Proliferation Of Hodgkin Lymphoma Cells Research Articles

High Expression of FGF2/SDC1 Signaling Contributes to Poor Clinical Outcome in Hodgkin Lymphoma Patients and SDC1 siRNA Suppresses Proliferation, Migration, and Invasion in Hodgkin Lymphoma Cells

Our recent studies have shown that Fibroblast Growth Factor-2 (FGF2) and Syndecan-1 (SDC1) expressions are upregulated, by 45- and 24-fold respectively, in CD30+ cells from tissues of relapsed versus relapsed-free Hodgkin lymphoma (HL) patients (p<0.005; Fig. 1A). Normal control and relapsed-free groups showed similar FGF2 and SDC1 expression profiles in tissue samples. The circulating HL cells in peripheral blood lymphocytes (PBL) have demonstrated that FGF2 and SDC1 mRNA were upregulated, by nearly 7000- and 9000-fold respectively, in relapsed versus relapsed-free HL patient samples (p<0.0001; Fig. 1B). Preliminary Kaplan-Meier analyses indicate that the HL patient group (n=4) with elevated co-expression of FGF2 and SDC1 had shortened survival by six-fold when compared to the majority of patients (n=11) with significantly lowered expression of FGF2/SDC1 (p < 0.05). This suggests that high levels of FGF2/SDC1 contribute to poor prognosis in HL patients. Western blot analyses showed that shed SDC1 regulate the HL cell division rate by a negative feedback mechanism. However, secretion of FGF2 and an abundance of four FGF receptor (FGFR) proteins were not associated with the cell division rate.Among eleven HL cell lines examined, HD-MyZ represented the best model for relapsed/refractory HL due to its highest cell proliferation rate (35 hour doubling time) and overexpression of FGF2 and SDC1, by 60- and 150-fold respectively, when compared with the B lymphoblast cell line, C1RB7. Immunofluorescence and co-immunoprecipitation studies in normal fibroblasts and HL cells demonstrate that FGF2 and SDC1 are co-localized on the cell membrane and bind together as ligand and receptor. Additionally, FGF2 binds to FGFRs in HL cells with lower affinity than in normal fibroblasts with predominant binding to FGFR1, suggesting that FGF2 signaling largely involves both SDC1 and FGFR1. Fluorescence microscopy studies showed that FGFR1 is constitutively phosphorylated in HL cells, and it stimulates cell proliferation by an autocrine pathway. Treatment with exogenous FGF2 in the absence of serum resulted in phosphorylation of both AKT and ERK, thereby activating FGF2-mediated cell proliferation. FGF2 induction also upregulated mRNA expressions of SDC1 (12-fold), proliferation marker- CCND1 (242-fold) and metastatic marker- MMP9 (67-fold) compared to unstimulated cells.To examine the molecular mechanism, whereby SDC1 is involved in the proliferation and metastasis of HL cells, SDC1 knockdown (KD) was achieved near 100% with 1nM siRNA after 48 hours of transfection. The SDC1 KD condition did not affect cell morphology but significantly attenuated cell proliferation by two-fold (p < 0.05; Fig. 2A) by downregulating BCL2 (87%), CCND1 (74%), and CDK4 (64%), indicating that SDC1 has pivotal functional roles in proliferation of HL cells. Transwell chamber assays showed that SDC1 KD reduced migration of HL cells by two-fold (p < 0.05; Fig. 2B) and invasion by three-fold (p < 0.005; Fig. 2C) by downregulating metastatic markers MMP9, ADAM9, ITGB3, and VEGF . The flow cytometry data demonstrated that 50% of HL cells expressed residual SDC1 on the cell membrane after SDC1 KD for 48 hours, which potentially maintains the normal HL cells morphology and cellular functions. However, four days of prolonged SDC1 KD caused an induction of apoptosis by upregulating CASP3, and continuous KD for 17 days caused near eradication of surviving HL cells.In conclusion, our results provide evidence that increased FGF2/SDC1 expression and signaling may drive and maintain the hyperproliferation of HL cells, thereby contributing to disease progression and poor prognosis in HL patients. Therefore, a novel method to inhibit altered FGF2/SDC1 signaling offers an excellent potential for treating high-risk and refractory HL patient populations. [Display omitted] DisclosuresGoy:Genentech: Research Funding; Acerta: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pharmacyclics / J&J: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Seattle Genetics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Feldman:Janssen: Speakers Bureau; Celgene: Speakers Bureau; AbbVie: Speakers Bureau; Bristol-Myers Squibb: Consultancy; Kite Pharma: Speakers Bureau; Seattle Genetics: Honoraria, Research Funding, Speakers Bureau; Pharmacyclics: Speakers Bureau. Leslie:KITE pharma: Speakers Bureau; celgene: Speakers Bureau; seattle genetics: Speakers Bureau. Pecora:COTA: Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties; Caladrius Biosciences: Equity Ownership, Membership on an entity's Board of Directors or advisory committees.

Long non-coding RNA NEAT1 regulates Hodgkin's lymphoma cell proliferation and invasion via miR-448 mediated regulation of DCLK1.

To explore whether long non-coding RNA nuclear enriched abundant transcript 1 (lncRNA NEAT1) could regulate Hodgkin's lymphoma (HL) cell proliferation and invasion through miR-448, which could target doublecortin like kinase 1 (DCLK1) and mediate DCLK1 expression. Expressions of NEAT1, miR-448 and DCLK1 were evaluated by qRT-PCR or Western blot assay. Cell Counting Kit-8 (CCK-8) and transwell assay were utilized to detect cell proliferation and invasion capability in L428 cells respectively. The target relationship between NEAT1, miR-448 and DCLK1 was confirmed by Luciferase reporter assay. QRT-PCR results showed that NEAT1 expressed higher in HL tissues and cell lines than that in controls. In vitro experiments, NEAT1 downregulation could decrease cell proliferation and invasion capability in L428 cells. NEAT1 directly interacted with miR-448 and negatively regulated it. Moreover, DCLK1 was confirmed as a target of miR-448. DCLK1 expression was increased in L428 cells and positively regulated by NEAT1. NEAT1 overexpression upregulated the protein level of DCLK1 in L428 cells according to Western blot analysis. Additionally, DCLK1 overexpression could reverse the suppression on cell proliferation and invasion capability induced by NEAT1 knockdown or miR-448 overexpression. NEAT1 might be contributed to HL progression by promoting cell proliferation and invasion capability via miR-448 mediated DCLK1 expression.

Targeting Cancer Cells and Tumor Microenvironment in Preclinical and Clinical Models of Hodgkin Lymphoma Using the Dual PI3Kδ/γ Inhibitor RP6530.

Tumor-associated macrophages (TAMs) and the hyperactivation of the PI3K/AKT pathway are involved in the pathogenesis of Hodgkin lymphoma and affect disease outcome. Because the δ and γ isoforms of PI3K are overexpressed in Hodgkin/Reed-Sternberg (HRS) cells and the tumor microenvironment (TME), we propose that the PI3Kδ/γ inhibitor RP6530 might affect both HRS cells and TME, ultimately leading to an enhanced antitumor response. Hodgkin lymphoma cell lines (L-540, KM-H2, and L-428) and primary human macrophages were used to investigate the activity of RP6530 in vitro and in vivo in Hodgkin lymphoma cell line xenografts. In vitro, RP6530 besides killing and inhibiting the proliferation of Hodgkin lymphoma cells, downregulated lactic acid metabolism, switching the activation of macrophages from an immunosuppressive M2-like phenotype to a more inflammatory M1-like state. By RNA sequencing, we define tumor glycolysis as a specific PI3Kδ/γ-dependent pathway implicated in the metabolic reprogramming of cancer cells. We identify the metabolic regulator pyruvate kinase M2 as the main mediator of tumor-induced immunosuppressive phenotype of macrophages. Furthermore, we show in human tumor xenografts that RP6530 repolarizes TAMs into proinflammatory macrophages and inhibits tumor vasculature, leading to tumor regression. Interestingly, patients with Hodgkin lymphoma experiencing objective responses (complete response and partial response) in a phase I trial using RP6530 showed a significant inhibition of circulating myeloid-derived suppressor cells and an average mean reduction in serum thymus and activation-regulated chemokine levels of 40% (range, 4%-76%). Our results support PI3Kδ/γ inhibition as a novel therapeutic strategy that targets both malignant cells and the TME to treat patients with Hodgkin lymphoma.

MicroRNA-24-3p regulates Hodgkin's lymphoma cell proliferation, migration and invasion by targeting DEDD.

Hodgkin's lymphoma (HL) is a common hematologic tumor, and the incidence is increasing. At present, it is considered that miRNAs are closely related to HL. Substantial attention has been paid to the effects of miRNA on the pathophysiological process of HL. This study was focused on the potential role of miR-24-3p in HL by targeting DEDD. The reverse transcription-quantitative PCR (RT-qPCR) results demonstrated that miR-24-3p expression was highly elevated and DEDD expression reduced inversely in HL tissues compared to adjacent tissues. According to the results of CKK-8 assays, miR-24-3p was able to accelerate HL cell proliferation. In addition, the results of the Transwell assays also indicated that miR-24-3p promoted the invasion and migration abilities of HL cells. Moreover, the results demonstrated that miR-24-3p inhibited DEDD expression. Hence, the present study revealed that miR-24-3p could accelerate HL development through inhibiting DEDD.

Targeted Repression of Overexpressed CD30 Downregulates NF-κB and ERK1/2 Pathway in Hodgkin Lymphoma Cell Lines

We previously reported that CD30 is induced during lymphocyte transformation and that overexpressed CD30 can transduce ligand-independent signals in Hodgkin lymphoma (HL) cells. However, its biological consequence is not fully addressed. In this study, we examined the effects of targeted repression of overexpressed CD30 on cell signaling and proliferation using small-interfering RNAs (siRNAs) in HL cell lines. Repression of CD30 inhibited cellular proliferation through reduced activation of IkappaB kinase (IKK) and extracellular signal-regulated kinase (ERK) 1/2 in both B- and T-HL cell lines. These HL cell lines bear one or more defects in negative regulators of nuclear factor (NF)-kappaB signaling, including A20, cylindromatosis tumor suppressor protein (CYLD), and IkappaBalpha, and when CD30 is repressed, they show reduced activation of the canonical NF-kappaB pathway. This suggests that CD30 governs NF-kappaB and ERK1/2 signaling pathways, and is involved in the proliferation of HL cells. Defective mutations in negative regulators of NF-kappaB signaling appear to promote CD30-initiated basal NF-kappaB activation. These results indicate that CD30 is involved in the tumorigenic process of HL, and that it may be useful as a therapeutic target for the treatment of HL.

STAT3 is essential for Hodgkin lymphoma cell proliferation and is a target of tyrphostin AG17 which confers sensitization for apoptosis

STAT3 is essential for Hodgkin lymphoma cell proliferation and is a target of tyrphostin AG17 which confers sensitization for apoptosis