Tobacco-induced hyperglycemia promotes lung cancer progression via cancer cell-macrophage interaction through paracrine IGF2/IR/NPM1-driven PD-L1 expression.

Tobacco smoking (TS) is implicated in lung cancer (LC) progression through the development of metabolic syndrome. However, direct evidence linking metabolic syndrome to TS-mediated LC progression remains to be established. Our findings demonstrate that 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone and benzo[a]pyrene (NNK and BaP; NB), components of tobacco smoke, induce metabolic syndrome characteristics, particularly hyperglycemia, promoting lung cancer progression in male C57BL/6 J mice. NB enhances glucose uptake in tumor-associated macrophages by increasing the expression and surface localization of glucose transporter (GLUT) 1 and 3, thereby leading to transcriptional upregulation of insulin-like growth factor 2 (IGF2), which subsequently activates insulin receptor (IR) in LC cells in a paracrine manner, promoting its nuclear import. Nuclear IR binds to nucleophosmin (NPM1), resulting in IR/NPM1-mediated activation of the CD274 promoter and expression of programmed death ligand-1 (PD-L1). Restricting glycolysis, depleting macrophages, or blocking PD-L1 inhibits NB-mediated LC progression. Analysis of patient tissues and public databases reveals elevated levels of IGF2 and GLUT1 in tumor-associated macrophages, as well as tumoral PD-L1 and phosphorylated insulin-like growth factor 1 receptor/insulin receptor (pIGF-1R/IR) expression, suggesting potential poor prognostic biomarkers for LC patients. Our data indicate that paracrine IGF2/IR/NPM1/PD-L1 signaling, facilitated by NB-induced dysregulation of glucose levels and metabolic reprogramming of macrophages, contributes to TS-mediated LC progression.

Direct and Indirect Chimeric Antigen Receptor T-Cell Imaging with PET/MRI in a Tumor Xenograft Model.

Background Chimeric antigen receptor (CAR) T cells are a promising cancer therapy; however, reliable and repeatable methods for tracking and monitoring CAR T cells in vivo remain underexplored. Purpose To investigate direct and indirect imaging strategies for tracking the biodistribution of CAR T cells and monitoring their therapeutic effect in target tumors. Materials and Methods CAR T cells co-expressing a tumor-targeting gene (anti-CD19 CAR) and a human somatostatin receptor subtype 2 (hSSTr2) reporter gene were generated from human peripheral blood mononuclear cells. After direct labeling with zirconium 89 (89Zr)-p-isothiocyanatobenzyl-desferrioxamine (DFO), CAR T cells were intravenously injected into immunodeficient mice with a CD19-positive and CD19-negative human tumor xenograft on the left and right flank, respectively. PET/MRI was used for direct in vivo imaging of 89Zr-DFO-labeled CAR T cells on days 0, 1, 3, and 7 and for indirect cell imaging with the radiolabeled somatostatin receptor-targeted ligand gallium 68 (68Ga)-DOTA-Tyr3-octreotide (DOTATOC) on days 6, 9, and 13. On day 13, mice were euthanized, and tissues and tumors were excised. Results The 89Zr-DFO-labeled CAR T cells were observed on PET/MRI scans in the liver and lungs of mice (n = 4) at all time points assessed. However, they were not visualized in CD19-positive or CD19-negative tumors, even on day 7. Serial 68Ga-DOTATOC PET/MRI showed CAR T cell accumulation in CD19-positive tumors but not in CD19-negative tumors from days 6 to 13. Notably, 68Ga-DOTATOC accumulation in CD19-positive tumors was highest on day 9 (mean percentage injected dose [%ID], 3.7% ± 1.0 [SD]) and decreased on day 13 (mean %ID, 2.6% ± 0.7) in parallel with a decrease in tumor volume (day 9: mean, 195 mm3 ± 27; day 13: mean, 127 mm3 ± 43) in the group with tumor growth inhibition. Enhanced immunohistochemistry staining of cluster of differentiation 3 (CD3) and hSSTr2 was also observed in excised CD19-positive tumor tissues. Conclusion Direct and indirect cell imaging with PET/MRI enabled in vivo tracking and monitoring of CAR T cells in an animal model. © RSNA, 2024 Supplemental material is available for this article. See also the editorial by Bulte in this issue.

18F-ASEM PET/MRI targeting alpha7-nicotinic acetylcholine receptor can reveal skeletal muscle denervation.

BACKGROUND: The increased expression of the nicotinic acetylcholine receptor (nAChR) in muscle denervation is thought to be associated with electrophysiological acetylcholine supersensitivity after nerve injury. Hence, we investigated the utility of the 18F-ASEM alpha7-nAChR targeting radiotracer as a new diagnostic method by visualizing skeletal muscle denervation in mouse models of sciatic nerve injury. METHODS: Ten-week-old C57BL/6 male mice were utilized. The mice were anesthetized, and the left sciatic nerve was resected after splitting the gluteal muscle. One week (n = 11) and three weeks (n = 6) after the denervation, 18F-ASEM positron emission tomography/magnetic resonance imaging (PET/MRI) was acquired. Maximum standardized uptake values (SUVmax) of the tibialis anterior muscle were measured for the denervated side and the control side. Autoradiographic evaluation was performed to measure the mean counts of the denervated and control tibialis anterior muscles at one week. In addition, immunohistochemistry was used to identify alpha7-nAChR-positive areas in denervated and control tibialis anterior muscles at one week (n = 6). Furthermore, a blocking study was conducted with methyllycaconitine (MLA, n = 5). RESULTS: 18F-ASEM PET/MRI showed significantly increased 18F-ASEM uptake in the denervated tibialis anterior muscle relative to the control side one week and three weeks post-denervation. SUVmax of the denervated muscles at one week and three weeks showed significantly higher uptake than the control (P = 0.0033 and 0.0277, respectively). The relative uptake by autoradiography for the denervated muscle was significantly higher than in the control, and immunohistochemistry revealed significantly greater alpha7-nAChR expression in the denervated muscle (P = 0.0277). In addition, the blocking study showed no significant 18F-ASEM uptake in the denervated side when compared to the control (P = 0.0796). CONCLUSIONS: Our results suggest that nAChR imaging with 18F-ASEM has potential as a noninvasive diagnostic method for peripheral nervous system disorders.

Direct Differentiation of Bone Marrow Mononucleated Cells Into Insulin-Producing Cells Using 4 Specific Soluble Factors.

Bone marrow-derived stem cells are self-renewing and multipotent adult stem cells that differentiate into several types of cells. Here, we investigated a unique combination of 4 differentiation-inducing factors (DIFs), including putrescine (Put), glucosamine (GlcN), nicotinamide, and BP-1-102, to develop a differentiation method for inducing mature insulin-producing cells (IPCs) and apply this method to bone marrow mononucleated cells (BMNCs) isolated from mice. BMNCs, primed with the 4 soluble DIFs, were differentiated into functional IPCs. BMNCs cultured under the defined conditions synergistically expressed multiple genes, including those for PDX1, NKX6.1, MAFA, NEUROG3, GLUT2, and insulin, related to pancreatic beta cell development and function. They produced insulin/C-peptide and PDX1, as assessed using immunofluorescence and flow cytometry. The induced cells secreted insulin in a glucose-responsive manner, similar to normal pancreatic beta cells. Grafting BMNC-derived IPCs under kidney capsules of mice with streptozotocin (STZ)-induced diabetes alleviated hyperglycemia by lowering blood glucose levels, enhancing glucose tolerance, and improving glucose-stimulated insulin secretion. Insulin- and PDX1-expressing cells were observed in the IPC-bearing graft sections of nephrectomized mice. Therefore, this study provides a simple protocol for BMNC differentiation, which can be a novel approach for cell-based therapy in diabetes mellitus.

MR Template-Based Individual Brain PET Volumes-of-Interest Generation Neither Using MR nor Using Spatial Normalization.

For more anatomically precise quantitation of mouse brain PET, spatial normalization (SN) of PET onto MR template and subsequent template volumes-of-interest (VOIs)-based analysis are commonly used. Although this leads to dependency on the corresponding MR and the process of SN, routine preclinical/clinical PET images cannot always afford corresponding MR and relevant VOIs. To resolve this issue, we propose a deep learning (DL)-based individual-brain-specific VOIs (i.e., cortex, hippocampus, striatum, thalamus, and cerebellum) directly generated from PET images using the inverse-spatial-normalization (iSN)-based VOI labels and deep convolutional neural network model (deep CNN). Our technique was applied to mutated amyloid precursor protein and presenilin-1 mouse model of Alzheimer's disease. Eighteen mice underwent T2-weighted MRI and 18F FDG PET scans before and after the administration of human immunoglobin or antibody-based treatments. To train the CNN, PET images were used as inputs and MR iSN-based target VOIs as labels. Our devised methods achieved decent performance in terms of not only VOI agreements (i.e., Dice similarity coefficient) but also the correlation of mean counts and SUVR, and CNN-based VOIs was highly accordant with ground-truth (the corresponding MR and MR template-based VOIs). Moreover, the performance metrics were comparable to that of VOI generated by MR-based deep CNN. In conclusion, we established a novel quantitative analysis method both MR-less and SN-less fashion to generate individual brain space VOIs using MR template-based VOIs for PET image quantification. Supplementary Information: The online version contains supplementary material available at 10.1007/s13139-022-00772-4.

Unified Deep Learning-Based Mouse Brain MR Segmentation: Template-Based Individual Brain Positron Emission Tomography Volumes-of-Interest Generation Without Spatial Normalization in Mouse Alzheimer Model.

Although skull-stripping and brain region segmentation are essential for precise quantitative analysis of positron emission tomography (PET) of mouse brains, deep learning (DL)-based unified solutions, particularly for spatial normalization (SN), have posed a challenging problem in DL-based image processing. In this study, we propose an approach based on DL to resolve these issues. We generated both skull-stripping masks and individual brain-specific volumes-of-interest (VOIs-cortex, hippocampus, striatum, thalamus, and cerebellum) based on inverse spatial normalization (iSN) and deep convolutional neural network (deep CNN) models. We applied the proposed methods to mutated amyloid precursor protein and presenilin-1 mouse model of Alzheimer's disease. Eighteen mice underwent T2-weighted MRI and 18F FDG PET scans two times, before and after the administration of human immunoglobulin or antibody-based treatments. For training the CNN, manually traced brain masks and iSN-based target VOIs were used as the label. We compared our CNN-based VOIs with conventional (template-based) VOIs in terms of the correlation of standardized uptake value ratio (SUVR) by both methods and two-sample t-tests of SUVR % changes in target VOIs before and after treatment. Our deep CNN-based method successfully generated brain parenchyma mask and target VOIs, which shows no significant difference from conventional VOI methods in SUVR correlation analysis, thus establishing methods of template-based VOI without SN.

PET-Based Radiogenomics Supports mTOR Pathway Targeting for Hepatocellular Carcinoma.

PURPOSE: This work aimed to explore in depth the genomic and molecular underpinnings of hepatocellular carcinoma (HCC) with increased 2[18F]fluoro-2-deoxy-d-glucose (FDG) uptake in PET and to identify therapeutic targets based on this imaging-genomic surrogate. EXPERIMENTAL DESIGN: We used RNA sequencing and whole-exome sequencing data obtained from 117 patients with HCC who underwent hepatic resection with preoperative FDG-PET/CT imaging as a discovery cohort. The primary radiogenomic results were validated with transcriptomes from a second cohort of 81 patients with more advanced tumors. All patients were allocated to an FDG-avid or FDG-non-avid group according to the PET findings. We also screened potential drug candidates targeting FDG-avid HCCs in vitro and in vivo. RESULTS: High FDG avidity conferred worse recurrence-free survival after HCC resection. Whole transcriptome analysis revealed upregulation of mTOR pathway signals in the FDG-avid tumors, together with higher abundance of associated mutations. These clinical and genomic findings were replicated in the validation set. A molecular signature of FDG-avid HCCs identified in the discovery set consistently predicted poor prognoses in the public-access datasets of two cohorts. Treatment with an mTOR inhibitor resulted in decreased FDG uptake followed by effective tumor control in both the hyperglycolytic HCC cell lines and xenograft mouse models. CONCLUSIONS: Our PET-based radiogenomic analysis indicates that mTOR pathway genes are markedly activated and altered in HCCs with high FDG retention. This nuclear imaging biomarker may stimulate umbrella trials and tailored treatments in precision care of patients with HCC.

PET Imaging of System xC - in Immune Cells for Assessment of Disease Activity in Mice and Patients with Inflammatory Bowel Disease.

We aimed to explore whether the imaging of antiporter system xC - of immune cells with (4S)-4-(3-18F-fluoropropyl)-l-glutamate (18F-FSPG) PET can assess inflammatory bowel disease (IBD) activity in murine models and patients (NCT03546868). Methods: 18F-FSPG PET imaging was performed to assess IBD activity in mice with dextran sulfate sodium-induced and adoptive T-cell transfer-induced IBD and a cohort of 20 patients at a tertiary care center in South Korea. Immunohistochemical analysis of system xC - and cell surface markers was also studied. Results: Mice with experimental IBD showed increased intestinal 18F-FSPG uptake and xCT expression in cells positive (+) for CD11c, F4/80, and CD3 in the lamina propria, increases positively associated with clinical and pathologic disease activity. 18F-FSPG PET studies in patients, most of whom were clinically in remission or had mildly active IBD, showed that PET imaging was sufficiently accurate in diagnosing endoscopically active IBD and remission in patients and bowel segments. 18F-FSPG PET correctly identified all 9 patients with superficial or deep ulcers. Quantitative intestinal 18F-FSPG uptake was strongly associated with endoscopic indices of IBD activity. The number of CD68+xCT+ and CD3+xCT+ cells in 22 bowel segments from patients with ulcerative colitis and the number of CD68+xCT+ cells in 7 bowel segments from patients with Crohn disease showed a significant positive association with endoscopic indices of IBD activity. Conclusion: The assessment of system xC - in immune cells may provide diagnostic information on the immune responses responsible for chronic active inflammation in IBD. 18F-FSPG PET imaging of system xC - activity may noninvasively assess the IBD activity.

Smart Vitamin Micelles as Cancer Nanomedicines for Enhanced Intracellular Delivery of Doxorubicin.

Chemotherapy is one of the most effective treatments for cancer. However, intracellular delivery of many anticancer drugs is hindered by their hydrophobicity and low molecular weight. Here, we describe highly biocompatible and biodegradable amphiphilic vitamin conjugates comprising hydrophobic vitamin E and hydrophilic vitamin B labeled with dual pH and glutathione-responsive degradable linkages. Vitamin-based micelles (vitamicelles), formed by self-assembly in aqueous solutions, were optimized based on their stability after encapsulation of doxorubicin (DOX). The resulting vitamicelles have great potential as vehicles for anticancer drugs because they show excellent biocompatibility (>94% after 48 h of incubation) and rapid biodegradability (>90% after 2.5 h). Compared with free DOX, DOX-loaded vitamicelles showed a markedly enhanced anticancer effect as they released the drug rapidly and inhibited drug efflux out of cells efficiently. By exploiting these advantages, this study not only provides a promising strategy for circumventing existing challenges regarding the delivery of anticancer drugs but also extends the utility of current DOX-induced chemotherapy.

Dual pH- and GSH-Responsive Degradable PEGylated Graphene Quantum Dot-Based Nanoparticles for Enhanced HER2-Positive Breast Cancer Therapy.

Dual stimuli-responsive degradable carbon-based nanoparticles (DS-CNPs) conjugated with Herceptin (HER) and polyethylene glycol (PEG) have been designed for the treatment of HER2-positive breast cancer. Each component has been linked through disulfide linkages that are sensitive to glutathione in a cancer microenvironment. ß-cyclodextrin (ß-CD) on the surface of DS-CNPs formed an inclusion complex (DL-CNPs) with doxorubicin (DOX) at a high loading capacity of 5.3 ± 0.4%. In response to a high level of glutathione (GSH) and low pH in a tumor environment, DL-CNPs were rapidly degraded and released DOX in a controlled manner via disruption of host-guest inclusion. These novel DL-CNPs exhibited high cellular uptake with low toxicity, which induced the efficient inhibition of antitumor activity both in vitro and in vivo. Cell viability, confocal laser scanning microscopy, and animal studies indicate that DL-CNPs are a great platform with a synergistically enhanced antitumor effect from the dual delivery of HER and DOX in DL-CNPs.

Feasibility of real-time in vivo 89Zr-DFO-labeled CAR T-cell trafficking using PET imaging.

INTRODUCTION: Chimeric antigen receptor (CAR) T-cells have been recently developed and are producing impressive outcomes in patients with hematologic malignancies. However, there is no standardized method for cell trafficking and in vivo CAR T-cell monitoring. We assessed the feasibility of real-time in vivo 89Zr-p-Isothiocyanatobenzyl-desferrioxamine (Df-Bz-NCS, DFO) labeled CAR T-cell trafficking using positron emission tomography (PET). RESULTS: The 89Zr-DFO radiolabeling efficiency of Jurkat/CAR and human peripheral blood mononuclear cells (hPBMC)/CAR T-cells was 70%-79%, and cell radiolabeling activity was 98.1-103.6 kBq/106 cells. Cell viability after radiolabeling was >95%. Cell proliferation was not significantly different during the early period after radiolabeling, compared with unlabeled cells; however, the proliferative capacity decreased over time (day 7 after labeling). IL-2 or IFN-γ secretion was not significantly different between unlabeled and labeled CAR T-cells. PET/magnetic resonance imaging in the xenograft model showed that most of the 89Zr-DFO-labeled Jurkat/CAR T-cells were distributed in the lung (24.4% ± 3.4%ID) and liver (22.9% ± 5.6%ID) by one hour after injection. The cells gradually migrated from the lung to the liver and spleen by day 1, and remained stable in these sites until day 7 (on day 7: lung 3.9% ± 0.3%ID, liver 36.4% ± 2.7%ID, spleen 1.4% ± 0.3%ID). No significant accumulation of labeled cells was identified in tumors. A similar pattern was observed in ex vivo biodistributions on day 7 (lung 3.0% ± 1.0%ID, liver 19.8% ± 2.2%ID, spleen 2.3% ± 1.7%ID). 89Zr-DFO-labeled hPBMC/CAR T-cells showed a similar distribution, compared with Jurkat/CAR T-cells, on serial PET images. CAR T cell distribution was cross-confirmed by flow cytometry, Alu polymerase chain reaction, and immunohistochemistry. CONCLUSION: Real-time in vivo cell trafficking is feasible using PET imaging of 89Zr-DFO-labeled CAR T-cells. This can be used to investigate cellular kinetics, initial in vivo biodistribution, and safety profiles in future CAR T-cell development.

Diagnostic accuracy and safety of 16α-[18F]fluoro-17ß-oestradiol PET-CT for the assessment of oestrogen receptor status in recurrent or metastatic lesions in patients with breast cancer: a prospective cohort study.

BACKGROUND: A biopsy of first recurrence or metastatic disease is recommended to re-evaluate oestrogen receptor status in patients with breast cancer and to select appropriate treatment. However, retesting for oestrogen receptor status with rebiopsy is not always feasible, depending on lesion location and the risk associated with biopsy, and in these cases clinicians continue to treat patients according to the oestrogen receptor status of the primary tumour. Consequently suboptimal therapy might be offered to these patients. We assessed the diagnostic accuracy and safety of 16α-[18F]fluoro-17ß-oestradiol (18F-FES) PET-CT to assess oestrogen receptor status in patients with recurrent or metastatic breast cancer. METHODS: We did a prospective cohort study at the Asan Medical Center, Seoul, South Korea. Eligible patients had breast cancer, with first recurrence or metastatic disease at presentation, were 19 years or older, and had an Eastern Cooperative Oncology Group performance status of 0-2. The primary objective was to show the agreement between qualitative 18F-FES PET-CT interpretation and the results of oestrogen receptor expression by immunohistochemical assay, a non-reference standard test. Whole-body 18F-FES PET-CT imaging was done after intravenous injection of 111-222 MBq of 18F-FES, with dosing primarily determined by radiation dosimetry analysis. 18F-FES uptake above background intensity was interpreted as positive. Efficacy was assessed in all patients with histologically confirmed recurrent or metastatic breast cancer who received 18F-FES and had PET-CT images available (intention-to-diagnose analysis), and safety was assessed in all patients who received 18F-FES. This study is registered with ClinicalTrials.gov, number NCT01986569. FINDINGS: Between Nov 27, 2013, and Nov 10, 2016, 93 patients were enrolled. Of the 85 patients included in the efficacy analysis, 47 (55%) were oestrogen receptor-positive and 38 (45%) were oestrogen receptor-negative. Positive status percent agreement between the 18F-FES PET-CT results and oestrogen receptor status by immunohistochemical assay was 76·6% (95% CI 62·0-87·7) and the negative status percent agreement was 100·0% (90·8-100·0). Patients who were oestrogen receptor-positive and had a positive 18F-FES PET-CT result had a significantly higher progesterone receptor expression than those who were oestrogen receptor-positive and had a negative 18F-FES PET-CT result (23 [68%] of 34 patients vs 0 of 11 patients; p<0·0001). The most common adverse event was procedural pain in nine (10%) of 90 patients injected with 18F-FES. No adverse events were related to the study drug except injection site pain in one (1%) patient. No serious adverse events were recorded. INTERPRETATION: The high negative percent agreement between 18F-FES PET-CT and oestrogen receptor status by immunohistochemical assay in this cohort suggests that positive 18F-FES uptake by recurrent or metastatic oestrogen receptor-positive breast cancer lesions could be an alternative to oestrogen receptor assays in this setting. Staging assessment should include 18F-FES PET-CT when retesting oestrogen receptor status is not feasible. FUNDING: Asan Institute for Life Sciences, Ministry of Health and Welfare, South Korea.

Tartary Buckwheat Extract Attenuated the Obesity-Induced Inflammation and Increased Muscle PGC-1a/SIRT1 Expression in High Fat Diet-Induced Obese Rats.

Obesity is intimately related to a chronic inflammatory state, with augmentation of macrophage infiltration and pro-inflammatory cytokine secretion in white adipose tissue (WAT) and mitochondrial dysfunction in skeletal muscle. The specific aim of this study is to evaluate effects of tartary buckwheat extract (TB) on obesity-induced adipose tissue inflammation and muscle peroxisome proliferator-activated receptor-γ coactivator (PGC)-1α/sirtulin 1 (SIRT1) pathway in rats fed a high-fat diet. Sprague-Dawley rats were divided into four groups and fed either a normal diet (NOR), 45% high-fat diet (HF), HF + low dose of TB (TB-L; 5 g/kg diet), or HF + high dose of TB (TB-H; 10 g/kg diet) for 13 weeks. TB significantly reduced adipose tissue mass with decreased adipogenic gene expression of PPAR-γ and aP2. Serum nitric oxide levels and adipose tissue macrophage M1 polarization gene markers, such as iNOS, CD11c, and Arg1, and pro-inflammatory gene expression, including TNF-α, IL-6, and MCP-1, were remarkably downregulated in the TB-L and TB-H groups. Moreover, TB supplementation increased gene expression of PGC-1α and SIRT1, involved in muscle biogenesis and function. These results suggested that TB might attenuate obesity-induced inflammation and mitochondrial dysfunction by modulating adipose tissue inflammation and the muscle PGC-1α/SIRT1 pathway.

Formulation and Characterization of Quercetin-loaded Oil in Water Nanoemulsion and Evaluation of Hypocholesterolemic Activity in Rats.

Due to poor water solubility and high susceptibility to chemical degradation, the applications of quercetin have been limited. This study investigated the effects of pH on the formation of quercetin-loaded nanoemulsion (NQ) and compared the hypocholesterolemic activity between quercetin and NQ to utilize the quercetin as functional food ingredient. NQ particle size exhibited a range of 207⁻289 nm with polydispersity index range (<0.47). The encapsulation efficiency increased stepwise from 56 to 92% as the pH increased from 4.0 to 9.0. Good stability of NQ was achieved in the pH range of 6.5⁻9.0 during 3-month storage at 21 and 37 °C. NQ displayed higher efficacy in reducing serum and hepatic cholesterol levels and increasing the release of bile acid into feces in rats fed high-cholesterol diet, compared to quercetin alone. NQ upregulated hepatic gene expression involved in bile acid synthesis and cholesterol efflux, such as cholesterol 7 alpha-hydroxylase (CYP7A1), liver X receptor alpha (LXRα), ATP-binding cassette transporter A1 (ABCA1) and ATP-binding cassette sub-family G member 1 (ABCG1). These results suggest at least partial involvement of hepatic bile acid synthesis and fecal cholesterol excretion in nanoemulsion quercetin-mediated beneficial effect on lipid abnormalities.

Clusterin contributes to early stage of Alzheimer's disease pathogenesis.

While clusterin is reportedly involved in Alzheimer's disease (AD) pathogenesis, how clusterin interacts with amyloid-ß (Aß) to cause Aß neurotoxicity remains unclear in vivo. Using 5×FAD transgenic mice, which develop robust AD pathology and memory deficits when very young, we detected interactions between clusterin and Aß in the mouse brains. The two proteins were concurrently upregulated and bound or colocalized with each other in the same complexes or in amyloid plaques. Neuropathology and cognitive performance were assessed in the progeny of clusterin-null mice crossed with 5×FAD mice, yielding clu-/- ;5×FAD and clu+/+ ;5×FAD. We found far less of the various pools of Aß proteins, most strikingly soluble Aß oligomers and amyloid plaques in clu-/- ;5×FAD mice at 5 months of age. At that age, those mice also had higher levels of neuronal and synaptic proteins and better motor coordination, spatial learning and memory than age-matched clu+/+ ;5×FAD mice. However, at 10 months of age, these differences disappeared, with Aß and plaque deposition, neuronal and synaptic proteins and impairment of behavioral and cognitive performance similar in both groups. These findings demonstrate that clusterin is necessarily involved in early stages of AD pathogenesis by enhancing toxic Aß pools to cause Aß-directed neurodegeneration and behavioral and cognitive impairments, but not in late stage.

Schedule-dependent synergistic effects of 5-fluorouracil and selumetinib in KRAS or BRAF mutant colon cancer models.

Combination of MEK inhibitor and 5-FU had showed limited efficacy in clinical trials. We previously reported that acquired resistance to 5-FU was related with continued activation of salvage pathway. Here we investigated whether combination of 5-FU and a MEK inhibitor had treatment sequence-dependent synergistic effects in KRAS or BRAF mutant colon cancer models. Treatment with 5-FU followed by selumetinib (FS) induced highest cell death and synergy compared with reverse (SF) and concomitant (cFS) treatment in six cell lines. SF or cFS combination induced synergy in 1 or 2 cell lines, respectively, in which the synergy was less than that by FS combination. FS enhanced apoptosis and decreased anchorage-independent growth. Induction of thymidine kinase 1, a rate-limiting enzyme in salvage pathway, by 5-FU was abrogated by subsequent treatment with selumetinib, and ERK reactivation after selumetinib was prohibited by pretreatment with 5-FU. FS altered mRNA expression in groups of genes distinct from SF. Administration of 5-FU (10 or 30 mg/kg/day) for 7 days, followed by selumetinib (10 or 25 mg/kg/day) for another 7 days, in colo205 and HCT8 xenograft models significantly decreased tumor growth compared with a single agent. However, co-administration in the reverse sequence did not show the difference in tumor size compared with the treatment of single agent. Decreased expression of Ki67 was observed in tumors from mice treated with FS. Our results suggest that sequential administration of 5-FU plus selumetinib would be a promising strategy for patients having KRAS or BRAF mutant colon cancers.

Panax ginseng as an adjuvant treatment for Alzheimer's disease.

Longevity in medicine can be defined as a long life without mental or physical deficits. This can be prevented by Alzheimer's disease (AD). Current conventional AD treatments only alleviate the symptoms without reversing AD progression. Recent studies demonstrated that Panax ginseng extract improves AD symptoms in patients with AD, and the two main components of ginseng might contribute to AD amelioration. Ginsenosides show various AD-related neuroprotective effects. Gintonin is a newly identified ginseng constituent that contains lysophosphatidic acids and attenuates AD-related brain neuropathies. Ginsenosides decrease amyloid ß-protein (Aß) formation by inhibiting ß- and γ-secretase activity or by activating the nonamyloidogenic pathway, inhibit acetylcholinesterase activity and Aß-induced neurotoxicity, and decrease Aß-induced production of reactive oxygen species and neuroinflammatory reactions. Oral administration of ginsenosides increases the expression levels of enzymes involved in acetylcholine synthesis in the brain and alleviates Aß-induced cholinergic deficits in AD models. Similarly, gintonin inhibits Aß-induced neurotoxicity and activates the nonamyloidogenic pathway to reduce Aß formation and to increase acetylcholine and choline acetyltransferase expression in the brain through lysophosphatidic acid receptors. Oral administration of gintonin attenuates brain amyloid plaque deposits, boosting hippocampal cholinergic systems and neurogenesis, thereby ameliorating learning and memory impairments. It also improves cognitive functions in patients with AD. Ginsenosides and gintonin attenuate AD-related neuropathology through multiple routes. This review focuses research demonstrating that ginseng constituents could be a candidate as an adjuvant for AD treatment. However, clinical investigations including efficacy and tolerability analyses may be necessary for the clinical acceptance of ginseng components in combination with conventional AD drugs.

Ginger Extract Ameliorates Obesity and Inflammation via Regulating MicroRNA-21/132 Expression and AMPK Activation in White Adipose Tissue.

Ginger is a plant whose rhizome is used as a spice or folk medicine. We aimed to investigate the effect of ginger root extract on obesity and inflammation in rats fed a high-fat diet. Sprague-Dawley rats were divided into three groups and fed either a 45% high-fat diet (HF), HF + hot-water extract of ginger (WEG; 8 g/kg diet), or HF + high-hydrostatic pressure extract of ginger (HPG; 8 g/kg diet) for 10 weeks. The HPG group had lower body weight and white adipose tissue (WAT) mass compared to the HF group. Serum and hepatic lipid levels of HPG group were lower, while fecal lipid excretion of the HPG group was higher than that of the HF group. In the WAT of the WEG and HPG groups, mRNA levels of adipogenic genes were lower than those of the HF group. Moreover, HPG group had lower mRNA levels of pro-inflammatory cytokines than did the HF group. MicroRNA (miR)-21 expression was down-regulated by both WEG and HPG. Additionally, miR-132 expression was down-regulated by HPG. The adenosine monophosphate-activated protein kinase (AMPK) activity of HPG group was greater than that of the HF group. HPG may have beneficial effects on obesity and inflammation, partially mediated by regulation of miR-21/132 expression and AMPK activation in WAT.

[18F]fluorothymidine PET Informs the Synergistic Efficacy of Capecitabine and Trifluridine/Tipiracil in Colon Cancer.

In cancer therapy, enhanced thymidine uptake by the salvage pathway can bypass dTMP depletion, thereby conferring resistance to thymidylate synthase inhibition. We investigated whether sequential combination therapy of capecitabine and trifluridine/tipiracil (TAS-102) could synergistically enhance antitumor efficacy in colon cancer xenograft models. We also examined 3'-deoxy-3'-[18F]fluorothymidine ([18F]FLT) PET as a means to predict therapeutic response to a sequential combination of capecitabine and trifluridine/tipiracil. [3H]FLT uptake after 5-fluorouracil treatment in vitro and [18F]FLT uptake after capecitabine (360 mg/kg/day) in athymic nude mice (Balb/c-nu) with xenografts (n = 10-12 per group) were measured using eight human colon cancer cell lines. We determined the synergistic effects of sequential combinations of 5-fluorouracil and trifluridine in vitro as well as the sequential combination of oral capecitabine (30-360 mg/kg) and trifluridine/tipiracil (trifluridine 75 or 150 mg/kg with tipiracil) in six xenograft models (n = 6-10 per group). We observed significant increases in [3H]FLT uptake in all cell lines and [18F]FLT uptake in five xenograft models after 5-fluorouracil and capecitabine treatment, respectively. Increased [18F]FLT uptake after capecitabine followed by extinction of uptake correlated strongly with tumor growth inhibition (ρ = -0.81, P = 0.02). The effects of these combinations were synergistic in vitro A synergy for sequential capecitabine and trifluridine/tipiracil was found only in mouse xenograft models showing increased [18F]FLT uptake after capecitabine. Our results suggest that the sequential combination of capecitabine and trifluridine/tipiracil is synergistic in tumors with an activated salvage pathway after capecitabine treatment in mice, and [18F]FLT PET imaging may predict the response to capecitabine and the synergistic antitumor efficacy of a sequential combination of capecitabine and trifluridine/tipiracil. Cancer Res; 77(24); 7120-30. ©2017 AACR.