the specific mechanism still needs further study. Furthermore, we have observed a significant reduction in survivin expression in tumor vascular endothelial cells in bevacizumab- or bevacizumab/adriamycin-treated groups. These findings imply that bevacizumab improved the therapy effect of adriamycin partly by reducing survivin expression in tumors, especially in endothelial cells. There is now considerable evidence suggesting that survivin enhances angiogenesis by inhibiting apoptosis in endothelial cells (Kawasaki et al. 2001), and targeting survivin should play an important role in halting tumor progression, including blocking angiogenesis (Blanc-Brude et al. 2003; Tu et al. 2005; Xiang et al. 2005), which makes combination of bevacizumab and survivin antagonists not a good choice, considering the overlap effect between them.

The mechanisms of the chemosensitizing activity of antiangiogenic drugs are complicate and involved in many factors except inhibit survivin expression. Normalization of the tumor vasculature induced by antiangiogenic therapy might contribute to distribution of chemotherapeutic agents across the capillaries to the tumor cells (Jain 2005). In our results (Fig. 3c), the vessels’ phenotype was obviously normalized after bevacizumab treatment compare with control group, while in adriamycin group the vessels still abnormal although the vessel density was decreased too. Moreover, because bevacizumab alone can suppress tumor cell proliferation (Fig. 3c), the combination of a VEGF-targeting agent with chemotherapy would be expected to have an additive suppressive effect on tumor growth. Furthermore, antiangiogenic therapy can enhance the efficacy of chemotherapy by reducing circulating endothelial progenitor cells (EPCs) (Shaked et al. 2005) and tumor stem-like cell fraction (Folkins et al. 2007).

In conclusion, this study demonstrated that bevacizumab enhances the chemosensitivity of HUVECs by inhibiting the VEGF-PI3 K/Akt-survivin signaling cascade. Combination of bevacizumab with adriamycin therapy resulted in better outcomes, associated with reduced survivin expression in tumor tissues, including tumor endothelial cells. Furthermore, knockdown of survivin expression before treating with bevacizumab and adriamycin will help to clarify this mechanism.

Acknowledgments

This work was supported by National Natural Science Foundation of China (30731160005, 30872504), China National “211” Project for Higher Education, and National Key Science and Technology Specific Project (2008ZX10002-019, 021).

Conflict of interest statement

None.

Footnotes

Y.-Q. Xiong and H.-C. Sun contributed equally to this work.

References

  1. Altieri DC (2008) Survivin, cancer networks and pathway-directed drug discovery. Nat Rev Cancer 8:61–70 [DOI] [PubMed] [Google Scholar]
  2. Altznauer F, Martinelli S, Yousefi S, Thurig C, Schmid I, Conway EM, Schoni MH, Vogt P, Mueller C, Fey MF, Zangemeister-Wittke U, Simon HU (2004) Inflammation-associated cell cycle-independent block of apoptosis by survivin in terminally differentiated neutrophils. J Exp Med 199:1343–1354 [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bergers G, Benjamin LE (2003) Tumorigenesis and the angiogenic switch. Nat Rev Cancer 3:401–410 [DOI] [PubMed] [Google Scholar]
  4. Blanc-Brude OP, Mesri M, Wall NR, Plescia J, Dohi T, Altieri DC (2003) Therapeutic targeting of the survivin pathway in cancer: initiation of mitochondrial apoptosis and suppression of tumor-associated angiogenesis. Clin Cancer Res 9:2683–2692 [PubMed] [Google Scholar]
  5. Bruix J, Llovet JM (2002) Prognostic prediction and treatment strategy in hepatocellular carcinoma. Hepatology 35:519–524 [DOI] [PubMed] [Google Scholar]
  6. Caldas H, Fangusaro JR, Boue DR, Holloway MP, Altura RA (2007) Dissecting the role of endothelial SURVIVIN DeltaEx3 in angiogenesis. Blood 109:1479–1489 [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Cosgrave N, Hill AD, Young LS (2006) Growth factor-dependent regulation of survivin by c-myc in human breast cancer. J Mol Endocrinol 37:377–390 [DOI] [PubMed] [Google Scholar]
  8. Ferrara N (2005) The role of VEGF in the regulation of physiological and pathological angiogenesis. EXS 94:209–231 [DOI] [PubMed] [Google Scholar]
  9. Fields AC, Cotsonis G, Sexton D, Santoianni R, Cohen C (2004) Survivin expression in hepatocellular carcinoma: correlation with proliferation, prognostic parameters, and outcome. Mod Pathol 17:1378–1385 [DOI] [PubMed] [Google Scholar]
  10. Finn RS, Zhu AX (2009) Targeting angiogenesis in hepatocellular carcinoma: focus on VEGF and bevacizumab. Expert Rev Anticancer Ther 9:503–509 [DOI] [PubMed] [Google Scholar]
  11. Folkins C, Man S, Xu P, Shaked Y, Hicklin DJ, Kerbel RS (2007) Anticancer therapies combining antiangiogenic and tumor cell cytotoxic effects reduce the tumor stem-like cell fraction in glioma xenograft tumors. Cancer Res 67:3560–3564 [DOI] [PubMed] [Google Scholar]
  12. Hurwitz H, Fehrenbacher L, Novotny W, Cartwright T, Hainsworth J, Heim W, Berlin J, Baron A, Griffing S, Holmgren E, Ferrara N, Fyfe G, Rogers B, Ross R, Kabbinavar F (2004) Bevacizumab plus irinotecan, fluorouracil, and leucovorin for metastatic colorectal cancer. N Engl J Med 350:2335–2342 [DOI] [PubMed] [Google Scholar]
  13. Ito T, Shiraki K, Sugimoto K, Yamanaka T, Fujikawa K, Ito M, Takase K, Moriyama M, Kawano H, Hayashida M, Nakano T, Suzuki A (2000) Survivin promotes cell proliferation in human hepatocellular carcinoma. Hepatology 31:1080–1085 [DOI] [PubMed] [Google Scholar]
  14. Jain RK (2005) Normalization of tumor vasculature: an emerging concept in antiangiogenic therapy. Science 307:58–62 [DOI] [PubMed] [Google Scholar]
  15. Kawasaki H, Toyoda M, Shinohara H, Okuda J, Watanabe I, Yamamoto T, Tanaka K, Tenjo T, Tanigawa N (2001) Expression of survivin correlates with apoptosis, proliferation, and angiogenesis during human colorectal tumorigenesis. Cancer 91:2026–2032 [DOI] [PubMed] [Google Scholar]
  16. Kerbel RS (2006) Antiangiogenic therapy: a universal chemosensitization strategy for cancer? Science 312:1171–1175 [DOI] [PubMed] [Google Scholar]
  17. Le Gouill S, Podar K, Amiot M, Hideshima T, Chauhan D, Ishitsuka K, Kumar S, Raje N, Richardson PG, Harousseau JL, Anderson KC (2004) VEGF induces Mcl-1 up-regulation and protects multiple myeloma cells against apoptosis. Blood 104:2886–2892 [DOI] [PubMed] [Google Scholar]
  18. Li F, Ling X (2006) Survivin study: an update of “what is the next wave”? J Cell Physiol 208:476–486 [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Marusawa H, Matsuzawa S, Welsh K, Zou H, Armstrong R, Tamm I, Reed JC (2003) HBXIP functions as a cofactor of survivin in apoptosis suppression. EMBO J 22:2729–2740 [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Mesri M, Morales-Ruiz M, Ackermann EJ, Bennett CF, Pober JS, Sessa WC, Altieri DC (2001) Suppression of vascular endothelial growth factor-mediated endothelial cell protection by survivin targeting. Am J Pathol 158:1757–1765 [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Miller K, Wang M, Gralow J, Dickler M, Cobleigh M, Perez EA, Shenkier T, Cella D, Davidson NE (2007) Paclitaxel plus bevacizumab versus paclitaxel alone for metastatic breast cancer. N Engl J Med 357:2666–2676 [DOI] [PubMed] [Google Scholar]
  22. Moriai R, Tsuji N, Moriai M, Kobayashi D, Watanabe N (2008) Survivin plays as a resistant factor against tamoxifen-induced apoptosis in human breast cancer cells. Breast Cancer Res Treat 117:261–271 [DOI] [PubMed] [Google Scholar]
  23. Nakagawa Y, Abe S, Kurata M, Hasegawa M, Yamamoto K, Inoue M, Takemura T, Suzuki K, Kitagawa M (2006) IAP family protein expression correlates with poor outcome of multiple myeloma patients in association with chemotherapy-induced overexpression of multidrug resistance genes. Am J Hematol 81:824–831 [DOI] [PubMed] [Google Scholar]
  24. O’Connor DS, Schechner JS, Adida C, Mesri M, Rothermel AL, Li F, Nath AK, Pober JS, Altieri DC (2000) Control of apoptosis during angiogenesis by survivin expression in endothelial cells. Am J Pathol 156:393–398 [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Podar K, Tonon G, Sattler M, Tai YT, Legouill S, Yasui H, Ishitsuka K, Kumar S, Kumar R, Pandite LN, Hideshima T, Chauhan D, Anderson KC (2006) The small-molecule VEGF receptor inhibitor pazopanib (GW786034B) targets both tumor and endothelial cells in multiple myeloma. Proc Natl Acad Sci USA 103:19478–19483 [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Riedel F, Gotte K, Goessler U, Sadick H, Hormann K (2004) Targeting chemotherapy-induced VEGF up-regulation by VEGF antisense oligonucleotides in HNSCC cell lines. Anticancer Res 24:2179–2183 [PubMed] [Google Scholar]
  27. Sakurai T, Maeda S, Chang L, Karin M (2006) Loss of hepatic NF-kappa B activity enhances chemical hepatocarcinogenesis through sustained c-Jun N-terminal kinase 1 activation. Proc Natl Acad Sci U S A 103:10544–10551 [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Shaked Y, Bertolini F, Man S, Rogers MS, Cervi D, Foutz T, Rawn K, Voskas D, Dumont DJ, Ben-David Y, Lawler J, Henkin J, Huber J, Hicklin DJ, D’Amato RJ, Kerbel RS (2005) Genetic heterogeneity of the vasculogenic phenotype parallels angiogenesis; Implications for cellular surrogate marker analysis of antiangiogenesis. Cancer Cell 7:101–111 [DOI] [PubMed] [Google Scholar]
  29. Singh RP, Dhanalakshmi S, Agarwal C, Agarwal R (2005) Silibinin strongly inhibits growth and survival of human endothelial cells via cell cycle arrest and downregulation of survivin, Akt and NF-kappaB: implications for angioprevention and antiangiogenic therapy. Oncogene 24:1188–1202 [DOI] [PubMed] [Google Scholar]
  30. Tran J, Master Z, Yu JL, Rak J, Dumont DJ, Kerbel RS (2002) A role for survivin in chemoresistance of endothelial cells mediated by VEGF. Proc Natl Acad Sci U S A 99:4349–4354 [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Tu SP, Cui JT, Liston P, Xiao HJ, Xu R, Lin MC, Zhu YB, Zou B, Ng SS, Jiang SH, Xia HH, Wong WM, Chan AO, Yuen MF, Lam SK, Kung HF, Wong BC (2005) Gene therapy for colon cancer by adeno-associated viral vector-mediated transfer of survivin Cys84Ala mutant. Gastroenterology 128:361–375 [DOI] [PubMed] [Google Scholar]
  32. Virrey JJ, Guan S, Li W, Schonthal AH, Chen TC, Hofman FM (2008) Increased survivin expression confers chemoresistance to tumor-associated endothelial cells. Am J Pathol 173:575–585 [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Xiang R, Mizutani N, Luo Y, Chiodoni C, Zhou H, Mizutani M, Ba Y, Becker JC, Reisfeld RA (2005) A DNA vaccine targeting survivin combines apoptosis with suppression of angiogenesis in lung tumor eradication. Cancer Res 65:553–561 [PubMed] [Google Scholar]
  34. Xin H, Zhang C, Herrmann A, Du Y, Figlin R, Yu H (2009) Sunitinib inhibition of Stat3 induces renal cell carcinoma tumor cell apoptosis and reduces immunosuppressive cells. Cancer Res 69:2506–2513 [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Yamamoto H, Ngan CY, Monden M (2008) Cancer cells survive with survivin. Cancer Sci 99:1709–1714 [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Yang L, Cao Z, Li F, Post DE, Van Meir EG, Zhong H, Wood WC (2004) Tumor-specific gene expression using the survivin promoter is further increased by hypoxia. Gene Ther 11:1215–1223 [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Zaffaroni N, Daidone MG (2002) Survivin expression and resistance to anticancer treatments: perspectives for new therapeutic interventions. Drug Resist Updat 5:65–72 [DOI] [PubMed] [Google Scholar]
  38. Zaffaroni N, Pennati M, Daidone MG (2005) Survivin as a target for new anticancer interventions. J Cell Mol Med 9:360–372 [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Zhu AX, Blaszkowsky LS, Ryan DP, Clark JW, Muzikansky A, Horgan K, Sheehan S, Hale KE, Enzinger PC, Bhargava P, Stuart K (2006) Phase II study of gemcitabine and oxaliplatin in combination with bevacizumab in patients with advanced hepatocellular carcinoma. J Clin Oncol 24:1898–1903 [DOI] [PubMed] [Google Scholar]

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