Platinum-Acridine Agents with High Activity in Cancers Expressing the Solute Carrier MATE1 (SLC47A1).

Platinum-acridine anticancer agents (PAs) containing acyclic (1 and 3) and heterocyclic (R)-3-aminopiperidine (2) and 2-iminopyrrolidine (4) based linker moieties were studied. Similar to 1, rigidified 2 shows a strong positive correlation between potency and SLC47A1 (multidrug and toxin extrusion protein 1, MATE1) gene expression levels across the NCI-60 panel of cancer cell lines. All derivatives show nanomolar activity in HepG2 (liver), NCI-H460 (lung), and MDA-MB-436 (breast), which express high levels of SLC47A1 (Cancer Cell Line Encyclopedia, CCLE). The PAs are up to 350-fold more potent than cisplatin. In a MATE1 inhibition assay, a significant reduction in activity is observed in the three cancer cell lines (4000-fold lower for HepG2). Molecular docking experiments provide insight into the compatibility of the structurally diverse set of PAs with MATE1-mediated transport. MATE1 is a predictive marker and actionable target that sensitizes cancer cells regardless of the tissue of origin to PAs.

Tryptophan residue of plasmid-encoded Pgp3 is important for Chlamydia muridarum to induce hydrosalpinx in mice.

The crucial role of plasmid-encoded protein Pgp3 in Chlamydia pathogenesis has been demonstrated in various animal models. Previous studies have revealed that the Pgp3-deficient C. muridarum mutant fails to induce hydrosalpinx after vaginal inoculation in mice. Structural analysis of C. trachomatis Pgp3 trimer has indicated that Trp234 may play a critical role in trimeric crystal packing interactions and that Tyr197 is involved at predominant cation-binding sites. In this study, we constructed C. muridarum transformants harboring Pgp3, Trp234, or Tyr197 point mutations (Pgp3W234A and Pgp3Y197A). C3H/HeJ mice infected with Pgp3W234A mutant failed to induce severe hydrosalpinx in the oviduct tissue, which largely phenocopied the full-length Pgp3-deficient C. muridarum. The Pgp3Y197A variant induced an intermediate severity of pathology. The attenuated pathogenicity caused by the Pgp3W234A mutant may be due to its decreased survival in the lower genital tracts of mice, reduced ascension to the oviduct, and milder induction of inflammatory cell infiltration in the oviduct tissue. Thus, our results point to an important amino acid residue involved in Pgp3 virulence, providing a potential therapeutic target for chlamydial infection.

Chemosensitivity-Gene Expression Correlations and Functional Enrichment Analysis Provide Insight into the Mechanism of Action of a Platinum-Acridine Anticancer Agent.

NCI-60 growth inhibition and gene expression profiles were analyzed using Pearson correlation and functional enrichment computational tools to demonstrate critical mechanistic differences between a nucleolus-targeting platinum-acridine anticancer agent (PA) and other DNA-directed chemotherapies. The results support prior experimental data and are consistent with DNA being a major target of the hybrid agent based on the negative correlations observed between its potency and expression levels of genes implicated in DNA double-strand break (DSB) repair. Gene ontology terms related to RNA processing, including ribosome biogenesis, are also negatively enriched, suggesting a mechanism by which these processes render cancer cells more resistant to the highly cytotoxic agent. The opposite trend is observed for oxaliplatin and other DNA-targeted drugs. Significant functional interactions exist between genes/gene products involved in ribosome biogenesis and DSB repair, including the ribosomal protein (RPL5)-MDM2-p53 surveillance pathway, as a response to the nucleolar stress produced by PAs.

Prevalence and Genotype Distribution of Human Papillomavirus Among Attendees at a Sexually Transmitted Diseases Clinic in Urban Tianjin, China.

BACKGROUND: Human papillomavirus (HPV) is linked to various cancers in males and females. The prevalence and genotype distribution of HPV vary depending on geographical region and the immunity provided by vaccines. Investigation of HPV epidemiology is of great meaning for the development of prevention programs. METHODS: From January 2017 to September 2019, using PCR-reverse dot blot hybridisation, we determined the HPV subtypes in 2801 patients 17-89 years old at the sexually transmitted diseases (STD) clinic of Tianjin Medical University General Hospital. RESULTS: The HPV infection rate was 50.79% in males and 50.64% in females. The most common HPV genotype in males and females was HPV6 (30.15% and 30.43%), followed by HPV16 (18.76% and 20.65%) and HPV11 (14.61% and 15.67%). Infection with a single HPV subtype predominated in both males and females, and the rate in males was higher than in females. By contrast, the rate of high-risk HPV (hrHPV) and low-risk HPV (lrHPV) mixed infection in females was higher than in males. Most HPV-positive patients were 20-39 years of age. The prevalence of infection with only hrHPV differed among the age groups; the peak age was 50 to 59 years. CONCLUSION: The HPV prevalence was higher among the STD clinic outpatients than the general population. Therefore, a large-scale survey of high-risk populations is needed. It is anticipated that HPV vaccines, regular education and physical examinations may have a significant impact on the prevention of HPV-related diseases in high-risk groups.

Efficacy of minocycline in the treatment of early syphilis.

Syphilis is the third prevalent infectious disease in China, caused by the spirochete bacterium Treponema pallidum. Minocycline is a derivative of tetracycline used as an alternative treatment for syphilis, but there are few studies in this field. In this research, we compared the efficacy of benzathine penicillin and minocycline in the treatment of early syphilis patients and analyzed some of the factors affecting the efficacy of minocycline. A total of 276 eligible patients treated between January 2011 and December 2017 were retrospectively analyzed, and 158 patients received 100 mg of minocycline orally, twice daily for 28 days, while 118 patients received benzathine penicillin, 2.4 million units intramuscular injections, once a week, 1-2 times in all. All patients accepted rapid plasma regain (RPR) serological tests and followed up for 24 months to evaluate serological treatment responses. After comparison, the serological cure rate of the minocycline treatment group (85.44%) was similar to the benzathine penicillin treatment group (88.14%). Besides, patients in the minocycline treatment group with higher initial RPR titer (≥1:32) exhibited better treatment effect. In addition, during the 24-month follow-up, the serological cure rate of primary syphilis patients after minocycline treatment was significantly higher than that of secondary and early latent syphilis patients. Therefore, minocycline may be an effective alternative treatment to early syphilis when benzathine penicillin is not available.

GlgA plays an important role in the induction of hydrosalpinx by Chlamydia muridarum.

While glycogen synthase A deficiency can reduce the growth and proliferation of Chlamydia muridarum, the effect of glycogen synthase A on the pathogenic process of C. muridarum remains unclear. To characterize the effect of glycogen synthase A deficiency on the pathogenicity of C. muridarum in the genital tract, BALB/c mice were intravaginally inoculated with wild-type, plasmid-free and glycogen synthase A-deficient C. muridarum, and the genital tract tissue was isolated to assess the severity of hydrosalpinx and the levels of oviduct dilatation at day 60 after infection. The glycogen storage capacity and in vitro infection ability of different C. muridarum strains were analyzed by periodic acid-Schiff staining and quantification of progeny elementary body(EB) formation. The tissue homogenate was used to determine the recovery of different C. muridarum strains. The results show that glycogen synthase A-deficient C. muridarum induced reduction of hydrosalpinx and attenuated the extent of oviduct dilatation in mice, and exhibited reduced growth and proliferation in the mouse lower genital tract. In addition, glycogen synthase A point mutations at different sites reduced the glycogen storage capacity and in vitro infectivity of C. muridarum to different degrees. Glycogen synthase A deficiency also reduced the host inflammatory reaction and ascending infection of C. muridarum.

Identification of differentially expressed circular RNAs in HeLa cells infected with Chlamydia trachomatis.

Non-coding circular RNAs (circRNAs) have been shown to have important roles in many diseases; however, no study has indicated circRNAs are involved in Chlamydia trachomatis infection. In this study, we used circRNA microarray to measure the global circRNA expression profiles in HeLa cells with or without C. trachomatis serovar E (Ct.E) infection. CircRNA/miRNA/mRNA interactions were predicted and bioinformatics analyses were performed. The differentially expressed circRNAs were selected according to our criterion for validation by reverse-transcription and quantitative polymerase chain reaction (RT-qPCR). The mRNA microarray was used to detect the mRNA expression profiles after Ct.E infection. Among 853 differentially expressed circRNAs, 453 were upregulated and 400 were downregulated after Ct.E infection. Target miRNAs and miRNA-targeted mRNAs of these circRNAs were predicted. RT-qPCR analysis indicated hsa_circRNA_001226, hsa_circRNA_007046 and hsa_circRNA_400027 were elevated similar to those determined in the circRNA microarray analysis. The mRNA microarray results showed 915 genes were upregulated and 619 genes were downregulated after Ct.E infection. Thirty-four differentially expressed genes overlapped in the bioinformatics and mRNA microarray results. KEGG pathway analysis revealed several signaling pathways, including endocytosis, MAPK and PI3P-Akt signaling pathways, that were targeted by circRNAs may play important roles in Chlamydia infection. This study provides evidence that circRNAs in host cells are involved in the process of Chlamydia infection.

Decitabine enhances bortezomib treatment in RPMI 8226 multiple myeloma cells.

The present study investigated the interactions between decitabine (DAC) and bortezomib (BTZ) in RPMI 8226 multiple myeloma (MM) cells. Cells were exposed to DAC alone and in combination with BTZ for 48 h. A Cell Counting Kit­8 assay was performed to assess the rate of proliferation inhibition in the cells. Cell apoptosis was investigated by Annexin V-fluorescein isothiocyanate and propidium iodide staining. Flow cytometry was used to detect the different cell cycle stages. Western blotting was performed to analyze the protein expression levels of poly(ADP­ribose) polymerase 1 (PARP­1), caspase­3, ­9 and DNA (cytosine­5­)­methyltransferase 1 (DNMT1). Reverse transcription­quantitative polymerase chain reaction was used to assess DNMT1 gene expression. The combination of DAC and BTZ increased the proliferation inhibition, apoptotic rate and G0­G1 arrest compared with use of a single therapeutic agent. In addition, the combination treatment enhanced PARP­1 cleavage, caspase­3 and caspase­9 activation and downregulated the protein and mRNA expression levels of DNMT1. Therefore, the current study determined that the combination of BTZ and the epigenetic agent DAC may be a novel therapeutic strategy to improve the efficacy of BTZ in patients with MM.

[Comparison of allelic frequencies of 15 short tandem repeat loci between chronic myeloid leukemia patients and non-related healthy individuals].

OBJECTIVE: To compare allelic frequencies of 15 short tandem repeat (STR) loci (D8S1179, D21S11, D7S820, CSF1PO, D3S1358, TH01, D13S317, D16S539, D2S1338, D19S433, vWA, TPOX, D18S51, D5S818 and FGA) between chronic myeloid leukemia (CML) patients and non-related healthy individuals from Changzhou region in order to predict genes related with the CML. METHODS: Blood samples were collected from 745 healthy subjects and 132 CML patients with complete remission. Genotypes were determined with gene scan technology and multiplex PCR with fluorescence-labeled primers. Allelic polymorphisms of 15 STR loci were compared between the two groups. Potential genes related with CML were predicted with statistical analysis of differences in allelic frequencies. RESULTS: Allelic frequencies of 3 loci, including CSF1PO, vWA and TPOX, showed a significant difference (P<0.05) between the two groups. CONCLUSION: CSF1PO, vWA and TPOX loci may be related with CML, albeit that the exact biologic mechanisms is unclear.

[Effects of simvastatin on PI3K/AKT signaling pathway in human acute monocytic leukemia cell line SHI-1].

To investigate the effects of 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor simvastatin (SV) on proliferation, apoptosis and the PI3K/AKT signaling pathway in human acute monocytic leukemia cell line SHI-1. SHI-1 cells were incubated with different concentrations of SV (5, 10, 15 µmol/L). Otherwise, SHI-1 cells without any treatment were used as control. Cells in different groups were collected at 24, 48 and 72 h after incubation for further detection. MTT method was used to assay the growth inhibition rate and flow cytometry was used to detect the early stage apoptosis ratio. The human PI3K-AKT Signaling Pathway RT(2) Profiler(TM) PCR Array was used to detect the expression of 84 genes involved in PI3K-AKT signaling. The results indicated that the SV inhibited the proliferation and inducted the apoptosis of SHI-1 cells in time- and dose-dependent manners significantly. The growth inhibition rates of SHI-1 cells treated with 15 µmol/L SV for 24, 48 and 72 h were 26.82, 47.09 and 63.92, respectively; and their early stage apoptosis ratios were 5.75, 13.25 and 15.59, respectively. Compared with the control group, expression levels of 39 genes were changed in the group of 15 µmol/L SV at 48 h, among them 26 genes were down-regulated and 13 genes were up-regulated. It is concluded that the SV can inhibit proliferation and induce apoptosis of SHI-1 cells, and the mechanism may be associated with the changes of gene expression level in PI3K-AKT signaling pathway regulated by SV.

[Effects of simvastatin plus all-trans retinoic acid on WT1/hDMP1 gene expression profiles of human promyelocytic leukemia cell line NB4].

OBJECTIVE: To investigate the effects of simvastatin (SV) plus all-trans retinoic acid (ATRA) on the proliferation, differentiation, apoptosis and WT1/hDMP1 gene expression profiles of human promyelocytic leukemia cell line NB4. METHODS: The NB4 cell was incubated with simvastatin and ATRA alone or in combination. And the NB4 cell without any treatment was adopted as a normal control. The cells of different groups were collected at 24, 48 and 72 h post-incubation. Their morphological changes were observed after Wright staining. The method of MTT was employed to assay the growth inhibition rate and flow cytometry was used to detect the early-stage ratios of apoptosis and cell necrosis. Real-time quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) was used to detect the WT1/hDMP1 gene expression levels. RESULTS: The cell inhibition rates increased gradually (F = 7.15, P = 0.000) at 15, 10 and 5 µmol/L SV respectively. And so did the expression levels of CD11b (F = 3.41, P = 0.014) and Annexin-V (F = 43.38, P = 0.000). However the expression levels of WT1 decreased gradually (F = 5.35, P = 0.001) reversely with the elevated levels of hDMP1 (F = 22.61, P = 0.000). Furthermore the NB4 cell exhibited the most significant changes at 15 µmol/L SV. After a 72-hour incubation, the expression levels of CD11b (89.46% ± 9.13%)and hDMP1 (626.9 ± 56.9) in NB4 cells at 15 µmol/L SV plus 0.5 µmol/L ATRA were significantly higher than those with ATRA(71.27% ± 7.27%, P = 0.000 and 421.8 ± 38.3, P = 0.003 in each) and SV alone(62.41% ± 6.37%, P = 0.003 and 241.4 ± 21.9, P = 0.003 in each). A combination of 15 µmol/L SV with 0.5 µmol/L ATRA displayed obvious interactions with the expressions of CD11b and hDMP1 (F = 4.09, P = 0.025 and F = 29.58, P = 0.000 in each). And there was no significant interaction for cell inhibition rates and Annexin-V expression. CONCLUSION: Simvastatin in vitro inhibits the proliferation of NB4 cell, induces its differentiation and promotes its apoptosis. And the lowered expression of WT1 has a dose-dependent correlation with the elevated expression of hDMP1. It indicates that simvastatin has the synergistic in vitro anti-promyelocytic potency.

[The comparison of inducing specific CTL by two methods of leukemia cell antigen loading of DCs and fusion of DCs with leukemia cells].

AIM: To compare the potency of the fusion of DCs with leukemia cells and freeze-thawed leukemia antigen-loading DCs in inducing antigen-specific CTLs. METHODS: Peripheral blood mononuclear cells (PBMC) isolated by HES-Ficoll two-step method were incubated 2 hours to select adherent monocytes. The adherent monocytes were then cultured in the presence of GM-CSF and IL-4 for 5 days and then divided into four groups: DCs were fused with K562 cells or CML cells from CML patients in the presence of 500 g/L PEG-100 mL/L DMSO (Group A), DCs were loaded with lysates from K562 cells or CML cells (group B), DCs were co-cultured with K562 cells or CML cells (group C) and DCs were cultured alone (group D). Before cell fusion, K562 cells were labeled using a red fluorescent dye, PKH26. After fusion, flow cytometry was used to detect hybrids labeled by both PKH26 and FITC-conjugated anti-HLA-ABC mAb to assess the efficiency of fusion. On day 6, TNF-alpha was added to induce the terminal maturation of DCs. Then each group DCs were co-cultured with autologous T cells respectively. The cytotoxicity of CTLs of each group against different target cells was measured using MTT coloremetry. RESULTS: DCs induced by GM-CSF+IL-4 and TNF-alpha had DC-classical phenotypic characteristics. The efficiency of cell fusion ranged from 17.33% to 29.94%. Both DC-K562 hybrids and DC loaded with leukemic freeze-thaw lysates could stimulate specific CTL responses against target cells. Furthermore, the cytotoxicity induced by the former DCs were much stronger than that induced by lysates-loaded DCs at the same E/T ratio. CONCLUSION: Compared with freeze-thaw lysates loaded DCs, DC-leukemic hybrids showed higher efficiency in antigen presentation and the stimulation of leukemia-specific CTLs.

[In vitro stimulation of specific antileukemia T-cell response by dendritic cells derived from CD14+ acute monocytic leukemia cells].

BACKGROUND & OBJECTIVE: Dendritic cells (DCs) or DC-like cells had been successfully induced in vitro from leukemia cells, which may provide a promising immunotherapeutic protocol for leukemia. This study was designed to investigate the efficiency of in vitro generation of dendritic cells from CD14+ acute myelomonocytic (M4) or monocytic (M5) leukemia cells and their ability of stimulating specific antileukemia T-cell response. METHODS: Bone marrow mononuclear cells (BMMNCs) were isolated from 5 M4/M5 leukemia patients with high CD14 expression, and then divided into 3 groups: adherent leukemia cells, nonadherent blasts, and total unfractioned blasts. CD14 expression of the 3 groups was evaluated by flow cytometry (FCM). When cultured with or without granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-4 (IL-4) and tumor necrosis factor alpha (TNF-alpha) for 7-10 days, monocytic leukemia cell-derived dendritic cells (Mo-LDCs) were identified through morphologic observation and immunophenotype analysis using FCM. The immune function of Mo-LDCs was detected through allogeneic mixed lymphocyte reaction (Allo-MLR) and cytotoxicity assay of antileukemia cytotoxic T lymphocytes (CTLs). The leukemic origin of Mo-LDCs was confirmed by chromosomal karyotype analysis combined with the aberrant expression of myeloid antigens. RESULTS: The amount of CD14+ cells, which could differentiate into CD83+ mature DCs under induction of the cytokine combination, was higher in adherent leukemia cells than in nonadherent blasts and total unfractioned blasts. Regarding each 3 cell groups of the same patient or the unfractioned blasts of various patients, initial CD14 expression was positively related to the yield of CD83+ DCs after induction (r = 0.967, P = 0.007). Mo-LDCs exhibited typical morphology and phenotype as mature DCs, induced potent proliferation of homogeneous T cells in Allo-MLR, stimulated the expansion of leukemia-specific CTLs, and continued to possess the cytogenetic abnormalities of the original leukemia, as well as the aberrant expression of myeloid antigens. CONCLUSIONS: In M4/M5 subtype of AML, CD14+ cells could differentiate into immune-competent Mo-LDCs under the induction of the cytokine combination. CD14 expression level may predict the DCs differentiation ability of monocytic leukemia. Mo-LDCs, which possess the classical phenotype and function of DCs, as well as the abnormal leukemic antigens, may be useful for the immunotherapy of M4/M5 AML.