María Suárez-Lledó

Self-Renewing Human Bone Marrow Mesenspheres Promote Hematopoietic Stem Cell Expansion

Strategies for expanding hematopoietic stem cells (HSCs) include coculture with cells that recapitulate their natural microenvironment, such as bone marrow stromal stem/progenitor cells (BMSCs). Plastic-adherent BMSCs may be insufficient to preserve primitive HSCs. Here, we describe a method of isolating and culturing human BMSCs as nonadherent mesenchymal spheres. Human mesenspheres were derived from CD45- CD31- CD71- CD146+ CD105+ nestin+ cells but could also be simply grown from fetal and adult BM CD45--enriched cells. Human mesenspheres robustly differentiated into mesenchymal lineages. In culture conditions where they displayed a relatively undifferentiated phenotype, with decreased adherence to plastic and increased self-renewal, they promoted enhanced expansion of cord blood CD34+ cells through secreted soluble factors. Expanded HSCs were serially transplantable in immunodeficient mice and significantly increased long-term human hematopoietic engraftment. These results pave the way for culture techniques that preserve the self-renewal of human BMSCs and their ability to support functional HSCs.

Effect of meropenem administration in extended infusion on the clinical outcome of febrile neutropenia: a retrospective observational study

OBJECTIVES Information on the efficacy of extended meropenem administration in neutropenic patients is scarce. Our objective was to determine whether the administration of meropenem in a 4 h extended infusion (EI) leads to a better clinical outcome in patients with febrile neutropenia than the conventional short infusion (SI). METHODS This was a retrospective observational study. The subjects were neutropenic patients who presented with fever after receiving haematopoietic stem-cell transplantation or induction chemotherapy for acute myeloid leukaemia. The primary endpoint was the success of treatment after 5 days of meropenem therapy, defined as follows: the disappearance of fever leading to a maintained (≥ 24 h) feverless state; the resolution or improvement of the clinical signs and symptoms of infection; the absence of persistent or breakthrough bacteraemia; and no additional antibiotics prescribed because of an unsatisfactory clinical evolution. RESULTS Eighty-eight patients received meropenem (1 g/8 h) in SI and 76 received the same dose in EI. Treatment success on day 5 was superior in the EI group [52/76 (68.4%) versus 36/88 (40.9%); P<0.001]. Meropenem administered in EI was independently associated with success (OR 3.13, 95% CI 1.61-6.10). Fewer additional antibiotics were prescribed in the EI group during the first 5 days of treatment [20/76 (26.3%) versus 44/88 (50.0%); P=0.002]. Using Kaplan-Meier survival analysis a more prompt defervescence and a faster decrease in C-reactive protein concentration were observed in the EI group (P=0.021 and P=0.037, respectively). There were no significant differences in the length of hospital stay and in the mortality rate. CONCLUSIONS Meropenem administration in EI results in a better clinical outcome for febrile neutropenia episodes, with fewer additional antibiotics needed.

Autologous Haematopoietic Stem Cell Transplantation for Refractory Crohn’s Disease: Efficacy in a Single-Centre Cohort

Background Haematopoietic stem cell transplantation [HSCT] is considered a therapeutic option for patients with severe Crohns disease [CD] unresponsive to currently available therapies. Methods Autologous HSCT was considered for CD patients with active disease, unresponsive or intolerant to approved medications and unsuitable for surgery. After HSCT, patients were closely followed up every 6 weeks during the first 2 years and every 6 months thereafter up to 5 years. Colonoscopy and/or magnetic resonance imaging were performed at Months 6, 12, 24, and 48 after HSCT. Results From December 1, 2007 to December 31, 2015, 37 CD patients were assessed for HSCT. Of these, 35 patients [13 within the ASTIC trial] underwent mobilisation. Six patients did not complete the transplant for various reasons and 29 patients were finally transplanted. Patients were followed up during a median of 12 months [6-60]. At 6 months, 70% of patients achieved drug-free clinical remission (Crohns Disease Index of Severity [CDAI] < 150). The proportion of patients in drug-free remission (CDAI < 150, Simple Endoscopic activity Score [SES]-CD < 7] was 61% at 1 year, 52% at 2 years, 47% at 3 years, 39% at 4 years, and 15% at 5 years. Patients who relapsed were re-treated and 80% regained clinical remission. Six out of the 29 [21%] required surgery. One patient died due to systemic cytomegalovirus infection 2 months after transplant. Conclusions HSCT is a salvage therapy for patients with extensive and refractory CD. Although relapse occurs in a majority of patients within 5 years after transplant, drug responsiveness is regained and clinical remission achieved in 80% of cases.

Allogeneic hematopoietic SCT in multiple myeloma: long-term results from a single institution

The role of allogeneic hematopoietic SCT (allo-HCT) in multiple myeloma (MM) remains controversial. A total of 58 patients received an allo-HCT (25 of them with myeloablative conditioning—allo-MAC—and 33 with reduced-intensity conditioning—allo-RIC) at our institution over a 28-year period. The CR rate for allo-MAC was 36%. The incidence of grade III–IV acute GVHD (aGVHD) and chronic GVHD (cGVHD) was 28% and 39%, respectively The TRM at any time was 60% and the main causes of death were aGVHD or infectious complications not directly related to GVHD. The estimated PFS and OS at 15 years were 8% and 15%, respectively. The CR rate with allo-RIC was 45%. The incidence of grade III–IV aGVHD and cGVHD were 24% and 41%, respectively. The TRM at any time was 33% and was mainly related to aGVHD. The estimated PFS and OS at 5 years were 22% and 38%, respectively. Despite its high TRM, a proportion of patients with high-risk myeloma (early relapse and newly diagnosed ultrahigh risk) may obtain long-term disease control with allo-HCT. New approaches aimed at decreasing the incidence of aGVHD, and consequently to decrease the TRM, are needed.

A variant in IRF3 impacts on the clinical outcome of AML patients submitted to Allo-SCT

Allo-SCT has a strong curative potential for AML patients mainly due to a GVL effect. Unfortunately, GvL and GVHD are intimately linked. IFN regulatory factor-3 (IRF3), by modulating innate immune reactions, could impact on the incidence and intensity of GVL and GVHD. We analyzed two gene variants in IRF3 (rs7251 and rs2304205) on the clinical outcome of 249 AML patients submitted to HLA-identical sibling allo-SCT. Patients with a donor carrying the dominant GG gene variant in rs7251 had, as compared with GC and CC variants, a lower acute GVHD (aGVHD) III–IV incidence (4% vs 11% vs 27%; P=0.0078), a higher relapse incidence (49% vs 35% vs 26%; P=0.018), and lower TRM (7% vs 24% vs 18%; P=0.0065). In functional studies, the GG variant was associated with lower production of IFN-γ, decreased lymphocyte proliferation after antigen presentation by DCs, and lower cytotoxic response of mature natural killer cells. Patients carrying the AA dominant variant in rs2304205 had higher relapse incidence (50% vs 39% vs 18%, P=0.0068). The presence of both variants (GG in rs7251 and AA in rs2304205) in donors and patients resulted in a stronger clinical impact.

A constitutional variant in the transcription factor EP300 strongly influences the clinical outcome of patients submitted to allo-SCT

An adequate response of the innate immune system after allo-SCT is crucial for the clinical outcome of patients submitted to this procedure. EP300 is one of the key genes of the innate immune system (IIS). We evaluated the influence of gene variant A>G rs20551 in EP300 in donor and/or recipient on clinical results after HLA-identical sibling allo-SCT. Patients with AA gene variant had a lower relapse incidence (31 vs 48%, P=0.025; odds ratio (OR)=1.6, P=0.05), attained better disease-free survival (AA: 53% vs AG+GG: 24%, P=0.001; OR=1.8, P=0.01), and better OS (AA: 53% vs AG+GG: 34%, P=0.001; OR=1.9, P=0.007). This beneficial association was more evident when AA gene variant was present in both donor and patient. In healthy individuals, AA gene variant was associated with lower IL2 production after a mitogenic stimuli, higher CD4+ cell response after CMV infection, and higher expression of innate immune genes (IRF-3 and MIF), cell cycle genes (AURKB, CCNA2 and CCNB1), lymphocyte survival genes (NFAT5 and SLC38A2), and with a lower expression of P53 compared with recessive GG gene variant. These findings suggest a beneficial effect of the AA gene variant in rs20551 on clinical outcome after allo-SCT.

Innovative strategies minimize engraftment syndrome in multiple myeloma patients with novel induction therapy following autologous hematopoietic stem cell transplantation

Autologous stem cell transplantation (PBSCT) is standard for young patients in MM and its TRM has decreased after the 2000s. Bortezomib and immunomodulatory agents (IMiDs) in MM have improved the outcome. However, they seem to boost pro-inflammatory stage increasing the incidence of engraftment syndrome (ES). Favorable factors in PBSCT such as G-CSF could increase inflammatory stage during transplant. Corticosteroids have shown an excellent response of ES and some authors propose them as prophylaxis for ES. The aim was to analyze the impact of G-CSF avoidance and corticosteroids’ prophylaxis in 170 patients diagnosed of MM treated with bortezomib/IMiDs that underwent PBSCT. We established three groups: Group-I [(G-CSF_administration), 60 patients (35%)], group-II [(nonG-CSF), 60 patients (35%)] and group-III [(nonG-CSF plus corticosteroid’s prophylaxis), 50 patients (30%)]. A decreased ES incidence among groups was observed: 62, 42, and 22% (P < 0.0001). The incidence of symptoms mimicking a capillary leak syndrome associated with ES dropped: 43, 32, and 0% (P = 0.03). The G-CSF avoidance and corticosteroids had impact over admission 24, 21, and 20 days (P = 0.001). The most important variables related to ES were HCT-CI >2 (p < 0.0001; HR 8.5) and risk groups (p < 0.0001; HR 7.2). Hence, G-CSF avoidance and corticosteroid’s prophylaxis decrease morbidity in patients undergoing PBSCT with MM treated with bortezomib/IMiDs.

At-home autologous stem cell transplantation in multiple myeloma with and without G-CSF administration: a comparative study

Severe neutropenia remains the main cause of morbidity and mortality after autologous stem cell transplantation (ASCT). Improvements in this procedure, such as the use of peripheral blood progenitor cells and the administration of an oral antibiotic prophylaxis, have reduced the incidence of infections during the time of neutropenia and have made ASCT safer. The use of G-CSF after ASCT with the intention of reducing the duration of neutropenia and minimising the risk of severe infection is a common practice in most transplantation centres. In addition, because of a faster neutrophil recovery, G-CSF administration may also be associated with a reduction in the incidence and severity of oral and intestinal mucositis after ASCT. However, it has been reported that G-CSF administration after ASCT does not reduce the incidence or duration of febrile neutropenia or the infectious mortality rate. Moreover, G-CSF administration has been associated with a higher incidence of clinical complications such as engraftment syndrome (ES) and capillary leak syndrome. For these reasons, its use after ASCT is controversial. Thus, in May 2011, our centre discontinued the administration of post-transplant G-CSF in patients with multiple myeloma (MM). We hypothesised that the use of G-CSF in these patients does not provide any significant clinical benefit as compared with not using G-CSF. Here we report the results of this policy by comparing two cohorts of patients; one cohort received post-transplant G-CSF, whereas the other cohort did not receive G-CSF. One hundred ninety-five patients with MM underwent an ASCT between November 2000 and October 2014. Of them, 131 (67%) were not included in an at-home ASCT programme because of ECOG 42 (35%), patient decision (27%), geographic distance (21%) and no caregiver available (17%). Finally, 64 (33%) patients were included. The hospital ethics committee approved this programme and all patients signed a written informed consent. Patient-, diseaseand transplant-related variables are described in Table 1. All patients received high-dose melphalan (200 mg/m) and a peripheral blood stem cell infusion in the hospital and were discharged on day +1. Specialised nurses in Hematology looked after the patients daily, either at home or by phone. They checked patients’ vital signs, oral intake, central venous catheter status and presence of mucositis; they administered IV medications and they took blood samples. If necessary, medical assistance was provided in the hospital by a designated physician or by the haematologist on call. Patients who underwent an ASCT between November 2000 and April 2011 (n= 32) received G-CSF (5 mcg/kg per day) starting on day +7 until their ANC reached 1× 10/L for two consecutive days. The second cohort beginning in May 2011 (n= 32), received no G-CSF. All patients received antimicrobial prophylaxis with quinolone, fluconazole, aerosolized pentamidine and low-dose acyclovir if herpes serology was positive. Patients also received ceftriaxone (1 g per day IV) from day +1 until the first day of febrile neutropenia or until attaining an ANC of 1 × 10/L. Transfusions were administered when the haemoglobin concentration and platelet count were below 8 g/dL and 10× 10/L, respectively.

Deleterious Effect of Steroids on Cytomegalovirus Infection Rate after Allogeneic Stem Cell Transplantation Depends on Pretransplant Cytomegalovirus Serostatus of Donors and Recipients

This study examined the impact of prednisone (PDN) on cytomegalovirus (CMV) infection after allogeneic stem cell transplantation (allo-SCT) according to donor and recipient CMV serostatus. Seventy-five patients underwent allo-SCT from June 2010 to July 2012. The risk of CMV infection according to donor and recipient serostatus was defined as follows: high risk (HR; D-/R+), intermediate risk (IR; D+/R+ and D+/R-), and low risk (D-/R-). Forty-five patients (60%) developed CMV infection, and 46 patients (61%) received steroids (PDN ≥ 1 mg/kg/day) to treat acute graft-versus-host disease. CMV infection was more common in those treated with steroids than in those not treated with steroids (70% versus 44%, respectively, P < .05). Overall, 40% of patients had recurrent CMV infection (50% PDN versus 24% no PDN, P < .05). Steroids had no impact on the incidence of CMV infection or its recurrence in HR patients; however, steroids did prolong the need for antiviral treatment. The incidence of CMV infection in IR patients was higher in those receiving PDN (80% PDN versus 41% no PDN, P = .01); recurrence rates were also higher (55% PDN versus 18% no PDN, P = .02). We analyzed CMV-specific immune reconstitution in the first 22 patients of the series and observed that patients on steroids had lower levels of CMV-specific lymphocytes TCD8 (P < .05 on days +60, +100, and +180) and that CMV-specific immune reconstitution (defined as lymphocytes CD8/IFN ≥ 1 cell/µL) was achieved later (after day +100 post-SCT) in the steroid group.

Single Antigen–Mismatched Unrelated Hematopoietic Stem Cell Transplantation Using High-Dose Post-Transplantation Cyclophosphamide Is a Suitable Alternative for Patients Lacking HLA-Matched Donors

The optimal prophylaxis regimen for graft-versus-host disease (GVHD) in the setting of mismatched unrelated donor (MMUD) allogeneic hematopoietic stem cell transplantation (alloHSCT) is not defined. The use of high-dose post-transplant cyclophosphamide (PTCy) in haploidentical transplantation has proven feasible and effective in overcoming the negative impact of HLA disparity on survival. We hypothesized that PTCy could also be effective in the setting of MMUD transplantation. We retrospectively analyzed 86 consecutive adult recipients of alloHSCT in our institution, comparing 2 contemporaneous groups: PTCy MMUD (n = 26) versus matched unrelated donor (MUD) (n = 60). Graft source was primarily peripheral blood (92%). All PTCy MMUD were HLA 7/8 (differences in HLA class I loci in 92% of patients) and received PTCy plus tacrolimus ± mofetil mycophenolate as GVHD prophylaxis. No differences were observed between PTCy MMUD and MUD in the 100-day cumulative incidence of acute GVHD grades II to IV (31% versus 22%, respectively; P = .59) and III to IV (8% versus 10%, P = .67). There was a trend for a lower incidence of moderate to severe chronic GVHD at 1 year after PTCy MMUD in comparison with MUD (22% versus 41%, P = .098). No differences between PTCy MMUD and MUD were found regarding nonrelapse mortality (25% versus 18%, P = .52) or relapse rate (11% versus 19%, P = .18). Progression-free survival and overall survival at 2 years were similar in both cohorts (67% versus 54% [HR, .84; 95% CI, .38 to 1.88; P = .68] and 72% versus 57% [HR, .71; 95% CI, .31 to 1.67; P = .44], respectively). The 2-year cumulative incidence of survival free of moderate to severe chronic GVHD and relapse tended to be higher in the PTCy MMUD group (47% versus 24%; HR, .60; 95% CI, .31 to 1.14; P = .12). We conclude that HLA 7/8 MMUD transplantation using PTCy plus tacrolimus is a suitable alternative for those patients who lack a MUD.