• Delineating the Role of NKT cell Activation in B cell Lymphoma

      Lee, Michael; Webb, Tonya J. (2020)
      Natural Killer T (NKT) cells play an important role in cancer surveillance and can reduce lymphoma burden in vivo; however, a hallmark of cancer is its ability to evade immune surveillance. Our goals were to elucidate novel mechanisms utilized by B cell lymphoma to evade NKT cell-mediated immune surveillance and determine the prognostic potential of assessing NKT cell function in lymphoma patients. We found that knockdown of sphingosine kinase 1 (SK1) in human lymphoma cells results in a significant increase in CD1d-mediated NKT cell activation. Lipidomic and co-culture studies identified cardiolipin as being upregulated in SK1 knockdown cells and implicated cardiolipin as an NKT cell-specific cancer neoantigen. We also sought to determine the efficacy of NKT cell-based therapy on survival and the induction of anti-tumor immune responses in a mouse model of B cell lymphoma. We found that activation of NKT cells via early administration of α-galactosylceramide (α-GalCer) only provided modest protection. Our data suggest that the lack of protection is due, at least in part, to the expansion of myeloid-derived suppressor cells in α-GalCer-treated tumor bearing mice. Lastly, we sought to identify novel immunological biomarkers in lymphoma patients. It was found that lymphoma patients have a reduction in NKT cell function compared to healthy donors. Furthermore, lymphoma patients have significantly higher levels of both pro- and anti-inflammatory cytokines in their sera compared to healthy donors. In addition, lymphoma patients who experience relapse have significantly reduced NKT cell function in the blood, compared to lymphoma patients who did not relapse. Collectively, our studies demonstrate the multifaceted role NKT cells play in immune responses to B cell lymphoma and will help inform the next generation of cancer immunotherapy.
    • Expression, Regulation, and Inhibition of Matriptase in Human B-cell Neoplasm

      Chou, Feng-Pai; Lin, Chen-Yong (2012)
      Matriptase, a type II transmembrane serine protease, possesses potent oncogenic activity when an imbalance to its cognate inhibitor, hepatocyte activator inhibitor-1 (HAI-1), is established in a transgenic mouse model. The oncogenic activity of matriptase is in part attributed to its role in activation of other oncogenic molecules, such as urokinase plasminogen activator (uPA) and hepatocyte growth factor (HGF). These matriptase substrates play important roles in extracellular matrix-degrading, cellular motility, cell growth, and angiogenesis. All of these biological events are important for the development and progression of human cancers. While most of matriptase studies have been focused on epithelial and carcinoma cells in which HAI-1 plays a pivotal role in regulation and inhibition of matriptase, growing evidence showed that matriptase may be also important in blood malignancies. In this dissertation, I examined matriptase and HAI-1 expression in human blood tumor specimens and a variety of hematological cell lines. Significant imbalanced expression of matriptase to HAI-1 was determined in aggressive type B-cell malignancies in vitro and in vivo. In contrast, the protease was not detected in several indolent B-cell lymphomas and hyperactive B-cells residing in lymph nodes. The absence of HAI-1 expression in aggressive B-cell cancers has impact to the current dogma of matriptase-HAI-1 pairing in epithelial cells. Additionally, I discovered hypoxia initiated matriptase zymogen activation in blood tumors, producing inhibitor free, enzymatic matriptase to shed into extracellular milieu. Increased reactive oxygen species (ROSs) may be attributed to hypoxia-induced matriptase activation while the zymogen activation can be reduced by pre-treating cells with ROSs scavengers. To further define the roles of matriptase in the development and progression of B-cell tumors, stable matriptase knockdown populations were created in lymphoma and myeloma lines, respectively. The matriptase knockdown cells showed to negatively affect cell growth in vitro and colony formation in methylcellulose-based culture. Besides, in mice xenograft model, matriptase-deficient cells grew in significantly slow rate and formed less tumor volume than the control group. In summary, deregulated matriptase play important roles to affect the tumor microenvironment which is favoring the growth and progression of B-cell lymphoma and multiple myeloma.
    • Regulation of translation initiation upon B-cell receptor activation

      Steinhardt, James John; Gartenhaus, Ronald B. (2014)
      Eukaryotic initiation factor 4A (eIF4A) promotes the translation initiation of genes harboring 5' untranslated regions (UTRs) with complex secondary structures. eIF4A activity is modulated by its inhibitor, programmed cell death 4 (PDCD4), which is regulated by the AKT pathway in B-cells. The release and degradation of PDCD4 frees eIF4A to bind to eIF4G, which together with the m7GTP cap-binding protein, eIF4E, forms the eIF4F cap-binding complex. Human diffuse large B-cell lymphomas (DLBCLs) are derived from antigen experienced B-cells and often display high expression of oncogenes that contain complex secondary structures in their 5' UTRs. We hypothesized that activated B-cells such as GC B-cells require enhanced eIF4A activity through B-cell receptor (BCR) stimulation. We found that eIF4A cap-binding activity is in fact greatly enhanced upon in vitro activation of human splenic B-cells after treatment with anti-BCR, while the pharmacological inhibitor of eIF4A, Silvestrol, potently inhibited BCR-induced protein translation. Upon antigen-induced activation, naïve, IgM+ B-cells differentiate within the germinal centers (GC) and undergo isotype switching to IgG. We found that IgG expressing human splenic B-cells displayed more robust eIF4A cap-binding activity and protein translation than IgM expressing B-cells after isotype specific BCR-stimulation. We also performed translational profiling to determine which mRNAs are preferentially loaded onto the polysome upon BCR activation and depleted upon pharmacological inhibition of eIF4A. Among the top differentially modulated genes was caspase recruitment domain family member 11 (CARD11). We found that CARD11, along with the B-cell CLL/lymphoma 10 (BCL10) and mucosa associated lymphoid tissue lymphoma translocation gene 1 (MALT1) harbor complex 5'UTRs unlike the majority of other components in the BCR signaling cascade. Importantly, these proto-oncogenes form the CARD11-BCL10-MALT1 (CBM) signalosome, which mediates BCR activation of nuclear factor of kappa light chain gene enhancer in B-cells (NF-κB) signaling. Both constitutive activation of NF-κB and the aberrant expression of CBM components are associated with B-cell lymphomas in humans. Collectively, these data indicate that eIF4A-mediated control of oncogene translation may be a critical component for B-cell lymphoma progression and suggest that pharmacological targeting of eIF4A may be an attractive therapeutic approach in the management of human B-cell lymphomas.