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ABSTRACT: Reactive oxygen species (ROS) are important in regulating normal cellular processes, but deregulated ROS contribute to the development of various human diseases including cancers. Autophagy is one of the first lines of defense against oxidative stress damage. The autophagy pathway can be induced and upregulated in response to intracellular ROS or extracellular oxidative stress. This leads to selective lysosomal self-digestion of intracellular components to maintain cellular homeostasis. Hence, autophagy is the survival pathway, conferring stress adaptation and promoting viability under oxidative stress. However, increasing evidence has demonstrated that autophagy can also lead to cell death under oxidative stress conditions. In addition, altered autophagic signaling pathways that lead to decreased autophagy are frequently found in many human cancers. This review discusses the advances in understanding of the mechanisms of ROS-induced autophagy and how this process relates to tumorigenesis and cancer therapy.
Free radical biology & medicine 07/2012; 53(7):1399-410. · 5.42 Impact Factor
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ABSTRACT: Autophagy is a cellular process that maintains the homeostasis of the normal cell. It not only allows for cell survival in times of metabolic stress with nutrient recycling but also is able to lead to cell death when required. During malignant transformation the cell is able to proliferate and survive. This is due to altered cell metabolism and the presence of altered genetic changes that maintain the cell survival. Metabolism was considered an innocent bystander that was a consequence of the increased nutrient requirement for the survival and proliferation of haematological malignancies. The interdependency of metabolism and cellular mechanisms such as autophagy are becoming more evident and important. This interdependence contributes to increased cancer progression and drug resistance. In this paper we aim to discuss autophagy, how it pertains to metabolism in the context of hematologic malignancies, and the implications for therapy.
International Journal of Cell Biology 01/2012; 2012:595976.
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ABSTRACT: Hematological malignancies are characterized by the accumulation of lymphoid and myeloid cell types due to selective proliferation and survival in blood, bone marrow and lymph nodes. Treatments of hematological malignancies are often effective but eventually relapse, and drug resistance occurs. A better understanding of the mechanism of action of both chemotherapeutic drugs and drug resistance is required. Autophagy has been shown to regulate both cell survival and cell death, leading to both cancer development and tumor suppression. In addition, many chemotherapeutic drugs induce autophagy, leading to either drug resistance or cell death. Autophagy is regulated by signaling pathways such as p53 and by the production of reactive oxygen species (ROS). This review focuses on the regulation of autophagy in human hematologic malignancy leading to either cell survival or death. In addition, the role that ROS play in regulating autophagy and its implication for hematological cancers is discussed.
Leukemia & lymphoma 07/2011; 53(1):26-33. · 2.40 Impact Factor
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ABSTRACT: Tumor necrosis related apoptosis inducing ligand receptor 1 (TRAIL-R1, death receptor 4 [DR4]) and TRAIL-R2 (DR5) have been proposed as targets for cancer therapy, but which death receptor to target for chemotherapy in chronic lymphocytic leukemia (CLL) is uncertain. Herein, we discovered that Burkitt lymphoma B cell line, BJAB, CLL-like cell line, I-83, and pre-acute lymphocytic leukemia B cell line, NALM-6, underwent apoptosis following TRAIL, whereas a CLL-like cell line, JMV-3, and primary CLL cells failed to undergo apoptosis. In TRAIL resistant CLL cells, only activation of DR4 provided an increase in fludarabine induced apoptosis. This was mediated in part by the localization of DR4 but not DR5 in lipid rafts following TRAIL and fludarabine treatment. This preference for DR4 activation leading to increased fludarabine induced apoptosis was also observed following SAHA, PS-341, and chlorambucil treatment in primary CLL cells. Thus, CLL cells selectively activate DR4 partially mediated through its localization to lipid rafts leading to apoptosis when combined with chemotherapeutic drugs.
Leukemia & lymphoma 07/2011; 52(7):1290-301. · 2.40 Impact Factor
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ABSTRACT: Cucurbitacin-I (JSI-124) is potent inhibitor of JAK/STAT3 signaling pathway and has anti-tumor activity in a variety of cancer including B cell leukemia. However, other molecular targets of JSI-124 beyond the JAK/STAT3 pathway are not fully understood.
BJAB, I-83, NALM-6 and primary CLL cells were treated with JSI-124 as indicated. Apoptosis was measured using flow cytometry for accumulation of sub-G1 phase cells (indicator of apoptosis) and Annexin V/PI staining. Cell cycle was analyzed by FACS for DNA content of G1 and G2 phases. Changes in phosphorylation and protein expression of p38, Erk1/2, JNK, c-Jun, and XIAP were detected by Western blot analysis. STAT3 and c-Jun genes were knocked out using siRNA transfection. VEGF expression was determined by mRNA and protein levels by RT-PCR and western blotting. Streptavidin Pull-Down Assay was used to determine c-Jun binding to the AP-1 DNA binding site.
Herein, we show that JSI-124 activates c-Jun N-terminal kinase (JNK) and increases both the expression and serine phosphorylation of c-Jun protein in the B leukemic cell lines BJAB, I-83 and NALM-6. JSI-124 also activated MAPK p38 and MAPK Erk1/2 albeit at lower levels than JNK activation. Inhibition of the JNK signaling pathway failed to effect cell cycle arrest or apoptosis induced by JSI-124 but repressed JSI-124 induced c-Jun expression in these leukemia cells. The JNK pathway activation c-Jun leads to transcriptional activation of many genes. Treatment of BJAB, I-83, and NALM-6 cells with JSI-124 lead to an increase of Vascular Endothelial Growth Factor (VEGF) at both the mRNA and protein level. Knockdown of c-Jun expression and inhibition of JNK activation significantly blocked JSI-124 induced VEGF expression. Pretreatment with recombinant VEGF reduced JSI-124 induced apoptosis.
Taken together, our data demonstrates that JSI-124 activates the JNK signaling pathway independent of apoptosis and cell cycle arrest, leading to increased VEGF expression.
BMC Cancer 06/2011; 11:268. · 3.01 Impact Factor
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ABSTRACT: BNIP3 (Bcl-2/adenovirus E1B Nineteen Kilodalton Interacting Protein) is a pro-cell death member of the Bcl-2 family of proteins. Its expression is induced by the transcription factor Hypoxia Inducible Factor-1 (HIF-1) under conditions of low oxygen (hypoxia) and is found over expressed in hypoxic regions of many tumors. When over expressed, BNIP3 induces cell death through induction of mitochondrial dysfunction that is dependent on the presence of BNIP3's TM domain. Herein, we have determined that the SkOv3 ovarian cancer cell line expresses a truncated BNIP3 protein, which results in the elimination of the transmembrane domain. Truncation that eliminates all four domains of BNIP3 protein also inhibits hypoxia-induced cell death in SkOv3, HEK293, U251 and MCF-7 cells. Three different mutations in a BNIP3 expression vector that lead to a truncated BNIP3 protein, lacking TM domain only, or lacking CD, BH3, and TM domains resulted in inhibition of hypoxia-induced cell death when transfected into HEK293 cells. We found that truncated BNIP3 failed to associate with the mitochondria and the truncated BNIP3 lacking all four domains can bind to wild type BNIP3. Taken together, truncation of BNIP3 could be a novel mechanism for cancer cells to avoid hypoxia-induced cell death mediated by BNIP3 over expression.
Biochimica et Biophysica Acta 03/2011; 1812(3):302-11. · 4.66 Impact Factor
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ABSTRACT: RNA species produced during virus replication are pathogen-associated molecular patterns (PAMPs) triggering cellular innate immune responses including induction of type I interferon expression and apoptosis. Pattern recognition receptors (PRRs) for these RNAs include the retinoic acid-inducible gene I (RIG-I) like receptors (RLRs) RIG-I and melanoma differentiation associated gene 5 (MDA5) and the dsRNA dependent protein kinase (PKR). Currently, poxvirus PAMPs and their associated PRRs are not well characterized. We report that RNA species generated in vaccinia infected cells can activate MDA5 or RIG-I dependent interferon-β (IFN-β) gene transcription in a cell type-specific manner. These RNA species also induce the activation of apoptosis in a PKR dependent, but MDA5 and RIG-I independent, manner. Collectively our results demonstrate that RNA species generated during vaccinia virus replication are major PAMPs activating apoptosis and IFN-β gene transcription. Moreover, our results delineate the signaling pathways involved in the recognition of RNA-based poxvirus PAMPs.
Virology 02/2011; 413(2):183-93. · 3.35 Impact Factor
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ABSTRACT: Phosphorylation of STAT3 on serine 727 regulates gene expression and is found to be elevated in many B-leukemia cells including chronic lymphocytic leukemia (CLL). It is, however, unclear whether targeting STAT3 will be an effective antileukemia therapy. In this study, we assessed in vitro antileukemia activity of the STAT3 inhibitor JSI-124 (cucurbitacin I). JSI-124 potently induces apoptosis in 3 B-leukemia cell lines (BJAB, I-83, and NALM-6) and in primary CLL cells and was associated with a reduction in serine 727 phosphorylation of STAT3. Similarly, knockdown of STAT3 expression induced apoptosis in these leukemia cells. In addition, we found that JSI-124 and knockdown of STAT3 decreased antiapoptotic protein XIAP expression and overexpression of XIAP blocked JSI-124-induced apoptosis. Furthermore, we found that combined treatment of JSI-124 and TRAIL increased apoptosis associated with an increase in death receptor 4 expression. Besides apoptosis, we found that JSI-124 also induced cell-cycle arrest prior to apoptosis in B-leukemia cells. This corresponded with reduced expression of the cell-cycle regulatory gene, cdc-2. Thus, we present here for the first time that JSI-124 induced suppression of serine 727 phosphorylation of STAT3, leading to apoptosis and cell-cycle arrest through alterations in gene transcription in B-leukemia cells.
Molecular Cancer Therapeutics 12/2010; 9(12):3302-14. · 5.23 Impact Factor
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Spencer B Gibson
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ABSTRACT: Reactive oxygen species (ROS) have been implicated in many biological functions and diseases. Often their role is counterintuitive, where ROS can either promote cell survival or cell death depending on the cellular context. Similarly, autophagy is involved in many biological functions and diseases where it can either promote cell survival or cell death. There is now a growing consensus that ROS controls autophagy in multiple contexts and cell types. Furthermore, alterations in ROS and autophagy regulation contribute to cancer initiation and progression. However, how ROS and autophagy contribute to cancer and how to target either for cancer treatment is controversial. Blocking ROS generation could prevent cancer initiation, whereas blockage of autophagy seems to be required for initiation of cancer. In cancer progression, high levels of ROS correspond with increased metabolism and under metabolic stress autophagy is required to maintain cellular integrity. In cancer treatment, therapeutic drugs that increase ROS and autophagy have been implicated in their mechanism for cell death, such as 2-methoxyestrodial (2-ME) and arsenic trioxide (As(2)O(3)), whereas other therapeutic drugs that induce ROS and autophagy seem to have a protective effect. This has led to different approaches to treat cancer patients where autophagy is either activated or inhibited. Both views of ROS and autophagy are valid and reflect the balance within a cell to either survive or die. Understanding this balancing act within a cell is essential to determine whether to block or activate ROS-controlled autophagy for cancer therapy.
Autophagy 10/2010; 6(7):835-7. · 7.45 Impact Factor
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ABSTRACT: Autophagy is a regulated degradation pathway functioning in both cell survival and cell death. Its role in cancer is controversial because autophagy can be either protective or destructive to tumor cells, depending on individual genetic signatures and treatment conditions. Hypoxia is common in solid tumors, correlating with chemoresistance and poor prognosis. We have detected autophagic cell death in hypoxic cancer cells occurring independently of apoptosis through a mechanism involving the hypoxia-inducible protein, Bcl-2/E1B-nineteen kilodalton interacting protein (BNIP3). Loss of BNIP3 was protective against hypoxia-induced autophagy and cell death. Unexpectedly, BNIP3 ablation also caused differential cell cycle progression in vitro and increased cellularity in vivo. Collectively, these results support the emerging theory that autophagy could be effectively targeted as an alternative cell death pathway in hypoxic and/or apoptosis-resistant tumors. Furthermore, our data suggest that BNIP3 may be a potential target molecule in this pathway.
Annals of the New York Academy of Sciences 10/2010; 1210:8-16. · 3.15 Impact Factor
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ABSTRACT: Autophagy is an intracellular lysosomal degradation process, which in the case of macroautophagy, is characterized by the formation of double-membraned autophagosomes. Enhanced under stress conditions, autophagy can function to promote cell survival or cell death depending on the type of cellular stress. Interest in autophagy has increased substantially in the past several years as new research implicates this "self-eating" pathway in cell growth, development, and many human diseases. Various methods have been developed for detecting autophagy; however, the implementation of these methods and the interpretation of the results often vary between studies, and a more standardized approach is required. In this review, we summarize the current methods available for detecting autophagy and for determining its contribution to cell death. Furthermore, we discuss the critical points for the successful application of these methods based on experiences from our laboratory and from other research groups.
Canadian Journal of Physiology and Pharmacology 03/2010; 88(3):285-95. · 1.95 Impact Factor
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ABSTRACT: Lysophosphatidic acid (LPA) protects chronic lymphocytic leukemia (CLL) cells from apoptosis. Vascular endothelial growth factor (VEGF) also protects CLL cells against apoptosis. The mechanism for LPA protection against apoptosis in CLL cells is unknown. Herein, we show CLL cells express LPA receptors LPA(1-5) but in normal B cells, LPA(1) was rarely expressed and LPA(3,) LPA(4,) and LPA(6) were undetectable whereas the other LPA receptors were expressed. LPA plasma levels are similar in patients with CLL compared to healthy controls. In contrast, plasma levels of VEGF are elevated in patients with CLL compared to healthy controls and LPA treatment induced VEGF secretion in CLL cells. CLL cells also express VEGF receptors and LPA protection against Flu induced apoptosis is blocked by inhibition of VEGF receptor activation. These results indicate that LPA protects CLL cells from apoptosis through higher expression of LPA receptors and autocrine production of VEGF.
Leukemia & lymphoma 10/2009; 50(12):2038-48. · 2.40 Impact Factor
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British Journal of Haematology 09/2009; 147(3):396-8. · 4.94 Impact Factor
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ABSTRACT: Tandem pleckstrin homology domain proteins (TAPPs) are recruited to the plasma membrane via binding to phosphoinositides produced by phosphoinositide 3-kinases (PI3Ks). Whereas PI3Ks are critical for B-cell activation, the functions of TAPP proteins in B cells are unknown. We have identified 40 potential interaction partners of TAPP2 in B cells, including proteins involved in cytoskeletal rearrangement, signal transduction and endocytic trafficking. The association of TAPP2 with the cytoskeletal proteins utrophin and syntrophin was confirmed by Western blotting. We found that TAPP2, syntrophin, and utrophin are coexpressed in normal human B cells and B-chronic lymphocytic leukemia (B-CLL) cells. TAPP2 and syntrophin expression in B-CLL was variable from patient to patient, with significantly higher expression in the more aggressive disease subset identified by zeta-chain-associated protein kinase of 70 kDa (ZAP70) expression and unmutated immunoglobulin heavy chain (IgH) genes. We examined whether TAPP can regulate cell adhesion, a known function of utrophin/syntrophin in other cell types. Expression of membrane-targeted TAPP2 enhanced B-cell adhesion to fibronectin and laminin, whereas PH domain-mutant TAPP2 inhibited adhesion. siRNA knockdown of TAPP2 or utrophin, or treatment with PI3K inhibitors, significantly inhibited adhesion. These findings identify TAPP2 as a novel link between PI3K signaling and the cytoskeleton with potential relevance for leukemia progression.
Blood 09/2009; 114(21):4703-12. · 9.90 Impact Factor
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ABSTRACT: The Bcl-2 19 kDa interacting protein (BNIP3) is a pro-cell-death BH3-only member of the Bcl-2 family. We previously found that BNIP3 is localized to the nucleus in the majority of glioblastoma multiforme (GBM) tumors and fails to induce cell death. Herein, we have discovered that nuclear BNIP3 binds to the promoter of the apoptosis-inducing factor (AIF) gene and represses its expression. BNIP3 associates with PTB-associating splicing factor (PSF) and HDAC1 (histone deacetylase 1) contributing to transcriptional repression of the AIF gene. This BNIP3-mediated reduction in AIF expression leads to decreased temozolomide-induced apoptosis in glioma cells. Furthermore, nuclear BNIP3 expression in GBMs correlates with decreased AIF expression. Together, we have discovered a novel transcriptional repression function for BNIP3 causing reduced AIF expression and increased resistance to apoptosis. Thus, nuclear BNIP3 may confer a survival advantage to glioma cells and explain, in part, why BNIP3 is expressed at high levels in solid tumors, especially GBM.
Journal of Neuroscience 05/2009; 29(13):4189-99. · 7.11 Impact Factor
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ABSTRACT: The dual c-abl/Src kinase inhibitor, dasatinib, utilized to treat chronic myeloid leukaemia (CML) when used at clinically attainable sublethal concentrations, synergistically sensitized primary chronic lymphocytic leukaemia (CLL) lymphocytes to chlorambucil and fludarabine. In contrast, dasatinib alone demonstrated toxicity to CLL lymphocytes at concentrations that are generally not clinically attainable. Dasatinib resistance and poorer dasatinib-mediated sensitization to chlorambucil and fludarabine was associated with higher expression of c-abl protein levels. In contrast, chlorambucil and fludarabine resistance correlated with basal p53 protein levels. Moreover, Western blot analysis after in vitro treatment of primary CLL lymphocytes with dasatinib, chlorambucil and/or fludarabine, showed that dasatinib: (i) inhibited c-abl function (e.g. downregulation of c-abl protein levels and decreased the phosphorylation of a c-abl downstream target, Dok2), (ii) decreased chlorambucil/fludarabine induced accumulation of p53 protein levels, (iii) altered the response to chlorambucil/fludarabine induced DNA-damage as evidenced by an increase in chlorambucil/fludarabine-induced H2AX phosphorylation, and (iv) accentuated the c-abl downregulation induced by chlorambucil/fludarabine. Our results suggest that dasatinib in combination with chlorambucil or fludarabine may improve the therapy of CLL.
British Journal of Haematology 01/2009; 143(5):698-706. · 4.94 Impact Factor
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ABSTRACT: Reactive oxygen species (ROS) have been identified as signaling molecules in various pathways regulating both cell survival and cell death. Autophagy, a self-digestion process that degrades intracellular structures in response to stress, such as nutrient starvation, is also involved in both cell survival and cell death. Alterations in both ROS and autophagy regulation contribute to cancer initiation and progression, and both are targets for developing therapies to induce cell death selectively in cancer cells. Many stimuli that induce ROS generation also induce autophagy, including nutrient starvation, mitochondrial toxins, hypoxia, and oxidative stress. Some of these stimuli are under clinical investigation as cancer treatments, such as 2-methoxyestrodial and arsenic trioxide. Recently, it was demonstrated that ROS can induce autophagy through several distinct mechanisms involving Atg4, catalase, and the mitochondrial electron transport chain (mETC). This leads to both cell-survival and cell-death responses and could be selective toward cancer cells. In this review, we give an overview of the roles ROS and autophagy play in cell survival and cell death, and their importance to cancer. Furthermore, we describe how autophagy is mediated by ROS and the implications of this regulation to cancer treatments.
Antioxidants & Redox Signaling 11/2008; 11(4):777-90. · 8.20 Impact Factor
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ABSTRACT: NAD(P)H:quinoneoxidoreductase 1 (NQO1) inhibits some cancers and increases p53 and apoptosis in cells. Due to an inactivating polymorphism, 10% of humans have no NQO1 activity. A case:control study suggested that chronic lymphocytic leukemia (CLL) patients may have an increased incidence of the NQO1 null genotype compared with controls. NQO1 genotype did not correlate with various CLL prognostic factors, but we observed a trend toward lower drug response in patients with the NQO1 null genotype. Inhibiting NQO1 activity decreased p53 levels and drug induced apoptosis in CLL cells. These results raise the possibility that the NQO1 polymorphism may be a risk factor for CLL and a predictor of response to chemotherapy.
Leukemia Research 09/2008; 33(1):74-81. · 2.92 Impact Factor
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ABSTRACT: Histone deacetylases (HDACs) catalyze the removal of acetyl groups from histones and contribute to transcriptional repression. In addition, the HDAC inhibitors induce apoptosis in cancer cells through alterations in histone acetylation and activation of the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) apoptotic pathway. Lysophosphatidic acid (LPA) is a growth factor that promotes survival of cancer cells through activation of G protein-coupled receptors. Here we show that HDAC inhibitors can induce apoptosis through activation of the TRAIL apoptotic pathway, and LPA prevented HDAC inhibitor-induced apoptosis and increased TRAIL receptor DR4 (death receptor 4) protein expression. This was associated with increased HDAC1 recruitment to the DR4 promoter following LPA treatment and a reduction in HDAC inhibitor-induced histone acetylation in the DR4 promoter. In addition, LPA induces HDAC enzyme activity in a dose- and time-dependent manner, and this is associated with HDAC1 activation and increased binding of HDAC1 to HDAC2. Reducing the expression of HDAC1 significantly lowered LPA-induced HDAC activity and increased histone acetylation. LPA induction of HDAC activity was blocked by the LPA receptor antagonist, Ki16425, or by inhibiting receptor activation with pertussis toxin. Reducing the expression of the LPA receptor LPA(1) also blocked LPA-induced HDAC activation. In addition, LPA reduced histone acetyltransferase enzymatic activity. Finally, LPA attenuated the ability of the HDAC inhibitor to reduce HDAC activity. Thus, LPA enhances survival of cancer cells by increasing HDAC activity and reducing histone acetylation.
Journal of Biological Chemistry 07/2008; 283(24):16818-29. · 4.77 Impact Factor
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ABSTRACT: Vascular endothelial growth factor (VEGF) is a survival and angiogenesis factor that is a target for therapy in a variety of cancers. In many hematological malignancies, VEGF production is increased leading to cell survival responses. Herein, we demonstrate that lysophosphatidic acid (LPA) induces mRNA expression of VEGF in the multiple myeloma cell line, U266, the Burkitt's lymphoma cell line, BJAB, and the chronic lymphocytic leukemia (CLL)-like cell line, I-83. This increase in mRNA levels of VEGF corresponded with increased luciferase activity of the VEGF promoter in BJAB and I-83 cells and increased protein levels in I-83 cells. Secretion of VEGF was also increased in these cells following LPA treatment. LPA treatment also caused the activation of both VEGFR1 and VEGFR2. The increase in VEGF expression by LPA is mediated by the activation of c-Jun N-terminal Kinase (JNK) and transcription factor NFkappaB since blocking JNK or NFkappaB activation inhibited LPA induced VEGF expression. Furthermore, we have demonstrated that LPA protects cells from apoptosis and blocking activation of both VEGFR1 and VEGFR2 using a VEGF receptor kinase inhibitor prevented LPA survival responses. Knocking down expression of VEGFR1 and inhibiting activation of NFkappaB and JNK also blocked LPA induced protection against apoptosis. Taken together, this indicates that LPA contributes to VEGF production in B cell malignancies leading to cell survival.
Cellular Signalling 07/2008; 20(6):1198-208. · 4.06 Impact Factor
