MicroRNAs 15a and 16 regulate tumor proliferation in multiple myeloma.

Detailed genomic studies have shown that cytogenetic abnormalities contribute to multiple myeloma (MM) pathogenesis and disease progression. Nevertheless, little is known about the characteristics of MM at the epigenetic level and specifically how microRNAs regulate MM progression in the context of the bone marrow milieu. Therefore, we performed microRNA expression profiling of bone marrow derived CD138(+) MM cells versus their normal cellular counterparts and validated data by qRT-PCR. We identified a MM-specific microRNA signature characterized by down-expression of microRNA-15a/-16 and overexpression of microRNA-222/-221/-382/-181a/-181b (P < .01). We investigated the functional role of microRNA-15a and -16 and showed that they regulate proliferation and growth of MM cells in vitro and in vivo by inhibiting AKT serine/threonine-protein-kinase (AKT3), ribosomal-protein-S6, MAP-kinases, and NF-kappaB-activator MAP3KIP3. Moreover, miRNA-15a and -16 exerted their anti-MM activity even in the context of the bone marrow milieu in vitro and in vivo. These data indicate that microRNAs play a pivotal role in the biology of MM and represent important targets for novel therapies in MM.

microRNA expression in the biology, prognosis, and therapy of Waldenström macroglobulinemia.

Multilevel genetic characterization of Waldenström macroglobulinemia (WM) is required to improve our understanding of the underlying molecular changes that lead to the initiation and progression of this disease. We performed microRNA-expression profiling of bone marrow-derived CD19(+) WM cells, compared with their normal cellular counterparts and validated data by quantitative reverse-transcription-polymerase chain reaction (qRT-PCR). We identified a WM-specific microRNA signature characterized by increased expression of microRNA-363*/-206/-494/-155/-184/-542-3p, and decreased expression of microRNA-9* (ANOVA; P < .01). We found that microRNA-155 regulates proliferation and growth of WM cells in vitro and in vivo, by inhibiting MAPK/ERK, PI3/AKT, and NF-kappaB pathways. Potential microRNA-155 target genes were identified using gene-expression profiling and included genes involved in cell-cycle progression, adhesion, and migration. Importantly, increased expression of the 6 miRNAs significantly correlated with a poorer outcome predicted by the International Prognostic Staging System for WM. We further demonstrated that therapeutic agents commonly used in WM alter the levels of the major miRNAs identified, by inducing downmodulation of 5 increased miRNAs and up-modulation of patient-downexpressed miRNA-9*. These data indicate that microRNAs play a pivotal role in the biology of WM; represent important prognostic marker; and provide the basis for the development of new microRNA-based targeted therapies in WM.

RhoA and Rac1 GTPases play major and differential roles in stromal cell-derived factor-1-induced cell adhesion and chemotaxis in multiple myeloma.

The interaction of multiple myeloma (MM) cells with the bone marrow (BM) milieu plays a crucial role in MM pathogenesis. Stromal cell-derived factor-1 (SDF1) regulates homing of MM cells to the BM. In this study, we examined the role of RhoA and Rac1 GTPases in SDF1-induced adhesion and chemotaxis of MM. We found that both RhoA and Rac1 play key roles in SDF1-induced adhesion of MM cells to BM stromal cells, whereas RhoA was involved in chemotaxis and motility. Furthermore, both ROCK and Rac1 inhibitors reduced SDF1-induced polymerization of actin and activation of LIMK, SRC, FAK, and cofilin. Moreover, RhoA and Rac1 reduced homing of MM cells to BM niches. In conclusion, we characterized the role of RhoA and Rac1 GTPases in SDF1-induced adhesion, chemotaxis, and homing of MM cells to the BM, providing the framework for targeting RhoA and Rac1 GTPases as novel MM therapy.

CXCR4 inhibitor AMD3100 disrupts the interaction of multiple myeloma cells with the bone marrow microenvironment and enhances their sensitivity to therapy.

The interaction of multiple myeloma (MM) cells with their microenvironment in the bone marrow (BM) provides a protective environment and resistance to therapeutic agents. We hypothesized that disruption of the interaction of MM cells with their BM milieu would lead to their sensitization to therapeutic agents such as bortezomib, melphalan, doxorubicin, and dexamethasone. We report that the CXCR4 inhibitor AMD3100 induces disruption of the interaction of MM cells with the BM reflected by mobilization of MM cells into the circulation in vivo, with kinetics that differed from that of hematopoietic stem cells. AMD3100 enhanced sensitivity of MM cell to multiple therapeutic agents in vitro by disrupting adhesion of MM cells to bone marrow stromal cells (BMSCs). Moreover, AMD3100 increased mobilization of MM cells to the circulation in vivo, increased the ratio of apoptotic circulating MM cells, and enhanced the tumor reduction induced by bortezomib. Mechanistically, AMD3100 significantly inhibited Akt phosphorylation and enhanced poly(ADP-ribose) polymerase (PARP) cleavage as a result of bortezomib, in the presence of BMSCs in coculture. These experiments provide a proof of concept for the use of agents that disrupt interaction with the microenvironment for enhancement of efficacy of cytotoxic agents in cancer therapy.

Targeting NF-kappaB in Waldenstrom macroglobulinemia.

The nuclear factor-kappaB (NF-kappaB) path-way has been implicated in tumor B-cell survival, growth, and resistance to therapy. Because tumor cells overcome single-agent antitumor activity, we hypothesized that combination of agents that target differentially NF-kappaB pathway will induce significant cytotoxicity. Therapeutic agents that target proteasome and Akt pathways should induce significant activity in B-cell malignancies as both pathways impact NF-kappaB activity. We demonstrated that perifosine and bortezomib both targeted NF-kappaB through its recruitment to the promoter of its target gene IkappaB using chromatin immunoprecipitation assay. This combination led to synergistic cytotoxicity in Waldenstrom macroglobulinemia (WM) cells that was mediated through a combined reduction of the PI3K/Akt and ERK signaling pathways, found to be critical for survival of WM cells. Moreover, a combination of these drugs with the CD20 monoclonal antibody rituximab further increased their cytotoxic activity. Thus, effective WM therapy may require combination regimens targeting the NF-kappaB pathway.

3D Printed Stem-Cell-Laden, Microchanneled Hydrogel Patch for the Enhanced Release of Cell-Secreting Factors and Treatment of Myocardial Infarctions.

Over the past several years, biomaterials loaded with mesenchymal stem cells (MSCs) have increasingly been used to reduce the myocardial fate of postinfarction collagen deposition and scar tissue formation. Despite successful gains, therapeutic efficacy has remained limited because of restricted transport of cell-secreting factors at the site of implantation. We hypothesized that an MSC-laden hydrogel patch with multiple microchannels would retain transplanted cells on target tissue and support transport of cell-secreting factors into tissue. By doing so, the gel patch will improve the therapeutic potential of the cells and minimize the degradation of myocardial tissue postinfarction. To examine this hypothesis, a stereolithographic apparatus (SLA) was used to introduce microchannels of controlled diameters (e.g., 500 and 1000 µm) during in situ cross-linking reaction of poly(ethylene glycol)dimethacrylate solution suspended with cells. Placement of the MSC-laden, microchanneled gel patch on the occluded left coronary artery in a murine model showed significant improvement in the ejection fraction, fractional shortening, and stroke volume, compared with gel patches without MSCs and MSC-laden gel patches without microchannels. In particular, the microchannels significantly reduced the number of cells required to recover cardiac function, while minimizing cardiac remodeling. In sum, the microchanneled gel patch would provide a means to prevent abnormal fibrosis resulting from acute ischemic injury.