Multicenter clinical radiomics-integrated model based on [18F]FDG PET and multi-modal MRI predict ATRX mutation status in IDH-mutant lower-grade gliomas.

OBJECTIVES: To develop a clinical radiomics-integrated model based on 18 F-fluorodeoxyglucose positron emission tomography ([18F]FDG PET) and multi-modal MRI for predicting alpha thalassemia/mental retardation X-linked (ATRX) mutation status of IDH-mutant lower-grade gliomas (LGGs). METHODS: One hundred and two patients (47 ATRX mutant-type, 55 ATRX wild-type) diagnosed with IDH-mutant LGGs (CNS WHO grades 1 and 2) were retrospectively enrolled. A total of 5540 radiomics features were extracted from structural MR (sMR) images (contrast-enhanced T1-weighted imaging, CE-T1WI; T2-weighted imaging, and T2WI), functional MR (fMR) images (apparent diffusion coefficient, ADC; cerebral blood volume, CBV), and metabolic PET images ([18F]FDG PET). The random forest algorithm was used to establish a clinical radiomics-integrated model, integrating the optimal multi-modal radiomics model with three clinical parameters. The predictive effectiveness of the models was evaluated by receiver operating characteristic (ROC) and decision curve analysis (DCA). RESULTS: The optimal multi-modal model incorporated sMR (CE-T1WI), fMR (ADC), and metabolic ([18F]FDG) images ([18F]FDG PET+ADC+ CE-T1WI) with the area under curves (AUCs) in the training and test groups of 0.971 and 0.962, respectively. The clinical radiomics-integrated model, incorporating [18F]FDG PET+ADC+CE-T1WI, three clinical parameters (KPS, SFSD, and ATGR), showed the best predictive effectiveness in the training and test groups (0.987 and 0.975, respectively). CONCLUSIONS: The clinical radiomics-integrated model with metabolic, structural, and functional information based on [18F]FDG PET and multi-modal MRI achieved promising performance for predicting the ATRX mutation status of IDH-mutant LGGs. KEY POINTS: • The clinical radiomics-integrated model based on [18F]FDG PET and multi-modal MRI achieved promising performance for predicting ATRX mutation status in LGGs. • The study investigated the value of multicenter clinical radiomics-integrated model based on [18F]FDG PET and multi-modal MRI in LGGs regarding ATRX mutation status prediction. • The integrated model provided structural, functional, and metabolic information simultaneously and demonstrated with satisfactory calibration and discrimination in the training and test groups (0.987 and 0.975, respectively).

Gut microbiota is associated with response to 131I therapy in patients with papillary thyroid carcinoma.

PURPOSE: Radioactive iodine (131I) therapy is a conventional post-surgery treatment widely used for papillary thyroid carcinoma (PTC). Since 131I is orally administered, we hypothesize that it may affect gut microbiome. This study aims to investigate alterations of intestinal microbiome caused by 131I therapy in PTC patients and explore its association with response to 131I therapy. METHODS: Fecal samples of 60 PTC patients pre- and post-131I therapy were collected to characterize the 131I therapy-induced gut microbiota alterations using 16S rRNA gene sequencing. According to the inclusion criteria, sequence data of 40 out of the 60 patients, divided into excellent response (ER) group and non-excellent response (NER) group, were recruited to investigate the possible connection between gut microbiota and response to 131I therapy. Multivariate binary logistic regression was employed to construct a predictive model for response to 131I therapy. RESULTS: Microbial richness, diversity, and composition were tremendously altered by 131I therapy. A significant decline of Firmicutes to Bacteroides (F/B) ratio was observed post-131I therapy. 131I therapy also led to changes of gut microbiome-related metabolic pathways. Discrepancies in ß diversity were found between ER and NER groups both pre- and post-131I therapy. Furthermore, a predictive model for response to 131I therapy with a p value of 0.003 and an overall percentage correct of 80.0% was established, with three variables including lymph node metastasis, relative abundance of g_Bifidobacterium and g_Dorea. Among them, g_Dorea was identified to be an in independent predictor of response to 131I therapy (p = 0.04). CONCLUSION: For the first time, the present study demonstrates the gut microbial dysbiosis caused by 131I therapy in post-surgery PTC patients and reveals a previously undefined role of gut microbiome as predictor for 131I ablation response. G_Dorea and g_Bifidobacterium may be potential targets for clinical intervention to improve response to 131I in post-operative PTC patients. TRIAL REGISTRATION: ChiCTR2100048000. Registered 28 June 2021.

Early diagnosis of breast cancer lung metastasis by nanoprobe-based luminescence imaging of the pre-metastatic niche.

BACKGROUND: Early detection of breast cancer lung metastasis remains highly challenging, due to few metastatic cancer cells at an early stage. Herein we propose a new strategy for early diagnosis of lung metastasis of breast cancer by luminescence imaging of pulmonary neutrophil infiltration via self-illuminating nanoprobes. METHODS: Luminescent nanoparticles (LAD NPs) were engineered using a biocompatible, neutrophil-responsive self-illuminating cyclodextrin material and an aggregation-induced emission agent. The chemiluminescence resonance energy transfer (CRET) effect and luminescence properties of LAD NPs were fully characterized. Using mouse peritoneal neutrophils, in vitro luminescence properties of LAD NPs were thoroughly examined. In vivo luminescence imaging and correlation analyses were performed in mice inoculated with 4T1 cancer cells. Moreover, an active targeting nanoprobe was developed by surface decoration of LAD NPs with a neutrophil-targeting peptide, which was also systemically evaluated by in vitro and in vivo studies. RESULTS: LAD NPs can generate long-wavelength and persistent luminescence due to the CRET effect. In a mouse model of 4T1 breast cancer lung metastasis, we found desirable correlation between neutrophils and tumor cells in the lungs, demonstrating the effectiveness of early imaging of the pre-metastatic niche by the newly developed LAD NPs. The active targeting nanoprobe showed further enhanced luminescence imaging capability for early detection of pulmonary metastasis. Notably, the targeting nanoprobe-based luminescence imaging strategy remarkably outperformed PET/CT imaging modalities in the examined mouse model. Also, preliminary tests demonstrated good safety of LAD NPs. CONCLUSIONS: The neutrophil-targeting imaging strategy based on newly developed luminescence nanoparticles can serve as a promising modality for early diagnosis of lung metastasis of breast cancers.

NIS and epithelial-mesenchymal transition marker expression of circulating tumor cells for predicting and monitoring the radioactive iodine-131 therapy effect in differentiated thyroid cancers.

Current methods, such as serum thyroglobulin measurement and medical imaging, have limitations in the routine monitoring of the disease status and treatment response of patients with differentiated thyroid cancers (DTCs), and additional methods remain to be explored. The aim of this study was to investigate the clinical value of the sodium/iodide symporter (NIS) expression and epithelial-mesenchymal transition (EMT) phenotypes of circulating tumor cells (CTCs) in monitoring the disease status and treatment response of DTC. Blood samples were obtained from DTC patients before (1 to 3 months after total thyroidectomy) and 4 to 6 months after radioactive iodine-131 (RAI) therapy for the CTC assessments. The number, NIS expression, and EMT phenotypes of CTCs were enumerated and characterized with CanPatrol™ CTC enrichment and mRNA in situ hybridization. Postoperative NIS high expression was independently correlated with a better response to first RAI therapy and good treatment efficacy. Postoperative NIS-/epithelial-/mesenchymal+ CTCs presence was independently correlated with a worse response to first RAI therapy. The numbers of total NIS+ CTCs and NIS+/epithelial+/mesenchymal+ CTCs after first RAI therapy were negatively correlated with a better response to RAI therapy only in univariate analyses. Univariate and multivariate analyses showed that a decreased or unchanged number of total NIS+ CTCs after RAI therapy may denote good efficacy and effective RAI therapy. These preliminary data suggest that assessment of the NIS expression and EMT phenotypes of CTCs may serve as potential adjuncts for predicting and monitoring the curative effect of RAI therapy in DTC patients and avoid ineffective treatment. Further validation is warranted.

Solute Carrier Organic Anion Transporter Family Member 3A1 Is a Bile Acid Efflux Transporter in Cholestasis.

BACKGROUND & AIMS: Bile acid transporters maintain bile acid homeostasis. Little is known about the functions of some transporters in cholestasis or their regulatory mechanism. We investigated the hepatic expression of solute carrier organic anion transporter family member 3A1 (SLCO3A1, also called OATP3A1) and assessed its functions during development of cholestasis. METHODS: We measured levels of OATP3A1 protein and messenger RNA and localized the protein in liver tissues from 22 patients with cholestasis and 21 patients without cholestasis, using real-time quantitative polymerase chain reaction, immunoblot, and immunofluorescence analyses. We performed experiments with Slco3a1-knockout and C57BL/6J (control) mice. Mice and Sprague-Dawley rats underwent bile duct ligation (BDL) or a sham operation. Some mice were placed on a 1% cholic acid (CA) diet to induce cholestasis or on a control diet. Serum and liver tissues were collected and analyzed; hepatic levels of bile acids and 7-α-C4 were measured using liquid chromatography/mass spectrometry. Human primary hepatocytes and hepatoma (PLC/PRF/5) cell lines were used to study mechanisms that regulate OATP3A1 expression and transport. RESULTS: Hepatic levels of OATP3A1 messenger RNA and protein were significantly increased in liver tissues from patients with cholestasis and from rodents with BDL or 1% CA diet-induced cholestasis. Levels of fibroblast growth factor 19 (FGF19, FGF15 in rodents) were also increased in liver tissues from patients and rodents with cholestasis. FGF19 signaling activated the Sp1 transcription factor and nuclear factor κB to increase expression of OATP3A1 in hepatocytes; we found binding sites for these factors in the SLCO3A1 promoter. Slco3a1-knockout mice had shorter survival times and increased hepatic levels of bile acid, and they developed more liver injury after the 1% CA diet or BDL than control mice. In hepatoma cell lines, we found OATP3A1 to take prostaglandin E2 and thyroxine into cells and efflux bile acids. CONCLUSIONS: We found levels of OATP3A1 to be increased in cholestatic liver tissues from patients and rodents compared with healthy liver tissues. We show that OATP3A1 functions as a bile acid efflux transporter that is up-regulated as an adaptive response to cholestasis.

Novel small peptides derived from VEGF125-136: potential drugs for radioactive diagnosis and therapy in A549 tumor-bearing nude mice.

Vascular endothelial growth factor receptor (VEGFR) is a critical factor in tumor angiogenesis and has been considered a potential target for receptor-mediated radionuclide imaging and therapy. In this study, we identified two peptides (QKRKRKKSRKKH and RKRKRKKSRYIVLS) derived from VEGF125-136 that displayed high binding affinities to VEGFR and strong inhibition of A549 cell growth. 99mTc- and 188Re-labeled peptides displayed high labeling efficiency and favorable stability in saline and human plasma. At the cellular level, the radiolabeled peptides could bind with A549 cells and be internalized via the VEGFR-1-mediated pathway. 99mTc/188Re-labeled peptide was significantly accumulated at xenograft tumors, as observed with single-photon emission computed tomography (SPECT) planar imaging. Moreover, 188Re-labeled peptides significantly inhibited tumor growth, prolonged the survival time of the tumor-bearing nude mice and resulted in much more necrotic regions and apoptotic cells in the A549 xenograft tumors. These results demonstrated that these two peptides as candidate drugs for radionuclide imaging and tumor therapy.

Molecular imaging of fibrosis using a novel collagen-binding peptide labelled with 99mTc on SPECT/CT.

Fibrosis, closely related to chronic various diseases, is a pathological process characterised by the accumulation of collagen (largely collagen type I). Non-invasive methods are necessary for the diagnosis and follow-up of fibrosis. This study aimed to develop a collagen-targeted probe for the molecular imaging of fibrosis. We identified CPKESCNLFVLKD (CBP1495) as an original collagen-binding peptide using isothermal titration calorimetry and enzyme-linked immunosorbent assay. CBP1495 effectively bound to collagen type I (K d = 861 nM) and (GPO)9 (K d = 633 nM), a collagen mimetic peptide. Western blot and histochemistry validated CBP1495 targeting collagen in vitro and ex vivo. (Gly-(D)-Ala-Gly-Gly) was introduced to CBP1495 for coupling 99mTc. Labelling efficiency of 99mTc-CBP1495 was 95.06 ± 1.08 %. The physico-chemical properties, tracer kinetics and biodistribution of 99mTc-CBP1495 were carried out, and showed that the peptide stably chelated 99mTc in vitro and in vivo. SPECT/CT imaging with 99mTc-CBP1495 was performed in rat fibrosis models, and revealed that 99mTc-CBP1495 significantly accumulated in fibrotic lungs or livers of rats. Finally, 99mTc-CBP1495 uptake and hydroxyproline (Hyp), a specific amino acid of collagen, were quantitatively analysed. The results demonstrated that 99mTc-CBP1495 uptake was positvely correlated with Hyp content in lungs (P < 0.0001, r 2 = 0.8266) or livers (P < 0.0001, r 2 = 0.7581). Therefore, CBP1495 is a novel collagen-binding peptide, and 99mTc-labelled CBP1495 may be a promising radiotracer for the molecular imaging of fibrosis.

LDA-SVM-based EGFR mutation model for NSCLC brain metastases: an observational study.

Epidermal growth factor receptor (EGFR) activating mutations are a predictor of tyrosine kinase inhibitor effectiveness in the treatment of non-small-cell lung cancer (NSCLC). The objective of this study is to build a model for predicting the EGFR mutation status of brain metastasis in patients with NSCLC. Observation and model set-up. This study was conducted between January 2003 and December 2011 in 6 medical centers in Southwest China. The study included 31 NSCLC patients with brain metastases. Eligibility requirements were histological proof of NSCLC, as well as sufficient quantity of paraffin-embedded lung and brain metastases specimens for EGFR mutation detection. The linear discriminant analysis (LDA) method was used for analyzing the dimensional reduction of clinical features, and a support vector machine (SVM) algorithm was employed to generate an EGFR mutation model for NSCLC brain metastases. Training-testing-validation (3 : 1 : 1) processes were applied to find the best fit in 12 patients (validation test set) with NSCLC and brain metastases treated with a tyrosine kinase inhibitor and whole-brain radiotherapy. Primary and secondary outcome measures: EGFR mutation analysis in patients with NSCLC and brain metastases and the development of a LDA-SVM-based EGFR mutation model for NSCLC brain metastases patients. EGFR mutation discordance between the primary lung tumor and brain metastases was found in 5 patients. Using LDA, 13 clinical features were transformed into 9 characteristics, and 3 were selected as primary vectors. The EGFR mutation model constructed with SVM algorithms had an accuracy, sensitivity, and specificity for determining the mutation status of brain metastases of 0.879, 0.886, and 0.875, respectively. Furthermore, the replicability of our model was confirmed by testing 100 random combinations of input values. The LDA-SVM-based model developed in this study could predict the EGFR status of brain metastases in this small cohort of patients with NSCLC. Further studies with larger cohorts should be carried out to validate our findings in the clinical setting.

Modulation, bioinformatic screening, and assessment of small molecular peptides targeting the vascular endothelial growth factor receptor.

Vascular endothelial growth factor (VEGF) and VEGF receptor (VEGFR) are important factors in tumor growth and metastasis. Molecular probes or drugs designed to target VEGF/VEGFR interactions are crucial in tumor molecular imaging and targeted therapy. Bioinformatic methods enable molecular design based on the structure of bio-macromolecules and their interactions. This study was aimed to identify tumor-targeting small-molecule peptides with high affinity for VEGFR using bioinformatics screening. The VEGFR extracellular immunoglobulin-like modules Ig1-Ig3 were used as the target to systematically alter the primary peptide sequence of VEGF125-136. Molecular docking and surface functional group interaction methods were combined in an in silico screen for polypeptides, which in theory, would have higher affinities for VEGFR. In vitro receptor competition binding assays were used to assess the affinity of the putative VEGFR-binding polypeptides. Rhodamine-conjugated peptides were used to label and visualize peptide-binding sites on A549 cells. Using bioinformatic screening, we identified 20 polypeptides with potentially higher affinity for VEGFR. The polypeptides were capable of inhibiting the binding of (125)I-VEGF to VEGFR in a dose-dependent manner. The IC50 values of QKRKRKKSRKKH and RKRKRKKSRYIVLS (80 and 185 nmol/L, respectively) were significantly lower than that of VEGF125-136 (464 nmol/L); thus, the affinity of these peptides for VEGFR was 6- and 2.5-fold higher, respectively, than that of VEGF125-136. Rhodamine labeling of A549 cells revealed peptide binding mainly on the plasma membrane and in the cytoplasm. Bioinformatic approaches hold promise for the development of molecular imaging probes. Using this approach, we designed two peptides that showed higher affinity toward VEGFR. These polypeptides may be used as molecular probes or drugs targeting VEGFR, which can be utilized in molecular imaging and targeted therapy of certain tumors.

Screening of a specific peptide binding to VPAC1 receptor from a phage display peptide library.

BACKGROUND/PURPOSE: The VPAC1 receptor, a member of the vasoactive intestinal peptide receptors (VIPRs), is overexpressed in the most frequently occurring malignant tumors and plays a major role in the progression and angiogenesis of a number of malignancies. Recently, phage display has become widely used for many applications, including ligand generation for targeted imaging, drug delivery and therapy. In this work, we developed a panning procedure using a phage display peptide library to select a peptide that specifically binds to the VPAC1 receptor to develop a novel targeted probe for molecular imaging and therapy. METHODS: CHO-K1 cells stably expressing VPAC1 receptors (CHO-K1/VPAC1 cells) were used to select a VPAC1-binding peptide from a 12-mer phage peptide library. DNA sequencing and homologous analysis of the randomly selected phage clones were performed. A cellular ELISA was used to determine the most selectively binding peptide for further investigation. Binding specificity to the VPAC1 receptor was analyzed by competitive inhibition ELISA and flow cytometry. The binding ability of the selected peptide to CHO-K1/VPAC1 cells and colorectal cancer (CRC) cell lines was confirmed using fluorescence microscopy and flow cytometry. RESULTS: A significant enrichment of phages that specifically bound to CHO-K1/VPAC1 cells was obtained after four rounds of panning. Of the selected phage clones, 16 out of 60 shared the same peptide sequence, GFRFGALHEYNS, which we termed the VP2 peptide. VP2 and vasoactive intestinal peptide (VIP) competitively bound to the VPAC1 receptor. More importantly, we confirmed that VP2 specifically bound to CHO-K1/VPAC1 cells and several CRC cell lines. CONCLUSION: Our results demonstrate that the VP2 peptide could specifically bind to VPAC1 receptor and several CRC cell lines. And VP2 peptide may be a potential candidate to be developed as a useful diagnostic molecular imaging probe for early detection of CRC.

Detection of invasive Candida albicans infection using a specific (99m)Tc-labeled monoclonal antibody for the C. albicans germ tube.

Accurate diagnosis is critical for effective treatment of the invasive infection by Candida albicans. Here, we investigated whether a (99m) technetium (Tc)-labeled Fab' fragment of the monoclonal antibody specific for the C. albicans germ tube could specifically identify an invasive C. albicans infection. The germ tube of C. albicans was used as an immunogen to obtain monoclonal antibodies and the Fab' fragment of MAb03.2 C1-C2 with highest affinity and specificity was labeled with (99m)Tc. In vitro binding assays showed that the labeled Fab' preferentially bound to the germ tubes of C. albicans (4.23 ± 0.17 × 10(2) Bq per 1 × 10(7) cells). These values were significantly higher than those for blastospores of C. albicans, blastospores of heat-killed C. albicans, Aspergillus fumigatus, Staphylococcus aureus, and Escherichia coli (P < 0.05). By using in vivo biodistribution and planar imaging with single photon emission computed tomography, we demonstrated a significant specific accumulation of radioactivity in C. albicans-infected tissues. In summary, (99m)Tc-MAb03.2 C1-C2 Fab' is able to specifically accumulate in C. albicans-infected tissues, but not in tissue infected with A. fumigatus or bacteria or in a sterile inflammation. This study provides a new and specific radiopharmaceutical for the diagnosis of invasive C. albicans infections.

Imaging targeted at tumor with (188)Re-labeled VEGF(189) exon 6-encoded peptide and effects of the transfecting truncated KDR gene in tumor-bearing nude mice.

INTRODUCTION: Planar imaging of (188)Re-labeled vascular endothelial growth factor (VEGF)(189) exon 6-encoded peptide (QKRKRKKSRYKS) with single photon emission computed tomography (SPECT) in tumor-bearing nude mice and effects of the transfecting truncated KDR gene on its imaging were investigated, so as to provide a basis for further applying the peptide to tumor-targeted radionuclide treatment. METHODS: QKRKRKKSRYKS, coupling with mercaptoacetyltriglycine (MAG(3)) chelator was labeled with (188)Re; then in vivo distribution, planar imaging with SPECT and blocking experiment in tumor-bearing nude mice were analyzed. Recombinant adenovirus vectors carrying the truncated KDR gene were constructed to transfect tumor tissues to evaluate the effects of truncated KDR on the in vivo distribution and tumor planar imaging of (188)Re-MAG(3)-QKRKRKKSRYKS in tumor-bearing nude mice. RESULTS: The labeled peptide exhibited a sound receptor binding activity. Planar imaging with SPECT demonstrated significant radioactivity accumulation in tumor 1 h after injection of the labeled peptide and disappearance of radioactivity 3 h later. Significant radioactivity accumulation was also observed in the liver, intestines and kidneys but was not obvious in other tissues. An hour after injection of the labeled peptide, the percentage of the injected radioactive dose per gram (%ID/g) of tumor and tumor/contralateral muscle tissues ratio were 1.98+/-0.38 and 2.53+/-0.33, respectively, and increased to 3.08+/-0.84 and 3.61+/-0.59 in the group transfected with the truncated KDR gene, respectively, and radioactivity accumulation in tumor with planar imaging also increased significantly in the transfection group. CONCLUSION: (188)Re-MAG(3)-QKRKRKKSRYKS can accumulate in tumor tissues, which could be increased by the transfection of truncated KDR gene. This study provides a basis for further applying the peptide to tumor targeted radionuclide imaging and treatment.

Expression changes of ERK1/2, STAT3 and SHP-2 in bone marrow cells from gamma-ray induced leukemia mice.

The aim is to clarify expression changes of ERK1/2, STAT3 and SHP-2 in bone marrow cells from gamma-ray induced leukemia mice. A mouse model of gamma-ray induced leukemia was produced, and by means of quantitative real-time PCR, immunoprecipitation, Western blotting and electrophoretic mobility shift assays (EMSA), the expression of mRNA and protein, phosphorylation level, and protein activity of ERK1/2, STAT3 and SHP-2 in bone marrow cells were investigated in these mice. The results indicated that mRNA and protein expressions of ERK1/2 were upregulated, with significant increase of phosphorylation level and protein activity, but with insignificant differences in mRNA and protein expressions, phosphorylation level and protein activity of STAT3 and SHP-2 in bone marrow cells from gamma-ray induced leukemia mice compared to the radiation/tumor-free or control mice. It is concluded that in the pathogenesis of gamma-ray induced leukemia in Balb/C mice, activated ERK1/2 pathway may play a role, without involving STAT3 pathway; meanwhile, SHP-2 exerts no regulative effect on pathways of Ras-ERK1/2 and JAK-STAT.