scholarly journals Inhibition of Colon Cancer in Mice by Microencapsulated Probiotic

2016 ◽  
Author(s):  
◽  
Frederick Oluwasheyi Odun-Ayo
Keyword(s):  
Colon Cancer ◽  
Mouse Model ◽  
Dietary Fibre ◽  
Drug Carrier ◽  
Control Group ◽  
Rrna Gene ◽  
High Morbidity ◽  
Citrus Pectin ◽  
Probiotic Treatment

Colon cancer is the third most common cancer worldwide with a high morbidity and mortality rate. Therapies are less effective during metastasis, therefore prevention and earlier detection is key to reducing the risk of colon cancer. Increased dietary fibre and probiotic intake is known to lower the risk of colon cancer. Probiotics are defined as “live microorganisms which when administered orally in an adequate amount confer a health benefit on the host”. The International Dairy Federation recommends a viable minimum level of 6–7 log10cfu/g in a probiotic product being consumed. Different biopolymer matrices have been used for encapsulation of probiotics; however, loss of viability is still a major challenge. Citrus pectin is a dietary fibre polysaccharide broken down into smaller fragments to form modified citrus pectin (MCP). The unique bioactivity of MCP against carcinogenesisis is linked to its sugar β-galactose inhibiting the cell signalling protein marker, galectin-3 (gal-3), which is intimately involved in endothelial cell morphogenesis. The vascular endothelial growth factor (VEGF) signalling, which invariably drives angiogenesis can be activated when gal-3 binds to integrins. The bioactivity and uptake of MCP may be improved through a novel approach if conjoined with a supplement for example probiotic. Therefore, the synergistic inhibitory effect of modified citrus pectin alginate (MCPA) probiotic microbeads on gal-3 and VEGF in an azoxymethane (AOM) induced colon carcinogenesis Balb/c mouse model was investigated. A microencapsulation process was used to produce a MCPA microbead containing probiotic, Lactobacillus acidophilus ATCC 4356. Efficiency of the microbead was evaluated in vitro (simulated conditions) and in vivo (Balb/c mouse model). Genomic identification of faecal lactobacilli samples from the treated mice was analyzed. Optimization of AOM dose-time with 10 and 15 mg/kg AOM intraperitoneal (ip) administered to Balb/c mice for 2 and 4 weeks were performed. The optimal AOM dose was initiated prior to intake of MCPA, AP (alginate calcium) probiotic microbeads and MCP in Balb/c mice for 16 weeks; samples were analyzed for colon histopathology and immunohistochemistry. The MCPA probiotic microbeads significantly enhanced the viability of L. acidophilus ATCC 4356 compared to the AP microbeads in vitro (p< 0.05). Exposure of the MCPA probiotic microbeads to 3 h of simulated gastric juice (SGJ) resulted in 82.7% survival of L. acidophilus ATCC 4356. Also, the faecal lactobacilli count in the MCPA probiotic treated mice significantly increased after 28 days by 10.2% compared to the AP probiotic, MCP treated and control mice (p< 0.0001). A total of 4DNA encoding 16S rRNA gene closest to the genera namely Lactobacillus, Bacillus, Enterococcus and Bifidobacterium were identified from faecal samples of the colon cancer-induced Balb/c mice. Azoxymethane at 15 mg/kg for 4 weeks induced optimal gal-3 and VEGF immunoexpression. Furthermore, MCPA probiotic treatment significantly reduced gal-3 immunoexpression in the colon of AOM induced cancer Balb/c mice compared to the control mice (p< 0.0001). The immunoexpresion of VEGF in the MCPA and AP probiotic treated groups was weakly positive and significantly reduced when compared to the control group (p<0.05), while the MCP treated group was barely positive (p< 0.001). Modified citrus pectin alginate is a novel effective means of oral delivery of bacterial cells and bioactive compounds. It has a good biodegradability, inexpensive, non-toxic, proven efficiency, and stability at low temperatures warranting its use as a drug carrier by pharmaceuticals. Modified citrus pectin alginate probiotic microbeads increase bioactivity and chemoprevention against colon pre-cancerous lesions and adenocarcinoma through inhibition of gal-3 and VEGF in the mouse model. Modified citrus pectin alginate can be used in probiotic therapy, which may improve the prevention of colon cancer.

scholarly journals Taurine Attenuates Carcinogenicity in Ulcerative Colitis-Colorectal Cancer Mouse Model

2020 ◽  
Vol 2020 ◽  
pp. 1-7 ◽  
Author(s):  
Guifeng Wang ◽  
Ning Ma ◽  
Feng He ◽  
Shosuke Kawanishi ◽  
Hatasu Kobayashi ◽  
...  
Keyword(s):  
Colon Cancer ◽  
Drinking Water ◽  
Mouse Model ◽  
Histopathological Examination ◽  
Tumor Formation ◽  
Water Model ◽  
Control Group ◽  
Ki 67

Taurine (2-aminoethane-sulfonic acid) is a type of amino acids and has numerous physiological and therapeutic functions, including anti-inflammation. However, there are few studies on the anticancer action of taurine. Our previous studies have demonstrated that taurine exhibits an apoptosis-inducing effect on human nasopharyngeal carcinoma cells in vitro. In this study, we have investigated whether taurine has an anticancer effect, using azoxymethane (AOM)/sulfate sodium (DSS)- induced mouse model for colon carcinogenesis. All mice, except those in control group, received a single intraperitoneal injection of AOM and DSS in the drinking water for 7 days twice, with 1-week interval. After the first DSS treatment, mice were given distilled water (model group) or taurine in the drinking water (taurine group) ad libitum. No tumor was observed in the control group. Taurine significantly suppressed AOM+DSS-induced tumor formation. Histopathological examination revealed AOM/DSS treatment induced colon cancer in all mice (8/8, 100%), and taurine significantly inhibited the progression of colon cancer (4/9, 44.4%). Taurine significantly attenuated cell proliferation in cancer tissues detected by Ki-67 staining. Taurine significantly increased the levels of an apoptosis marker cleaved caspase-9 and tumor suppressor protein PTEN. This is the first study that demonstrated that taurine significantly reduced carcinogenicity in vivo using AOM/DSS-induced colon cancer mouse model.

scholarly journals Cell-Free Supernatant Odoribacter splanchnicus Isolated From Human Feces Exhibits Anti-colorectal Cancer Activity

2021 ◽  
Vol 12 ◽  
Author(s):  
Byeong Seob Oh ◽  
Won Jung Choi ◽  
Ji-Sun Kim ◽  
Seoung Woo Ryu ◽  
Seung Yeob Yu ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Colon Cancer ◽  
Mouse Model ◽  
Proliferative Activity ◽  
Control Group ◽  
Healthy Individuals ◽  
Gut Microbes ◽  
Cancer Activity

The gut microbiota (GM) has been shown to be closely associated with the development of colorectal cancer (CRC). However, the involvement of GM is CRC has mainly been demonstrated by metagenomic profiling studies showing the compositional difference between the GM of healthy individuals and that of CRC patients and not by directly studying isolated gut microbes. Thus, to discover novel gut microbes involved in CRC, we isolated the GM from the feces of healthy individuals and evaluated its anti-CRC activity in vitro and in vivo. After GM isolation, cell-free supernatants (CFSs) were prepared from the isolated gut microorganisms to efficiently screen a large amount of the GM for anti-proliferative ability in vitro. Our results showed that the CFSs of 21 GM isolates had anti-proliferative activity against human colon cancer HCT 116 cells. Of these 21 GM isolates, GM07 was chosen for additional study because it had the highest anti-cancer activity against mouse colon cancer CT 26 cells in vitro and was further evaluated in a CT 26 allograft mouse model in vivo. GM07 was identified as Odoribacter splanchnicus through phylogenetic analysis based on 16S rRNA gene sequencing. Further investigation determined that the CFS of O. splanchnicus (OsCFS) induced anti-proliferative activity via apoptosis, but not cell cycle arrest. Moreover, GC/MS analysis suggested that the putative active molecule in OsCFS is malic acid. Finally, in the CRC mouse model, peri-tumoral injection of OsCFS significantly decreased CRC formation, compared to the control group. Altogether, these findings will provide valuable information for the discovery of potential probiotic candidates that inhibit CRC.

Reduction of Factor VIII Inhibitor Formation with F.VIII-Pulsed Tolerogenic Dendritic Cells in the Murine Hemophilia A Model.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 213-213 ◽  
Author(s):  
Margaret V. Ragni ◽  
Wenhu Wu ◽  
Xiaoyan Liang ◽  
Lina Lu
Keyword(s):  
Bone Marrow ◽  
Dendritic Cells ◽  
T Cell ◽  
Hemophilia A ◽  
Binding Activity ◽  
Control Group ◽  
High Morbidity ◽  
Gm Csf

Abstract Inhibitor formation is a severe complication of hemophilia, occurring in up to 25% and associated with poor response to factor replacement, uncontrolled bleeding, and high morbidity. Preventing inhibitor formation is, thus, a major goal of hemophilia management. The role of dendritic cells (DC) in regulating immune response has been increasingly recognized: immature DC (imDC) induce T regulatory cells in vitro and promote Ag-specific tolerance in vivo. We, therefore, studied the role of imDC propagated from bone marrow with GM-CSF + TGFβ to prevent inhibitor formation in the hemophilia A murine model. Following tail vein injection of recombinant F.VIII (Advate, Baxter) 2.5 U (0.2 μg) on days 0, 2, and 4 in hemophilia A exon 16 KO C57Bl/6 mice, anti-VIII antibodies were detected by semi-quantitative APTT (scored 1-4), peaking on day 6. On rechallenge with F.VIII 2.5 U on days 12, 14, and 16, anti-VIII was detected, peaking on day 17. Anti-VIII production was associated with high level splenic T cell proliferation in response to F.VIII stimulation in vitro, measured by 3H-thymidine incorporation in mixed lymphocyte reaction (MLR). By contrast, there was no antibody formation in F.VIII-treated Wt C57Bl/6 mice: the latter was associated with low T cell response to F.VIII in vitro. Functionally immature DC (imDC) were propagated from the bone marrow of hemophilia A mice with GM-CSF (4ng/ml) and TGFβ (0.2ng/ml). For comparison, functionally mature dendritic cells (mDC) were propagated with GM-CSF (4ng/ml) and IL-4 (1000U/ml).The former (imDC) demonstrated deficient NF-kB binding activity in nuclear protein as detected by gel shifting assay and expressed low level of costimulatory molecules CD80, CD86; by contrast, the latter (mDC) demonstrated enhanced NF-kB binding activity and high levels of co-stimulatory molecules. Administration of 2x106 F.VIII-pulsed imDC (20U/ml x 24h) 7 days before F.VIII dosing on days 0, 2, and 4, led to reduction in inhibitor formation on day 6 (score 1.6 vs. 2.3 in control group) which was further reduced on day 8 (score 1.0 vs. 2.0 in control group). The inhibitor could not be detected on day 8 in 2 of 4 mice pretreated with F.VIII-pulsed imDC. By contrast, high levels of inhibitor were detected in mice pretreated with F.VIII-pulsed mDC (score 3.3). Rechallenge with F.VIII on day 10 in imDC-treated mice resulted in no increase in the reduced or absent anti-VIII effect on day 12. Splenic T cells (CD3+) from the imDC-pretreated mice showed lower proliferative capacity when restimulated in vitro with F.VIII, suggesting that imDC induced F.VIII unresponsiveness. These studies show that FVIII-pulsed imDC reduce the intensity of inhibitor formation, and suggest the potential role of modified DC in preventing or reducing F.VIII inhibitor formation.

FXIIa Differentially Regulates Thrombin Generation during Steady State and Vaso-Occlusive Crisis in Sickle Cell Mice

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 162-162 ◽  
Author(s):  
Erica M Sparkenbaugh ◽  
Camille Faes ◽  
Denis Noubouossie ◽  
Daniel K. Kirchhofer ◽  
András Gruber ◽  
...  
Keyword(s):  
Steady State ◽  
Mouse Model ◽  
Vascular Permeability ◽  
Sickle Cell ◽  
Plasma Levels ◽  
Thrombin Generation ◽  
Conflicts Of Interest ◽  
Control Group ◽  
Liver And Kidney

Abstract Sickle cell disease (SCD) is associated with chronic activation of coagulation. Previously, we demonstrated that inhibition of tissue factor (TF) attenuates thrombin generation (measured by plasma levels of thrombin-antithrombin complexes [TAT]) in a mouse model of SCD during steady state. Furthermore, we showed that neither inhibition of FXIIa-dependent activation of FXI (using 14E11 antibody) nor FXI deficiency reduces thrombin generation (TG) in sickle mice. In contrast, genetic deficiency of FXII or kininogen (HK) reduced plasma TAT levels. These data suggest that during steady state, FXIIa contributes to TG in sickle mice via activation of the kallikrein/HK pathway, but not FXI. In the present study, we further investigated the mechanisms of HK-induced TG at steady state, and increased TG observed during vaso-occlusive crisis (VOC). All experiments were performed using 4-5 month old Townes SS (sickle) and AA (control) mice. Kallikrein cleaves HK into HK fragments (HKFs) and bradykinin (BK). First, we investigated whether a BK-mediated increase in vascular permeability contributes to TG by exposing perivascular TF. This hypothesis was disproved by data demonstrating no difference in vascular permeability (measured by the extravasation of Evans blue in the heart, lung, liver and kidney) between AA (n=8) and SS (n=10) mice. HKFs were shown to induce leukocyte TF expression in vitro via binding to CD11b/CD18 (Mac-1). Therefore, we investigated whether Mac-1 inhibition affects TG in SS mice. AA and SS mice were treated with an inhibitory anti Mac-1 (M1/70) or IgG control antibody on days 0, 3 and 6 (i.p. 1 mg/kg) and TG was analyzed 1 day after the last injection. In the control group, SS mice demonstrated higher plasma TAT levels compared to AA mice (8.1±1.6 vs 4.2±0.6 ng/mL, n=10-11, p<0.05), but inhibition of Mac-1 significantly reduced plasma TAT levels in SS mice (4.6±0.7 ng/mL, n=11, p<0.05). These data suggest that HK might contribute to TG during steady state via Mac-1-dependent induction of monocyte TF. The steady state of SCD is interspersed with acute periods of VOC. Clinical data demonstrate that compared to the steady state, plasma levels of cell free DNA (cfDNA), activation of the contact system, and TG are further enhanced during VOC. To determine the mechanism of increased TG during VOC, we used the previously characterized mouse model of TNFα -induced VOC. Townes AA and SS mice were injected with recombinant TNFα (2 µg/g body weight) or the same volume of PBS, and plasma was collected 5 hours later. TNFα not only dramatically increased plasma levels of cfDNA in SS mice (14.78 ± 1.64 vs 679 ± 300 ng/mL; p<0.01), but also further increased plasma TAT levels compared to those observed in PBS-treated SS mice (2.9 fold, p<0.001, n=8). Importantly, there was a significant positive correlation between cfDNA and TAT in SS mice (r2 =0.65, p<0.001). Since cfDNA can activate FXII, we determined whether FXIIa-dependent activation of FXI contributes to TG during VOC. AA and SS mice received 14E11 or IgG control (4 mg/kg) 30 minutes before TNFα (2 μg/g) or PBS injection, and plasma TAT was assessed 5 hours later. Strikingly, 14E11 attenuated the increased TAT level in TNFα-treated SS mice, to the level observed in SS mice injected with PBS and IgG (IgG/SS/PBS: 9 ng/mL ± 1.8 vs. IgG/SS/TNF: 18.9 ± 3.6, p<0.001; 14E11/SS/TNF: 9.86 ± 0.72, p<0.05 vs. IgG/SS/TNF). We also determined if TF activity is required for the increased TG observed during VOC. Interestingly, inhibition of TF with an inhibitory 1H1 antibody (25 or 75 mg/kg injected i.p. 1 or 18 hours prior to TNFα, respectively) had no effect on the increased TG observed in TNFα treated SS mice. In aggregate, our data suggest that during the steady state of SCD, FXII-dependent TG is not FXI-dependent, but instead is mediated by a pathway involving HK, Mac-1 integrin and leukocyte TF. Furthermore, we propose that during VOC the massive release of cfDNA results in FXIIa-dependent FXI activation and enhances TG independently of TF. This study provides mechanistic insight into the initiators of TG in SCD. Moreover, it implicates FXIIa as a potential therapeutic target to reduce the prothrombotic state in SCD, during both steady state and VOC. Disclosures No relevant conflicts of interest to declare.

scholarly journals Evaluation of recombinant Brachyspira pilosicoli oligopeptide-binding proteins as vaccine candidates in a mouse model of intestinal spirochaetosis

2010 ◽  
Vol 59 (3) ◽  
pp. 353-359 ◽  
Author(s):  
Abdolreza Movahedi ◽  
David J. Hampson
Keyword(s):  
Mouse Model ◽  
Binding Proteins ◽  
Control Group ◽  
Cumulative Number ◽  
Igg Antibody ◽  
Brachyspira Pilosicoli ◽  
Distribution Study ◽  
Different Strains

The anaerobic intestinal spirochaete Brachyspira pilosicoli colonizes the large intestine of humans, and various species of animals and birds, in which it may induce a mild colitis and diarrhoea. The aim of the current study was to evaluate the use of putative oligopeptide-binding proteins of B. pilosicoli as vaccine components. A partial genome sequence of B. pilosicoli porcine strain 95/1000 was subjected to bioinformatics analysis, and six genes predicted to encode oligopeptide-binding proteins were selected. Following a PCR-based distribution study of the genes across different strains of the spirochaete, they were amplified from B. pilosicoli human strain WesB and cloned in Escherichia coli. The recombinant histidine-tagged proteins were purified and subjected to in vitro and in vivo immunogenicity analysis. Recombinant products (P-1 and P-3) from two genes that were immunogenic and recognized by sera from pigs that had recovered from B. pilosicoli infections were tested in a mouse model of intestinal spirochaetosis. For each recombinant protein, groups of 12 C3H/HeJ mice were vaccinated subcutaneously with 100 μg protein emulsified in Freund's incomplete adjuvant, twice with a 2 week interval. Two weeks later the vaccinated and non-vaccinated control animals were challenged orally with B. pilosicoli strain WesB. Both proteins induced systemic and local colonic IgG antibody responses, and, following experimental infection, the cumulative number of colonization days was significantly (P<0.001) less in both groups of vaccinated mice compared to the control mice. There were significantly (P=0.012) fewer mice colonized in the group vaccinated with P-1 than in the non-vaccinated control group. The results suggest that oligopeptide-binding proteins may have potential for use as components of vaccines for B. pilosicoli.

scholarly journals Potency of omadacycline against Mycobacteroides abscessus clinical isolates in vitro and in a mouse model of pulmonary infection

2021 ◽  
Author(s):  
Danielle A. Nicklas ◽  
Emily C. Maggioncalda ◽  
Elizabeth Story-Roller ◽  
Benjamin Eichelman ◽  
Chavis Tabor ◽  
...  
Keyword(s):  
Mouse Model ◽  
Opportunistic Pathogen ◽  
Current Treatment ◽  
Clinical Isolates ◽  
Control Group ◽  
Chronic Infections ◽  
Human Dose ◽  
Time Kill

The incidence of nontuberculous mycobacterial diseases in the US is rising and has surpassed tuberculosis. Most notable among the nontuberculous mycobacteria is Mycobacteroides abscessus , an emerging environmental opportunistic pathogen capable of causing chronic infections. M. abscessus disease is difficult to treat and the current treatment recommendations include repurposed antibiotics, several of which are associated with undesirable side effects. In this study, we have evaluated the activity of omadacycline, a new tetracycline derivative, against M. abscessus using in vitro and in vivo approaches. Omadacycline exhibited an MIC 90 of 0.5 μg/ml against a panel of 32 contemporary M. abscessus clinical isolates several of which were resistant to antibiotics that are commonly used for treatment of M. abscessus disease. Omadacycline when combined with clarithromycin, azithromycin, cefdinir, rifabutin or linezolid also exhibited synergism against several M. abscessus strains and did not exhibit antagonism when combined with an additional nine antibiotics also commonly considered to treat M. abscessus disease. Concentration-dependent activity of omadacycline was observed in time-kill assessments. Efficacy of omadacycline was evaluated in a mouse model of lung infection against four M. abscessus strains. A dose equivalent to the 300 mg standard oral human dose was used. Compared to the untreated control group, within four weeks of treatment, 1 to 3 log 10 fewer M. abscessus colony forming units were observed in the lungs of mice treated with omadacycline. Treatment outcome was biphasic, with bactericidal activity observed after the first two weeks of treatment against all four M. abscessus strains.

scholarly journals Selection of probiotic bifidobacteria for lambs

2009 ◽  
Vol 54 (No. 12) ◽  
pp. 552-565 ◽  
Author(s):  
E. Vlková ◽  
M. Grmanová ◽  
V. Rada ◽  
I. Homutová ◽  
S. Dubná
Keyword(s):  
Antimicrobial Activities ◽  
Control Group ◽  
In Vitro Test ◽  
Faecal Samples ◽  
Probiotic Treatment ◽  
Wild Strains ◽  
Vitro Test ◽  
Gradient Plate ◽  
Selection Of

Twenty-six bifidobacteria were isolated from faecal samples of lambs. The isolates were identified, functional properties (survival ability at low pH and bile conditions) and antimicrobial activities against potential pathogens were determined. From the isolates with suitable properties (13 strains) rifampicin-resistant mutants were prepared by gradient plate techniques. This property enabled us to differentiate the administered organism from wild strains because resistance to rifampicin is rare among bifidobacteria. Rifampicin-resistant bifidobacteria (RRBifs) were administered to 3-days-old lambs in two trials. In the first trial the strain <i>B. ruminantium</i> L29 was applied to 3 lambs and was detected in faecal samples at high counts (6 log CFU/g on average) for one week. In the second trial 3 lambs received a “cocktail” of 12 strains and RRBifs survived in the intestinal tract at counts of about 6 log CFU/g for 25 days. The control group without probiotic treatment consisted of 6 animals. In both treated groups RRBifs dominated among bifidobacteria after their administration. Total bifidobacterial counts (5.64–7.32 log CFU/g) were significantly higher (<i>P</i> < 0.05) in treated groups compared to 2.31–2.85 log CFU/g detected in the control group during the first month of lamb life. Lactobacilli counts were also significantly higher (<i>P</i> < 0.05) in treated groups compared to the control. The administered bifidobacteria did not affect any other monitored bacterial groups. On the basis of in vitro test results, suitable probiotic bifidobacterial strains for lambs were chosen. Some of them survived for 30 days in the gastrointestinal tract of treated lambs, but no tested strain was able to colonise the lamb’s tract permanently. The administration of bifidobacterial “cocktail” and consequent identification of the best survived strain seems to be an effective method for selection of potential probiotics.

Preclinical Immunogenicity Assessment of Baxter‘s Longer-Acting FVIII Candidate BAX 855 Using Novel Preclinical Models,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3323-3323
Author(s):  
Frank M Horling ◽  
Sandra Schwele ◽  
Christian Lubich ◽  
Rafi U Ahmad ◽  
Markus Weiller ◽  
...  
Keyword(s):  
Immune System ◽  
Mouse Model ◽  
Mouse Models ◽  
Mhc Class Ii ◽  
Immune Tolerance ◽  
Class Ii ◽  
Control Group ◽  
Potential Impact ◽  
Buffer Control

Abstract Abstract 3323 Baxter is developing a PEGylated recombinant human factor VIII conjugate (BAX 855) based on the modification of full-length FVIII with polyethylene glycol (PEG). The FVIII molecule is derived from a CHO cell line using a plasma-protein-free method. Any chemical modification of FVIII might alter its immunogenic potential. Therefore, careful preclinical immunogenicity studies of PEGylated FVIII in comparison to unmodified FVIII are important. We evaluated the immunogenicity of BAX 855 in comparison to ADVATE, Baxter‘s unmodified recombinant full-length FVIII concentrate. For this purpose, we used different in vitro and in vivo models to s assess the potential impact of BAX 855 and ADVATE on both the innate and the adaptive immune system. The assessment of the potential impact of BAX 855 and ADVATE on the human innate immune system was based on their potential to induce the release of pro- inflammatory cytokines (IL-1β, IL-6, IL-8 and TNF-α) in an in vitro human whole blood assay and on its potential to activate the human complement system in human plasma in vitro. The assessment of the potential impact of BAX 855 and ADVATE on the adaptive immune system was based on their potential to induce antibodies in 3 different hemophilic mouse models and in cynomolgous monkeys. The hemophilic moue models included a hemophilic mouse model with a knockout of the murine FVIII that express a human F8 cDNA as a transgene, a hemophilic mouse model that expresses the human MHC-class II protein HLA-DRB1*1501 on the background of a knockout of the murine MHC-class II complex and a conventional hemophilic mouse model with a knockout of the murine FVIII gene. Hemophilic mice that express a human F8 cDNA as a transgene are immunologically tolerant to human FVIII and develop antibodies against human FVIII only if the immune tolerance breaks down. Using this model, we asked if BAX 855 is able to maintain immune tolerance to human FVIII. The hemophilic mouse models that expresses the human MHC-class II protein HLA-DRB1*1501 mimics the situation in an important fraction of hemophilia A patients with FVIII inhibitors. The human MHC-class II haplotype HLA-DRB1*1501 was previously shown to be associated with an increased risk for patients to develop FVIII inhibitors. Using this model, we asked if BAX 855 expresses an immunogenicity profile similar to ADVATE. BAX 855 and ADAVTE induced similar low levels of cytokine release (IL-1β, IL-6, IL-8 and TNF-α) that were similar to the cytokine release observed in the buffer control group after incubation with human whole blood in vitro. In addition, BAX 855 and ADVATE induced similar low levels of complement activation that were not different from the buffer control group after incubation with human plasma in vitro. Importantly, BAX 855 and ADVATE induced similar levels and incidences of antibodies against FVIII and against PEG-FVIII in all mouse models. In addition, immune tolerance to FVIII was maintained in hemophilic mice expressing the human F8 cDNA. Finally, no major differences in antibody titers were seen after treatment of cynomolgus monkeys with BAX 855 and ADVATE. We conclude that results obtained in comparative immunogenicity studies in vitro and in vivo demonstrate that BAX 855 and ADVATE have a similar immunogenicity profile. Therefore, we expect that BAX 855 will express a similar safety profile as ADVATE in patients. Disclosures: Horling: Baxter Innovations GmbH: Employment. Schwele:Baxter Innovations GmbH: Employment. Lubich:Baxter BioScience: Employment. Ahmad:Baxter Innovations GmbH: Employment. Weiller:Baxter BioScience: Employment. Spatzenegger:Baxter Innovation GmbH: Employment. Schwarz:Baxter Innovations GmbH: Employment. Reipert:Baxter Innovations GmbH: Employment.

HDAC Inhibitors Lead to Significant Survival Benefit in a Novel Fusion Protein TBLR1-Rarα Induced Murine Promyelocytic Leukemia Model

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1558-1558 ◽  
Author(s):  
Shouyun Li ◽  
Shuang Liu ◽  
Shuying Chen ◽  
Yirui Chen ◽  
Ying Wang ◽  
...  
Keyword(s):  
Body Weight ◽  
Mouse Model ◽  
Survival Time ◽  
Hdac Inhibitors ◽  
Therapeutic Targets ◽  
Promyelocytic Leukemia ◽  
Leukemia Cells ◽  
Control Group

Abstract Introduction: TBLR1-RARα is the tenth fusion gene of acute promyelocytic leukemia (APL) first identified in a rare case of APL with t(3;17)(q26;q21) chromosomal translocation in our previous study. The characteristics of its basic structure and functions had been clarified in our previous study. In this study, we successfully established a novel TBLR1-RARα leukemia mouse model (TR mouse) which fully recapitulated the most relevant features of human APLs. The therapeutic effects of retinoic acid (ATRA), arsenic trioxide (As2O3), cytarabine (Ara-C) and histone deacetylase inhibitors (HDACi) on TR mice were examined. The differentially expressed genes (DEGs) between TR mice and normal mice were compared to explore the possible mechanisms and better therapeutic targets for this kind of APL. Methods: pMSCV-TBLR1-RARα-Flag-IRES-GFP (MSCV-TR) and pMSCV-IRES-GFP (vehicle) retroviral plasmids were constructed and transfected 293T packaging cells to produce retroviruses. Lin- cells from C57BL/6 mice bone marrow were purified and infected with MSCV-TR and vehicle retroviral supernatant. For in vitro assay, the GFP+ lin- cells sorted and incubated with or without different concentrations of ATRA were analyzed for the differentiation and proliferation capacity by cell morphology, myeloid markers (CD11b and GR-1) and colony formation assay. For the in vivo experiment, GFP+ lin- cells transfected with indicated retroviral vectors were injected intravenously to lethally irradiated C57BL/6 mice to establish an APL mouse model. Cell surface markers were analyzed by flow cytometry. In treatment assays, GFP+ spleen cells from TR leukemia mice were injected intravenously into recipient mice. The mice were randomly separated into groups and received different treatment with ATRA, As2O3, As2O3 in combination with ATRA, Ara-C, Ara-C in combination with ATRA, chidamide and NL101, respectively. The percentage of GFP+ cells in peripheral blood and body weight were measured dynamically. The survival time of every group was recorded and compared. RNA-seq assay was used to identify DEGs between TR mice and normal mice. Results: In vitro assays indicated that TBLR1-RARα could either block the differentiation of HSCs at a relatively early stage or enhanced the clonogenic potential of cells. The TBLR1-RARα leukemia mouse model was successfully established. During the ten-month observational period, 3 out of 15 mice transplanted with TBLR1-RARα expressing cells developed an APL-like disease. Development of leukemia was not observed in any of the mice in control group. All the leukemia mice had a body weight loss as well as splenomegaly and hepatomegaly. The phenotype analysis revealed that the progenitor markers Sca-1, CD34 and C-kit were positive, the myeloid lineage markers Gr-1 and CD11b were also positive, erythroid lineage marker Ter119 was weekly positive, but the lymphatic lineage marker B220, CD3,CD4 and CD8 were all negative. TR mice treated with 1.5-2.5 mg/kg ATRA alone or together with 2.0 mg/kg As2O3 didn't survive longer than that of control group, although in vitro differentiation experiment showed that the leukemia cells were sensitive to ATRA. Leukemic mice receiving Ara-C treatment had a much longer survive time. Surprisingly, HDAC inhibitors (12.5 and 25 mg/kg chidamide and 30 mg/kg NL-101) could significantly prolong the survival time of TR mice. Thousands of DEGs had been identified between TR mice and wild type mice, which were widely involved in multiple pathways and participated in various biological functions. Conclusion: The TBLR1-RARα leukemia mouse model was successfully established for the first time, and its main characteristics were clarified. Although the leukemia cells were sensitive to ATRA in vitro, TR mice didn't benefit from ATRA or As2O3 treatment in vivo. Besides Ara-C, HDAC inhibitors, such as chidamide and NL-101 exhibited potency therapeutic values for TR mice, which provided a new strategy for this kind of refractory APL. What' more, lots of genes that might be related with the process of leukemogenesis and new therapeutic targets for TR leukemia were identified. This model would serve as a versatile tool to study the mechanisms of leukemogenesis and help to design better strategies for APLs in further studies. Disclosures No relevant conflicts of interest to declare.

scholarly journals Inhibition of vitamin D analog eldecalcitol on hepatoma in vitro and in vivo

Open Medicine ◽  
2020 ◽  
Vol 15 (1) ◽  
pp. 663-671
Author(s):  
Limin Ye ◽  
Liyi Zhu ◽  
Jinglin Wang ◽  
Fei Li
Keyword(s):  
Cell Cycle ◽  
Vitamin D ◽  
Cell Apoptosis ◽  
Control Group ◽  
High Morbidity ◽  
Rt Pcr ◽  
Vitamin D Analog ◽  
E Cadherin ◽  
Cycle Detection

AbstractHepatoma is a serious liver cancer with high morbidity and mortality. Eldecalcitol (ED-71), a vitamin D analog, is extensively used as anti-cancer agent in vitro. Hepatocellular carcinoma cell, SMMC-7721 cell lines were used in this study. Transwell assay, cell apoptosis and cell cycle detection assays were investigated after treatment with ED-71 and phosphate buffered saline (PBS) as control. Sizes of tumors were measured after ED-71 treatment in a mouse model. E-cadherin and Akt gene expressions were detected by real-time PCR (RT-PCR). The results showed that cell invasion and migration were decreased markedly after ED-71 treatment compared to control group. Cell cycle detection showed that the G2 stage was 13.18% and total S-stage was 41.16% in the ED-71 group and G2 stage: 22.88%, total S-stage: 27.34% in the control group. Cell apoptosis rate was promoted in the ED-71 group. Size of the tumors reduced more after the ED-71 treatment than the PBS treatment in mice. ED-71 markedly inhibited the expression of Akt and E-cadherin, either detected by immunohistochemistry or RT-PCR. ED-71 treatment can inhibit the hepatoma agent proliferation by increasing the E-cadherin expression and decreasing Akt expression. Therefore, these findings provide novel evidence that ED-71 can be used as an anti-hepatoma agent.

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