Chiara Capelli

Autologous Mesenchymal Stromal Cells and Kidney Transplantation: A Pilot Study of Safety and Clinical Feasibility

BACKGROUND AND OBJECTIVES Mesenchymal stromal cells (MSCs) abrogate alloimmune response in vitro, suggesting a novel cell-based approach in transplantation. Moving this concept toward clinical application in organ transplantation should be critically assessed. DESIGN, SETTING, PARTICIPANTS & MEASUREMENTS A safety and clinical feasibility study (ClinicalTrials.gov, NCT00752479) of autologous MSC infusion was conducted in two recipients of kidneys from living-related donors. Patients were given T cell-depleting induction therapy and maintenance immunosuppression with cyclosporine and mycophenolate mofetil. On day 7 posttransplant, MSCs were administered intravenously. Clinical and immunomonitoring of MSC-treated patients was performed up to day 360 postsurgery. RESULTS Serum creatinine levels increased 7 to 14 days after cell infusion in both MSC-treated patients. A graft biopsy in patient 2 excluded acute graft rejection, but showed a focal inflammatory infiltrate, mostly granulocytes. In patient 1 protocol biopsy at 1-year posttransplant showed a normal graft. Both MSC-treated patients are in good health with stable graft function. A progressive increase of the percentage of CD4+CD25highFoxP3+CD127- Treg and a marked inhibition of memory CD45RO+RA-CD8+ T cell expansion were observed posttransplant. Patient T cells showed a profound reduction of CD8+ T cell activity. CONCLUSIONS Findings from this study in the two patients show that MSC infusion in kidney transplant recipients is feasible, allows enlargement of Treg in the peripheral blood, and controls memory CD8+ T cell function. Future clinical trials with MSCs to look with the greatest care for unwanted side effects is advised.

Transfer of Growth Factor Receptor mRNA Via Exosomes Unravels the Regenerative Effect of Mesenchymal Stem Cells

Bone marrow-mesenchymal stem cells (BM-MSC) ameliorate renal dysfunction and repair tubular damage of acute kidney injury by locally releasing growth factors, including the insulin-like growth factor-1 (IGF-1). The restricted homing of BM-MSC at the site of injury led us to investigate a possible gene-based communication mechanism between BM-MSC and tubular cells. Human BM-MSC (hBM-MSC) released microparticles and exosomes (Exo) enriched in mRNAs. A selected pattern of transcripts was detected in Exo versus parental cells. Exo expressed the IGF-1 receptor (IGF-1R), but not IGF-1 mRNA, while hBM-MSC contained both mRNAs. R- cells lacking IGF-1R exposed to hBM-MSC-derived Exo acquired the human IGF-1R transcript that was translated in the corresponding protein. Transfer of IGF-1R mRNA from Exo to cisplatin-damaged proximal tubular cells (proximal tubular epithelial cell [PTEC]) increased PTEC proliferation. Coincubation of damaged PTEC with Exo and soluble IGF-1 further enhanced cell proliferation. These findings suggest that horizontal transfer of the mRNA for IGF-1R to tubular cells through Exo potentiates tubular cell sensitivity to locally produced IGF-1 providing a new mechanism underlying the powerful renoprotection of few BM-MSC observed in vivo.

Life-Sparing Effect of Human Cord Blood-Mesenchymal Stem Cells in Experimental Acute Kidney Injury

In search for new sources of mesenchymal stem cells (MSCs) for renal repair in acute kidney injury (AKI), we investigated the potential of human cord blood (CB)‐MSCs to cure mice with AKI. Infusion of CB‐MSCs in immunodeficient mice with cisplatin‐induced AKI ameliorated both renal function and tubular cell injury, and prolonged survival. Transplanted CB‐MSCs localized in peritubular areas, limited capillary alterations and neutrophil infiltration. Apoptosis reduced and tubular cell proliferation increased by virtue of stem cell capacity to produce growth factors. The reno‐protective effect of CB‐MSCs was further confirmed by their ability to inhibit oxidative damage and to induce the prosurvival factor Akt in tubular cells. The evidence that CB‐MSCs in vitro increased the production of growth factors and inhibited IL‐1β and TNFα synthesis when cocultured with damaged proximal tubular cells indicates a regenerative and anti‐inflammatory action of stem cell treatment. Altogether these results highlight the potential of human CB‐MSCs as future cell therapy for testing in human AKI. STEM CELLS 2010;28:513–522

Treatment of Graft versus Host Disease with Mesenchymal Stromal Cells: A Phase I Study on 40 Adult and Pediatric Patients

This phase I multicenter study was aimed at assessing the feasibility and safety of intravenous administration of third party bone marrow-derived mesenchymal stromal cells (MSC) expanded in platelet lysate in 40 patients (15 children and 25 adults), experiencing steroid-resistant grade II to IV graft-versus-host disease (GVHD). Patients received a median of 3 MSC infusions after having failed conventional immunosuppressive therapy. A median cell dose of 1.5 × 10(6)/kg per infusion was administered. No acute toxicity was reported. Overall, 86 adverse events and serious adverse events were reported in the study, most of which (72.1%) were of infectious nature. Overall response rate, measured at 28 days after the last MSC injection, was 67.5%, with 27.5% complete response. The latter was significantly more frequent in patients exhibiting grade II GVHD as compared with higher grades (61.5% versus 11.1%, P = .002) and was borderline significant in children as compared with adults (46.7 versus 16.0%, P = .065). Overall survival at 1 and 2 years from the first MSC administration was 50.0% and 38.6%, with a median survival time of 1.1 years. In conclusion, MSC can be safely administered on top of conventional immunosuppression for steroid resistant GVHD treatment. Eudract Number 2008-007869-23, NCT01764100.

Mesenchymal stromal cells and kidney transplantation: Pretransplant infusion protects from graft dysfunction while fostering immunoregulation

Bone marrow‐derived mesenchymal stromal cells (MSC) have emerged as useful cell population for immunomodulation therapy in transplantation. Moving this concept towards clinical application, however, should be critically assessed by a tailor‐made step‐wise approach. Here, we report results of the second step of the multistep MSC‐based clinical protocol in kidney transplantation. We examined in two living‐related kidney transplant recipients whether: (i) pre‐transplant (DAY‐1) infusion of autologous MSC protected from the development of acute graft dysfunction previously reported in patients given MSC post‐transplant, (ii) avoiding basiliximab in the induction regimen improved the MSC‐induced Treg expansion previously reported with therapy including this anti‐CD25‐antibody. In patient 3, MSC treatment was uneventful and graft function remained normal during 1 year follow‐up. In patient 4, acute cellular rejection occurred 2 weeks post‐transplant. Both patients had excellent graft function at the last observation. Circulating memory CD8+ T cells and donor‐specific CD8+ T‐cell cytolytic response were reduced in MSC‐treated patients, not in transplant controls not given MSC. CD4+FoxP3+Treg expansion was comparable in MSC‐treated patients with or without basiliximab induction. Thus, pre‐transplant MSC no longer negatively affect kidney graft at least to the point of impairing graft function, and maintained MSC‐immunomodulatory properties. Induction therapy without basiliximab does not offer any advantage on CD4+FoxP3+Treg expansion (ClinicalTrials.gov number: NCT 00752479).

Toward MSC in Solid Organ Transplantation: 2008 Position Paper of the MISOT Study Group

The following position paper summarizes the recommendations for early clinical trials and ongoing basic research in the field of mesenchymal stem cell-induced solid organ graft acceptance—agreed upon on the first meeting of the Mesenchymal Stem Cells In Solid Organ Transplantation (MISOT) study group in late 2008.

Feasibility and Safety of Adoptive Immunotherapy with CIK Cells after Cord Blood Transplantation

Five patients with aggressive acute leukemias who had relapsed after cord blood transplantation were treated with cord blood derived cytokine-induced killer (CIK) cells. These were obtained by ex vivo expansion, using as starting material the washouts of the cord blood units, left over at the end of the transplant. We did not observe any acute or delayed adverse event, and observed 1 partial response in 1 patient concomitantly with the development of acute grade III graft-versus-host disease (GVHD). These observations show the relatively low toxicity of cord blood-derived CIK cells and, more importantly, the feasibility of this immunotherapy program for patients who could not otherwise benefit from donor lymphocyte infusions.

Transplanted Umbilical Cord Mesenchymal Stem Cells Modify the In Vivo Microenvironment Enhancing Angiogenesis and Leading to Bone Regeneration

Umbilical cord mesenchymal stem cells (UC-MSCs) show properties similar to bone marrow mesenchymal stem cells (BM-MSCs), although controversial data exist regarding their osteogenic potential. We prepared clinical-grade UC-MSCs from Whartons Jelly and we investigated if UC-MSCs could be used as substitutes for BM-MSCs in muscoloskeletal regeneration as a more readily available and functional source of MSCs. UC-MSCs were loaded onto scaffolds and implanted subcutaneously (ectopically) and in critical-sized calvarial defects (orthotopically) in mice. For live cell-tracking experiments, UC-MSCs were first transduced with the luciferase gene. Angiogenic properties of UC-MSCs were tested using the mouse metatarsal angiogenesis assay. Cell secretomes were screened for the presence of various cytokines using an array assay. Analysis of implanted scaffolds showed that UC-MSCs, contrary to BM-MSCs, remained detectable in the implants for 3 weeks at most and did not induce bone formation in an ectopic location. Instead, they induced a significant increase of blood vessel ingrowth. In agreement with these observations, UC-MSC-conditioned medium presented a distinct and stronger proinflammatory/chemotactic cytokine profile than BM-MSCs and a significantly enhanced angiogenic activity. When UC-MSCs were orthotopically transplanted in a calvarial defect, they promoted increased bone formation as well as BM-MSCs. However, at variance with BM-MSCs, the new bone was deposited through the activity of stimulated host cells, highlighting the importance of the microenvironment on determining cell commitment and response. Therefore, we propose, as therapy for bone lesions, the use of allogeneic UC-MSCs by not depositing bone matrix directly, but acting through the activation of endogenous repair mechanisms.

The washouts of discarded bone marrow collection bags and filters are a very abundant source of hMSCs

BACKGROUND AIMS Human multipotent mesenchymal stromal cells (hMSC) are considered good candidates for a growing spectrum of cell therapies. We have validated a protocol that makes use of the washouts of discarded collection sets, left over at the end of the filtration of bone marrow (BM) explants performed for hematopoietic stem cell (HSC) transplantation. METHODS The method consists of direct plating of cells without density-gradient isolation followed by two detachment steps and expansion in 5% human platelet lysate (hPL). RESULTS In a median of 26 days, 14 bags for adult patients and nine bags for pediatric patients for a standard dose of 1x10(6) hMSC/kg body weight could be prepared from the expansion of a fraction of the cells recovered from seven independent washouts. Moreover, 151 vials could be frozen from the remaining cells. The theoretical full expansion of all the frozen vials (validated by the expansion of two independent vials) could have allowed the production of 173 bags for adults and 348 bags for pediatric patients. CONCLUSIONS The washouts of discarded bags and filters left over at the end of routine BM explants filtration are a very abundant source of hMSC precursors that can be easily utilized for clinical applications.

Frequent occurrence of non-malignant genetic alterations in clinical grade mesenchymal stromal cells expanded for cell therapy protocols

Human bone marrow mesenchymal stromal cells (BM-MSC) represent one of the most investigated “advanced therapeutic medicinal products”.[1][1] Recent safety concerns have focused attention on the possible malignant transformation due to mutations acquired during their large-scale in vitro