Prognostic impact of δ-like ligand 4 and Notch1 in acute myeloid leukemia
- Authors:
- Published online on: July 31, 2012 https://doi.org/10.3892/or.2012.1943
- Pages: 1503-1511
Abstract
Introduction
Angiogenesis, the process of new blood vessel formation from an existing network of vasculature, plays a key role in the growth and metastatic spread of solid tumors (1,2). Emerging evidence underscores the potential role of bone marrow (BM) vascularization, indicative of angiogenesis, in the progression and chemosensitivity of hematological malignancies, including acute myeloid leukemia (AML) (3,4). Moreover, the BM microenvironment has been demonstrated to regulate the angiogenic switch and give rise to novel vascularization by controlling the balance between pro- and anti-angiogenic factors (5,6). Among them, the vascular endothelial growth factor (VEGF) and angiopoietin (Ang) are 2 of the most important families involved in the regulation of angiogenesis (7).
The VEGF family is essential for the initiation of vascular development, while the Ang family is responsible for vascular maturation and stabilization (8). VEGF-A (from here on termed VEGF) is one of the most prominent angiogenic factors and it induces a pro-angiogenic and permeability-enhancing signal via the tyrosine kinase receptor, VEGFR-2 (9,10). However, signaling through VEGFR-1 and the regulation of angiogenesis by this receptor is more complex (9,10). Additionally, Ang-1 and -2 can bind to the Tie2 receptor, although they have opposite effects on Tie2 activation (11,12). Ang-1 functions as a stabilizing signal for mature vasculature, whereas Ang-2 may be regarded as a regulator of vessel plasticity (11). Dysregulation of the VEGF and Ang families is suggested to have a major impact on leukemic growth and has been established as a crucial step in the development of AML (13,14). Indeed, elevated levels of VEGF and Ang-2 have been reported in patients with de novo AML and appear to be negative prognostic factors in AML (15–18).
The Notch pathway is an evolutionarily conserved intercellular pathway affecting a myriad of cellular activities, such as cell survival, proliferation, migration and invasion (19), and thereby may contribute to leukemogenesis (20,21). Recently, the Notch pathway, particularly the vascular-specific δ-like ligand 4 (Dll4), has been identified as another pivotal pathway in the regulation of vascular development (22). The activation of the Notch pathway is initiated by ligand binding to Notch receptors mainly between adjacent cells, and there seems to be an important Notch cross-signaling between tumor and endothelial cells that promotes angiogenesis (23). Haploinsufficiency of Dll4 is essential for embryonic vascular development, and Dll4-mediated Notch signaling has a unique role in regulating endothelial cell proliferation and differentiation (24,25). Studies have demonstrated that the Notch pathway displays either increased or decreased angiogenic processes, such as endothelial proliferation, migration and tube formation in vitro, depending on the cell type and context (22,23). Furthermore, the prognostic impact of the Notch pathway in different malignancies has demonstrated a diverging pattern (26), and thus exploring the specific prognostic impact in AML angiogenesis is of substantial interest.
Previous studies have shown that VEGF and Ang act coordinately during vascular growth and remodeling (27). Moreover, the VEGF pathway acts as a potent upstream activating stimulus for angiogenesis, while the Notch/Dll4 pathway contributes to guide cell fate decisions that shape the activation appropriately (28,29). In addition, Notch1 signaling activates the Ang pathway, presenting that angiogenesis of endothelial cells is mediated in part through a Notch-dependent Ang-1/Tie2 pathway (30). In turn, Ang-1/Tie2 signaling potentiates basal Notch1 signaling controlling vascular quiescence by upregulating Dll4 (31). Since BM neovascularization depends on the mutual and coordinated interaction of various angiogenic factors from endothelial cells and leukemia cells (4,6,32), and many of these factors that are present in significant amounts in the BM microenvironment may function in a synchronized fashion, it will be of interest to comprehensively analyze the correlation among these factors and further assess the clinical implications.
In this study, we investigated the expression of 8 angiogenesis-related factors (Dll4, Notch1, VEGF, VEGFR-1, VEGFR-2, Ang-1, Ang-2 and Tie2) in BM mononuclear cells of adult patients with untreated AML. Furthermore, we assessed the prognostic impact of Dll4 and Notch1 expression levels and their association with the VEGF and Ang families.
Materials and methods
Patient samples
Sixty adult patients with untreated AML (31 males and 29 females; median age, 45 years; range, 14–73 years) were enrolled in this study. Diagnoses were established according to the French-American-British (FAB) criteria (33). All the patients with non-M3 AML subtypes underwent standard induction chemotherapy with one of the anthracyclines (doxorubicin or idarubicin) for 3 days and cytarabine for 7 days. The patients with acute promyelocytic leukemia (subtype M3) received all-trans retinoic acid with or without concurrent induction chemotherapy. After the patients achieved complete remission (CR), they underwent consolidation chemotherapy with a conventional dose of cytarabine and one anthracycline or with a high dose of cytarabine. The control group consisted of 40 healthy adult donors (22 males and 18 females; median age, 44 years; range, 24–68 years), with normal BM morphology as demonstrated by cytological and histological analyses. Chromosomal analyses for newly diagnosed patients were conducted on BM cells after 1–3 days of unstimulated culture and karyotyped according to the International System for Human Cytogenetic Nomenclature (ISCN). The median follow-up duration was 25 months. This study protocol was approved by the Medical Ethics Committee of the Affiliated Qilu Hospital of Shandong University, Jinan, China, and written informed consent was obtained from all patients. The clinical information for the 60 AML patients is presented in Table I.
Real-time reverse transcription-PCR (RT-PCR) analysis
Total RNA was isolated using TRIzol (Invitrogen, Carlsbad, CA, USA) and cDNA was prepared using M-MLV Reverse Transcriptase (Promega, Madison, WI, USA) according to the manufacturer's instructions. Real-time RT-PCR was carried out using an ABI Prism 7500 sequence detection system (Applied Biosystems, Foster City, CA, USA) and performed with SYBR-Green PCR Master Mix (Toyobo, Osaka, Japan) in a 20 μl reaction volume as described previously (34). The specificity of the desired products was documented with melting curves analysis and examined using electrophoresis on a 2% agarose gel. Each sample was tested twice, while PCR-grade water instead of template cDNA was used as the negative controls. A comparative CT method (2−ΔΔCT) was used to analyze the relative changes in gene expression. The results were expressed relative to the number of GAPDH transcripts used as the internal control. The sequences of primers of different angiogenic factors are listed in Table II.
Western blot analysis for Dll4 and Notch1 protein
The cells were lysed in lysis buffer [50 mmol/l Tris (pH 7.5), 100 mmol/l NaCl, 1 mmol/l EDTA, 0.5% NP40, 0.5% Triton X-100, 2.5 mmol/l sodium orthovanadate, 10 μl/ml protease inhibitor cocktail, 1 mmol/l phenylmethylsulfonyl fluoride] for 30 min at 4°C. Total proteins were fractionated using SDS-PAGE and were then transferred onto nitrocellulose membranes. The membranes were then incubated with the appropriate primary antibodies (Dll4 and Notch1), followed by incubation with a secondary HRP-conjugated antibody (Jingmei Co., Ltd., Beijing, China). The probed proteins were detected using enhanced chemiluminescent reagents (Amersham Pharmacia Biotech, Piscataway, NJ, USA). The two-dimensional optical densities of proteins on the film were quantified and analyzed using the Quantity One software (Bio-Rad, Hercules, CA, USA).
Statistical analysis
The Mann-Whitney U test, analysis of variance (ANOVA) and the Kruskal-Wallis test were used to analyze the differences in the expression of angiogenic factors between the AML and control groups, as well as differences in age, gender, FAB subtype and karyotype subgroups of the AML patients. Spearman's test was used to evaluate the correlation between the individual expression of the genes studied, and the association of gene expression with clinical features. Furthermore, overall survival (OS) was defined as the number of days from the date of the first diagnosis to death from any cause. Patients were categorized into high and low angiogenic factors expressing subgroups using the median value as the cut-off. Kaplan-Meier estimation was applied to plot survival curves, and a log-rank test was used to compare survival between groups. Univariate Cox proportional hazards regression models were used to evaluate the predictive effect of each factor alone on survival. Factors statistically significant in the univariate models at a 5% level were included in the multivariate models. Subsequently, multivariate models were reduced one factor at a time such that all factors remaining in the model were statistically significant at a 5% significance level. Multivariate Cox models estimated the hazard ratio in terms of relative risk (RR) for death, and 95% confidence interval (CI) to determine independent risk factors associated with survival. Two-sided P<0.05 were considered statistically significant. All statistical analyses were performed with the SPSS 13 software (SPSS Inc., Chicago, IL, USA).
Results
Aberrant expression profile of angiogenic factors in AML patients
As a ratio with the expression of the housekeeping gene GAPDH, BM levels of the 8 angiogenesis-related factors were compared between the untreated AML patients and the healthy controls. In spite of the wide range of individual values, median levels of Notch1 (P=0.012), Dll4 (P=0.001), VEGF (P=0.005), VEGFR-2 (P=0.009) and Ang-2 (P=0.006) were significantly elevated in the AML patients compared with the controls, while the median levels of VEGFR-1 (P=0.439), Ang-1 (P=0.123) and Tie2 (P=0.733) were similar in AML and control patients (Fig. 1A).
It is important to note that the active functional form of Notch1 is the Notch1 intracellular domain. Therefore, we focused our study on the Notch1 intracellular domain at the protein level and Notch1 in the figure legends indicates the active functional form of Notch1. In concordance with the real-time RT-PCR data, western blot analysis showed that the protein expression levels of Notch1 and Dll4 were upregulated in the untreated AML patients compared to the controls (P<0.001, Fig. 1B). Taken together, these data indicate that the Notch/Dll4 pathway is aberrantly activated in AML.
It has been reported that BM vascularization is regulated by the relative balance between angiogenic activators and inhibitors (4,35). Therefore, we investigated the correlation between the angiogenic factors studied in AML patients. Significantly positive correlations were observed between Notch1 and Dll4 (r=0.326; P=0.013), VEGF and VEGFR-2 (r=0.354; P=0.007) and Ang-1 and Tie2 (r=0.463; P=0.001). Furthermore, Dll4 was closely associated with VEGF (r=0.663; P=0.001) and Notch1 correlated well with Ang-2 (r=0.324; P=0.012).
VEGF receptors expressed in leukemic cells are functional and convey distinct signals, such as increasing proliferation, metalloproteinase activation and trans-membrane migration (9). The degree of Tie2 receptor activation depends on the balance between agonistic and antagonistic ligands (11). Thus, we further calculated the VEGFR-2:VEGFR-1 and Ang-2:Ang-1 expression ratio in normal and leukemic cells. The median VEGFR-2:VEGFR-1 expression ratio was 3.61-fold higher in the AML patients than in the controls (P=0.04), suggesting that the pathway enhanced by the VEGF/VEGFR-2 interaction may be essential for the pathogenesis of AML. The median Ang-2:Ang-1 expression ratio was 3.18-fold higher in the AML patients than in the controls (P=0.015), indicating a dominant influence of Ang-2 in the neoplastic BM and Ang-1 in the normal BM microenvironment.
Correlation of angiogenic factor expression with clinical features
The association between BM levels of these 8 angiogenic factors in the 60 AML patients and clinical features were investigated, and significant differences were not observed in the gender and FAB subtypes (Table III). However, the Tie-2 level was higher in AML patients <45 years old than those >45 years old (P=0.03), and the levels of Dll4 and VEGF were significantly different among the karyotype subgroups (P=0.001). The BM of the AML patients studied was highly infiltrated by leukemic blasts. The median (range) percentage of blasts was 70% (range 40–98%). Significantly positive associations of the percentage of leukemic blast infiltration were observed with the individual expression of Notch1 (r=0.428; P=0.001), Dll4 (r=0.288; P=0.026), VEGF (r=0.428; P=0.001) and Ang-2 (r=0.474; P<0.001).
Association between angiogenic factor expression and clinical outcome
We subsequently investigated the correlation between pre-therapeutic angiogenic factor expression and prognosis in AML. The median OS was 25 months (range, 7.0–45.9 months). Univariate analysis of the factors associated with OS revealed a significantly shorter survival in the patients with the unfavorable karyotype, higher Notch1 expression, higher Dll4 expression, higher VEGF expression or higher Ang-2 expression (Table IV). Variables, such as age, gender, hemoglobin levels, white blood cell (WBC) counts, platelet counts, VEGFR-1, VEGFR-2, Ang-1 and Tie2 expression, as well as VEGFR-2:VEGFR-1 and Ang-2:Ang-1 expression ratio had no impact. Cox proportional hazards multivariate analysis of the univariate predictors identified karyotype (RR, 3.19; 95% CI, 1.18–8.63; P=0.22) and gene expression of Notch1 (RR, 5.04; 95% CI, 1.62–15.69; P=0.005), Dll4 (RR, 8.09; 95% CI, 1.72–37.95; P=0.008), VEGF (RR, 5.25; 95% CI, 1.17–23.48; P=0.03) and Ang-2 (RR, 11.25; 95% CI, 3.09–41.01; P=0.001) as prognostic factors for OS. The Kaplan-Meier curves for OS stratified according to Notch1, Dll4, VEGF and Ang-2 expression in BM leukemia cells are shown in Fig. 2A-D.
In order to further illustrate the impact of Notch1 and Dll4 expression on OS, the Kaplan-Meier survival curves of 3 cytogenetic-risk groups were analyzed. Dll4 expression had a similar impact on OS in all the patients and those with the intermediate-risk karyotype, whereas Notch1 expression lost its predictive value for survival in patients with the intermediate-risk karyotype (Fig. 2E and F). However, there was no survival difference between patients with a low and high expression of Notch1 or Dll4 in the favorable-risk group and unfavorable-risk group.
Finally, we performed subgroup analyses to investigate potential interactions between the Notch/Dll4 pathway and other angiogenic factors on OS. Patients were divided into 2 groups with a low or high expression (respectively, below or above the median level) of VEGF and Ang-2. Survival estimates were calculated for each group according to the Notch1 and Dll4 expression levels. The survival difference between patients with a low and high Notch1 expression was pronounced in the subgroups with high VEGF levels (P=0.001) and high Ang-2 levels (P=0.001), respectively, but not evident in the patients with low values (Fig. 3A and B). However, the outcome was insignificant in patients with a high compared to a low Dll4 expression in the presence of high VEGF expression (P=0.001, Fig. 3C). The impact of the Dll4 expression on OS was equally prominent in the subgroups of AML patients expressing low and high levels of Ang-2 (P=0.001, Fig. 3D).
Discussion
During the past few years, it has been proposed that AML may be an angiogenesis-dependent disease with progressive recruitment of blood vessels in the BM microenvironment (35). Previous studies have demonstrated a close correlation between leukemogenesis and dysregulation of angiogenesis, including increased BM vascularity (3) and elevated levels of numerous angiogenic factors (7,13). Our present study clearly demonstrates an elevated expression of Notch1, Dll4, VEGF, VEGFR-2 and Ang-2 in the BM leukemia cells of patients with untreated AML, compared with the controls. Cox regression analyses revealed that the expression levels of Notch1, Dll4, VEGF and Ang-2 were predictors of poor OS, independent of cytogenetics. Furthermore, the prognostic relevance of Dll4 expression was more evident in patients with the intermediate-risk karyotype (62.5% of patients with low Dll4 expression vs. 33.3% with high level in this subgroup survived, P=0.001). Therefore, the Dll4 expression may serve as a potential biomarker for prognosis prediction, particularly in the intermediate-risk cytogenetic group.
Aberrant Notch signaling contributes to the genesis of diverse cancers (26). The fact that constitutively active Notch1 transforms cells in vitro and causes mice to develop leukemia indicates the Notch pathway involvement in the development of leukemia (21). Conversely, Notch is a tumor suppressor in certain epithelial cancers where its normal function is to promote terminal differentiation (36). Our present study shows that patients with untreated AML express higher levels of Notch1 and Dll4 compared to healthy controls, as assessed by real-time RT-PCR and western blot analysis. Moreover, the high expression levels of Notch1 and Dll4 represent a significant feature of AML patients. Patients with high Notch1 and Dll4 expression levels show several adverse prognostic indicators, such as high BM blast counts and unfavorable karotype. Recent studies have shown that reciprocal Notch signaling may be necessary for the proliferation and survival of AML cells, possibly through the maintenance of c-Myc and Bcl2, as well as the phosphorylation of Rb protein (37,38). Furthermore, the activation of the Notch pathway has been reported to be associated with an adverse outcome in various solid cancers, such as lung (39), breast (40) and bladder cancer (41). Consistent with these findings, our study also demonstrates that there is an inverse correlation between Notch1/Dll4 expression and OS in AML. Collectively, our data provide evidence for the critical role of the Notch/Dll4 pathway activation in the pathogenesis and prognosis of AML.
Elegant genetic and molecular studies have suggested that the Notch/Dll4 pathway interacts extensively with the VEGF pathway in endothelial cells (42,43). Indeed, Dll4 expression may be induced by VEGF and hypoxia (42). Conversely, Dll4 is considered as a selective inhibitor of VEGF biological activity by downregulating VEGFR-2 (43). Recently, emphasis has been placed on the interaction of the Notch/Dll4 and VEGF pathways in tumor angiogenesis (28,29). Our study shows that Dll4 positively correlates with VEGF in AML patients. Moreover, the poor prognosis in patients with high Notch1 and Dll4 expression levels was only evident in the presence of higher (not lower) levels of VEGF, suggesting that the angiogenic effect of Notch/Dll4 pathway may be dependent on VEGF. Donnem et al also reported that Notch1 and Dll4 were independent prognostic factors for non-small cell lung cancer, and the influence of Notch1 was enhanced in the presence of high VEGF expression (39). Thus, our results and published reports confirm the hypothesis of a cross-talk between Notch/Dll4 and VEGF pathways in tumor angiogenesis.
Notch1 signaling has been shown to activate the Ang pathway, and Ang-1/Tie2 signaling augments basal Notch signaling by inducing Dll4 expression through the Akt-mediated activation of β-catenin (30,31). The fact that Notch1 lost its impact on survival in the patients with low Ang-2 expression, as revealed in our study, suggests these 2 factors may function in a synergetic fashion; however, the mechanism involved remains unclear. The grade of Tie2 phosphorylation is determined by the balance between Ang-1 and its naturally occurring antagonist Ang-2 (11). Owing to low Ang-2 expression, Ang-1-mediated Tie2 activation was relatively enhanced, resulting in the activation of the Akt pathway, which is associated with increased survival and the anti-apoptotic effect (44,45). Therefore, the prognostic impact of Notch1 expression may be lessened in the presence of low Ang-2 expression.
Previous studies have led to the proposal that VEGFR-2 is the critical receptor for transmitting cellular signals for the proliferation and differentiation of endothelial cells, whereas VEGFR-1 may be more important for vascular remodeling and monocyte migration (9). Additionally, several studies have shown that Ang-1 mediates vascular stability, while Ang-2 induces vascular instability by overriding Ang-1-mediated Tie2 activation (11). Therefore, it is important to elucidate the balance between VEGFR-1 and VEGFR-2, Ang-1 and Ang-2 in the BM microenvironment. In this study, we observed a difference in the VEGFR-2:VEGFR-1 and Ang-2:Ang-1 expression ratio between AML patients and healthy controls, indicating that the angiogenic balance is altered in favor of VEGFR-2 and Ang-2 in AML. Indeed, a high ratio of Ang-2:Ang-1 expression in association with high levels of VEGF, bFGF and other mitogenic angiogenic factors stimulates angiogenesis in a wide range of solid tumors (11). Thus, our data provide evidence that the alteration of the angiogenic balance in favor of Ang-2 acting in concert with VEGFR-2 may be essential for BM neovascularization in AML. However, we could not predict the prognosis of the AML patients based on the VEGFR-2:VEGFR-1 and Ang-2:Ang-1 expression ratio.
In conclusion, BM angiogenesis in AML is a complex process involving the interplay of various angiogenic factors. Most investigators have attempted to elucidate the impact of a single angiogenic factor on the pathogenesis or prognosis of hematologic malignancies. Our present study not only investigated the prognostic role of the angiogenesis-related Dll4 and Notch1, but also assessed their association with the VEGF and Ang families. Our results provide evidence that the activation of the Notch/Dll4 pathway may indicate an unfavorable prognosis in AML. Furthermore, the role of the Notch/Dll4 pathway in AML angiogenesis and its correlation with VEGF and Ang-2 establishes the Notch/Dll4 pathway as a potentially important target of anti-angiogenic therapy. However, further prospective and multi-centre studies as well as in-depth studies are required to verify the role of Notch/Dll4 pathway in AML angiogenesis.
Acknowledgements
This study was supported by grants from the National Natural Science Foundation of China [81070422, 30871088, 81070407, 81000223]; the Specialized Research Fund for the Doctoral Program of Higher Education [20100131110060]; the Shandong Technological Development Project [2009HD012, 2009GG20002020, BS2009SW014, ZR2010HQ030] and the Independent Innovation Fund of Shandong University [yzc10072, yzc10075].
References
|
Zhang Y, Tang H, Cai J, et al: Ovarian cancer-associated fibroblasts contribute to epithelial ovarian carcinoma metastasis by promoting angiogenesis, lymphangiogenesis and tumor cell invasion. Cancer Lett. 303:47–55. 2011. View Article : Google Scholar | |
|
Hao CY: Angiogenesis blockade as therapy for hepatocellular carcinoma: progress and challenges. J Gastroenterol Hepatol. 26:4–6. 2011. View Article : Google Scholar : PubMed/NCBI | |
|
Shih TT, Hou HA, Liu CY, et al: Bone marrow angiogenesis magnetic resonance imaging in patients with acute myeloid leukemia: peak enhancement ratio is an independent predictor for overall survival. Blood. 113:3161–3167. 2009. View Article : Google Scholar | |
|
Trujillo A, McGee C and Cogle CR: Angiogenesis in acute myeloid leukemia and opportunities for novel therapies. J Oncol. 2012:1286082012. View Article : Google Scholar : PubMed/NCBI | |
|
Payne SJ and Jones L: Influence of the tumor microenvironment on angiogenesis. Future Oncol. 7:395–408. 2011. View Article : Google Scholar : PubMed/NCBI | |
|
Ayala F, Dewar R, Kieran M and Kalluri R: Contribution of bone microenvironment to leukemogenesis and leukemia progression. Leukemia. 23:2233–2241. 2009. View Article : Google Scholar : PubMed/NCBI | |
|
Hou HA, Chou WC, Lin LI, et al: Expression of angiopoietins and vascular endothelial growth factors and their clinical significance in acute myeloid leukemia. Leuk Res. 32:904–912. 2008. View Article : Google Scholar : PubMed/NCBI | |
|
Carmeliet P and Jain RK: Molecular mechanisms and clinical applications of angiogenesis. Nature. 473:298–307. 2011. View Article : Google Scholar : PubMed/NCBI | |
|
Wegiel B, Ekberg J, Talasila KM, Jalili S and Persson JL: The role of VEGF and a functional link between VEGF and p27Kip1 in acute myeloid leukemia. Leukemia. 23:251–261. 2009. View Article : Google Scholar : PubMed/NCBI | |
|
Schmidt T and Carmeliet P: Angiogenesis: a target in solid tumors, also in leukemia? Hematology Am Soc Hematol Educ Program. 2011:1–8. 2011. View Article : Google Scholar : PubMed/NCBI | |
|
Tait CR and Jones PF: Angiopoietins in tumours: the angiogenic switch. J Pathol. 204:1–10. 2004. View Article : Google Scholar : PubMed/NCBI | |
|
Reikvam H, Hatfield KJ, Lassalle P, Kittang AO, Ersvaer E and Bruserud O: Targeting the angiopoietin (Ang)/Tie-2 pathway in the crosstalk between acute myeloid leukaemia and endothelial cells: studies of Tie-2 blocking antibodies, exogenous Ang-2 and inhibition of constitutive agonistic Ang-1 release. Expert Opin Investig Drugs. 19:169–183. 2010. View Article : Google Scholar | |
|
Maffei R, Martinelli S, Castelli I, et al: Increased angiogenesis induced by chronic lymphocytic leukemia B cells is mediated by leukemia-derived Ang2 and VEGF. Leuk Res. 34:312–321. 2010. View Article : Google Scholar : PubMed/NCBI | |
|
Keith T, Araki Y, Ohyagi M, et al: Regulation of angiogenesis in the bone marrow of myelodysplastic syndromes transforming to overt leukaemia. Br J Haematol. 137:206–215. 2007. View Article : Google Scholar : PubMed/NCBI | |
|
Aguayo A, Kantarjian HM, Estey EH, et al: Plasma vascular endothelial growth factor levels have prognostic significance in patients with acute myeloid leukemia but not in patients with myelodysplastic syndromes. Cancer. 95:1923–1930. 2002. View Article : Google Scholar | |
|
de Bont ES, Fidler V, Meeuwsen T, Scherpen F, Hahlen K and Kamps WA: Vascular endothelial growth factor secretion is an independent prognostic factor for relapse-free survival in pediatric acute myeloid leukemia patients. Clin Cancer Res. 8:2856–2861. 2002.PubMed/NCBI | |
|
Loges S, Heil G, Bruweleit M, et al: Analysis of concerted expression of angiogenic growth factors in acute myeloid leukemia: expression of angiopoietin-2 represents an independent prognostic factor for overall survival. J Clin Oncol. 23:1109–1117. 2005. View Article : Google Scholar | |
|
Schliemann C, Bieker R, Thoennissen N, et al: Circulating angiopoietin-2 is a strong prognostic factor in acute myeloid leukemia. Leukemia. 21:1901–1906. 2007. View Article : Google Scholar : PubMed/NCBI | |
|
Wu Y, Cain-Hom C, Choy L, et al: Therapeutic antibody targeting of individual Notch receptors. Nature. 464:1052–1057. 2010. View Article : Google Scholar : PubMed/NCBI | |
|
Demarest RM, Dahmane N and Capobianco AJ: Notch is oncogenic dominant in T-cell acute lymphoblastic leukemia. Blood. 117:2901–2909. 2011. View Article : Google Scholar : PubMed/NCBI | |
|
Leong KG and Karsan A: Recent insights into the role of Notch signaling in tumorigenesis. Blood. 107:2223–2233. 2006. View Article : Google Scholar : PubMed/NCBI | |
|
Phng LK and Gerhardt H: Angiogenesis: a team effort coordinated by notch. Dev Cell. 16:196–208. 2009. View Article : Google Scholar : PubMed/NCBI | |
|
Li JL and Harris AL: Notch signaling from tumor cells: a new mechanism of angiogenesis. Cancer Cell. 8:1–3. 2005. View Article : Google Scholar : PubMed/NCBI | |
|
Ridgway J, Zhang G, Wu Y, et al: Inhibition of Dll4 signalling inhibits tumour growth by deregulating angiogenesis. Nature. 444:1083–1087. 2006. View Article : Google Scholar : PubMed/NCBI | |
|
Noguera-Troise I, Daly C, Papadopoulos NJ, et al: Blockade of Dll4 inhibits tumour growth by promoting non-productive angiogenesis. Nature. 444:1032–1037. 2006. View Article : Google Scholar : PubMed/NCBI | |
|
Lobry C, Oh P and Aifantis I: Oncogenic and tumor suppressor functions of Notch in cancer: it's NOTCH what you think. J Exp Med. 208:1931–1935. 2011. View Article : Google Scholar | |
|
Lobov IB, Brooks PC and Lang RA: Angiopoietin-2 displays VEGF-dependent modulation of capillary structure and endothelial cell survival in vivo. Proc Natl Acad Sci USA. 99:11205–11210. 2002. View Article : Google Scholar : PubMed/NCBI | |
|
Li JL and Harris AL: Crosstalk of VEGF and Notch pathways in tumour angiogenesis: therapeutic implications. Front Biosci. 14:3094–3110. 2009.PubMed/NCBI | |
|
Thurston G and Kitajewski J: VEGF and Delta-Notch: interacting signalling pathways in tumour angiogenesis. Br J Cancer. 99:1204–1209. 2008. View Article : Google Scholar : PubMed/NCBI | |
|
Morrow D, Cullen JP, Cahill PA and Redmond EM: Ethanol stimulates endothelial cell angiogenic activity via a Notch- and angiopoietin-1-dependent pathway. Cardiovasc Res. 79:313–321. 2008. View Article : Google Scholar : PubMed/NCBI | |
|
Zhang J, Fukuhara S, Sako K, et al: Angiopoietin-1/Tie2 signal augments basal Notch signal controlling vascular quiescence by inducing delta-like 4 expression through AKT-mediated activation of beta-catenin. J Biol Chem. 286:8055–8066. 2011. View Article : Google Scholar | |
|
Kittang AO, Hatfield K, Sand K, Reikvam H and Bruserud O: The chemokine network in acute myelogenous leukemia: molecular mechanisms involved in leukemogenesis and therapeutic implications. Curr Top Microbiol Immunol. 341:149–172. 2010.PubMed/NCBI | |
|
Muller-Berndorff H, Haas PS, Kunzmann R, Schulte-Monting J and Lubbert M: Comparison of five prognostic scoring systems, the French-American-British (FAB) and World Health Organization (WHO) classifications in patients with myelodysplastic syndromes: Results of a single-center analysis. Ann Hematol. 85:502–513. 2006. View Article : Google Scholar | |
|
Ji M, Li J, Yu H, et al: Simultaneous targeting of MCL1 and ABCB1 as a novel strategy to overcome drug resistance in human leukaemia. Br J Haematol. 145:648–656. 2009. View Article : Google Scholar : PubMed/NCBI | |
|
Lee CY, Tien HF, Hu CY, Chou WC and Lin LI: Marrow angiogenesis-associated factors as prognostic biomarkers in patients with acute myelogenous leukaemia. Br J Cancer. 97:877–882. 2007.PubMed/NCBI | |
|
Ghaleb AM, Aggarwal G, Bialkowska AB, Nandan MO and Yang VW: Notch inhibits expression of the Kruppel-like factor 4 tumor suppressor in the intestinal epithelium. Mol Cancer Res. 6:1920–1927. 2008. View Article : Google Scholar : PubMed/NCBI | |
|
Li GH, Fan YZ, Liu XW, et al: Notch signaling maintains proliferation and survival of the HL60 human promyelocytic leukemia cell line and promotes the phosphorylation of the Rb protein. Mol Cell Biochem. 340:7–14. 2010. View Article : Google Scholar : PubMed/NCBI | |
|
Tohda S: Functional analysis of notch in the pathophysiology of leukemia. Rinsho Byori. 57:351–356. 2009.(In Japanese). | |
|
Donnem T, Andersen S, Al-Shibli K, Al-Saad S, Busund LT and Bremnes RM: Prognostic impact of Notch ligands and receptors in nonsmall cell lung cancer: coexpression of Notch-1 and vascular endothelial growth factor-A predicts poor survival. Cancer. 116:5676–5685. 2010. View Article : Google Scholar : PubMed/NCBI | |
|
Reedijk M, Odorcic S, Chang L, et al: High-level coexpression of JAG1 and NOTCH1 is observed in human breast cancer and is associated with poor overall survival. Cancer Res. 65:8530–8537. 2005. View Article : Google Scholar : PubMed/NCBI | |
|
Shi TP, Xu H, Wei JF, et al: Association of low expression of notch-1 and jagged-1 in human papillary bladder cancer and shorter survival. J Urol. 180:361–366. 2008. View Article : Google Scholar : PubMed/NCBI | |
|
Siekmann AF, Covassin L and Lawson ND: Modulation of VEGF signalling output by the Notch pathway. Bioessays. 30:303–313. 2008. View Article : Google Scholar : PubMed/NCBI | |
|
Williams CK, Li JL, Murga M, Harris AL and Tosato G: Up-regulation of the Notch ligand Delta-like 4 inhibits VEGF-induced endothelial cell function. Blood. 107:931–939. 2006. View Article : Google Scholar : PubMed/NCBI | |
|
Wakabayashi M, Miwa H, Shikami M, et al: Autocrine pathway of angiopoietins-Tie2 system in AML cells: association with phosphatidyl-inositol 3 kinase. Hematol J. 5:353–360. 2004. View Article : Google Scholar : PubMed/NCBI | |
|
Xu Q, Simpson SE, Scialla TJ, Bagg A and Carroll M: Survival of acute myeloid leukemia cells requires PI3 kinase activation. Blood. 102:972–980. 2003. View Article : Google Scholar : PubMed/NCBI |
