Discovery of an Imine Reductase for Reductive Amination of Carbonyl Compounds with Sterically Challenging Amines.

The synthesis of structurally diverse amines is of fundamental significance in the pharmaceutical industry due to the ubiquitous presence of amine motifs in biologically active molecules. Biocatalytic reductive amination for amine production has attracted great interest owing to its synthetic advantages. Herein, we report the direct synthesis of a wide range of sterically demanding secondary amines, including several important active pharmaceutical ingredients and pharmaceutical intermediates, via reductive amination of carbonyl substrates and bulky amine nucleophiles employing imine reductases. Key to success for this route is the identification of an imine reductase from Penicillium camemberti with unusual substrate specificity and its further engineering, which empowered the accommodation of a broad range of sterically demanding amine nucleophiles encompassing linear alkyl and (hetero)aromatic (oxy)alkyl substituents and the formation of final amine products with up to >99% conversion. The practical utility of the biocatalytic route has been demonstrated by its application in the preparative synthesis of the anti-hyperparathyroidism drug cinacalcet.

[The Role and Mechanism of Tfh and B Cell in Human Chronic Graft-Versus-Host-Disease].

AbstractObjective: To investigate the role and mechanism of circulating follicular helper T cells (cTfh), extrafollicular helper T cells, B cells and their subsets in chronic graft-versus-host disease (cGVHD) after transplantation. METHODS: Peripheral blood of cGVHD 64 patients after allogeneic hematopoietic stem cell transplantation (allo-HSCT) at the First Affiliated Hospital of Soochow University from 2016 to 2019 were collected. The percentage of cTfh cells, extrafollicular helper T cells, B cells and subsets were detected by flow cytometry. The healthy donors were detected as control. Percentage of each cell subpopulation between the two groups were compared by the two-tailed Students' T test. RESULTS: The percentage of circulating follicular helper T cells (cTfh, CD4+CXCR5+) was markedly decreased in patients with cGVHD as compared with that in the healthy donors (0.53%±0.10% vs 3.91%±0.60%, P<0.001). The percentage of extrafollicular helper T cells (CD44hiCD62LloPSGL-1loCD4+T) of the patients in cGVHD and the healthy donors were 8.86%±0.45% and 5.38%±0.79% (P=0.003). A significant change in B cell subsets was found in the patients with cGVHD. The two types of antigen-stimulate CD27+ B cell: the percentages of pre-GC B cells (CD19+IgD+CD38hiCD27+) and plasmablast/plasma cells (PB/PC, CD19+IgD-CD38hiCD27+) of patients with cGVHD were 20.91%±2.70% and 41.05%±5.00%, respectively, which were significantly higher than those in healthy donors (P=0.005, P=0.014). Meanwhile, the percentage of unstimulated CD27- B cells in patients with cGVHD was significantly reduced, especially the naive B cells (CD19+IgD+CD38loCD27-, 12.59%±2.63%, P=0.025). There was a positive correlation between the percentage of extrafollicular helper T cells and plasmablast/ plasma cells (PB/PC) in cGVHD patients (r=0.43). CONCLUSION: Compared with healthy donor, cTfh cells, extrafollicular helper T cells and B cell subsets in the peripheral blood of patients with cGVHD after transplantation changed in varying degress.

Muscleblind-like 1 antisense RNA 1 inhibits cell proliferation, invasion, and migration of prostate cancer by sponging miR-181a-5p and regulating PTEN/PI3K/AKT/mTOR signaling.

The present study aimed to investigate the role and underlying mechanisms of long non-coding RNA (lncRNA) muscleblind-like 1 antisense RNA 1 (MBNL1-AS1) in the progression of Prostate cancer (PCa). MBNL1-AS1 and microRNA (miR)-181a-5p expression in PCa tissues and several human PCa cell lines were analyzed, respectively, using StarBasev3.0 project and RT-qPCR assay. After MBNL1-AS1 overexpression, cell proliferation, invasion and migration were, respectively, evaluated using CCK-8, colony formation, transwell and wound healing assays. Dual luciferase assay were used for analysis of the interactions among MBNL1-AS1, miR-181a-5p, and phosphatase and tensin homolog (PTEN). Subsequently, the expression of PTEN and proteins in PI3K/AKT/mTOR signaling was examined using western blot analysis after transfection with miR-181a-5p mimic. The rescue assays were performed to investigate the effects of MBNL1-AS1 and miR-181a-5p on the functions of PCa cells and the expression of PTEN/PI3K/AKT/mTOR signaling by co-transfection with MBNL1-AS1 plasmid and miR-181a-5p mimic. Results indicated that MBNL1-AS1 was conspicuously downregulated while miR-181a-5p upregulating in PCa tissues and cell lines. MBNL1-AS1 overexpression decreased the abilities of cell proliferation, invasion, and migration. Further study revealed that MBNL1-AS1 acted as a sponge for miR-181a-5p and positively regulated PTEN by a sponge effect. Additionally, rescue assays proved that the effect of MBNL1-AS1-upregulation on the proliferation, invasion, and migration of PCa cells was dependent on miR-181a-5p. Furthermore, miR-181a-5p overexpression counteracted the expression of PTEN and proteins in PI3K/AKT/mTOR signaling exerted by MBNL1-AS1-upregulation in PCa cells. This study suggests that MBNL1-AS1 inhibits the progression of PCa via sponging miR-181a-5p and regulating PTEN/PI3K/AKT/mTOR pathway.

Low-dose decitabine for refractory prolonged isolated thrombocytopenia after HCT: a randomized multicenter trial.

Refractory prolonged isolated thrombocytopenia (RPIT) is an intractable complication after allogeneic hematopoietic cell transplantation (HCT), which often leads to poor prognosis. A clinical study was designed to validate the efficacy and safety of low-dose decitabine for RPIT after HCT and explore the related underlying mechanisms. Eligible patients were randomly allocated to receive 1 of 3 interventions: arm A, low-dose decitabine (15 mg/m2 daily IV for 3 consecutive days [days 1-3]) plus recombinant human thrombopoietin (300 U/kg daily); arm B, decitabine alone; or arm C, conventional treatment. The primary end point was the response rate of platelet recovery at day 28 after treatment. Secondary end points included megakaryocyte count 28 days after treatment and survival during additional follow-up of 24 weeks. Among the 91 evaluable patients, response rates were 66.7%, 73.3%, and 19.4% for the 3 arms, respectively (P < .001). One-year survival rates in arms A (64.4% ± 9.1%) and B (73.4% ± 8.8%) were similar (P = .662), and both were superior to that in arm C (41.0% ± 9.8%; P = .025). Megakaryocytes, endothelial cells (ECs), and cytokines relating to megakaryocyte migration and EC damage were improved in patients responding to decitabine. This study showed low-dose decitabine improved platelet recovery as well as overall survival in RPIT patients after transplantation. This trial was registered at www.clinicaltrials.gov as #NCT02487563.

Hybrid Molecular Dynamics for Elucidating Cooperativity Between Halogen Bond and Water Molecules During the Interaction of p53-Y220C and the PhiKan5196 Complex.

The cooperativity between hydrogen and halogen bonds plays an important role in rational drug design. However, mimicking the dynamic cooperation between these bonds is a challenging issue, which has impeded the development of the halogen bond force field. In this study, the Y220C-PhiKan5196 complex of p53 protein was adopted as a model, and the functions of three water molecules that formed hydrogen bonds with halogen atoms were analyzed by the simulation method governed by the hybrid quantum mechanical/molecular mechanical molecular dynamics. A comparison with the water-free model revealed that the strength of the halogen bond in the complex was consistently stronger. This confirmed that the water molecules formed weak hydrogen bonds with the halogen atom and cooperated with the halogen atom to enhance the halogen bond. Further, it was discovered that the roles of the three water molecules were not the same. Therefore, the results obtained herein can facilitate a rational drug design. Further, this work emphasizes on the fact that, in addition to protein pockets and ligands, the role of voids should also be considered with regard to the water molecules surrounding them.

[Effect of Additional Chromosomal Abnormalities on the Outcome of CML-CP Patients Receiving TKI Therapy].

OBJECTIVES: To explore the effect of additional chromosomal abnormalities on the prognosis and outcome of CML-CP patients receiving imatinib therapy. METHODS: The clinical and genetic data of 589 CML-CP patients receiving imatinib treatment between May 2009 and October 2014 in the 1st Affiliated Hospital of Soochow University were analyzed, the 589 patients were divided into 5 groups according to the karyotypes at the initial diagnosis. The OS(overall survival), PFS (progression-free survival), EFS (event-free survival), Cumulative MMR (major molecular remission) and Cumulative CCyR (complete cytogenetic remission) were calculated by using the Kaplan-Meier method and compared by using the log-rank text by Graphpad 6.0. The χ2 test was used to compare the frequency of optimal molecular response at 3, 6, 12 months among the 5 groups. RESULTS: There was significant difference about the frequency of optimal molecular response at 3 and 6 months between CML-CP patients with additional chromosomal abnormalities and those with classic t(9;22) ï¼»50%（12/24） vs. 73.94%(261 /353), P<0.05; 50%(10 /20) vs. 72.05%(232 /322) (P<0.05)ï¼½, and the same significant difference was found at 6 months between the group with variant translocations and that with classic t(9;22) ï¼»53.3% (16 /30) vs. 72.05%(232 /322) (P<0.05)ï¼½. The P values of cumulative CCyR (P<0.05) and EFS (P<0.01) for 4 years were statistically significant between CML-CP patients with additional chromosomal abnormalities and the other 4 groups. Compared one to another, there was the significant difference in cumulative CCyR and EFS for 4 years between CML-CP patients with additional chromosomal.abnormalities and those with classic t(9;22) (47.25% vs. 84.01%)(P<0.05); (75.03% vs. 90.01%)(P<0.01). CONCLUSION: The additional chromosomal abnormalities influence the outcome of CML-CP patients receiving imatinib treatment, which make poor prognosis.

[Risk Factors of Leukemia-free Survival in Ph+ ALL Patients with Allogeneic Hematopoietic Stem Cell Transplantation].

OBJECTIVE: To explore the effect of BCR-ABL gene transcripts on Leukemia-free survival (LFS) and prognosis of patients with Philadelphia chromosome positive acute lymphoblastic leukemia (Ph+ ALL) after allogeneic hematopoietic stem cell transplantation (allo-HSCT). METHODS: The clinical data of 107 cases of Ph+ B-ALL patients received allo-HSCT from July 2006 to November 2014 in the First Affiliated Hospital of Soochow University were collected and the relationship between the clinical characteristics and LFS after transplantation was analyzed. RESULTS: Out of 107 Ph+ ALL patients (64 males and 43 females) with a median age of 30(7 to 54)years old, 35.5% (38/107) cases relapsed after transplantation within a median time of 6.9 (1.5 to 40.7) months. A total of 39 (36.4%) cases died within a median time of 19.8 (3.6 to 83.7) months after HSCT, of which 51.3% (20/39) due to disease relapse and 25.6% (10/39) due to infection. BCR-ABL gene transcripts of 49 cases turn into negative before transplantation, of which the expected 5-year cumulative incidence of relapse (CIR), non-relapse mortality (NRM) and overall survival (OS) were 26.5%, 29.5% and 41.6%, respectively. Another 49 cases still had a positive BCR-ABL gene transcripts before transplantation, of which the life expectancy of 5 year CIR, NRM and OS were 64.4%,8.9% and 48.9%, respectively. Compared with BCR-ABL positive patients, BCR-ABL negative patients showed a lower CIR (P<0.001), a higher NRM (P=0.030) and a similar OS (41.6% versus 48.9%, P=0.497). Multivariate analysis showed that BCR-ABL positive (P=0.016) and a disease statusphase ≥CR2 (P<0.001) before HSCT were independent risk factors for LFS, while the age underwent HSCT was the principal element affecting prognosis (P<0.001). CONCLUSION: Both the relapse and infection are the main causes of death in the patients after transplantation. A disease status ≥CR2 and the BCR-ABL positive before transplantation are 2 independent risk factors of LFS in the patients with Ph+ ALL after allo-HSCT.

A wind energy powered wireless temperature sensor node.

A wireless temperature sensor node composed of a piezoelectric wind energy harvester, a temperature sensor, a microcontroller, a power management circuit and a wireless transmitting module was developed. The wind-induced vibration energy harvester with a cuboid chamber of 62 mm × 19.6 mm × 10 mm converts ambient wind energy into electrical energy to power the sensor node. A TMP102 temperature sensor and the MSP430 microcontroller are used to measure the temperature. The power management module consists of LTC3588-1 and LT3009 units. The measured temperature is transmitted by the nRF24l01 transceiver. Experimental results show that the critical wind speed of the harvester was about 5.4 m/s and the output power of the harvester was about 1.59 mW for the electrical load of 20 kΩ at wind speed of 11.2 m/s, which was sufficient to power the wireless sensor node to measure and transmit the temperature every 13 s. When the wind speed increased from 6 m/s to 11.5 m/s, the self-powered wireless sensor node worked normally.

[The prognosis analysis of novel influenza A (H1N1) in patients with hematologic disease].

OBJECTIVE: To explore disease progression and prognosis factors of novel influenza A (H1N1) in immunocompromised patients with or without hematologic disease. METHOD: A total of 76 confirmed novel influenza A (H1N1) infection patients from November 2009 to March 2010 included in the study, their clinical feature was analyzed, and the relationship between clinical feature and outcome was explored retrospectively by multivariate analysis method. RESULTS: The whole 76 patients were administrated of oseltamivir. Among the 76 patients, 46 were severe and 23 were critical. Of the 6 patients with immunocompromised hematologic disease, 2 were severe and 4 were critical, case-fatality rate was 66.67% (4/6). The case-fatality rate of patients with non-hematologic disease was 10.42% (5/48). Multivariable logistic-regression analysis showed that immunocompromised hematologic disease (P = 0.0008, odds ratio:75.368; 95% CI, 5.980 to 949.853) and age (P = 0.0380) were independent risk factors for death. And other variables such as chronical lung disease, interval time from the onset of illness and lymphocyte count had no statistical significance (P > 0.05). CONCLUSIONS: The novel influenza A (H1N1) patients with immunocompromised hematologic disease has a poor prognosis, they can deteriorate quickly and have high mortality, it may aid clinicians to pay high attention to these people.

[Genetic polymorphism of the killer cell immunoglobulin-like receptor genes in the Inner Mongolian population].

OBJECTIVE: To investigate the killer cell immunoglobulin-like receptor (KIR) gene frequencies and genotypes distributions in the Inner Mongolian population. METHODS: Ninety genomic DNA samples were extracted from blood samples of randomly chosen Mongolian individuals. Gene-specific PCR amplification was used to identify genes present or absent for 16 KIR loci. KIR genotype distributions were obtained and compared to that of 24 populations published in literatures using principal component analysis by SAS8.0 software. Genetic tree was constructed by the calculate Nei's genetic distance. RESULTS: (1) The frequency of KIR 2DL2, 2DS2 in Mongolian individual is higher than that in north Mongoloid and less than that in Caucasian. (2) Haplotype AA was identified in 37.78% of individuals, which is higher than that in north Mongoloid and lower than that in Caucasian. (3) Mongolian was considered between north Mongoloid and Caucasian by principal component and genetic tree analysis. CONCLUSION: Mongolian might be affected by the north Mongoloid and Caucasian, and showed intermediate between the two populations.

[Expressions of survivin and the splice variants survivin-2B and survivin-DeltaEx3 in bladder cancer and their clinical significance].

BACKGROUND AND OBJECTIVE: Studies on survivin over the past 5 years have shown that survivin participates in the genesis of several human cancers, including bladder cancer. Recent studies have indicated that survivin splice variants appeared to have unique subcellular localizations and functions as well. This study was to explore the roles of survivin and its two splice variants survivin-2B and survivin-DeltaEx3 in transitional cell carcinoma of bladder (BTCC). METHODS: The relative amount of survivin, survivin-2B, and survivin-DeltaEx3 mRNA of fresh carcinoma tissues from 60 patients with BTCC and 12 non-cancerous bladder tissues were detected by real-time quantitative reverse transcription polymerase chain reaction (RT-PCR), and the relationships of their expression levels in different pathologic grades to clinical stages of bladder cancer were analyzed. The time of follow-up was 4-24 months. RESULTS: Survivin, survivin-2B, and survivin-DeltaEx3 mRNA were detected in all BTCC tissues, and their relative expressions were 0.333+/-0.163, 0.056+/-0.017, and 0.124+/-0.096, respectively. In the control group,three and four samples expressed survivin and survivin-DeltaEx3 mRNA respectively, and all samples expressed survivin-2B mRNA. The expressions of survivin and survivin-DeltaEx3 mRNA were positively correlated with the pathologic grades and clinical stages (0 < r 's < 1,P < 0.05), however, survivin-2B mRNA was negatively correlated with those (-1 < r 's < 0, P < 0.05). CONCLUSION: Detecting the expression levels of survivin and its two splice variants survivin-2B and survivin-DeltaEx3 mRNA in BTCC by real-time PCR could have potential values to evaluate tumor progression and recurrence rate.

[Expression of survivin mRNA in urine exfoliated cells of patients with bladder transitional cell carcinoma detected by real-time PCR].

BACKGROUND AND OBJECTIVE: Bladder cancer is the most malignant cancer of the urinary system. However a noninvasive and sensitive method of the early diagnosis for bladder cancer has not been developed. This study was to explore the expression of survivin mRNA in urine exfoliated cells of preoperative and postoperative patients with bladder transitional cell carcinoma (BTCC), and to analyze its value in early diagnosis and postoperative monitoring. METHODS: Urine of 30 patients with initially diagnosed BTCC was collected before operation and one week, one month, six months and 15 months after operation. Urine of 10 healthy volunteers and 15 patients with cystitis was used as control. Expression of survivin mRNA in urine exfoliated cells was detected by real-time fluorescent quantitative polymerase chain reaction (real-time PCR). RESULTS: The relative copy number of survivin mRNA in the patients was (96.01+/-42.33) before operation, which was significantly higher than that of healthy volunteers and cystitis patients (P <0.05). The level of survivin mRNA was apparently declined one week after operation (25.30+/-1.51) compared with its preoperative level (P <0.05); and the level became as low as the control group after one month (13.20+/-1.49) and six months (13.90+/-1.36) (P>0.05). Patients were followed up for 15 months, and three patients recurred, whose survivin mRNA level (97.83+/-27.47) was significantly higher than that at six months after operation (P <0.05). CONCLUSION: Detecting survivin mRNA in urine exfoliated cells is sensitive for the diagnosis of BTCC. Detection survivin mRNA after operation can monitor recurrence.

[Studies on chemical constituents of Dendranthema indicum var. aromaticum].

OBJECTIVE: To study chemical constituents of Dendranthema indicum var. aromaticum. METHODS: The constituents were separated and purified by column chromatography with silica gel and Semi-preparative HPLC. Their structures were identified on the basis of physical-chemical properties and spectral data. RESULTS: Seven compounds were isolated and identified as acacetin, apignein, acacetin-7-O-beta-D-glucopy ranoside, apignein-7-O-beta-D-glucopy ranosids, luteolin, beta-sitosterol and daucosterol. CONCLUSION: Apigenin, acacetin-7-O-beta-D-glucopy ranoside and apignein-7-O-beta-D-glucopy ranoside are obtained from the plant for the first time.

[Analysis of the chemical constituents of essential oil from Magnolia biondii by GC-MS].

OBJECTIVE: The chemical components of essential oil from Magnolia biondii were analyzed by GC-MS. METHODS: Essential oil was extracted by steam distillation (SD). The chemical components of essential oil were analyzed by GC-MS. RESULTS: The chemical components in the oil were qualitatively and quantitatively analyzed by GC-MS. 63 components were separated and 50 components were identified. The main components were Eucalyptol (28.92%), P-pinene (12.39%), alpha-Terpineol (8.28%). CONCLUSION: This is the first time to adopt GC-MS to analyze the chemical components of volatile oil of Magnolia biondii, and this study can provide science basis for further research development of Magnolia biondii.