Counts In Spleen Research Articles

Inoculum effect of cefiderocol against NDM-1 producing Escherichia coli in vitro and in a murine model of peritonitis.

Cefiderocol is a siderophore cephalosporin active in vitro against carbapenemase-producing Enterobacterales, including New Delhi metallo-β-lactamases (NDM-1). A significant impact of the size of bacterial inoculum on its efficacy has been described in vitro, the clinical impact of which is unclear. Here, we analyse the inoculum effect of cefiderocol against E. coli-NDM-1 in vitro and in a murine peritonitis model. Escherichia coli 62-pTOPO and its isogenic variant expressing NDM-1, 62-pTOPO-NDM, were constructed from a clinical strain. MICs and bactericidal kinetics were determined at standard (105 cfu/mL) and high inoculum (107 cfu/mL). The in vivo effect was assessed in a severe murine peritonitis model, comparing low (106 cfu/mL) and high (108 cfu/mL) inoculum. Survival rates, organ sterilization and bacterial counts in spleen and peritoneal fluid were compared. Cefiderocol MICs for 62-pTOPO and 62-pTOPO-NDM at standard and high inoculum were 0.008, 2, 2 and 1024 mg/L, respectively. Bactericidal activity was not achieved in vitro for 62-pTOPO-NDM at high inoculum with high cefiderocol concentrations (16 mg/L). In vivo, for 62-pTOPO-NDM, no difference was found in survival, organ sterilization or bacterial counts between low and high inoculum. For 62-pTOPO, no difference was observed in survival, despite less organ sterilization and higher bacterial counts in organs with the high inoculum. A significant inoculum effect of cefiderocol was observed in vitro for 62-pTOPO and 62-pTOPO-NDM. However, the effectiveness of cefiderocol was not reduced in vivo with a high bacterial inoculum. In vitro inoculum effect of cefiderocol may not be clinically significant.

Platelet-to-albumin ratio and radiation-induced lymphopenia—prognostic biomarker for carcinoma esophagus

BackgroundEsophageal cancer has a poor survival outcome with 5-year OS at 16.7% despite treatment. Some inflammation-based prognostic indicators like the neutrophil-to-lymphocyte ratio (NLR) and platelet-to-lymphocyte ratio (PLR) have been previously studied as potential biomarker for predicting outcome in esophageal cancer. Recently, platelet-to-albumin ratio (PAR) has been reported as a promising prognostic factor in gastrointestinal malignancies.MethodsWe performed a retrospective analysis of prospectively treated patients of carcinoma esophagus to evaluate the prognostic significance of inflammation-based prognostic indicators—neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR), and a composite inflammation-nutrition index: platelet-to-albumin ratio (PAR) in esophageal cancer. Based on previous studies, the optimal cut-off value of PAR was kept at 5.7 × 10^9, and 2.62 for NLR.ResultsA total of 71 patients of locally advanced esophageal cancer treated between 2019 and 2022, with either neoadjuvant or definitive chemoradiotherapy, were included. Median follow-up time was 19 months [range: 7–44 months]. Median OS and PFS in our study cohort were 11.3 months [range: 7–23 months] and 7.8 months [range: 3–17 months], respectively. In univariate analysis, lower PAR was found to be significantly correlated with shorter survival time (HR = 2.41; 1.3–4.76; p = 0.047). There was no association found between the OS and the NLR [HR = 1.09; 0.95–1.26; p = 0.222]. Univariate and multivariate linear and logistic regressions found no association between V15, V10, V5, or V2 of spleen and nadir lymphocyte count or between Dmax or Dmean and nadir lymphocyte counts.ConclusionPresent analysis found a trend toward an inverse association between PAR and OS. PAR, in the not-so-distant future, may evolve as a novel, convenient, and inexpensive prognostic indicator in esophageal cancer.

Is the Eutos Long Term Survival (ELTS) Score a Useful Marker to Predict Outcome in Children with Newly Diagnosed Chronic Myeloid Leukemia (CML) in Chronic Phase (CP)? the Experience of the International Registry of Childhood CML

Aims. CML is a very rare disease in children. The present work aims to describe the characteristics of children with CML in chronic phase (CP) and to determine the outcome of this pediatric population. Methods. We retrospectively analyzed children with CML in CP registered in The International Registry of Childhood Chronic Myeloid Leukemia (I-CML-Ped Study; www.clinicaltrials.gov NCT01281735) which represents the largest cohort of children with CML. Results. Between January 2011 and March 2021, 576 CML patients less than 18 years of age at diagnosis were registered. We retrospectively selected 535 (92.8 %) children in CP according to the European leukemia net (ELN) criteria at diagnosis. Boys accounted for 58% of all patients with a boy to girl ratio of 1.25. The median age at diagnosis was 12.4 years (range, 8 months to 18 years). The most common symptoms at presentation were asthenia, weight loss and abdominal pain in 46%, 25% and 23% of the patients, respectively. CML in CP was diagnosed incidentally in 80 (16%) children when a blood assessment was performed for medical reasons. At diagnosis, thrombosis was observed in only one patient whilst 20% of the children presented with bleeding. Bleeding signs were associated with a higher frequency of palpable spleen (p=0.0007), higher leukocyte (p<0.0001) and platelets (p=0.02) counts, and a lower hemoglobin level (p=0.02). Clinical signs of leukostasis were observed in 88 (16.5%) patients. At diagnosis, spleen was palpable in 76% of the patients with a median of 9 cm (range, 1 to 32) below the costal margin. The median leukocyte count was 222 G/L (quartile 95-353). There is no statistical difference between boys and girls in terms of frequency of signs of leukostasis, frequency of splenomegaly, spleen size, median of the leucocyte count and hemoglobin level but the median of the platelet count was significantly higher in girls (p<0.0001). The majority (67%) of the children was allocated to the low-risk group according to the Eutos Long Term Survival (ELTS) score. First line therapy consisted of imatinib in 482 children with a median follow up of 4.6 years (IQR. 2.2-8.4). Among them, 208 (43%) children remained on imatinib with a median follow up of 2.9 years (IQR. 1.5-5.6). The main reason for switching (38.7% of the switches) was failure to achieve hematologic, cytogenetic or molecular response. Among the patients treated with imatinib first line, 15 (3.1%) children died and the probability of overall survival at 36 months was 97.4% (95% CI: 95-99%) (Figure 1). ELTS score was calculated for 406 of these patients: the 3-year survival rate was 98.1.% (95% CI: 96.3-100%), 94.4% (95% CI: 89.1-99.9%) and 95% (95% CI: 88.5-100%) in the low-, intermediate- and high-risk group, respectively (p logrank=0.009). Nineteen of these patients (3.9%) switched from imatinib to another treatment due to progression (accelerated or blastic phase). Progression free survival (PFS) rate at 3-years was 97.1% (95% CI: 94.9-99.2%), 91.7% (95% CI:84.5-98.3%) and 72.0% (95% CI: 59-87.9%) in the ELTS low-, intermediate- and high-risk group, respectively (p logrank<0.001) (Figure 2). Conclusion. The analysis of clinical and biological parameters of the largest cohort of children with CML in CP demonstrated that pediatric CML is characterized by a bulky disease (large splenomegaly, high leucocyte counts). However, these characteristics did not translate in a low rate of response and the outcome was similar to adults. While the ELTS score can identify children with the poorest outcome, a refined prognostic score specific for the pediatric population needs to be established and validated.

Salmonella Gallinarum mgtC mutant shows a delayed fowl typhoid progression in chicken

Salmonella Gallinarum (SG) provokes fowl typhoid, an infectious disease of acute clinical course that affects gallinaceous of any age and leads to high mortality rates. During the typhoid-like systemic infection of S. Typhimurium (STM) in mice, the bacterium expresses the mgtC gene, which is encoded in the Salmonella Pathogenecity Island – 3 (SPI-3). In this serovar, the function is linked to bacterial replication within macrophages, and its absence attenuates the pathogen. We hypothesized that deleting mgtC from SG genome would alter the microorganism pathogenicity in susceptible commercial poultry in a similar manner. Thus, the present study sought to elucidate the importance of mgtC on SG pathogenicity. For this, a mgtC-mutant lacking S. Gallinarum mutant was constructed (SG ΔmgtC). Its ability to replicate in medium that mimicries the mgtC-related intracellular environment of macrophages as well as in primary macrophages from chicken was evaluated. Moreover, the infection of susceptible chickens was performed to elucidate its pathogenicity and the elicited immune responses by measuring key interleukins by qRT-PCR and the population of macrophages and lymphocytes T CD4+ and CD8+ by means of immunohistochemistry. It was observed that mgtC was required for S. Gallinarum replication in acidified low-Mg2+ media and survival within macrophages. However, unlike its requirement for initial phase of STM infection in mice, lower bacterial counts were only observed at the late stage of macrophage infection without affecting the citotoxicity. Experiments showed that knocking-out the mgtC gene neither altered bacterial uptake by macrophages nor affects bacterial counts in liver and spleen and total chicken mortality. However, plotting a survival curve and analyzing the clinical-pathologic conditions, it was observed a slower progression of the disease in chickens infected by SG ΔmgtC compared to those challenged by the wild-type strain. Furthermore, the mRNA expression of IFN-γ and LITAF were similar between the infected chickens, but higher than in the uninfected group. The same was observed in macrophages and lymphocytes T CD4+ populations. On the other hand, the presence of lymphocytes T CD8+ was increased in the initial phase of the disease provoked by the wild-type strain over the mutant strain. We concluded that the role of mgtC in Fowl Typhoid in susceptible chickens differs from the role in typhoid-like infections in mammals. Thus, the deletion of mgtC gene from S. Gallinarum genome does not affect the overall pathogenicity, but slightly alters the pathogenesis.

The Idiosyncratic Efficacy of Spironolactone-Loaded PLGA Nanoparticles Against Murine Intestinal Schistosomiasis.

Schistosomiasis is a chronic debilitating parasitic disease accompanied with severe mortality rates. Although praziquantel (PZQ) acts as the sole drug for the management of this disease, it has many limitations that restrict the use of this treatment approach. Repurposing of spironolactone (SPL) and nanomedicine represents a promising approach to improve anti-schistosomal therapy. We have developed SPL-loaded poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) to enhance the solubility, efficacy, and drug delivery and hence decrease the frequency of administration, which is of great clinical value. The physico-chemical assessment was performed starting with particle size analysis and confirmed using TEM, FT-IR, DSC, and XRD. The antischistosomal effect of the SPL-loaded PLGA NPs against Schistosoma mansoni (S. mansoni)-induced infection in mice was also estimated. Our results manifested that the optimized prepared NPs had particle size of 238.00 ± 7.21 nm, and the zeta potential was -19.66 ± 0.98 nm, effective encapsulation 90.43±8.81%. Other physico-chemical features emphasized that nanoparticles were completely encapsulated inside the polymer matrix. The in vitro dissolution studies revealed that SPL-loaded PLGA NPs showed sustained biphasic release pattern and followed Korsmeyer-Peppas kinetics corresponding to Fickian diffusion (n<0.45). The used regimen was efficient against S. mansoni infection and induced significant reduction in spleen, liver indices, and total worm count (ρ<0.05). Besides, when targeting the adult stages, it induced decline in the hepatic egg load and the small intestinal egg load by 57.75% and 54.17%, respectively, when compared to the control group. SPL-loaded PLGA NPs caused extensive damage to adult worms on tegument and suckers, leading to the death of the parasites in less time, plus marked improvement in liver pathology. Collectively, these findings provided proof-of-evidence that the developed SPL-loaded PLGA NPs could be potentially used as a promising candidate for new antischistosomal drug development.

Effect of diets containing commercial bioactive compounds on Salmonella Heidelberg infection in broiler chicks.

Salmonella Heidelberg (SH) is responsible for economic losses in poultry farming and food infections in humans and is a serious public health problem. Recently, there has been an increase in the frequency of isolation of this serotype in batches of broilers raised in Brazil. It is necessary to find new ways to help control this pathogen. The present study aimed to evaluate the effect of diets containing the compound Original XPC, which is a prebiotic-like fermented compound (PFC), and/or Sangrovit, which is a sanguinarine-based phytobiotic (SAN), on SH infection in broiler chicks. For this purpose, SH colonization in the cecum and its invasion into the spleen and liver were evaluated, as were the histopathological changes caused in these organs. The lowest cecal SH counts were observed in birds that ingested SAN, followed by those fed PFC (P < 0.05), with no added effect when the two bioproducts were used together (SAN + PFC). The mean SH and liver spleen counts did not differ between groups (P > 0.05). In general, birds from all groups challenged with SH showed similar macroscopic changes, such as hemorrhagic areas, hepatomegaly, and splenomegaly, such changes being more intense in the infected control group. The microscopic changes observed in the liver included hepatocyte congestion, heterophil infiltration in the sinusoid capillaries, areas of necrosis, and mononuclear inflammation. In the cecum, heterophilic infiltrate and thickening of the lamina propria were observed. In the ileum, the most common changes were congestion and thickening of the lamina propria and atrophy of the villi and crypts. The microscopic changes were less intense in the supplemented birds than the infected control group, and those supplemented with SAN developed the least changes. As ideal conditions for histomorphometric parameters of the ileum, the villus:crypt ratio in birds should be high, the villi should be long, and the crypts should be shallow. In the present study, higher mean heights and villus areas were observed in uninfected control and SAN group birds, and the crypt depth was lower in birds in the negative control group. The lowest villus:crypt ratio was observed in the birds of the infected control group. Although additional studies are needed, the preliminary results of the current investigation indicated that the addition of bioproducts, especially SAN, to the diet of birds helped to control SH infection, reducing its count in the cecum and improving overall and intestinal health.

Immune Correlates of Disseminated BCG Infection in IL12RB1-Deficient Mice.

Interleukin-12 receptor β1 (IL12RB1)-deficient individuals show increased susceptibilities to local or disseminated BCG infection and environmental mycobacteria infection. However, the low clinical penetrance of IL12RB1 deficiency and low recurrence rate of mycobacteria infection suggest that protective immunity still exists in this population. In this study, we investigated the mechanism of tuberculosis suppression using the IL12RB1-deficient mouse model. Our results manifested that Il12rb1−/− mice had significantly increased CFU counts in spleens and lungs, especially when BCG (Danish strain) was inoculated subcutaneously. The innate TNF-a and IFN-γ responses decreased, while the IL-17 responses increased significantly in the lungs of Il12rb1−/− mice. We also found that PPD-specific IFN-γ release was impaired in Il12rb1−/− mice, but the specific TNF-a release was not compromised, and the antibody responses were significantly enhanced. Moreover, correlation analyses revealed that both the innate and PPD-specific IFN-γ responses positively correlated with CFU counts, whereas the innate IL-12a levels negatively correlated with CFU counts in Il12rb1−/− mice lungs. Collectively, these findings proved that the adaptive immunities against mycobacteria are not completely nullified in Il12rb1−/− mice. Additionally, our results imply that IFN-γ responses alone might not be able to contain BCGitis in the setting of IL12RB1 deficiency.

A Sri Lankan child with hypersplenism secondary to pre-hepatic portal hypertension, successfully managed with partial splenic artery embolization: a case report and review of the literature

BackgroundHypersplenism, one of the major complications of portal hypertension, is traditionally treated by splenectomy. However, partial splenic artery embolization is an evolving minimally invasive intervention to treat these patients effectively.Case presentationA 13-year-old girl was referred for further evaluation of isolated splenomegaly with pancytopenia. She did not have bleeding manifestations or features of anemia. She never had hematemesis or melena. On examination, she was pale. Abdominal examination revealed massive splenomegaly of 10 cm below the costal margin without hepatomegaly. Rest of the examination was unremarkable. Her investigations revealed a white cell count of 1700/mm3 (neutrophils 9.8% and lymphocytes 88.7%), hemoglobin 9.5 g/dL and platelet count 42,000/mm3. Blood picture showed pancytopenia without abnormal cells. Her reticulocyte count was 1.9%. Complete liver profile was normal. Abdominal ultrasonography revealed massive splenomegaly with the oblique length of 17 cm and normal echogenic liver with normal size. Cavernous transformation of portal vein with portal hypertension was evident. Mesenteric angiogram showed portal vein thrombosis and markedly tortuous splenic artery. Anti-nuclear antibodies and double-stranded DNA were negative. Ham test and urine for hemosiderin were negative. Clauss fibrinogen assay was normal. Hemoglobin high performance liquid chromatography for hemoglobin subtypes was normal. Anti-phospholipid antibodies were negative. JAK2 V617F mutation was not identified. Diagnosis of pre hepatic portal hypertension was made. Her upper gastrointestinal endoscopy was normal. Partial splenic artery coil embolization was done by interventional radiology team. Vaccines against capsulated organisms were given. Post-procedure contrast abdominal computed tomography revealed infarction of approximately 70% of the spleen and blood counts were improved. Index case is in the follow up for 3 years. She is on penicillin prophylaxis with regular blood count and annual upper gastrointestinal endoscopy monitoring.ConclusionsMinimally invasive interventions such as partial splenic artery embolization should be considered in managing the patients with hypersplenism secondary to portal hypertension.

P1060: REAL-WORLD CLINICAL OUTCOMES AFTER 3 AND 6 MONTHS OF TREATMENT WITH FEDRATINIB FOLLOWING RUXOLITINIB FAILURE

Background: Relief of symptoms and splenomegaly is an important treatment (Tx) goal for patients with myelofibrosis (MF). Fedratinib (FEDR) and ruxolitinib (RUX) are approved in the USA for Intermediate (Int)- or High-risk (HR) primary or secondary MF. Aims: To describe spleen size, and hematologic and MF-specific symptoms, during the first 6 months of FEDR therapy after prior Tx with RUX in US clinical practices. Methods: Adults with Int- or HR MF who initiated FEDR on or after August 16, 2019 (FEDR approval date) and with ≥90 days of follow-up were identified from community oncology practices. Eligible patients were those treated with RUX and then FEDR, who had spleen palpation at FEDR initiation, and who had completed ≥1 cycle of FEDR. Treating physicians completed electronic case report forms for the selected eligible patients, extracting patient characteristics, Tx pattern data, and clinical outcomes such as spleen size, MF-related symptoms per the Myelofibrosis Symptom Assessment Form version 4.0 (MFSAF v4.0), hemoglobin (Hb), and platelet and white blood cell (WBC) counts, at each visit in the first 6 months of FEDR Tx. Data were collected from February to March 2021. Mean spleen size, symptom count, Hb, and platelet and WBC counts at FEDR initiation vs at 3 and 6 months post-FEDR were compared via paired 2-sided t-test. Percentage reduction in spleen size and proportion of transfusion-dependent patients between FEDR initiation and 3 and 6 months post-FEDR were compared via 1-sided chi-square test. Only patients assessed at FEDR initiation (with palpable spleen or complete blood count available) and with a corresponding assessment at 3 or 6 months were included in the comparative analysis. Results: Among 150 eligible patients, the mean age at FEDR initiation was 68 years (range, 36–85). Median duration of RUX Tx prior to FEDR was 7.6 months (range, 0.7–65.5). At FEDR initiation, MF risk was categorized as International Prognostic Scoring System (IPSS)/Dynamic IPSS HR in 43% of patients, Int-2 in 37%, Int-1 in 9%, and unknown in 10%. A total of 74% of patients started FEDR at the recommended dosage of 400 mg daily. Median duration of post-FEDR follow-up was 5.1 months (IQR, 3.0–17.3) for patients initiated at FEDR 400 mg daily and 4.4 months (IQR, 3.0–14.3) for those initiating at <400 mg daily. At data cutoff, 55% of patients were still on FEDR. The primary reason for FEDR discontinuation was disease progression (43%). Mean duration of FEDR Tx was 4.7 months. Of 67 patients who discontinued FEDR, 44 were deceased at data cutoff. At FEDR initiation, 88% of patients had palpable spleen; mean spleen size was 16.0 cm (standard deviation [SD] 5.8) above the costal margin, which decreased to 13.2 cm (SD 7.9) at 3 months (P=0.0001) and 7.2 cm (SD 7.4) at 6 months (P=0.012). Mean number of MF-related symptoms declined significantly at 3 and 6 months; mean platelet count increased significantly at 6 months, from 160×109/L to 186×109/L (Table). Overall, 65.2% of patients experienced spleen size reduction, 18.8% had no change, and 16.0% had an increase. Mean best percentage change in spleen size was −29.2% (median −9.8%); complete resolution of palpable spleen occurred in 21% of patients. Image:Summary/Conclusion: This study illustrates the real-world effectiveness of FEDR after RUX failure in patients with Int- or HR MF. FEDR provided significant reductions in spleen size and reported number of MF-related symptoms after 3 months of Tx, including complete spleen responses. Greater benefits were observed with longer Tx duration.

Dietary Acrylamide Intake Alters Gut Microbiota in Mice and Increases Its Susceptibility to Salmonella Typhimurium Infection.

Acrylamide (AA) has been extensively examined for its potential toxicological effects on humans and animals, but its impacts on gut microbiota and effects on hosts’ susceptibility to enteric infection remain elusive. The present study was designed to evaluate the effect of AA on gut microbiota of mice and susceptibility of mice to S. Typhimurium infection. After four weeks’ intervention, mice fed with AA exhibited significantly decreased body weight. Meanwhile, 16S rRNA gene sequencing showed reduced relative abundance of Firmicutes and increased abundance of Bacteroidetes in AA-treated mice prior to infection. In addition, we observed high relative abundance of Burkholderiales and Erysipelotrichales, more specifically the genus Sutterella and Allobaculum, respectively, in AA-treated mice before infection. Subsequently, the mice were orally infected with S. Typhimurium. The histological changes, systemic dissemination of S. Typhimurium, and inflammatory responses were examined. Compared to mice fed with normal diet, mice fed AA exhibited higher level of bacterial counts in liver, spleen, and ileum, which was consistent with exacerbated tissue damage determined by histological analyses. In addition, higher expression of pro-inflammaroty cytokines, p-IκBα, and p-P65 and lower mRNA expressions of mucin2, occludin, zo-1, claudin-1, and E-cadherin were detected in AA-treated mice. These findings provide novel insights into the potential health impact of AA consumption and the detailed mechanism for its effect on S. Typhimurium infection merit further exploration.

Gene Expression Patterns and Functional Characterization of CD36 in Normal Hematopoietic Cells

Metabolic reprogramming is a hallmark of cancer development and progression. Lipid metabolism is among the most deregulated metabolic pathways in cancer contributing to cell survival and invasion. The fatty acid translocase CD36 has attracted attention in cancer research in recent years due to its role in fatty acid transport as a scavenger receptor. We have recently reported that upregulated APOC2 cooperates with CD36 contributing to leukemia growth by promoting the LYN-ERK signaling mediated metabolic activities of acute myeloid leukemia (AML) cells. Consistently, the knockdown of CD36 or treatment with anti-CD36 antibody reduced the leukemia progression and promoted the overall survival of AML xenograft mice. CD36 is also present on various types of normal cells, including monocytes, macrophages, endothelial cells, adipocytes, and platelets and contribute to their normal functions. Thus, whether targeting CD36 in normal hematopoietic cells poses any toxicity warrants investigation to bring the therapeutic strategy of targeting CD36 in AML to maturity. Here, we leveraged public data to assess the expression patterns of CD36 in normal hematopoietic cells. We found that CD36 expression is several folds higher in dendritic cells, monocytes, and NK cells compared with B cells, eosinophils, neutrophils, and T cells (ANOVA P < 0.001). To investigate the role of Cd36 in normal hematopoietic cells, we carried out gain- and loss-of-function approaches in murine normal hematopoietic primary cells. We engineered a tet-on inducible shRNA lentiviral plasmid targeting two different sequences of the mouse Cd36. In both healthy mouse primary spleen and bone marrow cells, the two pairs of shRNAs reduced the Cd36 mRNA levels compared with scramble control group (~70-90% reduction in mRNA expression). We also engineered two different lentivirus ectopic expression plasmid to overexpress murine Cd36 (PLVX-Cd36 virus and PCDH-Cd36). Lentiviral transduction of both spleen and bone marrow mouse cells confirmed by microscopy visualization of GFP-expressing cells showed several folds increase in Cd36 expression for both plasmids measured by qPCR and western blot. We assessed the effect of Cd36 knockdown on the expansion of spleen and bone marrow hematopoietic cells over five days of cell culture. Relative to the 1 st day of cell count, there was no significant decrease in the mean bone marrow cell and spleen count between the two pair of shRNA compared with scramble control group at day 3 and day 5 post doxycycline treatment (P > 0.05). Cells in the different groups showed about 2 folds expansion on day 5 relative to day 1. Our preliminary data suggest that Cd36 knockdown has limited impact on the survival of normal hematopoietic cells. In vivo experiments assessing the effect of Cd36 gain and loss of function on the engraftment of normal hematopoietic cells in mice are ongoing to further establish the functional impact of Cd36 manipulation on normal hematopoietic cells. Altogether, our studies will demonstrate whether CD36 is dispensable for the survival of normal hematopoietic cells and thus may potentially present a viable therapeutic approach in AML. DisclosuresYaghmour: Jazz: Speakers Bureau; Takeda: Consultancy, Speakers Bureau; Astellas: Speakers Bureau; Alexion: Speakers Bureau; BMS: Speakers Bureau; Novartis: Consultancy, Speakers Bureau; Agios: Consultancy, Speakers Bureau.

Real-World Utilization of Fedratinib for Myelofibrosis Post-Ruxolitinib: Patient Characteristics, Treatment Patterns, and Characterization of Ruxolitinib Failure

Real-World Utilization of Fedratinib for Myelofibrosis Post-Ruxolitinib: Patient Characteristics, Treatment Patterns, and Characterization of Ruxolitinib Failure

Clinical Outcomes at 3 and 6 Months of Fedratinib Therapy Following Prior Ruxolitinib Failure: Real-World Assessment of Spleen, Symptoms, and Hematologic Response

Clinical Outcomes at 3 and 6 Months of Fedratinib Therapy Following Prior Ruxolitinib Failure: Real-World Assessment of Spleen, Symptoms, and Hematologic Response

Systemic T Cell Exhaustion Dynamics Is Linked to Early High Mobility Group Box Protein 1 (HMGB1) Driven Hyper-Inflammation in a Polytrauma Rat Model.

We previously reported an early surge in high mobility group box protein 1 (HMGB1) levels in a polytrauma (PT) rat model. This study investigates the association of HMGB1 levels in mediating PT associated dysregulated immune responses and its influence on the cellular levels of receptor for advanced glycation end products (RAGE) and toll-like receptor 4 (TLR4). Using the same PT rat model treated with anti-HMGB1 polyclonal antibody, we evaluated changes in circulating inflammatory cytokines, monocytes/macrophages and T cells dynamics and cell surface expression of RAGE and TLR4 at 1, 3, and 7 days post-trauma (dpt) in blood and spleen. Notably, PT rats demonstrating T helper (Th)1 and Th2 cells type early hyper-inflammatory responses also exhibited increased monocyte/macrophage counts and diminished T cell counts in blood and spleen. In blood, expression of RAGE and TLR4 receptors was elevated on CD68+ monocyte/macrophages and severely diminished on CD4+ and CD8+ T cells. Neutralization of HMGB1 significantly decreased CD68+ monocyte/macrophage counts and increased CD4+ and CD8+ T cells, but not γδ+TCR T cells in circulation. Most importantly, RAGE and TLR4 expressions were restored on CD4+ and CD8+ T cells in treated PT rats. Overall, findings suggest that in PT, the HMGB1 surge is responsible for the onset of T cell exhaustion and dysfunction, leading to diminished RAGE and TLR4 surface expression, thereby possibly hindering the proper functioning of T cells.

A quantitative systems pharmacology model of hyporesponsiveness to erythropoietin in rats.

Recombinant human erythropoietin (rHuEPO) is effective in managing chronic kidney disease and chemotherapy-induced anemia. However, hyporesponsiveness to rHuEPO treatment was reported in about 10% of the patients. A decreased response in rats receiving a single or multiple doses of rHuEPO was also observed. In this study, we aimed to develop a quantitative systems pharmacology (QSP) model to examine hyporesponsiveness to rHuEPO in rats. Pharmacokinetic (PK) and pharmacodynamic (PD) data after a single intravenous dose of rHuEPO (100IU/kg) was obtained from a previous study (Yan et al. in Pharm Res, 30:1026-1036, 2013) including rHuEPO plasma concentrations, erythroid precursors counts in femur bone marrow and spleen, reticulocytes (RETs), red blood cells (RBCs), and hemoglobin (HGB) in circulation. Parameter values were obtained from literature or calibrated with experimental data. Global sensitivity analysis and model-based simulations were performed to assess parameter sensitivity and hyporesponsiveness. The final QSP model adequately characterizes time courses of rHuEPO PK and nine PD endpoints in both control and treatment groups simultaneously. The model indicates that negative feedback regulation, neocytolysis, and depletion of erythroid precursors are major factors leading to hyporesponsiveness to rHuEPO treatment in rats.

1635. Oral Delivery of Amikacin-Lipid NanoCrystal Formulations Safely and Effectively Treat Macrolide Resistant Mycobacteria Infections in a Mouse Model of Cystic Fibrosis

BackgroundIn the cystic fibrosis lung, infections by intracellular pathogens, such as Mycobacteria, are problematic to treat due to a thick buildup of mucous as well as the difficulty of many anti-microbial agents, such as amikacin, to penetrate across the plasma membranes of infected cells. Lipid NanoCrystals (LNCs), mediate oral bioavailability for injectable drugs, reduce toxicity, and significantly enhance targeting to mycobacterial infected cells followed by intracellular drug delivery.MethodsThe oral efficacy of Amikacin-LNC (AmK-LNC) was evaluated in the cystic fibrosis (B6CFTRtm1UNC/CFTRtm1UNC) chronic mouse model against each of the three NTM strains having high resistance to macrolide antibiotics (M. avium subsp intracellulare 25292, M. abscesus ssp abscessus 1513, and M. abscessus ssp bolletii 1948). Mice were infected with a pulmonary aerosol of 1x108 CFUs of the macrolide resistant strain and treated daily starting on day 28 for a total of 8 weeks with saline control, oral LP-4 CAmK Lyophilized 50 mg/kg BID, oral LP-4 CAMK Lyophilized 100 mg/kg BID, IP Amikacin (AMI) 150 mg/kg QD, or oral Clarithromycin 250 mg/kg QD. Bacterial burden was measured on day 1, 27, 42, 56 and 84 after infection by plating serial dilutions of organ homogenates on nutrient 7H11 and charcoal agar and counting CFUs after 25-30 days incubation at 32°C. Results represent the average of six experiments (n=5 mice per experiment) bacterial load was expressed as the average Log10 CFU (± SEM) cells (± SEM). ANOVA, saline control compared to drug-treated groups, * denotes the compound that resulted in the highest bacterial reduction, *p< 0.05.ResultsOral administration of AmK-LNCs safely and effectively treated all three macrolide resistant Mycobacteria infections. Colony counts showed that oral administration of AmK-LNC resulted in CFU lung, spleen and liver counts lower than treatment with IP amikacin or clarithromycin.Lung pathology showed that lesions were more numerous and larger in infected mice treated with clarithromycin or amikacin compared to the smaller lesions after treatment with oral AmK-LNC.Bacterial Counts in the Lungs (A), Spleens (B) and Livers (C) Lung Pathology ConclusionConclusionsOral administration of amikacin-LNCs safely and effectively treats macrolide resistant mycobacterial infections in a mouse model of Cystic Fibrosis.DisclosuresRuying Lu, n/a, Matinas BioPharma Inc. (Employee)Matinas BioPharma Inc. (Employee, Shareholder) Raphael J. Mannino, n/a, Matinas BioPharma Inc. (Employee, Shareholder)

Prolonged infection triggered by dormant Mycobacterium tuberculosis: Immune and inflammatory responses in lungs of genetically susceptible and resistant mice.

We developed an approach for substantial attenuation of Mycobacterium tuberculosis by prolonged culturing under gradually acidifying conditions. Bacteria subjected to acidification lost the capacity to form colonies on solid media, but readily resuscitated their growth in the murine host, providing a useful model to study in vivo development of infection mimicking latent and reactivation tuberculosis (TB) in humans. Here we characterize biomarkers of lung pathology and immune responses triggered by such attenuated bacteria in genetically TB-susceptible and resistant mice. In susceptible I/St mice, CFU counts in lungs and spleens were ~1.5-log higher than in resistant B6 mice, accompanied by diffuse pneumonia and excessive lung infiltration with highly activated CD44+CD62L- T-lymphocytes resulting in death between months 7–9 post challenge. B6 mice were characterized by development of local inflammatory foci, higher production of pro-inflammatory IL-6 and IL-11 cytokines and a more balanced T-cell activation in their lungs. CFU counts remained stable in B6 mice during the whole 18-mo observation period, and all mice survived. Thus, we established a mouse model of fatal reactivation TB vs. indefinite mycobacterial possession after identical challenge and characterized the features of immune responses in the lung tissue underlining these polar phenotypes.

Pharmacokinetic and pharmacodynamic profiling of generic amphotericin B colloidal dispersion in a rat model of invasive candidiasis

ObjectivesWe reported the pharmacokinetic/pharmacodynamic (PK/PD) targets of a biosimilar generic product of amphotericin B colloidal dispersion (G-ABCD) againstCandida albicans (MIC 1–2 μg/mL) in a rat model of invasive candidiasis (IC) to facilitate its precision administration. MethodsSingle-dose plasma PKs of G-ABCD was studied in a rat IC model following intravenous administration at doses of 0.0625–10 mg/kg. Amphotericin B concentrations were determined and PK parameters were calculated based on the concentrations in plasma. The efficacy of G-ABCD was evaluated after single administration by the log reduction of CFU counts in kidney, liver, spleen and lung. The relationship between G-ABCD PK/PD index and log CFU reduction in kidney was calculated. ResultsFollowing intravenous administration of G-ABCD at doses of 0.0625–10 mg/kg to rats, the maximum plasma concentration (Cmax) was 0.05–0.82 mg/L and the area under the concentration–time curve from 0 to 24 h (AUC0–24) was 0.50–5.29 mg h/L. G-ABCD showed potent antifungal activity against C. albicans C-13 with a maximum log CFU reduction of 2.1 in kidney. The mean AUC0–24/MIC target of G-ABCD against C. albicans was 0.97 for stasis, 1.40 for 1-log kill and 3.34 for 2-log kill, and the mean Cmax/MIC target was 0.063 for stasis, 0.097 for 1-log kill and 0.348 for 2-log kill. ConclusionsThe antifungal effect of G-ABCD was potent and correlated with AUC0–24/MIC and Cmax/MIC in this rat model of IC. The results of this study provide data for optimising G-ABCD dosing regimens and breakpoints for antifungals.

Dimercaptosuccinic acid in combination with carbapenems against isogenic strains of Escherichia coli producing or not producing a metallo-β-lactamase in vitro and in murine peritonitis.

Carbapenemase-producing Enterobacterales represent a major therapeutic challenge. MBLs, requiring zinc at their catalytic site, could be inhibited by meso-dimercaptosuccinic acid (DMSA), a heavy metal chelator already widely used for treating lead intoxication. To evaluate the activity of carbapenems alone or combined with DMSA against MBL-producing Escherichia coli in a severe murine peritonitis model. Isogenic strains of wild-type E. coli CFT073 producing the MBLs NDM-1, VIM-2 and IMP-1, and the control serine carbapenemases OXA-48 and KPC-3 were constructed. MIC determinations and time-kill assays were performed for imipenem, meropenem and ertapenem alone or in combination with DMSA. Infected mice were treated intraperitoneally for 24 h with imipenem, DMSA or their combination. Bacterial counts in peritoneal fluid and spleen were assessed at 24 h. DMSA in combination with each carbapenem caused a significant decrease in the MICs for all MBL-producing strains, in a concentration-dependent manner, but did not provide benefit against non-MBL strains. In mice infected with the NDM-1-producing strain, the combination of imipenem and DMSA significantly reduced bacterial counts in peritoneal fluid (P = 0.0006) and spleen (P < 0.0001), as compared with imipenem alone, with no benefit against the KPC-3-producing and CFT073 strains. DMSA concentrations in plasma of mice were comparable to those obtained in humans with a standard oral dose. DMSA restores the activity of carbapenems against MBL-producing strains, and its combination with carbapenems appears to be a promising strategy for the treatment of NDM-producing E. coli infections.

Analysis of Paradoxical Efficacy of Carbapenems against Carbapenemase-Producing Escherichia coli in a Murine Model of Lethal Peritonitis.

The clinical benefit of carbapenems against carbapenemase-producing Enterobacteriaceae (CPE) remains in question. MICs of imipenem (IMP) and ertapenem (ERT) against isogenic derivatives of the wild-type strain Escherichia coli CFT073 producing KPC-3, OXA-48, or NDM-1 were 0.25, 2, 16, and 64 mg/liter for IMP and 0.008, 0.5, 8, and 64 mg/liter for ERT, respectively. Swiss ICR-strain mice with peritonitis were treated for 24 h with IMP or ERT. Despite a limited duration of time during which free antibiotic concentrations were above the MIC (down to 0% for the NDM-1-producing strain), IMP and ERT significantly reduced bacterial counts in spleen and peritoneal fluid at 24 h (P < 0.005) and prevented mortality. Several possible explanations were investigated. Addition of 4% albumin or 50% normal human serum did not modify IMP activity. Bacterial fitness of resistant strains was not altered and virulence did not decrease with resistance. In the presence of subinhibitory concentrations of ERT, growth rates of OXA-48, KPC-3, and NDM-1 strains were significantly decreased and filamentation of the NDM-1 strain was observed. The expression of blaNDM-1 was not decreased in vivo compared to in vitro No zinc depletion was observed in infected mice compared with Mueller-Hinton broth. In conclusion, a paradoxical in vivo efficacy of IMP and ERT against highly resistant carbapenemase-producing E. coli was confirmed. Alternative mechanisms of antibacterial effects of subinhibitory concentrations of carbapenems may be involved to explain in vivo activity. These results are in agreement with a potential clinical benefit of carbapenems to treat CPE infections, despite high carbapenem MICs.