Haploidentical hematopoietic stem cell transplantation in the treatment of acquired aplastic anemia in children: a single center retrospective analysis / 中华儿科杂志

Objective@#To investigate the safety and efficacy of haploidentical hematopoietic stem cell transplantation with different intensity conditioning regimen in the treatment of childhood aplastic anemia (AA) .@*Methods@#Thirty-seven AA patients who underwent haploidentical transplantation in BaYi Children's Hospital Affiliated to PLA Army General Hospital from January 2013 to January 2017 were enrolled. According to the dosage of conditioning regimen, 34 patients excluding 3 other conditioning regimens were divided into high-dosage group (regimen 2, 22 cases) and low-dosage group (regimen 3, 12 cases). The data of Engraftment, graft-vs-host disease (GVHD), hematopoietic reconstitution, relapse, infection, overall survival (OS) were analyzed. The comparison between the two groups was tested by χ2 test.@*Results@#A total of 35 of 37 patients achieved primary engraftment; 2 cases died of regimen-related toxicity and severe infection before the infusing of the grafts. The activation rate of CMV and EBV was 60% (21/35) . Post-transplant lymphocyte disease (PTLD) of lung occurred in one case. The cumulative incidences of acute GVHD grade Ⅰ-Ⅳ and chronic GVHD were 29% (10/35) and 34% (12/35) respectively and the incidence of extensive chronic GVHD was 6% (2/35) . The median follow-up time was 18.8 (2.9-44.1) months, the OS was 92% (34/37) .All survived patients were no longer dependent on blood transfusion and none of them had recurrence. Comparing the rates of overall survival(86%(19/22) vs.100%(12/12)) and rates of chronic GVHD(40%(8/20) vs. 17%(2/12)) in regimen 2 and regimen 3 group, there were no significant difference (χ2=1.742, 1.841, all P>0.05) . Significant difference was found at the incidence of Ⅰ-Ⅳ acute GVHD (10% (2/20) vs. 50% (6/12) ,χ2=6.200, P=0.013).@*Conclusions@#Haploidentical hematopoietic stem cell transplantation is effective and safe. It is suitable for patients who are not eligible for matched donor transplantation. Application of reduced dose preconditioning in haploid transplantation is worth exploring.

Improved outcome of haploidentical transplantation in severe aplastic anemia using reduced-intensity fludarabine-based conditioning.

Significant improvements in hematopoietic stem cell transplantation (HSCT) with haploidentical family donors (HFD) have confirmed its therapeutic role in severe aplastic anemia (SAA) and led to the evolution of treatment algorithms. However, the optimal conditioning regimen for HFD-HSCT remains undefined, especially the dosage of cyclophosphamide (Cy). A total of 77 patients with SAA from two research centers, who received HFD-HSCT with reduced-intensity fludarabine + cyclophosphamide + thymoglobulin ± busulfan conditioning regimen plus third-party cells infusion were included in this study, of which 67 pairs had 4-5 loci mismatched. We were particularly interested in whether the dosage of Cy significantly impacted graft failure (GF) and overall survival (OS). All patients showed sustained hematopoietic engraftment without any increase in severe aGVHD and transplantation-related mortality (TRM). The incidences of grade II-IV aGVHD, grade III-IV aGVHD and extensive cGVHD were 18%, 10% and 7%, respectively. The probabilities of 1-year and 5-year OS were 93.1% and 87.9%, respectively. Furthermore, patient age <15 years, MNC cells >8×108/kg and donor age <45 years were associated with better survival (P=0.043, P=0.023, and P=0.037, respectively) and engraftment (P=0.019, P=0.008, and P=0.001, respectively). Our findings indicated that SAA patients lack MSD benefited the most if HFD-HSCT was performed with reduced-intensity fludarabine-based conditioning regimen. Improved outcomes with HFD-HSCT may lead to a salvaged therapy and an expanded direct role for SAA in the future.

Tumorablative conditioning regimen for haploidentical stem cell transplantation in 102 children with hematologic malignancies: a single-center experience.

Haploidentical hematopoietic stem cell transplantation (Haplo-HSCT) is widely carried out in China, and transplantation related complications decreased gradually with the transplant technology improving, and the overall survival(OS) increased year by year. However, relapse after transplantation is still one of the main causes of death in patients with hematological malignancy. In order to reduce the recurrence after HSCT, we set a tumorablative conditioning regimen (TAC ) regimen; the aim is as much as possible to eliminate the malignant clone to reduce the recurrence without increasing the conditioning toxicity. We retrospectively analyzed 102 cases of haplo-HSCT in our hospital from 2012 to 2017. Ninety-eight out of the 99 (99.0%) patients achieved primary engraftment. The 2-year OS and disease free survival (DFS) are 81.4% (83/102) and 77.45% (79/102). The cumulative incidence of leukemia relapse is 16.2% (16/99), Twenty-nine patients developed II-IV acute graft-versus-host disease (aGVHD) (29%) within 100 days and only nine patients have grade III-IV aGVHD (9%) in measurable 99 patients. The conditioning regimen was relatively well tolerated with limited regimen-related toxicity. The preliminary results show that TAC is safe and effective in haplo-HSCT of children with hematologic malignancies. This study will provide a clinical basis for the individualized conditioning regimen.

Construction and identification of lentiviral vectors carrying the connexin43 gene / 中国医师进修杂志

Objective To construct and identify lentiviral vectors carrying the connexin43 (CX43) gene. Methods Plasmids containing the CX43 gene and lentiviral vectors were digested using EcoRI/XbaI restriction enzymes, and the target gene fragments were cloned into the lentival vectors to result in CX43 recombinant lentiviral vectors (pHBLV-CMVIE-IRES-ZsGreen-CX43). CX43 recombinant lentiviral vectors were identified by restriction enzyme digestion and electrophoresis, and sequencing was carried on only for correct vectors after identification. The successfully-constructed CX43 recombinant lentiviral vectors and packaging plasmids were mixed and contransfected into 293FT cell for packaging and producting virus. Then the virus was collecting, concentrated and titrated in 293FT cells. Finally, the expression of CX43 gene was assessed by real-time polymerase chain reaction(PCR). Results The restriction enzyme digestion and electrophoresis and sequencing results proved that CX43 recombinant lentiviral vectors were constructed correctly. Lentiviral concentrated virus suspension titer was 1× 108/ml. The up-regulation expression of CX43 was detected correctly by real-time PCR in 293FT cells. Conclusions Stable lentiviral vectors expressing CX43 gene is constructed successfully.

Effects of adenovirus vector mediated Cx43 gene modification on GJIC function of acute leukemia bone marrow stromal cells / 第三军医大学学报

Objective To investigate the gene expression of connexin43(Cx43) and its effect on gap junction intercellular communication(GJIC) of acute leukemia bone marrow stromal cells(ALBMSCs).Methods After ALBMSCs were transfected by recombinant adenovirus Ad-Cx43-GFP,the expression of report gene GFP and the transfection efficiency were monitored by fluorescent microscopy.RT-PCR,Western blot and immunocytochemical method were used to detect the mRNA and protein expressions of Cx43.Dye transfer procedure was performed to examine the GJIC function.Results After transfected by Ad-Cx43-GFP for 24 h,the expression of GFP in ALBMSCs was detected by fluorescent microscopy and the transfection efficiency was(82.7?2.16)%;The mRNA and protein expressions of Cx43 in ALBMSCs transfected by Ad-Cx43-GFP were higher than those not transfected by Ad-Cx43-GFP(P