Beneficial Effect of Systemic Allogeneic Adipose Derived Mesenchymal Cells on the Clinical, Inflammatory and Immunologic Status of a Patient With Recessive Dystrophic Epidermolysis Bullosa: A Case Report.

Recessive dystrophic epidermolysis bullosa (RDEB) is an incurable inherited mucocutaneous fragility disorder characterized by recurrent blisters, erosions, and wounds. Continuous blistering triggers overlapping cycles of never-ending healing and scarring commonly evolving to chronic systemic inflammation and fibrosis. The systemic treatment with allogeneic mesenchymal cells (MSC) from bone marrow has previously shown benefits in RDEB. MSC from adipose tissue (ADMSC) are easier to isolate. This is the first report on the use of systemic allogeneic ADMSC, correlating the clinical, inflammatory, and immunologic outcomes in RDEB indicating long-lasting benefits. We present the case of an RDEB patient harboring heterozygous biallelic COL7A1 gene mutations and with a diminished expression of C7. The patient presented with long-lasting refractory and painful oral ulcers distressing her quality of life. Histamine receptor antagonists, opioid analgesics, proton-pump inhibitors, and low-dose tricyclic antidepressants barely improved gastric symptoms, pain, and pruritus. Concomitantly, allogeneic ADMSC were provided as three separate intravenous injections of 106 cells/kg every 21 days. ADMSC treatment was well-tolerated. Improvements in wound healing, itch, pain and quality of life were observed, maximally at 6-9 months post-treatment, with the relief of symptoms still noticeable for up to 2 years. Remarkably, significant modifications in PBL participating in both the innate and adaptive responses, alongside regulation of levels of profibrotic factors, MCP-1/CCL2 and TGF-ß, correlated with the health improvement. This treatment might represent an alternative for non-responding patients to conventional management. It seems critical to elucidate the paracrine modulation of the immune system by MSC for their rational use in regenerative/immunoregulatory therapies.

Combined adipose mesenchymal stromal cell advanced therapy resolved a recalcitrant leg ulcer in an 85-year-old patient.

Venous leg ulcers (VLU) represent an uphill economic, health and social burden, aggravated in the elderly. Best-practice care interventions are often insufficient and alternative therapies need to be explored. Herein, we have treated for the first time a chronic VLU in an elderly patient by combining cell therapy and tissue engineering in the context of a compassionate use. The administration of allogeneic adipose-derived mesenchymal stromal cells (MSCs) embedded in a plasma-based bioengineered dermis covering the ulcer bed and also injected into the ulcer margins led to the complete closure of a 10-year recalcitrant VLU in an 85-year-old patient. Regenerative properties of MSCs might be boosted by the use of bioengineered matrices for their delivery.

sRAGE as severe acute bronchiolitis biomarker, prospective observational study.

INTRODUCTION AND OBJECTIVES: Acute bronchiolitis (AB) is the leading cause of hospitalization in infants and around 5% require intensive care treatment. Early identification of children diagnosed with AB at a high risk of severe progression is of great interest. The receptor for advanced glycation end products (RAGE), highly expressed in lung tissue, regulates immune responses and inflammation, and its soluble form, sRAGE, is believed to have an anti-inflammatory role. We hypothesized serum sRAGE might be a major determinant of AB severity and prognosis. This study was conducted to measure serum sRAGE in infants with severe AB and to assess its correlation with clinical severity, immediate complications, and outcome. METHODS: Single-center, prospective, observational study of hospitalized children with severe bronchiolitis admitted to the Pediatric Intensive Care Unit (PICU), from September 2015 to September 2016. RESULTS: A total of 52 children and 27 controls were included. The cases age ranged from 11 days to 21 months, resulting in a significant age difference with controls (11.85 vs 4.84 months, P < .01). Serum levels of sRAGE were lower but not significant in severe AB patients than in controls (1350.93 vs 1450.42 pg/mL; P = .399). No correlation was found between serum sRAGE and causative viruses, clinical symptoms, Wood-Downes score (a clinical severity score) on admission, respiratory support, or length of hospital stay. Serum sRAGE was also lower in the cases having had a previous respiratory disease (1463.84 vs 1072.43 pg/mL; P = .049). However, it was higher in patients with any lung consolidation on the chest X-ray (1584.79 vs 1131.62 pg/mL; P = .044) and weakly positively correlated with classical biomarkers (maximum C-reactive protein, +0.295, P = .034; maximum procalcitonin, +0.309; P = .029). CONCLUSION: This single-center study reveals that sRAGE couldn't predict AB severity or outcome in children hospitalized at PICU. Nevertheless, it significantly increased in the presence of any lung consolidation and had a positive correlation with classical biomarkers. The utility of sRAGE in this population could be probably elucidated with a better understanding of AGE-RAGE axis.

Genetic and Immune Changes Associated with Disease Progression under the Pressure of Oncolytic Therapy in A Neuroblastoma Outlier Patient.

Little is known about the effect of oncolytic adenovirotherapy on pediatric tumors. Here we present the clinical case of a refractory neuroblastoma that responded positively to Celyvir (ICOVIR-5 oncolytic adenovirus delivered by autologous mesenchymal stem cells) for several months. We analyzed samples during tumor evolution in order to identify molecular and mutational features that could explain the interactions between treatment and tumor and how the balance between both of them evolved. We identified a higher adaptive immune infiltration during stabilized disease compared to progression, and also a higher mutational rate and T-cell receptor (TCR) diversity during disease progression. Our results indicate an initial active role of the immune system controlling tumor growth during Celyvir therapy. The tumor eventually escaped from the control exerted by virotherapy through acquisition of resistance by the tumor microenvironment that exhausted the initial T cell response.

First-in-Human, First-in-Child Trial of Autologous MSCs Carrying the Oncolytic Virus Icovir-5 in Patients with Advanced Tumors.

We present here the results of a first-in-human, first-in-child trial for patients with relapsed/refractory solid tumors using Celyvir, an advanced therapy medicine that combines autologous mesenchymal stem cells (MSCs) carrying an oncolytic adenovirus. Celyvir was manufactured from a bone marrow aspirate and then given intravenously. Patients received weekly infusions for 6 weeks at a dose of 2 × 106 cells/kg (children) or 0.5-1 × 106 cells/kg (adults), 2 × 104 viral particles per cell. Fifteen pediatric and 19 adult patients were recruited, but 18 were screen failures, mainly because rapid disease progression before Celyvir was available. No grade 2-5 toxicities were reported. Adenoviral replication detected by PCR was found in all but 2 pediatric patient and in none of the adult ones. Absolute numbers of circulating leukocytes suffered minor changes along therapy, but some subsets showed differences comparing the pediatric versus the adult cohorts. Two patients with neuroblastoma showed disease stabilization, and one of them continued on treatment for up to 6 additional weeks. Celyvir, the combination of MSCs and oncolytic adenovirus, is safe and warrants further evaluation in a phase 2 setting. The use of MSCs may be a strategy to increase the amount of oncolytic virus administered to patients, minimizing toxicities and avoiding direct tumor injections.

Molecular Scaffolds as Double-Targeting Agents For the Diagnosis and Treatment of Neuroblastoma.

The selective delivery of therapeutic and imaging agents to tumoral cells has been postulated as one of the most important challenges in the nanomedicine field. Meta-iodobenzilguanidine (MIBG) is widely used for the diagnosis of neuroblastoma (NB) due to its strong affinity for the norepinephrine transporter (NET), usually overexpressed on the membrane of malignant cells. Herein, a family of novel Y-shaped scaffolds has been synthesized, which have structural analogues of MIBG covalently attached at each end of the Y-structure. The cellular uptake capacity of these double-targeting ligands has been evaluated in vitro and in vivo, yielding one specific Y-shaped structure that is able to be engulfed by the malignant cells, and accumulates in the tumoral tissue, at significantly higher levels than the structure containing only one single targeting agent. This Y-shaped ligand can provide a powerful tool for the current treatment and diagnosis of this disease.

Off-label mesenchymal stromal cell treatment in two infants with severe bronchopulmonary dysplasia: clinical course and biomarkers profile.

BACKGROUND: Bronchopulmonary dysplasia (BPD) is the most prevalent sequelae of premature birth, for which therapeutic options are currently limited. Mesenchymal stromal cells (MSCs) are a potential therapy for prevention or reversal of BPD. SERIES OF CASES: We report on two infants with severe BPD in whom off-label treatment with repeated intravenous doses of allogeneic bone marrow-derived MSCs were administered. We analyzed the temporal profile of serum and tracheal cytokines and growth factors as well as safety, tolerability and clinical response. The administration of repeated intravenous doses of MSCs in two human babies with severe and advanced BPD was feasible and safe and was associated with a decrease of pro-inflammatory molecules and lung injury biomarkers. Both patients were at very advanced stages of BPD with very severe lung fibrosis and did not survive the disease. CONCLUSIONS: MSCs are a promising therapy for BPD, but they should be administered in early stages of the disease.

Circulating soluble RAGE and cell surface RAGE on peripheral blood mononuclear cells in healthy children.

BACKGROUND: The receptor for advanced glycation end products (RAGE) has a critical role in the pathogenesis of inflammation. In healthy children, its basal expression on the peripheral blood mononuclear cell (PBMC) and the basal circulating soluble RAGE (sRAGE) levels are unknown. The aim of this study was to describe both. METHODS: This is a monocentric, observational and descriptive study of samples obtained from healthy children. The RAGE expression on PBMC was analyzed using flow cytometry. The sRAGE values were determined with a specific sandwich enzyme-linked immunosorbent assay (ELISA) kit, later the relation between cellular RAGE and sRAGE was described. RESULTS: Forty-three children were included. The median sRAGE level was 849.0±579.0 pg/mL. The RAGE mean fluorescence intensity (MFI) was 1382±506 in monocytes and 792±506 in lymphocytes. There were no differences between genders. A negative correlation was found between sRAGE and RAGE MFI in lymphocytes (r=-0.3; p=0.04). CONCLUSIONS: We describe for the first time the RAGE surface levels on PBMC in children. It showed a negative correlation with sRAGE. The sRAGE circulating level is lower than the sRAGE level described in adult population or non-healthy children. Our findings should be confirmed in order to apply them as reference values for future investigations.

CD64 on monocytes and granulocytes in severe acute bronchiolitis: Pilot study on its usefulness as a bacterial infection biomarker.

The CD64 receptor has been described as a biomarker of bacterial infection. We speculated that CD64 surface expression on monocytes and granulocytes of children with severe acute bronchiolitis (SAB) could be altered in cases of probable bacterial infection (PBI) determined using classical biomarkers (procalcitonin and C-reactive protein, leukocyte count, and radiographic findings). A prospective observational pilot study was conducted from October 2015 to February 2016 in children admitted for pediatric critical care. A blood sample was taken in the first 24 hours of admission, and CD64 was measured by flow cytometry. The values obtained were analyzed and correlated with traditional biomarkers of PBI. Thirty-two children were included; a correlation was found between CD64 expression and the PBI criteria. CD64 surface expression was higher in children with PBI (area under the receiver operating characteristic curve of 0.73; P = 0.042) and the percentage of CD64+ granulocytes was higher in children with PBI. This is the first study to describe CD64 surface expression on monocytes and granulocytes in SAB, finding CD64 values to be higher in children with PBI. Larger clinical studies are needed to elucidate the real accuracy of CD64 as a biomarker of bacterial infection.

Influence of carrier cells on the clinical outcome of children with neuroblastoma treated with high dose of oncolytic adenovirus delivered in mesenchymal stem cells.

We report here our clinical experience of a program of compassionate use of Celyvir--autologous marrow-derived mesenchymal stem cells (MSCs) carrying an oncolytic adenovirus--for treating children with advanced metastatic neuroblastoma. Children received weekly doses of Celyvir with no concomitant treatments. The tolerance was excellent, with very mild and self-limited viral-related symptoms. Patients could be distinguished based on their response to therapy: those who had a clinical response (either complete, partial or stabilization) and those who did not respond. We found differences between patients who responded versus those who did not when analyzing their respective MSCs, at the expression levels of adhesion molecules (CCR1, CXCR1 and CXCR4) and in migration capacities in transwell assays, and in immune-related molecules (IFNγ, HLA-DR). These results suggest interpatient differences in the homing and immune modulation capacities of the therapy administered. In addition, the pretherapy immune T cell status and the T effector response were markedly different between responders and non-responders. We conclude that multidoses of Celyvir have an excellent safety profile in children with metastatic neuroblastoma. Intrinsic patients' and MSCs' factors appear to be related to clinical outcome.

Effects of local administration of allogenic adipose tissue-derived mesenchymal stem cells on functional recovery in experimental traumatic brain injury.

OBJECTIVE: Traumatic brain injury (TBI) is the leading cause of mortality and morbidity in paediatric patients after the first year of life. The aim of this study was to evaluate effects of locally administered allogeneic mesenchymal stem cells (MSC), in the acute period after a TBI. METHODOLOGY: MSC were isolated from peritoneal fat of healthy rats, expanded in vitro and labelled with the green fluorescent protein. Rats were placed in one of three experimental groups: (1) CONTROL: TBI, (2) IP-CONTROL: TBI + local saline and (3) IP-Treat: TBI + 2 × 10(5) MSC 24 hours after receiving a moderate, unilateral, controlled cortical impact. Motor and cognitive behavioural tests were performed to evaluate functional recovery. Histological examination and immunohistochemistry were used to identify cell distribution. MAIN RESULTS: Improved performance was found on motor tests in the MSC-treated group compared to control groups. MSC were found in the perilesional area and their number decreased with time after transplantation. MSC treatment increased the cell density in the hippocampus (CA3 pyramidal cells and granule cells in the dentate gyrus) and enhanced neurogenesis in this area. CONCLUSION: MSC cell therapy resulted in better recovery of motor function compared with the control group. This cellular therapy might be considered for patients suffering from TBI.

Chemokines and relapses in childhood acute lymphoblastic leukemia: A role in migration and in resistance to antileukemic drugs.

We studied whether chemokines may have a role in relapses in childhood acute lymphoblastic leukemia (ALL). We compared the levels of chemokine receptors in marrow samples from 82 children with ALL at diagnosis versus 15 at relapses, and quantified the levels of chemokines in central system fluid (CSF) samples. The functional role of specific chemokines was studied in vitro and in vivo. The expression of some chemokine receptors was upregulated upon leukemic relapse, both in B- and in T-ALL, and in cases of medullary and extramedullary involvement. CXCL10 induced chemotaxis in leukemic cell lines and in primary leukemic cells, depending upon the levels of CXCR3 expression. CXCL10 specifically diminished chemotherapy-induced apoptosis on ALL cells expressing CXCR3, partially inhibiting caspase activation and maintaining the levels of the antiapoptotic protein Bcl-2. Finally, immunodeficient mice engrafted with CXCR3-expressing human leukemic cells showed decreased infiltration of marrow, spleen, and CNS after receiving a CXCR3-antagonist molecule. CXCR3 signaling in ALL may have a dual function: chemotactic for the localisation of leukemic blasts in specific niches, and it may also confer resistance to chemotherapy, enhancing the chances for relapses.

The NFKB Inducing Kinase Modulates Hematopoiesis During Stress.

The genetic programs that maintain hematopoiesis during steady state in physiologic conditions are different from those activated during stress. Here, we show that hematopoietic stem cells (HSCs) with deficiencies in components of the alternative NFκB pathway (the NFκB inducing kinase, NIK, and the downstream molecule NFκB2) had a defect in response to stressors such as supraphysiological doses of cytokines, chemotherapy, and hematopoietic transplantation. NIK-deficient mice had peripheral blood and bone marrow leukocyte numbers within normal ranges (except for the already reported defects in B-cell maturation); however, HSCs showed significantly slower expansion capacity in in vitro cultures compared to wild-type HSCs. This was due to a delayed cell cycle and increased apoptosis. In vivo experiments showed that NIK-deficient HSCs did not recover at the same pace as controls when challenged with myeloablative chemotherapy. Finally, NIK-deficient HSCs showed a significantly decreased competitive repopulation capacity in vivo. Using HSCs from mice deficient in one of two downstream targets of NIK, that is, either NFκB2 or c-Rel, only NFκB2 deficiency recapitulated the defects detected with NIK-deficient HSCs. Our results underscore the role of NIK and the alternative NFκB pathway for the recovery of normal levels of hematopoiesis after stress.

Mesenchymal stem cells derived from low risk acute lymphoblastic leukemia patients promote NK cell antitumor activity.

Mesenchymal stem cells (MSCs) are key components of the bone marrow microenvironment which contribute to the maintenance of the hematopoietic stem cell niche and exert immunoregulatory functions in innate and adaptive immunity. We analyze the immunobiology of MSCs derived from acute lymphoblastic leukemia (ALL) patients and their impact on NK cell function. In contrast to the inhibitory effects on the immune response exerted by MSCs from healthy donors (Healthy-MSCs), we demonstrate that MSCs derived from low/intermediate risk ALL patients at diagnosis (ALL-MSCs) promote an efficient NK cell response including cytokine production, phenotypic activation and most importantly, cytotoxicity. Longitudinal studies indicate that these immunostimulatory effects of ALL-MSCs are progressively attenuated. Healthy-MSCs adopt ALL-MSC-like immunomodulatory features when exposed to leukemia cells, acquiring the ability to stimulate NK cell antitumor function. The mechanisms underlying to these functional changes of ALL-MSCs include reduced production of soluble inhibitory factors, differential expression of costimulatory and coinhibitory molecules, increased expression of specific TLRs and Notch pathway activation. Collectively our findings indicate that, in response to leukemia cells, ALL-MSCs could mediate a host beneficial immunomodulatory effect by stimulating the antitumor innate immune response.

A new targeting agent for the selective drug delivery of nanocarriers for treating neuroblastoma.

Novel targeting agents against neuroblastoma based on the meta-iodobenzylguanidine (MIBG) moiety were synthesized and biologically evaluated for nanocarrier vectorization. These compounds have been anchored on the surface of drug loaded mesoporous silica nanocarriers, resulting in the improved cellular uptake in tumoral cells. Neuroblastoma (NB) is the most frequent extracranial pediatric tumor. Advanced forms of the disease (metastatic and/or refractory) have a dismal prognosis despite the combination of chemotherapy, radiotherapy, surgery and bone narrow transplants. These treatments carry severe side effects and, in some cases, compromise the life of the patient. MIBG has been widely applied in the medical diagnosis of NB due to its affinity for tumor cells through the norepinephrine transporter (NET), which is expressed in 90% of NB tumors. The exclusive accumulation of MIBG in neuroblastoma has been widely studied; however, its properties have been never exploited as a targeting agent in nanocarrier drug delivery systems. Several structural analogues of MIBG have been prepared and attached on the surface of nanocarriers. Their selective internalization has been tested against human neuroblastoma cells, which show, in the best case, cellular uptake four times higher than that of the naked nanosystem. Furthermore, in vivo experiments showed preferential and selective accumulation and retention of the targeted nanosystem comparing with the naked and only PEGylated counterpart systems. This novel nanosystem could be easily applicable to all kinds of drug delivery nanocarriers, providing a universal tool for neuroblastoma chemotherapies that is superior to classical approaches through a novel nanosystem exclusively designed to target this terrible malignancy.

Patient-derived mesenchymal stem cells as delivery vehicles for oncolytic virotherapy: novel state-of-the-art technology.

Oncolytic virotherapy is gaining interest in the clinic as a new weapon against cancer. In vivo administration of oncolytic viruses showed important limitations that decrease their effectiveness very significantly: the antiviral immune response causes the elimination of the therapeutic effect, and the poor natural ability of oncolytic viruses to infect micrometastatic lesions significantly minimizes the effective dose of virus. This review will focus on updating the technical and scientific foundations of one of the strategies developed to overcome these limitations, ie, using cells as vehicles for oncolytic viruses. Among many candidates, a special type of adult stem cell, mesenchymal stem cells (MSCs), have already been used in the clinic as cell vehicles for oncolytic viruses, partly due to the fact that these cells are actively being evaluated for other indications. MSC carrier cells are used as Trojan horses loaded with oncoviruses, are administered systemically, and release their cargos at the right places. MSCs are equipped with an array of molecules involved in cell arrest in the capillaries (integrins and selectins), migration toward specific parenchymal locations within tissues (chemokine receptors), and invasion and degradation of the extracellular matrix (proteases). In addition to anatomical targeting capacity, MSCs have a well-recognized role in modulating immune responses by affecting cells of the innate (antigen-presenting cells, natural killer cells) and adaptive immune system (effector and regulatory lymphocytes). Therefore, carrier MSCs may also modulate the immune responses taking place after therapy, ie, the antiviral and the antitumor immune responses.

Serum sRAGE as a potential biomarker for pediatric bronchiolitis: a pilot study.

PURPOSE: Traditional inflammatory biomarkers are insufficient for the evaluation of bronchiolitis severity. Recent investigations have shown that the receptor for advanced glycation end product (RAGE) and its soluble isoforms (sRAGE) play a critical role in the pathogenesis of lung injury. Main objective was to assess the serum levels of sRAGE of children with severe bronchiolitis admitted to the pediatric intensive care unit (PICU). Secondary objective was to study sRAGE correlation with the evolution and traditional biomarkers. METHODS: Prospective, observational and descriptive study, 43 healthy controls and 37 patients (December 2011-February 2012) were enrolled. sRAGE levels were assessed and compared. In patients, the relation between sRAGE levels and clinical evolution, respiratory assistance, white blood cell count, absolute neutrophils count, serum C-reactive protein, and serum procalcitonin was analyzed. RESULTS: A statistical difference was found in the mean value of sRAGE at PICU admission between patients and controls (1,215.7 ± 535 vs 849 ± 579 pg/ml). Also a significant inverse correlation was found between sRAGE and the Wood-Downes Score at admission (p = 0.02). CONCLUSIONS: Serum sRAGE could be elevated in children with bronchiolitis. Larger clinical studies are necessary to elucidate its role as a bronchiolitis inflammatory and/or lung injury biomarker.

Optimal effector functions in human natural killer cells rely upon autocrine bone morphogenetic protein signaling.

Natural killer (NK) cells are critical for innate tumor immunity due to their specialized ability to recognize and kill neoplastically transformed cells. However, NK cells require a specific set of cytokine-mediated signals to achieve optimal effector function. Th1-associated cytokines promote effector functions that are inhibited by the prototypic Th2 cytokine IL4 and the TGFß superfamily members TGFß1 and activin-A. Interestingly, the largest subgroup of the TGFß superfamily are the bone morphogenetic proteins (BMP), but the effects of BMP signaling on NK cell effector functions have not been evaluated. Here, we demonstrate that blood-circulating NK cells express type I and II BMP receptors, BMP-2 and BMP-6 ligands, and phosphorylated isoforms of Smad-1/-5/-8, which mediate BMP family member signaling. In opposition to the inhibitory effects of TGFß1 or activin-A, autocrine BMP signaling was supportive to NK cell function. Mechanistic investigations in cytokine and TLR-L-activated NK cells revealed that BMP signaling optimized IFNγ and global cytokine and chemokine production, phenotypic activation and proliferation, and autologous dendritic cell activation and target cytotoxicity. Collectively, our findings identify a novel auto-activatory pathway that is essential for optimal NK cell effector function, one that might be therapeutically manipulated to help eradicate tumors. Cancer Res; 74(18); 5019-31. ©2014 AACR.

PEG-pHPMAm-based polymeric micelles loaded with doxorubicin-prodrugs in combination antitumor therapy with oncolytic vaccinia viruses.

An enzymatically activatable prodrug of doxorubicin was covalently coupled, using click-chemistry, to the hydrophobic core of poly(ethylene glycol)-b-poly[N-(2-hydroxypropyl)-methacrylamide-lactate] micelles. The release and cytotoxic activity of the prodrug was evaluated in vitro in A549 non-small-cell lung cancer cells after adding ß-glucuronidase, an enzyme which is present intracellularly in lysosomes and extracellularly in necrotic areas of tumor lesions. The prodrug-containing micelles alone and in combination with standard and ß-glucuronidase-producing oncolytic vaccinia viruses were also evaluated in vivo, in mice bearing A549 xenograft tumors. When combined with the oncolytic viruses, the micelles completely blocked tumor growth. Moreover, a significantly better antitumor efficacy as compared to virus treatment alone was observed when ß-glucuronidase virus treated tumor-bearing mice received the prodrug-containing micelles. These findings show that combining tumor-targeted drug delivery systems with oncolytic vaccinia viruses holds potential for improving anticancer therapy.

Mesenchymal stromal cells derived from the bone marrow of acute lymphoblastic leukemia patients show altered BMP4 production: correlations with the course of disease.

The relevance of tumor microenvironment for the development and progression of tumor cells in hematological malignancies has been extensively reported. Identification of factors involved in the information exchange between the malignant cells and the bone marrow mesenchymal stem cells (BM-MSCs) and the knowledge on their functioning may provide important information to eliminate leukemic cells from protective BM niches. We evaluated changes in BM-MSCs obtained from children with acute lymphoblastic leukemia (ALL) at different times in the course of disease. Whereas ALL-MSCs did not exhibit phenotypic changes compared to BM-derived MSCs isolated from healthy donors, they exhibited increased adipogenic capacity. In addition, the viability of healthy CD34+ hematopoietic progenitors was significantly reduced when co-cultured with ALL-MSCs. ALL-MSCs grow less efficiently, although gradually recover normal growth with treatment. Accordingly, proliferation is particularly low in MSCs obtained at diagnosis and in the first days of treatment (+15 days), recovering to control levels after 35 days of treatment. Correlating these results with bone morphogenetic protein 4 (BMP4) production, a molecule demonstrated to affect MSC biology, we found higher production of BMP4 in ALL-MSCs derived from patients over the course of disease but not in those free of leukemia. However, no significant differences in the expression of different members of the BMP4 signaling pathway were observed. Furthermore, an inverse correlation between high levels of BMP4 production in the cultures and MSC proliferation was found, as observed in MSCs derived from patients at diagnosis that produce high BMP4 levels. In addition, co-culturing ALL-MSC with the REH leukemia cell line, but not CD34+ hematopoietic progenitors, powerfully enhanced BMP4 production, suggesting an intimate crosstalk among ALL-MSCs isolated from BM colonized by ALL cells that presumably also occurs in situ conditions. Our data may support the participation of BMP4 in BM niche, but the mechanism remains to be elucidated.