Morphomics-informed population pharmacokinetic and physiologically-based pharmacokinetic modeling to optimize cefazolin surgical prophylaxis.

INTRODUCTION: Cefazolin is the leading antibiotic used to prevent surgical site infections worldwide. Consensus guidelines recommend adjustment of the cefazolin dose above and below 120 kg without regard to body composition. Algorithms exist to repurpose radiologic data into body composition (morphomics) and inform dosing decisions in obesity. OBJECTIVES: To compare the current standard of body weight to morphomic measurements as covariates of cefazolin pharmacokinetics and aid dose stratification of cefazolin in patients with obesity undergoing colorectal surgery. METHODS: This prospective study measured cefazolin plasma, fat, and colon tissue concentrations in colorectal surgery patients in order to develop a morphomics-informed population pharmacokinetic (PopPK) model to guide dose adjustments. A physiologically-based pharmacokinetic (PBPK) model was also constructed to inform tissue partitioning in morbidly obese patients (n = 21, body mass index ≥35 kg/m2 with one or more co-morbid conditions). RESULTS: Morphomics and pharmacokinetic data were available in 58 patients with a median [min, max] weight and age of 95.9 [68.5, 148.8] kg and 55 [25, 79] years, respectively. The plasma-to-subcutaneous fat partition coefficient was predicted to be 0.072 and 0.060 by the PopPK and PBPK models, respectively. The estimated creatinine clearance (eCLcr ) and body depth at the third lumbar vertebra (body depth_L3) were identified as covariates of cefazolin exposure. The probability of maintaining subcutaneous fat concentrations above 2 µg/mL for 100% of a 4-h surgical period was below 90% when eCLcr ≥105 mL/min and body depth_L3 ≥ 300 mm and less sensitive to the rate of infusion between 5 and 60 min. CONCLUSIONS: Kidney function and morphomics were more informative than body weight as covariates of cefazolin target site exposure. Data from more diverse populations, consensus on target cefazolin exposure, and comparative studies are needed before a change in practice can be implemented.

Prophylactic Mitigation of Acute Graft versus Host Disease by Novel 2-(Pyrrolidin-1-ylmethyl)pyrrole-Based Stimulation-2 (ST2) Inhibitors.

Hematopoietic cell transplantation (HCT) is a proven and potentially curable therapy for hematological malignancies and inherited hematological disease. The main risk of HCT is the development of graft versus host disease (GVHD) acquired in up to 50% of patients. Upregulation of soluble ST2 (sST2) is a key clinical biomarker for GVHD prognosis and was shown to be a potential therapeutic target for GVHD. Agents targeting sST2 to reduce the sST2 level after HCT have the potential to mitigate GVHD progression. Here, we report 32 (or XY52) as the lead ST2 inhibitor from our optimization campaign. XY52 had improved inhibitory activity and metabolic stability in vitro and in vivo. XY52 suppressed proinflammatory T-cell proliferation while increasing regulatory T cells in vitro. In a clinically relevant GVHD model, a 21-day prophylactic regimen of XY52 reduced plasma sST2 and IFN-γ levels and GVHD score and extended survival in mice. XY52 represented a significant improvement over our previous compound, iST2-1, and further optimization of XY52 is warranted. The small-molecule ST2 inhibitors can potentially be used as a biomarker-guided therapy for mitigating GVHD in future clinical applications.

Structure-Activity relationship of 1-(Furan-2ylmethyl)Pyrrolidine-Based Stimulation-2 (ST2) inhibitors for treating graft versus host disease.

An elevated plasma level of soluble ST2 (sST2) is a risk biomarker for graft-versus-host disease (GVHD) and death in patients receiving hematopoietic cell transplantation (HCT). sST2 functions as a trap for IL-33 and amplifies the pro-inflammatory type 1 and 17 response while suppressing the tolerogenic type 2 and regulatory T cells activation during GVHD development. We previously identified small-molecule ST2 inhibitors particularly iST2-1 that reduces plasma sST2 levels and improved survival in two animal models. Here, we reported the structure-activity relationship of the furanylmethylpyrrolidine-based ST2 inhibitors based on iST2-1. Based on the biochemical AlphaLISA assay, we improved the activity of iST2-1 by 6-fold (∼6 µM in IC50 values) in the inhibition of ST2/IL-33 and confirmed the activities of the compounds in a cellular reporter assay. To determine the inhibition of the alloreactivity in vitro, we used the mixed lymphocyte reaction assay to demonstrate that our ST2 inhibitors decreased CD4+ and CD8+ T cells proliferation and increased Treg population. The data presented in this work are critical to the development of ST2 inhibitors in future.

Albumin nanoparticle containing a PI3Kγ inhibitor and paclitaxel in combination with α-PD1 induces tumor remission of breast cancer in mice.

Immunomodulators that remodel the tumor immunosuppressive microenvironment have been combined with anti-programmed death 1 (α-PD1) or anti-programmed death ligand 1 (α-PDL1) immunotherapy but have shown limited success in clinical trials. However, therapeutic strategies to modulate the immunosuppressive microenvironment of lymph nodes have been largely overlooked. Here, we designed an albumin nanoparticle, Nano-PI, containing the immunomodulators PI3Kγ inhibitor (IPI-549) and paclitaxel (PTX). We treated two breast cancer mouse models with Nano-PI in combination with α-PD1, which remodeled the tumor microenvironment in both lymph nodes and tumors. This combination achieved long-term tumor remission in mouse models and eliminated lung metastases. PTX combined with IPI-549 enabled the formation of a stable nanoparticle and enhanced the repolarization of M2 to M1 macrophages. Nano-PI not only enhanced the delivery of both immunomodulators to lymph nodes and tumors but also improved the drug accumulation in the macrophages of these two tissues. Immune cell profiling revealed that the combination of Nano-PI with α-PD1 remodeled the immune microenvironment by polarizing M2 to M1 macrophages, increasing CD4+ and CD8+ T cells, B cells, and dendritic cells, decreasing regulatory T cells, and preventing T cell exhaustion. Our data suggest that Nano-PI in combination with α-PD1 modulates the immune microenvironment in both lymph nodes and tumors to achieve long-term remission in mice with metastatic breast cancer, and represents a promising candidate for future clinical trials.

Induction of glutathione biosynthesis by glycine-based treatment mitigates atherosclerosis.

Lower circulating levels of glycine are consistently reported in association with cardiovascular disease (CVD), but the causative role and therapeutic potential of glycine in atherosclerosis, the underlying cause of most CVDs, remain to be established. Here, following the identification of reduced circulating glycine in patients with significant coronary artery disease (sCAD), we investigated a causative role of glycine in atherosclerosis by modulating glycine availability in atheroprone mice. We further evaluated the atheroprotective potential of DT-109, a recently identified glycine-based compound with dual lipid/glucose-lowering properties. Glycine deficiency enhanced, while glycine supplementation attenuated, atherosclerosis development in apolipoprotein E-deficient (Apoe-/-) mice. DT-109 treatment showed the most significant atheroprotective effects and lowered atherosclerosis in the whole aortic tree and aortic sinus concomitant with reduced superoxide. In Apoe-/- mice with established atherosclerosis, DT-109 treatment significantly reduced atherosclerosis and aortic superoxide independent of lipid-lowering effects. Targeted metabolomics and kinetics studies revealed that DT-109 induces glutathione formation in mononuclear cells. In bone marrow-derived macrophages (BMDMs), glycine and DT-109 attenuated superoxide formation induced by glycine deficiency. This was abolished in BMDMs from glutamate-cysteine ligase modifier subunit-deficient (Gclm-/-) mice in which glutathione biosynthesis is impaired. Metabolic flux and carbon tracing experiments revealed that glycine deficiency inhibits glutathione formation in BMDMs while glycine-based treatment induces de novo glutathione biosynthesis. Through a combination of studies in patients with CAD, in vivo studies using atherosclerotic mice and in vitro studies using macrophages, we demonstrated a causative role of glycine in atherosclerosis and identified glycine-based treatment as an approach to mitigate atherosclerosis through antioxidant effects mediated by induction of glutathione biosynthesis.

A single dose of valproic acid improves neurologic recovery and decreases brain lesion size in swine subjected to an isolated traumatic brain injury.

BACKGROUND: We lack specific treatments for traumatic brain injury (TBI), which remains the leading cause of trauma-related morbidity and mortality. Treatment with valproic acid (VPA) improves outcomes in models of severe TBI with concurrent hemorrhage. However, it is unknown if VPA will have similar benefits after isolated nonlethal TBI, which is the more common clinical scenario. The goal of this study was to evaluate the effect of VPA treatment in a preclinical isolated TBI swine model on neurologic outcomes and brain lesion size and to perform detailed pharmacokinetic analyses for a future clinical trial. METHODS: Yorkshire swine (n = 10; 5/cohort) were subjected to TBI (8-mm controlled cortical impact). An hour later, we randomized them to receive VPA (150 mg/kg) or saline placebo (control). Neuroseverity scores were assessed daily (0 [normal] to 36 [comatose]), brain lesion size was measured on postinjury 3, and serial blood samples were collected for pharmacokinetic studies. RESULTS: Physiologic parameters and laboratory values were similar in both groups. Valproic acid-treated animals demonstrated significantly better neuroseverity scores on postinjury 1 (control, 9.2 ± 4.4; VPA, 0 ± 0; p = 0.001). Valproic acid-treated animals had significantly smaller brain lesion sizes (mean volume in microliter: control, 3,130 ± 2,166; VPA, 764 ± 208; p = 0.02). Pharmacokinetic data confirmed adequate plasma and tissue levels of VPA. CONCLUSION: In this clinically relevant model of isolated TBI, a single dose of VPA attenuates neurological impairment and decreases brain lesion size.

Elevated Carbon Dioxide and Chronic Warming Together Decrease Nitrogen Uptake Rate, Net Translocation, and Assimilation in Tomato.

The response of plant N relations to the combination of elevated CO2 (eCO2) and warming are poorly understood. To study this, tomato (Solanum lycopersicum) plants were grown at 400 or 700 ppm CO2 and 33/28 or 38/33 °C (day/night), and their soil was labeled with 15NO3- or 15NH4+. Plant dry mass, root N-uptake rate, root-to-shoot net N translocation, whole-plant N assimilation, and root resource availability (%C, %N, total nonstructural carbohydrates) were measured. Relative to eCO2 or warming alone, eCO2 + warming decreased growth, NO3- and NH4+-uptake rates, root-to-shoot net N translocation, and whole-plant N assimilation. Decreased N assimilation with eCO2 + warming was driven mostly by inhibition of NO3- assimilation, and was not associated with root resource limitations or damage to N-assimilatory proteins. Previously, we showed in tomato that eCO2 + warming decreases the concentration of N-uptake and -assimilatory proteins in roots, and dramatically increases leaf angle, which decreases whole-plant light capture and, hence, photosynthesis and growth. Thus, decreases in N uptake and assimilation with eCO2 + warming in tomato are likely due to reduced plant N demand.

The comparison of glycosphingolipids isolated from an epithelial ovarian cancer cell line and a nontumorigenic epithelial ovarian cell line using MALDI-MS and MALDI-MS/MS.

Glycosphingolipids (GSLs) are important biomolecules, which are linked to many diseases such as GSL storage disorders and cancer. Consequently, the expression of GSLs may be altered in ovarian cancer cell lines in comparison to apparently healthy cell lines. Here, differential expressions of GSLs in an epithelial ovarian cancer cell line SKOV3 and a nontumorigenic epithelial ovarian cell line T29 were studied using matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS) and MALDI-MS/MS. The isolation of GSLs from SKOV3 and T29 cell lines was carried out using Folch partition. GSLs were successfully detected by MALDI-MS, and structurally assigned by a comparison of their MALDI-MS/MS fragmentation patterns with MS/MS data found in SimLipid database. Additionally, LIPID MAPS was used to assign GSL ion masses in MALDI-MS spectra. Seventeen neutral GSLs were identified in Folch partition lower (chloroform/methanol) phases originating from both cell lines, while five globo series neutral GSLs were identified only in the Folch partition lower phase of SKOV3 cell line. Several different sialylated GSLs were detected in Folch partition upper (water/methanol) phases of SKOV3 and T29 cell lines. Overall, this study demonstrates the alteration and increased glycosylation of GSLs in an epithelial ovarian cancer cell line in comparison to a nontumorigenic epithelial ovarian cell line.