[Efficacy and prognostic factors of allogeneic hematopoietic stem cell transplantation in the treatment of secondary acute myeloid leukemia].

Objective: To evaluate the efficacy and prognostic factors of allogeneic hematopoietic stem cell transplantation (allo-HSCT) in patients with secondary acute myeloid leukemia (sAML) . Methods: In this multicenter, retrospective clinical study, adult patients aged ≥18 years who underwent allo-HSCT for sAML at four centers of the Zhejiang Hematopoietic Stem Cell Transplantation Collaborative Group from January 2014 to November 2022 were included, and the efficacy and prognostic factors of allo-HSCT were analyzed. Results: A total of 95 patients were enrolled; 66 (69.5%) had myelodysplastic syndrome-acute myeloid leukemia (MDS-AML) , 4 (4.2%) had MDS/MPN-AML, and 25 (26.3%) had therapy-related AML (tAML) . The 3-year CIR, LFS, and overall survival (OS) rates were 18.6% (95% CI 10.2%-27.0%) , 70.6% (95% CI 60.8%-80.4%) , and 73.3% (95% CI 63.9%-82.7%) , respectively. The 3-year CIRs of the M-AML group (including MDS-AML and MDS/MPN-AML) and the tAML group were 20.0% and 16.4%, respectively (P=0.430) . The 3-year LFSs were 68.3% and 75.4%, respectively (P=0.176) . The 3-year OS rates were 69.7% and 75.4%, respectively (P=0.233) . The 3-year CIRs of the groups with and without TP53 mutations were 60.0% and 13.7%, respectively (P=0.003) ; the 3-year LFSs were 20.0% and 76.5%, respectively (P=0.002) ; and the 3-year OS rates were 40.0% and 77.6%, respectively (P=0.002) . According to European LeukmiaNet 2022 (ELN2022) risk stratification, the 3-year CIRs of patients in the low-, intermediate-, and high-risk groups were 8.3%, 17.8%, and 22.6%, respectively (P=0.639) . The three-year LFSs were 91.7%, 69.5%, and 65.6%, respectively (P=0.268) . The 3-year OS rates were 91.7%, 71.4%, and 70.1%, respectively (P=0.314) . Multivariate analysis revealed that advanced disease at allo-HSCT and TP53 mutations were independent risk factors for CIR, LFS, and OS. Conclusion: There was no significant difference in the prognosis of patients who underwent allo-HSCT among the MDS-AML, MDS/MPN-AML, and tAML groups. Advanced disease at transplantation and TP53 mutations were poor prognostic factors. ELN2022 risk stratification had limited value for predicting the prognosis of patients with sAML following allo-HSCT.

[The impact of donor-to-recipient gender compatibility on outcomes of haploid hematopoietic stem cell transplantation in patients with hematological malignancies].

Objective: To investigate how gender differences between the donor and the recipient affect the effectiveness of antithymocyte globulin (ATG) and pure peripheral blood stem cell (PBSC) hematopoietic stem cell transplantation (haplo-HSCT) in the treatment of malignant hematological diseases. Methods: From February 2015 to September 2020, 648 hematological malignancies patients underwent myeloablative condition regimen haplo-HSCT treatment at the Bone Marrow Transplant Center of the First Affiliated Hospital of Zhejiang University. The median age was 32 (14-62) years, with 363 males (56.0% ) and 285 females (44.0% ) present. 242 cases of acute lymphoblastic leukemia (ALL) (37.3% ) , 293 cases of acute myeloid leukemia (AML) (45.2% ) , 56 cases of myelodysplastic syndrome (MDS) (8.7% ) , 27 cases of non-Hodgkin's lymphoma (NHL) (4.2% ) , and 30 cases of other hematological malignancies (4.6% ) . Results: ① The 3-year overall survival (OS) , DFS, the incidence of Ⅱ-Ⅳ grade acute graft-versus-host disease (aGVHD) , the incidence of Ⅲ-Ⅳ grade aGVHD, the 3-year incidence of moderate & severe chronic GVHD (cGVHD) , severe cGVHD, the 3-year incidence of relapse, and NRM of the whole group were (73.10±1.90) % , (70.80±1.90) % , (33.96±1.87) % , (13.08±1.33) % , (35.10±2.14) % , (10.66±1.38) % , (19.43±1.67) % , and (9.80±1.24) % , respectively. ②There was no statistically significant difference between the donor-recipient gender match and donor-recipient gender mismatch groups in the 28-day cumulative neutrophil engraftment rate, 28-day cumulative platelet engraftment rate, the incidence of Ⅱ-Ⅳ grade aGVHD, the incidence of Ⅲ-Ⅳ grade aGVHD, 3-year OS, 3-year DFS, the cumulative incidence of relapse, NRM, and incidence of moderate & severe cGVHD, severe cGVHD. ③The 28-day cumulative neutrophil engraftment rate did not differ statistically between the male-female, female-female, male-male, and female-male groups (P=0.148) . The incidence of Ⅱ-Ⅳ grade aGVHD, the incidence of Ⅲ-Ⅳ grade aGVHD, 3-year OS, 3-year DFS, cumulative relapse rate, and NRM, and the incidence of cGVHD were not statistically different among the four groups (P>0.05) . The 28-day cumulative platelet engraftment rate of the female-male group was significantly lower than male-female group, and the female-female group [ (91.45±2.63) % vs. (94.77±1.75) % , P=0.004; (91.45±2.63) % vs. (95.54±2.05) % , P=0.005]. No significant difference existed in the 28-day cumulative platelet engraftment rate between the female-male group and the male-male group [ (91.45±2.63) % vs. (95.08±1.41) % , P=0.284]. ④Among patients ≤35 years old, the 3-year incidence of severe cGVHD patients receiving sister donors and sibling donors were (26.71±5.90) % and (10.33±4.43) % , respectively (P=0.054) . Patients accepting daughter donors and son donors had a 3-year incidence of moderate and severe cGVHD that was 40.07% vs. 27.41% , respectively, among those over 35 (40.07±6.65) % vs. (27.41±4.54) % (P=0.084) . ⑤Female donors to male recipients had a significantly lower 28-day cumulative platelet engraftment rate compared to the other groups [ (91.45±2.63) % vs. (95.08±0.95) % , P=0.037]. ⑥ Female donors to male recipients had a significantly lower 28-day cumulative platelet engraftment rate than the other groups in the ATG-Fresenius (ATG-F) 10 mg/kg group [ (89.29±4.29) % vs. (94.49±1.45) % , P=0.037]. But when compared to the other groups in the Rabbit Antihuman Thymocyte Immunoglobulin (rATG-T) 6 mg/kg group, the 28-day cumulative platelet implantation rate between female donors and male recipients was not significantly different [ (93.44±3.38) % vs. (95.62±1.26) % , P=0.404]. Conclusion: The main clinical outcomes of patients with malignant blood diseases following transplantation are unaffected by the gender combination of the donor and patient in the haplo-HSCT mode based on ATG and PBSC sources. Female donors to male recipients have a lower 28-day cumulative platelet engraftment rate and longer platelet engraftment times.

Carfilzomib combined with ex vivo-expanded patient autologous natural killer cells for myeloma immunotherapy.

Natural killer (NK) cell-based immunotherapy is promising, as NK cells are in the first line of defense against cancer and capital of lysing tumor cells without pre-stimulation. However, NK cells from multiple myeloma (MM) patients are always deficient in numbers and the expression of certain activating receptors, disabling them in cytotoxicity against the cancer. Therefore, effective strategies to expand NK cells and increase NK cell-mediated cytotoxicity against MM are significant. Here, NK cells were efficiently expanded from peripheral blood mononuclear cells (PBMCs) of newly diagnosed MM patients after co-culture with irradiated K562 cells transfected with 41BBL and membrane-bound interleukin (IL)-15 (K562-mb15-41BBL) in the presence of 200 IU/ml human IL-2. The ex vivo-expanded NK cells were demonstrated to vigorously kill both MM cells and autologous primary MM cells without significant lysis of patient normal cells. Further exploration revealed a significant increase in cell surface expression of most activating receptors of NK cells and indicated that expanded NK (exp-NK) cell killing of MM cells was mediated by perforin/granzyme. NK cells are capital of lysing human leukocyte antigen (HLA) I-deficient tumor cells and carfizomib, a selective proteasome inhibitor approved for the treatment of relapsed/refractory MM patient, down-regulates the expression of HLA class I, thus enhancing NK cell-mediated lysis in MM. Here, we found for the first time that carfizomib dramatically augmented ex vivo exp-NK cell cytotoxicity against patient autologous MM cells, suggesting the use of exp-NK alone or in combination with the drug to treat MM patient.

[The effect of WT1 expression on the prognosis of allogeneic hematopoietic stem cell transplantation in acute leukemia].

Objective: To study the effect of WT1 expression on the prognosis of allogeneic hematopoietic stem cell transplantation (allo-HSCT) in acute leukemia (AL) and its significance as molecular marker to dynamically monitor minimal residual disease (MRD) . Methods: Retrospectively analyzed those AL patients who underwent allo-HSCT in the First Hospital Affiliated to Zhejiang University School of Medicine during Jan 2016 to Dec 2017, a total number of 314 cases, 163 males and 151 females, median age was 30 (9-64) years old. Comparing the difference of WT1 expression at diagnosed, pre-HSCT and after HSCT. Using the receiver operating characteristic (ROC) curve to determine the WT1 threshold at different time so as to predict relapse. The threshold of WT1 expression before transplantation was 1.010%, within 3 months after HSCT was 0.079% and 6 months after HSCT was 0.375%. According to these thresholds, WT1 positive patients were divided into low expression groups and high expression groups. Analyzed the relationship between overall survival (OS) , disease-free survival (DFS) , cumulative incidence of relapse (CIR) and WT1 expression. Results: The OS and DFS of high expression group pre-HSCT were lower than low expression group [69.2% (9/13) vs 89.1% (57/64) , χ(2)=4.086, P=0.043; 53.8% (7/13) vs 87.5% (56/64) , χ(2)=9.766, P=0.002], CIR was higher than low expression group [30.8% (4/13) vs 7.8% (5/64) , P=0.017]. There was no significant difference of OS and DFS between high expression and low expression group of 3 months after HSCT (P=0.558, P=0.269) . The OS and DFS of high expression group of 6 months after transplantation were both lower than low expression group (P=0.049, P=0.035) . Multivariate analysis showed that WT1>0.375% when 6 months after transplantation was the only independent prognostic factor for shorter DFS (P=0.022) . There was no statistically significant difference in CIR between the high-expression group and the low-expression group 3 months after transplantation and 6 months after transplantation (P=0.114, P=0.306) . Conclusion: High expression of WT1 before and after HSCT was an adverse prognosis factor. It is of clinical practical value to use WT1 as a transplant recommendation index for patients with acute leukemia and as a marker to monitor MRD dynamically.

Effects of acrylonitrile on lymphocyte lipid rafts and RAS/RAF/MAPK/ERK signaling pathways.

Acrylonitrile (ACN) is a widely used chemical in the production of plastics, resins, nitriles, acrylic fibers, and synthetic rubber. Previous epidemiological investigations and animal studies have confirmed that ACN affects the lymphocytes and spleen. However, the immune toxicity mechanism is unknown. Lipid rafts are cell membrane structures that are rich in cholesterol and involved in cell signal transduction. The B cell lymophoma-10 (Bcl10) protein is a joint protein that is important in lymphocyte development and signal pathways. This study was conducted to examine the in vitro effects of ACN. We separated lipid rafts, and analyzed Bcl10 protein and caveolin. Western blotting was used to detect mitogen-activated protein kinase (MAPK) and phosphorylated MAPK levels. The results indicated that with increasing ACN concentration, the total amount of Bcl10 remained stable, but was concentrated mainly in part 4 to part 11 in electrophoretic band district which is high density in gradient centrifugation. Caveolin-1 was evaluated as a lipid raft marker protein; caveolin-1 content and position were relatively unchanged. Western blotting showed that in a certain range, MAPK protein was secreted at a higher level. At some ACN exposure levels, MAPK protein secretion was significantly decreased compared to the control group (P < 0.05). These results indicate that ACN can cause immune toxicity by damaging lipid raft structures, causing Bcl10 protein and lipid raft separation and restraining Ras-Raf-MAPK-extracellular signal-regulated kinase signaling pathways.

Donor TLR9 gene tagSNPs influence susceptibility to aGVHD and CMV reactivation in the allo-HSCT setting without polymorphisms in the TLR4 and NOD2 genes.

Owing to ethnicity of the population, those best confirmed polymorphisms in the TLR (toll-like receptor)4 and NOD2 genes with significantly prognostic impact on allogeneic hematopoietic SCT (allo-HSCT) seem to be more applicable to Europeans and are nonpolymorphic in the Asian population. The influence of innate immunity gene polymorphisms on the outcomes of allo-HSCT in those populations has been questioned. We evaluated the influence of 10 candidate single nucleotide polymorphisms (SNPs) in the TLR1, TLR2, TLR3, TLR8 and TLR9 genes on the outcomes of allo-HSCT in a Chinese population including 138 pairs of patients and unrelated donors and a second cohort of 102 pairs of patients and HLA-identical sibling donors. We found that two tagSNPs in the TLR9 gene in the donor side, +1174 A/G (rs352139) and +1635 C/T (rs352140), influenced the risk of acute GVHD (aGVHD) and CMV reactivation. Furthermore, the presence of the susceptible haplotype (A-C) in donor may be an informative predicator of worse OS at 5 years compared with those with the G-C and G-T haplotypes (58% vs 82.9%, P=0.024). Our data suggested an unrecognized association between donor TLR9 tagSNPs and the risk of HSCT-related complications in a population without polymorphisms in the TLR4 and NOD2 genes.

Expression of multidrug resistance 1 and multidrug resistance-related protein 1 in C57BL/6 mice treated with benzene.

ATP-binding cassette super family (ABC) proteins are considered key to oncology and pharmacology studies. We examined the effect of benzene on ABC pump protein levels in C57BL/6 mouse bone marrow mononuclear cells. After a 2-week gavage (200 mg/kg, 5 days per week), the number of peripheral leukocytes, lymphocytes and basophils dropped significantly; there was also a significant decrease in MDR1 and MRP1 gene expression. A significant reduction in expression of P-gp was found; however, there was no significant decrease in the expression of MRP1 and NF-κB p65. We conclude that regulation of membrane efflux transport protein could be a factor in benzene hematotoxicity.

Haploidentical allogeneic haematopoietic stem cell transplantation as salvage therapy for engraftment failure after unrelated and autologous stem cell transplantation: a case report and review of the literature.

Engraftment failure is a rare but life-threatening complication of haematopoietic stem cell transplantation (HSCT) and treatment of this condition is often challenging. This case report describes a patient with acute myeloid leukaemia and engraftment failure after unrelated donor allogeneic stem cell transplantation. Rescue treatment with granulocyte-colony stimulating factor and reinfusion of autologous 'back-up' stem cells failed, but transplantation of haploidentical donor stem cells following a fludarabine and antithymocyte globulin (ATG)-based conditioning regimen resulted in haematological reconstitution and long-term disease-free survival. The use of haploidentical donor stem cell transplantation as salvage therapy after engraftment failure in adult patients has not, to the authors' knowledge, been previously reported. Additionally, a review of the relevant literature is presented. This case report and literature review suggest that reinfusion of cryopreserved 'back-up' haematopoietic stem cells is a safe and effective salvage therapy for engraftment failure after allogeneic HSCT. Haploidentical donor stem cell transplantation after a fludarabine and ATG-based conditioning regimen could provide effective second-line therapy in adult patients.

Relationship between TNFA, TNFB and TNFRII gene polymorphisms and outcome after unrelated hematopoietic cell transplantation in a Chinese population.

This study aimed to analyze the association between cytokine gene polymorphisms and outcome following allogeneic hematopoietic SCT (allo-HSCT). A total of 138 unrelated donor/recipient pairs who underwent allo-HSCT from 2001 to 2009 were tested for TNFA-1031 (T>C), -863 (C>A), -857 (C>T), -238 (G>A), TNFB+252 (A>G) and TNFRII codon 196 (T>G) single nucleotide polymorphisms by multiplex SnaPshot analysis. Transplantation involving recipients and/or donors with TNFA-857 C/C genotype or TNFB+252 G allele-positivity resulted in a higher incidence of acute GVHD (aGVHD), which was independent of HLA mismatching. In multivariate analysis, TNFA-857 C/C genotype donors (relative risk (RR)=2.29, P=0.006) and TNFB+252 G allele-positive recipients (RR=1.789, P=0.036) were found to be significantly associated with an increased incidence of aGVHD. TNFA-857 C/C genotype donors (RR=3.748, P=0.002) and TNFB+252 G allele-positive recipients (RR=1.823, P=0.063) were also associated with the development of grades II-IV aGVHD. TNFRII polymorphism in recipients was also related to relapse rate, but no significant associations were found between TNFA, TNFB or TNFRII 196 genotype and cGVHD, relapse or overall survival after transplantation. These results provide the first report of an association between TNFA, TNFB and TNFRII polymorphic features and outcome of allo-HSCT in a Chinese population, and suggest an interaction between TNFA-857 and TNFB+252 genotypes and risk of aGVHD.

The beneficial impact of missing KIR ligands and absence of donor KIR2DS3 gene on outcome following unrelated hematopoietic SCT for myeloid leukemia in the Chinese population.

The effect of natural killer (NK) cell alloreactivity on the outcome of unrelated hematopoietic SCT (HSCT) remains a topic of debate. NK cell alloreactivity after allogeneic HSCT is regulated by killer-cell Ig-like receptors (KIRs). To investigate the influence of KIRs on outcome after unrelated HSCT, we retrospectively analyzed the HLA and KIR genotypes of 116 donor-recipient pairs. We found that missing KIR ligands in recipients were significantly associated with a decreased leukemic relapse risk (P=0.019, HR=0.329), mainly in myeloid disease (P=0.003, HR=0.193). This beneficial effect was seen in AML/myelodysplastic syndrome and also in chronic myeloid leukemia. In myeloid disease, missing KIR ligands also improved 5-year OS (P=0.034, HR=0.430) and disease-free survival (DFS) (P=0.024, HR=0.445). Meanwhile, the presence of donor-activating KIR2DS3 gene was associated with increased relapse risk (P=0.003, HR=5.046), decreased OS (P=0.004, HR=3.181) and DFS (P=0.003, HR=2.919) in myeloid disease. No effect was seen in patients with lymphoid disease. Our study indicated that, in unrelated HSCT for myeloid leukemia, missing KIR ligands in recipients offered a lower relapse risk and a long-term survival advantage. The presence of KIR2DS3 in the donor was an important risk factor for myeloid leukemia.

A randomized trial of buprenorphine maintenance for heroin dependence in a primary care clinic for substance users versus a methadone clinic.

PURPOSE: Buprenorphine is an alternative to methadone for the maintenance treatment of heroine dependence and may be effective on a thrice weekly basis. Our objective was to evaluate the effect of thrice weekly buprenorphine maintenance for the treatment of heroin dependence in a primary care clinic on retention in treatment and illicit opioid use. SUBJECTS AND METHODS: Opioid-dependent patients were randomly assigned to receive thrice weekly buprenorphine maintenance in a primary care clinic that was affiliated with a drug treatment program (n = 23) or in a traditional drug treatment program (n = 23) in a 12-week clinical trial. Primary outcomes were retention in treatment and urine toxicology for opioids; secondary outcomes were opioid withdrawal symptoms and toxicology for cocaine. RESULTS: Retention during the 12-week study was higher in the primary care setting (78%, 18 of 23) than in the drug treatment setting (52%, 12 of 23; P = 0.06). Patients admitted to primary care had lower rates of opioid use based on overall urine toxicology (63% versus 85%, P < 0.01) and were more likely to achieve 3 or more consecutive weeks of abstinence (43% versus 13%, P = 0.02). Cocaine use was similar in both settings. CONCLUSIONS: Buprenorphine maintenance is an effective treatment for heroin dependence in a primary care setting.

Three methods of opioid detoxification in a primary care setting. A randomized trial.

BACKGROUND: Opioid detoxification in a primary care setting followed by ongoing substance abuse treatment may be appropriate for selected opioid-dependent patients. OBJECTIVE: To compare three pharmacologic protocols for opioid detoxification in a primary care setting. DESIGN: Randomized, double-blind clinical trial with random assignment to treatment protocols. SETTING: A free-standing primary care clinic affiliated with drug treatment programs. PATIENTS: 162 heroin-dependent patients. INTERVENTIONS: Three detoxification protocols: donidine, combined donidine and naltrexone, and buprenorphine. MEASUREMENTS: Successful detoxification (that is, when study participants received a full opioid-blocking dose [50 mg] of naltrexone), treatment retention (8 days), and withdrawal symptoms. RESULTS: Overall, 65% of participants (36 of 55) who received clonidine, 81% (44 of 54) who received combined clonidine and naltrexone, and 81% (43 of 53) who received buprenorphine were successfully detoxified. Retention did not differ significantly across the groups: 65% of participants (36 of 55) who received clonidine, 54% (29 of 54) who received combined clonidine and naltrexone, and 60% (32 of 53) who received buprenorphine. Participants who received buprenorphine had a significantly lower mean withdrawal symptom score than those who received clonidine or combined clonidine and naltrexone. CONCLUSIONS: Participants in the combined clonidine and naltrexone group and those in the buprenorphine group were more likely to complete detoxification, although retention at 8 days did not differ among the groups. Participants who were assigned to the buprenorphine group experienced less severe withdrawal symptoms than those assigned to the other two groups.

A pilot study of primary-care-based buprenorphine maintenance for heroin dependence.

The treatment of heroin dependence with opioid maintenance has traditionally employed methadone and more recently buprenorphine administered in traditional drug treatment settings. In this pilot study we evaluated buprenorphine maintenance for the treatment of heroin dependence in a program administered by primary-care providers in a primary-care setting. Seven patients were admitted to this nonblinded open-label pilot study and were offered 6 months of primary-care-based buprenorphine maintenance. Buprenorphine was administered in doses of 16 mg on Monday and Wednesday and 32 mg on Friday. Patients were seen weekly by primary-care providers and attended self-help meetings. Of the seven patients admitted to the study, five (71%) completed the 6-month pilot study and two (29%) were removed from the study. Urine toxicology data showed that the majority of urines tested were clear of opioids in four out of five patients who remained in treatment. These results suggest that primary-care-based opioid maintenance using buprenorphine shows promise as a new approach to the treatment of heroin dependence.