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Lentiviral vectors (LVs), derived from human immunodeficiency virus, are powerful tools for modifying the genes of eukaryotic cells such as hematopoietic stem cells and neural cells. With the extensive and in-depth studies on this gene therapy vehicle over the past two decades, LVs have been widely used in both research and clinical trials. For instance, third-generation and self-inactive LVs have been used to introduce a gene with therapeutic potential into the host genome and achieve targeted delivery into specific tissue. When LVs are employed in leukemia, the transduced T cells recognize and kill the tumor B cells; in β-thalassemia, the transduced CD34+ cells express normal β-globin; in adenosine deaminase-deficient severe combined immunodeficiency, the autologous CD34+ cells express adenosine deaminase and realize immune reconstitution. Overall, LVs can perform significant roles in the treatment of primary immunodeficiency diseases, hemoglobinopathies, B cell leukemia, and neurodegenerative diseases. In this review, we discuss the recent developments and therapeutic applications of LVs. The safe and efficient LVs show great promise as a tool for human gene therapy.

Pseudomonas aeruginosa infection continues to be a major threat to global public health, and new safe and efficacious vaccines are needed for prevention of infections caused by P. aeruginosa. X-ray irradiation has been used to prepare whole-cell inactivated vaccines against P. aeruginosa infection. However, the immunological mechanisms of X-ray-inactivated vaccines are still unclear and require further investigation. Our previous study found that an X-ray-inactivated whole-cell vaccine could provide protection against P. aeruginosa by boosting T cells. The aim of the present study was to further explore the immunological mechanisms of the vaccine. Herein, P. aeruginosa PAO1, a widely used laboratory strain, was utilized to prepare the vaccine, and we found nucleic acids and 8-hydroxyguanosine in the supernatant of X-ray-inactivated PAO1 (XPa). By detecting CD86, CD80, and MHCII expression, we found that XPa fostered dentritic cell (DC) maturation by detecting. XPa stimulated the cGAS-STING pathway as well as Toll-like receptors in DCs in vitro, and DC finally underwent apoptosis and pyroptosis after XPa stimulation. In addition, DC stimulated by XPa induced CD8+ T-cell proliferation in vitro and generated immunologic memory in vivo. Moreover, XPa vaccination induced both Th1 and Th2 cytokine responses in mice and reduced the level of inflammatory factors during infection. XPa protected mice in pneumonia models from infection with PAO1 or multidrug-resistant clinical isolate W9. Chronic obstructive pulmonary disease (COPD) mice immunized with XPa could resist PAO1 infection. Therefore, a new mechanism of an X-ray-inactivated whole-cell vaccine against P. aeruginosa infection was discovered in this study.

This study aimed to evaluate the antimicrobial activity of the novel monosulfactam 0073 against multidrug-resistant Gram-negative bacteria in vitro and in vivo and to characterize the mechanisms underlying 0073 activity. The in vitro activities of 0073, aztreonam, and the combination with avibactam were assessed by MIC and time-kill assays. The safety of 0073 was evaluated using 3-(4,5-dimethylthizol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and acute toxicity assays. Murine thigh infection and pneumonia models were employed to define in vivo efficacy. A penicillin-binding protein (PBP) competition assay and confocal microscopy were conducted. The inhibitory action of 0073 against β-lactamases was evaluated by the half-maximal inhibitory concentration (IC(50)), and resistance development was evaluated via serial passage. The monosulfactam 0073 showed promising antimicrobial activity against Enterobacteriaceae, Pseudomonas aeruginosa, and Acinetobacter baumannii isolates producing metallo-β-lactamases (MBLs) and serine β-lactamases. In preliminary experiments, compound 0073 exhibited safety both in vitro and in vivo. In the murine thigh infection model and the pneumonia models in which infection was induced by P. aeruginosa and Klebsiella pneumoniae, 0073 significantly reduced the bacterial burden. Compound 0073 targeted several PBPs and exerted inhibitory effects against some serine β-lactamases. Finally, 0073 showed a reduced propensity for resistance selection compared with that of aztreonam. The novel monosulfactam 0073 exhibited increased activity against β-lactamase-producing Gram-negative organisms compared with the activity of aztreonam and showed good safety profiles both in vitro and in vivo. The underlying mechanisms may be attributed to the affinity of 0073 for several PBPs and its inhibitory activity against some serine β-lactamases. These data indicate that 0073 represents a potential treatment for infections caused by β-lactamase-producing multidrug-resistant bacteria.

Ionizing irradiation kills pathogens by destroying nucleic acids without protein structure destruction. However, how pathogens respond to irradiation stress has not yet been fully elucidated. Here, we observed that Pseudomonas aeruginosa PAO1 could release nucleic acids into the extracellular environment under X-ray irradiation. Using scanning electron microscopy (SEM) and transmission electron microscopy (TEM), X-ray irradiation was observed to induce outer membrane vesicle (OMV) formation in P. aeruginosa PAO1. The size distribution of the OMVs of the irradiated PAO1 was similar to that of the OMVs of the non-irradiated PAO1 according to nanoparticle tracking analysis (NTA). The pyocin-related proteins are involved in OMV production in P. aeruginosa PAO1 under X-ray irradiation conditions, and that this is regulated by the key SOS gene recA. The OMV production was significantly impaired in the irradiated PAO1 Δlys mutant, suggesting that Lys endolysin is associated with OMV production in P. aeruginosa PAO1 upon irradiation stress. Meanwhile, no significant difference in OMV production was observed between PAO1 lacking the pqsR, lasR, or rhlR genes and the parent strain, demonstrating that the irradiation-induced OMV biosynthesis of P. aeruginosa was independent of the Pseudomonas quinolone signal (PQS).

Clustered regularly interspaced short palindromic repeats (CRISPR)-caspase 9 (Cas9) genome editing technology holds great promise for the field of human gene therapy. However, a lack of safe and effective delivery systems restricts its biomedical application. Here, a folate receptor-targeted liposome (F-LP) was used to deliver CRISPR plasmid DNA co-expressing Cas9 and single-guide RNA targeting the ovarian cancer-related DNA methyltransferase 1 (DNMT1) gene (gDNMT1). F-LP efficiently bound the gDNMT1 plasmid and formed a stable complex (F-LP/gDNMT1) that was safe for injection. F-LP/gDNMT1 effectively mutated endogenous DNMT1 in vitro, and then expressed the Cas9 endonuclease and downregulated DNMT1 in vivo. The tumor growth of both paclitaxel-sensitive and -resistant ovarian cancers were inhibited by F-LP/gDNMT1, which shows fewer adverse effects than paclitaxel injection. Therefore, CRISPR-Cas9-targeted DNMT1 manipulation may be a potential therapeutic regimen for ovarian cancer, and lipid-mediated delivery systems represent promising delivery vectors of CRISPR-Cas9 technology for precise genome editing therapeutics.

With the improvement of gene vectors, the rise of chimeric antigen receptor T cell immunotherapy and breakthroughs in the genome editing technology, gene therapy had once again returned to the central stage of disease treatment. It had brought new choices to clinical therapy of diseases such as tumors and genetic diseases, and had changed the status quo of treatment for monogenic disorders and diffuse large B-cell lymphoma. Until August 2019, 22 gene medicines had been approved by the drug regulatory agencies from various countries, but there were few relevant reviews of combing these drugs systematically. Consequently, this review summarizes the gene therapy drugs approved worldwide from 1998 to 2019 in details, including names, indications, dates of approval, companies, vectors, the applied technologies and mechanisms of gene therapy drugs, etc. Furthermore, the gene therapy drugs were classified and addressed in accordance with the employed vectors. Gene therapy had gradually been accepted by the government and the public since 1980s, and have become a new and important alternative to existing treatments for human diseases in the past few years. Therefore, gene therapy drugs, with safe vectors and advanced biotechnologies, would play a greater role in the prevention and treatment of human diseases in future.

Interleukin-15 has been implicated as a promising cytokine for cancer immunotherapy, while folate receptor α (FRα) has been shown to be a potentially useful target for colon cancer therapy. Herein, we developed F-PLP/pIL15, a FRα-targeted lipoplex loading recombinant interleukin-15 plasmid (pIL15) and studied its antitumor effects in vivo using a CT26 colon cancer mouse model. Compared with control (normal saline) treatment, F-PLP/pIL15 significantly suppressed tumor growth in regard to tumor weight (P < 0.001) and reduced tumor nodule formation (P < 0.001). Moreover, when compared to other lipoplex-treated mice, F-PLP/pIL15-treated mice showed higher levels of IL15 secreted in the serum (P < 0.001) and ascites (P < 0.01). These results suggested that the targeted delivery of IL15 gene might be associated with its in vivo antitumor effects, which include inducing tumor cell apoptosis, inhibiting tumor proliferation and promoting the activation of immune cells such as T cells and natural killer cells. Furthermore, hematoxylin and eosin staining of vital organs following F-PLP/pIL15 treatment showed no detectable toxicity, thus indicating that intraperitoneal administration may be a viable route of delivery. Overall, these results suggest that F-PLP/pIL15 may serve as a potential targeting preparation for colon cancer therapy.

Colorectal cancer (CRC) is a major cause of cancer-related mortality worldwide. Moreover, metastasis is one of the main causes of death in CRC patients. Nanotechnology-based gene therapy has shown significant therapeutic benefits in recent clinical trials for cancer treatment. Recent studies have shown that pigment epithelium-derived factor (PEDF) protein can inhibit tumor growth and metastasis by anti-angiogenesis and pro-apoptosis. In this study, we prepared a PEDF-DNA-loaded liposome for cancer-targeted gene therapy for metastatic CRC using an iRGD peptide. Our results showed that cancer-targeted PEDF-DNA liposomes (R-LP/PEDF) exhibited enhanced inhibitory effects on invasion, migration, and pro-apoptosis of CRC cells in vitro. In addition, it reduced metastasis tumor nodules in lung and prolonged the survival time in a mouse model of metastatic CRC.

We reported that perinatal exposure to bisphenol A (BPA) resulted in obesity in offspring female rats, and down-regulated zinc-alpha2-glycoprotein (ZAG) mRNA expression in subcutaneous adipose tissue maybe was associated with increased body weight in previous study. Considering that the mechanism of ZAG actions has not been clear, and ZAG gene expression levels are different with adipose tissues, we investigate the levels of ZAG mRNA and protein expression in visceral (perigonadal and perirenal) adipose tissue, and the mechanism of ZAG increasing body weight in BPA exposed rats. Pregnant rats were exposed to BPA in water at levels of either 1 μg/mL(low dose of BPA, LBPA) or 10 μg/mL (high dose of BPA, HBPA) from gestation day 6 to the end of lactation. ZAG mRNA and protein expression from visceral adipose tissue of offspring female rats on postnatal day 50 (PND50) were measured. The levels of fatty acid synthase (FAS) and hormone-sensitive lipase (HSL) gene expression, and peroxisome proliferator-activated receptors-γ (PPARγ) mRNA and protein in visceral adipose tissue were also determined. There was large size of adipocytes in perigonadal adipose tissues in BPA treated offspring female rats. Perinatal exposure to BPA could decrease the levels of ZAG and PPARγ mRNA and protein expression in visceral adipose tissues, and increase FAS gene expression, but decrease HSL gene expression. These findings indicated that BPA exposure during early development results in large size of adipocytes, which was associated with down-regulated ZAG gene. Low expression of ZAG gene increased lipogenesis and decreased lipolysis, which was associated with down-regulated PPARγ gene induced by BPA.

Ovarian cancers highly overexpress folate receptor α (FRα) and claudin3 (CLDN3), both of which are associated with tumor progression and poor prognosis of patients. Downregulation of FRα and CLDN3 in ovarian cancer may suppress tumor growth and promote benign differentiation of tumor. In this study, F-P-LP/CLDN3, a FRα targeted liposome loading with short hairpin RNA (shRNA) targeting CLDN3 was prepared and the pharmaceutical properties were characterized. Then, the antitumor effect of F-P-LP/CLDN3 was studied in an in vivo model of advanced ovarian cancer. Compared with Control, F-P-LP/CLDN3 promoted benign differentiation of tumor and achieved about 90% tumor growth inhibition. In the meantime, malignant ascites production was completely inhibited, and tumor nodule number and tumor weight were significantly reduced (p

Accumulating evidences indicate that bisphenol A (BPA) exposure during early development increases the risk of obesity and insulin resistance in later life. Despite growing public concern, it is not totally clear about the mechanism of BPA. This study aimed to evaluate the effect of perinatal exposure to BPA on body weight, glucose metabolism and adipokine expression in adolescent female offspring rats. Pregnant rats were exposed to BPA in water at levels of either 1 μg/mL (LBPA) or 10 μg/mL (HBPA) from gestation day 6 to the end of lactation. Body weight, the levels of fasting serum glucose, insulin and adiponectin (ADP), and Zinc-alpha2-glycoprotein (ZAG) and ADP mRNA and protein expression from adipose tissue in 7 weeks old female offspring were measured. There were high birth weight and high body weight at age of 7 weeks in BPA treated group. There were significantly higher levels of serum glucose and insulin, and lower levels of serum ADP and plasma ZAG protein, and lower ADP and ZAG mRNA and protein expression in BPA-exposed female offspring compared with controls. The results suggested that BPA-exposed during early development had long-term deleterious effects on body weight and glucose metabolism, which was probably associated with the down-regulated expression of ZAG and ADP gene in early developmental female rats.

Overexpression of folate receptor alpha (FRα) and high telomerase activity are considered to be the characteristics of ovarian cancers. In this study, we developed FRα-targeted lipoplexes loaded with an hTERT promoter-regulated plasmid that encodes a matrix protein (MP) of the vesicular stomatitis virus, F-LP/pMP(2.5), for application in ovarian cancer treatment. We first characterized the pharmaceutical properties of F-LP/pMP(2.5). The efficient expression of the MP-driven hTERT promoter in SKOV-3 cells was determined after an in-vitro transfection assay, which was significantly increased compared with a non-modified LP/pMP(2.5) group. F-LP/pMP(2.5) treatment significantly inhibited the growth of tumors and extended the survival of mice in a SKOV-3 tumor model compared with other groups. Such an anti-tumor effect was due to the increased expression of MP in tumor tissue, which led to the induction of tumor cell apoptosis, inhibition of tumor cell proliferation and suppression of tumor angiogenesis. Furthermore, a preliminary safety evaluation demonstrated a good safety profile of F-LP/pMP(2.5) as a gene therapy agent. Therefore, FRα-targeted lipoplexes with therapeutic gene expression regulated by an hTERT promoter might be a promising gene therapy agent and a potential translational candidate for the clinical treatment of ovarian cancer.

To identify susceptibility loci for schizophrenia, we performed a two-stage genome-wide association study (GWAS) of schizophrenia in the Han Chinese population (GWAS: 746 individuals with schizophrenia and 1,599 healthy controls; validation: 4,027 individuals with schizophrenia and 5,603 healthy controls). We identified two susceptibility loci for schizophrenia at 6p21-p22.1 (rs1233710 in an intron of ZKSCAN4, P(combined) = 4.76 × 10(-11), odds ratio (OR) = 0.79; rs1635 in an exon of NKAPL, P(combined) = 6.91 × 10(-12), OR = 0.78; rs2142731 in an intron of PGBD1, P(combined) = 5.14 × 10(-10), OR = 0.79) and 11p11.2 (rs11038167 near the 5' UTR of TSPAN18, P(combined) = 1.09 × 10(-11), OR = 1.29; rs11038172, P(combined) = 7.21 × 10(-10), OR = 1.25; rs835784, P(combined) = 2.73 × 10(-11), OR = 1.27). These results add to previous evidence of susceptibility loci for schizophrenia at 6p21-p22.1 in the Han Chinese population. We found that NKAPL and ZKSCAN4 were expressed in postnatal day 0 (P0) mouse brain. These findings may lead to new insights into the pathogenesis of schizophrenia.

Hongyan Zhang1, Huafang Li2, Liang Shu1, Niufan Gu2, Gang Wang3, Yongzhen Weng3, Shiping Xie4, Xinbao Zhang4, Ting Li5, Cui Ma5, Wei Yu6, Bruce Parsons7, Manjula Schou81Institute of Mental Health, Peking University, Beijing, China; 2Shanghai Mental Health Center, Shanghai, China; 3Capital Medical University, Beijing An Ding Hospital, Beijing, China; 4Nanjing Brain Hospital, Nanjing, China; 5Guangzhou Brain Hospital, Guangzhou, China; 6Pfizer China, Beijing, China; 7Pfizer Inc, New York, NY, USA; 8Pfizer Australia, Sydney, AustraliaBackground: The aim of the study was to evaluate the efficacy and safety of ziprasidone versus risperidone in Chinese subjects with acute exacerbation of schizophrenia.Methods: In patients meeting the Chinese Classification of Mental Disorders criteria for schizophrenia and with a Positive and Negative Syndrome Scale (PANSS) total score ≥60 were randomly assigned to six weeks of double-blind treatment with ziprasidone 40–80 mg twice daily or risperidone 1–3 mg bid, flexibly dosed. Noninferiority was demonstrated if the upper limit of the two-sided 95% confidence interval (CI) for the difference in PANSS total score improvement from baseline in the evaluable population was smaller than the prespecified noninferiority margin of 10 units.Results: The intent-to-treat population comprised 118 ziprasidone-treated and 121 risperidone-treated subjects. Improvement (reduction) from baseline to week 6 in PANSS total score was (-35.6 [95% CI: -38.6, -32.6]) for ziprasidone and (-37.1 [95% CI: -39.9, -34.4]) for risperidone. Noninferiority was demonstrated in the evaluable population with a difference score of 1.5 [95% CI: -2.5, 5.5]. Mean prolactin levels decreased at week 6 compared with baseline for ziprasidone (-3.5 ng/mL), but significantly increased for risperidone (61.1 ng/mL; P < 0.001). More risperidone-treated subjects (14.9%) than ziprasidone-treated subjects (4.2%) reported weight gain ≥7%. Akathisia and somnolence in the ziprasidone group and akathisia and insomnia in the risperidone group were the most common side effects. Treatment-related/treatment-emergent adverse events were reported by 79.7% and 71.1% of ziprasidone-treated and risperidone-treated subjects, respectively.Conclusion: In Chinese subjects, ziprasidone was as effective as risperidone, with less weight gain and less prolactin elevation.Keywords: ziprasidone, risperidone, schizophrenia

Here, three novel cholesterol (Ch)/low molecular weight polyethylene glycol (PEG) conjugates, termed α, ω-cholesterol-functionalized PEG (Ch2-PEGn), were successfully synthesized using three kinds of PEG with different average molecular weight (PEG600, PEG1000 and PEG2000). The purpose of the study was to investigate the potential application of novel cationic liposomes (Ch2-PEGn-CLs) containing Ch2-PEGn in gene delivery. The introduction of Ch2-PEGn affected both the particle size and zeta potential of cationic liposomes. Ch2-PEG2000 effectively compressed liposomal particles and Ch2-PEG2000-CLs were of the smallest size. Ch2-PEG1000 and Ch2-PEG2000 significantly decreased zeta potentials of Ch2-PEGn-CLs, while Ch2-PEG600 did not alter the zeta potential due to the short PEG chain. Moreover, the in vitro gene transfection efficiencies mediated by different Ch2-PEGn-CLs also differed, in which Ch2-PEG600-CLs achieved the strongest GFP expression than Ch2-PEG1000-CLs and Ch2-PEG2000-CLs in SKOV-3 cells. The gene delivery efficacy of Ch2-PEGn-CLs was further examined by addition of a targeting moiety (folate ligand) in both folate-receptor (FR) overexpressing SKOV-3 cells and A549 cells with low expression of FR. For Ch2-PEG1000-CLs and Ch2-PEG2000-CLs, higher molar ratios of folate ligand resulted in enhanced transfection efficacies, but Ch2-PEG600-CLs had no similar in contrast. Additionally, MTT assay proved the reduced cytotoxicities of cationic liposomes after modification by Ch2-PEGn. These findings provide important insights into the effects of Ch2-PEGn on cationic liposomes for delivering genes, which would be beneficial for the development of Ch2-PEGn-CLs-based gene delivery system.

Darier’s disease (DD) is an autosomal dominant genodermatology. Mutations in the ATP2A2 gene encoding sarco-endoplasmic reticulum calcium pumping ATPase type 2 (SERCA2) have been identified as the molecular basis of DD. The aim of this study was to report two Chinese pedigree of DD and to explore the genetic mutations. Polymerase chain reaction was carried out to amplify the exons and flanking intron boundaries of the ATP2A2 gene followed by direct sequencing. Two novel missense mutations were identified, a change of C203 to A (A68E) in exon 3 was found in one family and a change of C2759 to T (S920F) in exon 19 in the other, which were located within the transmembrane domain of SERCA2, highly conserved during evolution. The A68E and S920F mutations might be regarded as the causes of the disease in two Chinese families, but these were not tested functionally. Additional functional experiments are necessary to verify the relevance and suitability of these findings for future use in genetic counseling and prenatal diagnosis.