Immunotherapy and Allogeneic Bone Marrow Transplantation in B Acute Lymphoblastic Leukemia: How to Sequence?

Long-term disease control is achieved in 80-90% of patients with acute lymphoblastic leukemia of B origin (B-ALL). About half of adult and 10% of pediatric patients develop refractory or relapsed disease, whereas survival after relapse accounts about 10% in adults and 30-50% in children. Allogeneic bone marrow transplantation offers remarkable benefit in cases with unfavorable outcome. Nevertheless, novel immunotherapeutic options have been approved for patients with adverse prognosis. Immunotherapeutic agents, nowadays, are preferred over standard chemotherapy for patients with relapsed or refractory B-ALL The mode of action, efficacy and safety data of immunotherapeutic agents released, indications and sequence of those therapies over the course of treatment, are herein reviewed.

Dose individualization of intravenous busulfan in pediatric patients undergoing bone marrow transplantation: impact and in vitro evaluation of infusion lag-time.

OBJECTIVES: To apply therapeutic drug monitoring and dose-individualization of intravenous Busulfan to paediatric patients and evaluate the impact of syringe-pump induced Busulfan infusion lag-time after in vitro estimation. METHODS: 76 children and adolescents were administered 2 h intravenous Busulfan infusion every 6 h (16 doses). Busulfan plasma levels, withdrawn by an optimized sampling scheme and measured by a validated HPLC-PDA method, were used to estimate basic PK parameters, AUC, Cmax, kel, t1/2, applying Non-Compartmental Analysis. In vivo infusion lag-time was simulated in vitro and used to evaluate its impact on AUC estimation. KEY FINDINGS: Mean (%CV) Busulfan AUC, Cmax, clearance and t1/2 for pediatric population were found 962.3 µm × min (33.1), 0.95 mg/L (41.4), 0.27 L/h/kg (33.3), 2.2 h (27.8), respectively. TDM applied to 76 children revealed 6 (7.9%) being above and 25 (32.9%) below therapeutic-range (AUC: 900-1350 µm × min). After dose correction, all patients were measured below toxic levels (AUC < 1500 µm × min), no patient below 900 µm × min. Incorporation of infusion lag-time revealed lower AUCs with 17.1% more patients and 23.1% more younger patients, with body weight <16 kg, being below the therapeutic-range. CONCLUSIONS: TDM, applied successfully to 76 children, confirmed the need for Busulfan dose-individualization in paediatric patients. Infusion lag-time was proved clinically significant for younger, low body-weight patients and those close to the lower therapeutic-range limit.

Infectious complications following allogeneic stem cell transplantation by using anti-thymocyte globulin-based myeloablative conditioning regimens in children with hemoglobinopathies.

BACKGROUND: Anti-thymocyte globulin (ATG) has been used to prevent graft failure/rejection in the setting of allogeneic stem cell transplantation (allo-SCT) for hemoglobinopathies; however, epidemiology data for transplant-related infections in this population are scarce. METHOD: We retrospectively analyzed the epidemiology of bacterial, fungal, viral, and parasitic infections in a cohort of 105 children and adolescents with ß-thalassemia (n = 100) or sickle cell disease (n = 5) who underwent allo-SCT using human leukocyte antigen (HLA)-identical sibling (n = 96) or HLA-compatible unrelated donors (n = 9) in a single institution. All patients received an ATG-based conditioning regimen. RESULTS: The cumulative incidence of cytomegalovirus (CMV) viremia was 45.7% (95% confidence interval [CI] 33-55%), developing at a median of 48 (range 12-142) days without evidence of overt CMV disease. Herpes zoster developed in 8 patients at a median of 12 months post transplant, while 10 patients presented with late onset hemorrhagic cystitis at a median of 35 days post transplant. The cumulative incidence of bacteremia was 17.1% (95% CI 10.6-25%), occurring at a median of 74 (range 24-110) days. No patient developed probable or definite invasive fungal infection. Four deaths were recorded; 2 of them were attributed to infections (toxoplasmosis and Pneumocystis jirovecii pneumonia, respectively). CONCLUSION: The rate of infections after allo-SCT, using an ATG-containing preparative regimen, in our population of pediatric patients with hemoglobinopathies is comparable to that reported elsewhere with the use of non-ATG containing regimens.

HLA-matched sibling stem cell transplantation in children with ß-thalassemia with anti-thymocyte globulin as part of the preparative regimen: the Greek experience.

BU combined with CY, the preferred preparatory regimen for thalassemic patients, is associated with a substantial incidence of graft rejection especially in patients with advanced disease stage. This study retrospectively analyzes the outcome of 75 consecutive pediatric patients with ß-thalassemia who underwent HLA-matched sibling transplantation after anti-thymocyte globulin (ATG)-containing myeloablative conditioning regimens. With a median follow-up of 9 years (range 1-15 years), the overall survival (OS) and thalassemia free survival (TFS) rates were 96% and 92%, respectively. Both the estimated TRM and the cumulative incidence of rejection/failure were 4%. The cumulative incidences of acute GVHD grade II-III and grade III were 20% and 5.3%, respectively. No patient developed acute GVHD grade IV. Only two patients developed extensive chronic GVHD. The estimated OS and TFS for patients with Class 1 and 2 disease according to Pesaro criteria were 96.3% and 94.4%, whereas for patients with Class 3 disease they were 94.1% and 88.2%, respectively. In our series, the use of myeloablative conditioning regimens, which include ATG for the transplantation of thalassemic children from matched sibling donors, resulted in excellent outcomes with very low incidences of TRM and rejection.

Changes in free radical generation and antioxidant capacity during ultramarathon foot race.

BACKGROUND: Exhaustive exercise has been implicated in the generation of reactive oxygen species, resulting in oxidative stress. We studied the effect of a long-distance, endurance exercise on oxidative stress parameters in athletes who participated in the ultramarathon race Spartathlon (246 km). MATERIALS AND METHODS: This study included 18 runners (16 men and 2 women) aged 42.8 +/- 1.4 years. Blood samples were obtained 24 h before (prerace), at the end (postrace) and 48 h after the end of the race (48 h postrace). We measured oxidative stress indices, including red cell glutathione, malonyldialdehyde and 8-iso-prostaglandin F(2a), as well as the total antioxidant capacity. RESULTS: 8-Iso-prostaglandin F(2a) level increased significantly at the end of the race, compared to prerace levels (up to 914.7 +/- 61.4 pg mL(-1) from 197.6 +/- 8.4 pg mL(-1)), and remained 2.5-fold increased over the baseline 48 h after the race (532.0 +/- 54.2 pg mL(-1), P < 0.000). The total antioxidant capacity of the athletes increased from a baseline of 289.6 +/- 9.0 micromol L(-1) to 358.7 +/- 11.0 micromol L(-1) immediately after the race and remained elevated 48 h later (350.6 +/- 7.6 micromol L(-1)) (P < 0.001). CONCLUSIONS: Prolonged exercise induces a marked response of oxidative stress biomarkers, which in part is compensated by serum ability to scavenge free radicals. Whether these changes have long-term negative effects in the organism needs further investigation.

Dramatic reduction of erythrocyte glucose-6-phosphate dehydrogenase activity in athletes participating in the ultradistance foot race "Spartathlon".

OBJECTIVE: To investigate the effect of a long-distance endurance exercise "Spartathlon" on erythrocyte glucose-6-phosphate dehydrogenase (G(6)PD) activity. MATERIAL AND METHODS: The study comprised 15 male runners, median age 36.5 years. Blood samples were obtained in the 15 min before the race and again within 15 min after the end of the race. Erythrocyte glutathione (GSH and GSSG) and plasma malonyldialdehyde were measured with HPLC methods, and total antioxidant capacity (TAC), total hyperoxides and G(6)PD activity with commercial kits. Lipids, uric acid and total bilirubin were determined with a clinical chemistry analyser. RESULTS: Total hyperoxides were found statistically reduced, whereas total bilirubin was measured elevated post-race. Interestingly, GSSG levels were found increased (167.3+/-12.0 versus 219.5+/-20.3 micromol/L; p<0.005) as well as GSSG/GSH ratio (16.0+/-1.3 versus 20.60+/-1.65; p<0.05) post-race. In contrast, G(6)PD activity was found remarkably decreased (8.72+/-3.10 versus 3.8+/-2.5 U/g Hb; p<0.0001) pre versus post the event. CONCLUSION: Red blood cell G(6)PD activity in athletes may be reduced post-race as a consequence of the modulation of NADP/NADPH levels and elevation of the erythrocyte GSSG, and especially GSSG/GSH ratio, resulting in an impairment of the hexose monophosphate shunt.

Gonadal function of young patients with beta-thalassemia following bone marrow transplantation.

Bone marrow transplantation (BMT) can induce short- and long-term impairment of gonadal function. Patients with beta-thalassemia represent a special group, as their primary diagnosis and its treatment modalities are responsible for gonadal dysfunction. To address the effect of BMT on puberty and gonadal function, we investigated 25 patients (12 males) with thalassemia who received allogenic BMT during childhood or adolescence and at the post-transplant evaluation were at an age that the pubertal process should have started. Pubertal stage by Tanner of breast and pubic hair, as well as testicular volume were assessed pre-BMT once and post-BMT at least twice. Menstrual history was recorded. FSH, LH, testosterone and estradiol levels were also determined. The impact of BMT appears to be different in the two sexes. Males seem to have higher tolerance, as all males who were pubertal at the time of BMT had normal testosterone, and all but one normal gonadotropin levels. From those who were prepubertal at BMT, 62% proceeded to normal pubertal development. Post-menarcheal females seem to be an extremely sensitive group to the deleterious effect of the transplantation process, as 100% of the post-menarcheal females exhibited amenorrhea and elevated gonadotropin levels. These findings are important for pre- and post-BMT counseling.

Kinetics of quiescent cord blood stem/progenitor cells with high proliferative potential in stem-cell expansion culture.

BACKGROUND: The most primitive engrafting hematopoietic stem cell (HSC) resides mainly in a tumor growth factor-beta (TGF-beta)-dependent quiescent phase of the cell cycle. In this study, ex vivo expansion of UC blood (UCB) HSCs has been investigated, with the aim of showing whether quiescent HSCs can be recovered from expansion culture. METHODS: AC133(+) stem/progenitor cells from six full term-pregnancies UCB-samples were immunomagnetically selected, followed by ex vivo expansion culture in the presence of thrombopoietin (TPO), c-kit ligand (KL), flt-3 ligand (FL) and IL-6. Quiescent HSCs were detected by a clonogenic assay that allows the detection of multipotent and committed single- lineage quiescent stem/progenitor cells, named mHPP-Q and cHPP-Q, respectively, by means of a TGF-beta blocking Ab. RESULTS: Expansion culture of fresh selected AC133(+) cells for 1 week caused maintenance rather than expansion of mHPP-Q cells and a 1-fold increase in cHPP-Q cells. A further week culture initiated with 7-day expanded AC133(+) cells resulted in an additional 1.5-fold expansion of cHPP-Q while no mHPP-Q cells could be detected. Amplification of cHPP-Q cells in long-term expansion cultures initiated with 14-day expanded AC133(+) cells was observed for at least a further 4 weeks. DISCUSSION: A small proportion of HPP-Q cells recovered from 7-day expansion cultures retain their multilineage potential: longer culturing of these cells results in the loss of multilineage potential while they maintain quiescent behavior and high proliferative potential.

The oral manifestations of chronic graft-versus-host disease (cGVHD) in paediatric allogeneic bone marrow transplant recipients.

The oral manifestations of chronic graft-versus-host disease (cGVHD) in eight allogeneic bone marrow transplant (BMT) paediatric recipients were studied clinically, and lip biopsies were performed in seven of them. A prominent lichenoid reaction was observed in four patients, two with accompanying ulceration. Superficial mucoceles were present in three children. Clinically obvious xerostomia was seen in seven patients. Lip biopsies were positive and correlated with the clinical manifestations. Both clinical and histological findings confirmed the diagnosis of cGVHD. In three additional children, with systemic manifestations indicating cGVHD, the oral mucosa was clinically and histologically normal, and the systemic manifestations were, thus, attributed to drug reactions. The above findings indicate the high value of oral examination in diagnosing or confirming paediatric cGVHD. Superficial mucoceles, reported for the first time in paediatric recipients, seem to be important in the early diagnosis of cGVHD.

Combined umbilical cord blood and bone marrow transplantation in the treatment of beta-thalassemia major.

The authors report on three children with beta-thalassemia major, class II, III, and III according to the Pesaro classification, with a body weight of 16, 62, and 50 kg, respectively, who received grafts using both umbilical cord blood (UCB) and bone marrow (BM) stem cells from their HLA-matched siblings. The number of UCB nucleated cells collected was 2 x 10(7)/kg, 1.2 x 10(7)/kg, and 2.5 x 10(7)/kg, respectively, and was considered insufficient to secure engraftment. The authors increased the number of hematopoetic progenitors by harvesting BM from the same donors. All 3 patients showed prompt engraftment with neutrophil recovery on days 17, 18, and 17 post-transplant, respectively, and platelet recovery on days 19, 25, and 22 post-transplant, respectively. One patient had remarkably increased HbF of values 31, 19, and 12% at 3, 6, and 12 months post-transplant, respectively, which were accompanied by an increase in the G gamma/A gamma ratio, suggesting UCB-derived hematopoetic reconstitution. All patients are alive and transfusion independent 23, 18, and 16 months post-transplant, respectively. For patients with homozygous beta-thalassemia who are at high risk of graft failure, either because of major prior alloimmunization or an insufficient amount of UCB stem cells, combined transplantation with UCB and BM could offer a quick and safe alternative therapy.

A functional hierarchy among the CD34+ hematopoietic cells based on in vitro proliferative and differentiative potential of AC133+CD34(bright) and AC133(dim/)-CD34+ human cord blood cells.

The 5-transmembrane receptor AC133 is expressed on a subpopulation of human hematopoietic cells that includes the CD34(bright) cells. We evaluated the developmental potential of AC133+CD34(bright) and AC133(dim/-)CD34+ cells isolated from 5 cord blood (CB) samples by studying the in vitro proliferative and differentiative potential of each population in both progenitor and mature cell expansion cultures. Seven-day culture of AC133+CD34(bright) cells with a cytokine combination favoring primitive progenitor cells causes a significant increase in CD34+, CFU-C and noncycling stem/progenitor cells HPP-Q (High Proliferative Potential-Quiescent), whereas culture of AC133(dim/-)CD34+ cells shows a limited increase in committed progenitor cells only. HPP-Q cells were not found in freshly isolated AC133(dim/-)CD34+ nor in expanded CD34+ cells derived from AC133(dim/-)CD34+ cells. No statistically significant difference was observed between the 1-week expanded AC133+ and the initial AC133+CD34(bright) cells regarding their clonogenic efficiency (CE), while expanded CD34+ cells derived from AC133(dim/-)CD34+ cells exhibited a decreased CE. Subexpansion of the reselected AC133+ derived from AC133+CD34(bright) cells reveals a further increase of stem/progenitor cells and the 14-day expanded AC133+ cells reveal an unchanged CE. Subexpansion of reselected 7-day CD34+ cells derived from AC133(dim/-)CD34+ cells was not possible. Culture of AC133+CD34(bright) cells in cytokines that favor megakaryopoiesis or erythropoiesis resulted in a significant expansion of CD41+ and CD71+ cells, respectively; AC133(dim/-)CD34+, in comparison, showed a limited potential to megakaryocytic differentiation and a decreased production of erythroid cells. Our data indicate that early high proliferating stem/progenitor cells and early committed progenitors are present in AC133+CD34(bright) cells, but not in AC133(dim/-)CD34+ cells; the latter represent late committed progenitors with limited proliferative potential.