Bone Marrow Mesenchymal Stromal Cells in Multiple Myeloma: Their Role as Active Contributors to Myeloma Progression

Multiple myeloma (MM) is a hematological malignancy of plasma cells that proliferate and accumulate within the bone marrow (BM). Work from many groups has made evident that the complex microenvironment of the BM plays a crucial role in myeloma progression and response to therapeutic agents. Within the cellular components of the BM, we will specifically focus on mesenchymal stromal cells (MSCs), which are known to interact with myeloma cells and the other components of the BM through cell to cell, soluble factors and, as more recently evidenced, through extracellular vesicles. Multiple structural and functional abnormalities have been found when characterizing MSCs derived from myeloma patients (MM-MSCs) and comparing them to those from healthy donors (HD-MSCs). Other studies have identified differences in genomic, mRNA, microRNA, histone modification, and DNA methylation profiles. We discuss these distinctive features shaping MM-MSCs and propose a model for the transition from HD-MSCs to MM-MSCs as a consequence of the interaction with myeloma cells. Finally, we review the contribution of MM-MSCs to several aspects of myeloma pathology, specifically to myeloma growth and survival, drug resistance, dissemination and homing, myeloma bone disease, and the induction of a pro-inflammatory and immunosuppressive microenvironment.Funding: This study was funded by the Instituto de Salud Carlos III-FIS and co-financed by FEDER (PI19/01384 and PI18/01600); by the AECC (PROYE20047GUTI); the Network Center of Regenerative Medicine and Cellular Therapy of Castilla y León; and by the Gerencia Regional de Salud, Junta de Castilla y León (GRS 2066/A/19). P.M. (Patricia Maiso) was supported by a Miguel Servet fellowship from the ISCIII-FIS (CPII19/00028) co-financed by the European Social Fund; P.M. (Pedro Mogollón) is supported by a grant from IBSAL

Epigenetic modifications as key regulators of Waldenstrom's Macroglobulinemia biology

Waldenstrom's Macroglobulinemia is a low-grade B-cell lymphoma characterized by the presence of lymphoplasmacytic cells in the bone marrow and a monoclonal immunoglobulin M in the circulation. Recent evidences support the hypothesis that epigenetic modifications lead to Waldesntrom cell proliferation and therefore play a crucial role in the pathogenesis of this disease. Indeed, while cytogenetic and gene expression analysis have demonstrated minimal changes; microRNA aberrations and modification in the histone acetylation status of primary Waldenstrom Macroglobulinemia tumor cells have been described. These findings provide a better understanding of the underlying molecular changes that lead to the initiation and progression of this disease

The effect of the proteasome inhibitor bortezomib on acute myeloid leukemia cells and drug resistance associated with the CD34+ immature phenotype

[Background]: Proteasome inhibition represents a promising novel anticancer therapy, and bortezomib is a highly selective reversible inhibitor of the proteasome complex. Acute myeloid leukemia (AML) is an immnunophenotypically heterogeneous group of diseases, with CD34+ cases being associated with drug resistance and poor outcome. We investigated the effects of bortezomib on the growth and survival of AML cells. [Design and Methods]: We studied the in vitro activity and mechanism of action of bortezomib on both cell lines and fresh cells from 28 AML patients including CD34+ and CD34- cases. [Results]: Bortezomib showed potent anti-AML activity (IC50 < 50 nM), which was greater than that of conventional agents (doxorubicin, cytarabine and fludarabine). Moreover, synergistic effects were observed when bortezomib was adminstered in combination with doxorubicin and cytarabine. Mechanistically, bortezomib induced accumulation of cells in the G2/M phase, with up-regulation of p27, together with cell death through an increase in the mitochondrial outer membrane permeability involving caspase-dependent and -independent pathways. The apoptotic activity of bortezomib on fresh CD34 + blast cells from patients was similar to that observed on CD34 - blast cells. Importantly, bortezomib was significantly more active than doxorubicin in the immature CD34+ cells, while there were no differences in its action on CD34- cells. [Conclusions]: Bortezomib induces apoptosis in acute myeloid leukemia cells in vitro. Whether this drug might be useful in the treatment of patients with acute myeloid leukemia can be established only in ad hoc clinical trials. ©2008 Ferrata Storti Foundation.We thank Johnson and Johnson Pharmaceutical Research and Development (JJPRD).Peer Reviewe

Novel Tumor Suppressor Function of Glucocorticoid-Induced TNF Receptor GITR in Multiple Myeloma

Glucocorticoid-induced TNF receptor (GITR) plays a crucial role in modulating immune response and inflammation, however the role of GITR in human cancers is poorly understood. In this study, we demonstrated that GITR is inactivated during tumor progression in Multiple Myeloma (MM) through promoter CpG island methylation, mediating gene silencing in primary MM plasma cells and MM cell lines. Restoration of GITR expression in GITR deficient MM cells led to inhibition of MM proliferation in vitro and in vivo and induction of apoptosis. These findings were supported by the presence of induction of p21 and PUMA, two direct downstream targets of p53, together with modulation of NF-κB in GITR-overexpressing MM cells. Moreover, the unbalanced expression of GITR in clonal plasma cells correlated with MM disease progression, poor prognosis and survival. These findings provide novel insights into the pivotal role of GITR in MM pathogenesis and disease progression

Phenotypic and genomic analysis of multiple myeloma minimal residual disease tumor cells: a new model to understand chemoresistance

Persistence of chemoresistant minimal residual disease (MRD) plasma cells (PCs) is associated with inferior survival in multiple myeloma (MM). Thus, characterization of the minor MRD subclone may represent a unique model to understand chemoresistance, but to our knowledge, the phenotypic and genetic features of the MRD subclone have never been investigated. Here, we compared the antigenic profile of MRD vs diagnostic clonal PCs in 40 elderly MM patients enrolled in the GEM2010MAS65 study and showed that the MRD subclone is enriched in cells overexpressing integrins (CD11a/CD11c/CD29/CD49d/CD49e), chemokine receptors (CXCR4), and adhesion molecules (CD44/CD54). Genetic profiling of MRD vs diagnostic PCs was performed in 12 patients; 3 of them showed identical copy number alterations (CNAs), in another 3 cases, MRD clonal PCs displayed all genetic alterations detected at diagnosis plus additional CNAs that emerged at the MRD stage, whereas in the remaining 6 patients, there were CNAs present at diagnosis that were undetectable in MRD clonal PCs, but also a selected number of genetic alterations that became apparent only at the MRD stage. The MRD subclone showed significant downregulation of genes related to protein processing in endoplasmic reticulum, as well as novel deregulated genes such as ALCAM that is prognostically relevant in MM and may identify chemoresistant PCs in vitro. Altogether, our results suggest that therapy-induced clonal selection could be already present at the MRD stage, where chemoresistant PCs show a singular phenotypic signature that may result from the persistence of clones with different genetic and gene expression profiles. This trial was registered at www.clinicaltrials.gov as #NCT01237249

New drugs in multiple myeloma: mechanisms of action and phase I/II clinical findings

The outcome of multiple myeloma has substantially improved over the past decade, mainly due to recently approved drugs, such as thalidomide, lenalidomide, and bortezomib. Nevertheless, most patients still relapse and, therefore, drugs with new mechanisms of action are urgently needed to overcome this resistance. In this Review, we discuss some of the new targeted therapeutic strategies under assessment in preclinical and clinical studies in multiple myeloma. Unfortunately, the single-agent clinical activity of most of these new drugs has been limited; nevertheless, their effectiveness might be enhanced by their rational combination with each other or with conventional agents. © 2008 Elsevier Ltd. All rights reserved.This work was partially supported by the following grants: Scientifi c Foundation of Spanish Association against Cancer (AECC), Fondo de Investigación Sanitaria-FIS (RTICC-RD06/0020/0006 and RD06/0020/0041), and SaCyL (Ref 51-05). EMO is supported by the Plan Nacional de Investigación Científica, Desarrollo e Innovación Tecnológica (I+D+I) and by Instituto de Salud Carlos III-Fondo de Investigación Sanitaria with expedient number 4/00001. This work was also supported by a grant from the Ministry of Education and Science of Spain (BFU2006- 01813/BMC), and by an Institutional Cancer Center Network programme from the ISCIII. The Cancer Research Institute receives support from the European Community through the regional development funding program (FEDER). PM was supported by the FIS-FEDER through projects to JFSM, and a Spanish Myeloma Network Program (G03/136).Peer Reviewe

The synergy of panobinostat plus doxorubicin in acute myeloid leukemia suggests a role for HDAC inhibitors in the control of DNA repairSynergy of panobinostat and doxorubicin in AML

Acute myeloid leukemia (AML) is a clonal disorder characterized by the accumulation of myeloid blasts in the bone marrow. Here, we report the effects of the novel histone deacetylase inhibitor panobinostat (LBH589) in combination with doxorubicin on AML cells. Panobinostat exhibited potent anti-AML activity in all AML cell lines tested and in primary AML cells from patients (IC(50)<20 nM). In addition, panobinostat potentiated the action of several standard-of-care anti-AML compounds, particularly, doxorubicin. The molecular effects induced by panobinostat and doxorubicin treatment were investigated by analyzing gene expression, cell cycle, apoptosis and signaling pathways. Analyses of gene expression profiles identified 588 genes whose expression was exclusively affected by the combination of panobinostat and doxorubicin. The combination induced AML cell death by an increase in the mitochondrial outer membrane permeability and release of cytochrome c from the mitochondria, resulting in caspase-dependent apoptosis and accompanied by the upregulation of Bax, Bak and, particularly, Bad. The drug combination provoked a strong activation of a DNA damage response, indicating that this combination may trigger cell death by a mechanism that induced DNA double-strand breaks. These data indicate that the combination of panobinostat and doxorubicin may be an effective therapy for the treatment of AML.Peer Reviewe

Evaluación del 18F-FDG, 11C-Metionina y 11C- Colina para la detección de infiltración tumoral en Mieloma Múltiple: estudio traslacional en modelos preclínico y clínico en pacientes

El Mieloma Múltiple (MM) es la segunda neoplasia hematológica más frecuente y, a pesar de los avances terapéuticos, continúa siendo incurable. Se caracteriza por la infiltración tumoral ósea por células plasmáticas y, en algunos casos, lesiones extramedulares. Si bien el estándar para la evaluación de las lesiones óseas sigue siendo la radiología convencional, en los últimos años se han producido notables avances gracias a nuevas técnicas de imagen como la tomografía por emisión de positrones (PET)/Tomografía computarizada (TC) con glucosa marcada con flúor (18F-FDG) y se han explorado nuevos radiofármacos para mejorar el rendimiento diagnóstico de la FDG. Se comparó el rendimiento diagnóstico de tres radiofármacos (FDG, MET y COL) en la detección de infiltración tumoral de MM por PET/TC en un modelo murino mediante microPET e investigó la utilidad diagnóstica y el valor pronóstico de FDG y MET PET/TC en pacientes con mieloma. En el modelo preclínico, se comparó la captación de los trazadores FDG y MET explorando distintas líneas celulares in vitro, y también la sensibilidad y especificidad de FDG, MET y COL en un modelo murino de MM con tumor subcutáneo determinando la eficacia diagnóstica de FDG y MET en la valoración de respuesta al tratamiento con terapia convencional aislada (bortezomib), y en combinación (bortezomib, lenalidomida, dexametasona). En 52 pacientes, se realizó un estudio prospectivo utilizando FDG y MET comparando el rendimiento diagnóstico, analizando la correlación de los biomarcadores PET con parámetros analíticos de valor pronóstico conocido (β2 microglobulina, entre otros) así como el valor pronóstico de nuevos biomarcadores de imagen como el volumen metabólico tumoral (TMTV) y la carga tumoral metabólicamente activa con FDG (TLG) y MET (TLMU). La línea RPMI8226 demostró superior captación de MET que de FDG in vitro. En el modelo murino en los tumores RPMI8226 y MM1S, la COL mostró la menor sensibilidad. Tanto la FDG como la MET permitieron realizar un seguimiento adecuado de la progresión tumoral y reflejaron el efecto de los tratamientos empleados en mieloma, evidenciándose mayor control de la enfermedad con triple terapia que con bortezomib en monoterapia. La MET detectó infiltración tumoral en 6 pacientes (11%) con PET FDG negativo y detectó mayor número de lesiones que la FDG en 33/52 pacientes (63%). En el análisis univariante, los biomarcadores que se asociaron con la supervivencia libre de progresión (SLP) fueron: >3 lesiones focales (HR 2.65, IC 95% 0.84-8.41, p= 0.097), >10 lesiones focales (HR 3.01, IC 95% 1.04-8.68, p= 0.041), TMTV>210 cm3 (HR 3.69, IC 95% 1.07-12.66, p= 0.038) y enfermedad extramedular (EM) (HR 4.19, IC 95% 1.25-14.08, p= 0.02) en FDG PET/TC, así como valores de TMTV>366 cm3 (HR 4.58, IC 95% 1.25-16.82, p= 0.02), TLMU>1448 g (HR 4, IC 95% 1.09-14.64, p= 0.04) y EM (HR 4.19, IC 95% 1.25-14.08, p= 0.02) en MET PET/TC. En el análisis univariante para la supervivencia global (SG), solamente se encontró asociación pronóstica con la EM en FDG y MET PET/TC (HR 4.92, IC 95% 1.15-21.04, p= 0.03). En el análisis multivariante, únicamente la presencia de >10 lesiones focales (HR 3.81, IC 95% 1.11-13.05, p= 0.033) y TMTV>210 cm3 (HR 3.26, IC 95% 0.89-11.88, p= 0.073) en FDG PET/TC fueron factores predictores de supervivencia independientes. En base a resultados obtenidos, se desestima la COL para la investigación de MM en el modelo murino. Tanto la FDG como la MET permiten realizar estudio de extensión y valoración de respuesta al tratamiento. En pacientes con MM se confirma el valor diagnóstico y pronóstico de la FDG. La MET es un radiofármaco con rendimiento diagnóstico superior a la FDG, postulándose por primera vez el valor pronóstico de TMTV, TLMU. Sin embargo, son necesarios estudios con mayor número de pacientes y seguimiento para confirmar el valor pronóstico, tal y como se ha reportado con FDG PET/TC