Purpose
Patients with advanced stages of MCL have a poor prognosis after standard therapies. MCL cells in those patients often spread into tissues other than lymph nodes, such as the bone marrow. Apart from directed migration and homing, there is little understanding of the function of the CXCR4/SDF1 signaling axis in MCL. In this report, we aim to understand mechanisms of MCL cell survival in the bone marrow.
Experimental Design
For comprehensive analyses of MCL interactions with bone marrow stromal cells we have generated gene knock out cells using CRISPR-CAS9 system and gene knock down cells to reveal novel roles of the CXCR4/SDF-1 signaling.
Results
CXCR4 silencing in MCL cells led to a significant reduction in proliferation, cell adhesion to bone marrow stromal cells and colony formation in PHA-LCM methylcellulose medium, which were reversed upon the addition of SDF-1 neutralizing antibodies. In addition, tracking MCL cell engraftment in vivo revealed that quiescent MCL cells are significantly reduced in the bone marrow upon CXCR4 silencing, indicating that CXCR4/SDF-1 signaling is required for the survival and maintenance of the quiescent MCL cells. Further analysis revealed novel mechanisms of ROS induced CXCR4/SDF-1 signaling that stimulate autophagy formation in MCL cells for their survival.
Conclusions
Our data, for the first time, revealed new roles of the CXCR/SDF-1 signaling axis on autophagy formation in MCL, which further promoted their survival within the bone marrow microenvironment. Targeting the CXCR4/SDF-1/autophagy signaling axis may contribute to an enhanced efficacy of current therapies.
Here, we have prospectively isolated and characterized, for the first time, clonogenic cells with self-renewal capacities from mantle cell lymphoma (MCL), a particularly deadly form of Non-Hodgkin’s Lymphoma (NHL). Self-renewal and tumorigenic activities were enriched in MCL cell fractions that lacked expression of the prototypic B cell surface marker, CD19. CD45+CD19− cells represented a relatively small fraction of the total MCL tumor cells, however, they recapitulated the heterogeneity of original patient tumors upon transplantation into immunodeficient mice. As few as 100 of these cells displayed self-renewal capacities in secondary and tertiary recipient mice by in vivo limiting dilution assays. Similar to leukemic stem cells, CD45+CD19− MCL cells also displayed a quiescent status as determined by dye efflux assays. In summary, this study is the first to isolate subpopulations of MCL cells that have self-renewal and tumorigenic capacities. Identification and characterization of MCL-ICs is an important first step toward understanding how self-renewal and tumorigenicity are regulated in MCL and designing targeted therapies against MCL-ICs will ultimately lead to improved outcomes for MCL patients.
Expression of the transglutaminase TG2 has been linked to constitutive activation of NF-kB and chemotherapy resistance in mantle cell lymphoma (MCL) cells. TG2 forms complexes with NF-kB components, but mechanistic insights that could be used to leverage therapeutic responses has been lacking. In the present study, we address this issue with the discovery of an unexpected role for TG2 in triggering autophagy in drug-resistant MCL cells through induction of IL-6. CRISPR-mediated silencing of TG2 delayed apoptosis while overexpressing TG2 enhanced tumor progression. Under stress, TG2 and IL-6 mediate enhanced autophagy formation to promote MCL cell survival. Interestingly, the autophagy product ATG5 involved in autophagosome elongation positively regulated TG2/NF-kB/IL-6 signaling, suggesting a positive feedback loop. Our results uncover an interconnected network of TG2/NF-kB and IL-6/STAT3 signaling with autophagy regulation in MCL cells, the disruption of which may offer a promising therapeutic strategy.
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