Early Marker Of Response Research Articles

Bone Marrow (BM) Microenviroment Factors As Early Markers of Response in Patients with Newly Diagnosed Chronic Phase Chronic Myelogenous Leukemia (CML-CP) Treated with Nilotinib

IntroductionTyrosine Kinase Inhibitors (TKI) have completely changed the scenario of CML and dramatically improved the outcomes. Thus, early identification of patients expecting poor outcome is crucial to offer alternative TKI regimens or in some selected cases stem cell transplantation before disease progression may occur. The Evaluating Nilotinib Efficacy and Safety in Trial as First-Line Treatment (ENEST1st) is a phase 3b is an open-label study of nilotinib 300 mg twice daily (BID) in adults with newly diagnosed BCR-ABL positive CP-CML. Aim of the ENEST1st sub-study N10 was to investigate BM microenvironment markers that regulate leukemic stem cells in the bone marrow (BM) niche of Nilotinib-treated patients.MethodsThe study enrolled patients in 21 Italian ENEST1st participating centers. Response was based on ELN recommendations (Baccarani M, et al. Blood 2013 122:872-884). In an interim analysis, molecular and cytogenetic response by 24 months was assessed. Mononuclear cells were collected from BM and PB samples at the screening visit (V0) and after 3 months of treatment (V4). RT-qPCR for the expression of 10 genes (ARF, KIT, CXCR4, FLT3, LIF, NANOg, PML, PRAME, SET and TIE), involved in the stemness and hematopoietic stem cells survival signaling regulation was conducted. RT-qPCR data were normalized by the expression of GUS mRNA (normalized copy number, NCN). Plasma samples were collected at different time points from both BM or PB samples. Concentrations of 20 different analytes, including IL-1a, IL-3, M-CSF, SCF, SDF1-a, TRAIL, HGF, PDGF-bb, IL1b, IL-6, IL-7, IL-8, IL-10, IL-12, IL-15, G-CSF, GM-CSF, MIP-1a, TNF-a, and VEGF, were simultaneously evaluated using commercially available multiplex bead-based sandwich immunoassay kits.Results33 out of 37 patients enrolled were available for an interim molecular analysis at 24 months: an optimal response was achieved in 25 patients, a warning response in 5 patients and a failure response in 3 patients. We observed a significant correlation between the expression of two genes involved in the regulation of stem cell pluripotency (NANOg) or cytokine signaling (SET) and patient outcome. Indeed, NANOg and SET mRNA were significantly down-regulated in PB samples at diagnosis of patients with optimal response compared to patients with warning/failure response (NANOg mRNA: 0.3±0.25 NCN vs 0.6±0.7 NCN, respectively; p=0.05; SET mRNA: 0.2±0.3 NCN vs 2.3±4.2 NCN, respectively; p=0.03). We also investigated the plasma level of several factors involved in the hematopoietic stem cells (HSCs). Some of these markers showed a significant correlation with patient's outcome when evaluated at diagnosis in either PB or BM samples. Indeed, high level of IL12 (in the BM samples), or HGF, mCSF and SCF (in the PB samples) were associated to a worst prognosis markers, since significantly correlating with no MMR@12months (IL12, p=0.03), intermediate/high Socal score (mCSF, p=0.03; SCF, p=0.03), no reduction of MMR below to 1 at 3 month (SCF, p=0.04) or warning/failure response to Nilotinib treatment (HGF, p=0.03; SCF, p=0.04). Indeed, we find a lower levels of PDGFb, SDF1, TNFa, TRAIL (in the BM samples), and HGF, SDF1, TRAIL (in the PB samples) in those patients with intermediate/high Hasford or Sokal score (PDGFb, p=0.0007; SDF1, p=0.02), warning/failure response to Nilotinib treatment (HGF, p=0.03) or lacking of MMR4.0 (SDF1, p=0.01; TNFa, p=0.02; TRAIL, p=0.05).Conclusion/SummaryTaken together, our results suggest that the expression analysis of genes involved in cell pluripotency (NANOg) and/or cell signaling (SET) at baseline, may indicate early achievement of deep molecular response in shown CML-CP patients treated with nilotinib. In addition, in patients with optimal response to Nilotinib, high concentration of SDF-1, TRAIL (inversely correlated with BCR-ABL, and associated to an higher susceptibility to apoptosis in the leukemic blasts) were observed as well as BM TNF (cell-extrinsic and potent endogenous suppressor of HSC activity). A lower concentration of several factors associated to hematopoietic progenitor cell growth and survival (including HGF, SCF and IL12) were observed compared to patients failing to achieve response to Nilotinib. These data strongly suggest that stromal microenvironment supports the viability of BCR-ABL cells in BM niches through direct feeding, or environment releasing of survival factors. DisclosuresSoverini:Novartis, Briston-Myers Squibb, ARIAD: Consultancy. Martinelli:MSD: Consultancy; BMS: Speakers Bureau; Roche: Consultancy; ARIAD: Consultancy; Novartis: Speakers Bureau; Pfizer: Consultancy. Saglio:Bristol-Myers Squibb: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; ARIAD: Consultancy, Honoraria; Novartis Pharmaceutical Corporation: Consultancy, Honoraria. Galimberti:Novartis: Employment. Giles:Novartis: Consultancy, Honoraria, Research Funding. Hochhaus:Pfizer: Honoraria, Research Funding; Bristol-Myers Squibb: Honoraria, Research Funding; ARIAD: Honoraria, Research Funding; Novartis: Honoraria, Research Funding.

The National Lung Matrix Trial: translating the biology of stratification in advanced non-small-cell lung cancer

The management of NSCLC has been transformed by stratified medicine. The National Lung Matrix Trial (NLMT) is a UK-wide study exploring the activity of rationally selected biomarker/targeted therapy combinations. The Cancer Research UK (CRUK) Stratified Medicine Programme 2 is undertaking the large volume national molecular pre-screening which integrates with the NLMT. At study initiation, there are eight drugs being used to target 18 molecular cohorts. The aim is to determine whether there is sufficient signal of activity in any drug-biomarker combination to warrant further investigation. A Bayesian adaptive design that gives a more realistic approach to decision making and flexibility to make conclusions without fixing the sample size was chosen. The screening platform is an adaptable 28-gene Nextera next-generation sequencing platform designed by Illumina, covering the range of molecular abnormalities being targeted. The adaptive design allows new biomarker-drug combination cohorts to be incorporated by substantial amendment. The pre-clinical justification for each biomarker-drug combination has been rigorously assessed creating molecular exclusion rules and a trumping strategy in patients harbouring concomitant actionable genetic abnormalities. Discrete routes of pathway activation or inactivation determined by cancer genome aberrations are treated as separate cohorts. Key translational analyses include the deep genomic analysis of pre- and post-treatment biopsies, the establishment of patient-derived xenograft models and longitudinal ctDNA collection, in order to define predictive biomarkers, mechanisms of resistance and early markers of response and relapse. The SMP2 platform will provide large scale genetic screening to inform entry into the NLMT, a trial explicitly aimed at discovering novel actionable cohorts in NSCLC. 38344105.

Value of ultrasonography as a marker of early response to abatacept in patients with rheumatoid arthritis and an inadequate response to methotrexate: results from the APPRAISE study

ObjectivesTo study the responsiveness of a combined power Doppler and greyscale ultrasound (PDUS) score for assessing synovitis in biologic-naïve patients with rheumatoid arthritis (RA) starting abatacept plus methotrexate (MTX).MethodsIn this...

Longitudinal tumor hypoxia imaging with [18F]FAZA-PET provides early prediction of nanoliposomal irinotecan (nal-IRI) treatment activity

BackgroundNon-invasive measurement of tumor hypoxia has demonstrated potential for the evaluation of disease progression, as well as prediction and assessment of treatment outcome. [18F]fluoroazomycin arabinoside (FAZA) positron emission tomography (PET) has been identified as a robust method for quantification of hypoxia both preclinically and clinically. The goal of this investigation was to evaluate the feasibility and value of repeated FAZA-PET imaging to quantify hypoxia in tumors that received multi-dose chemotherapy.MethodsFAZA-PET imaging was conducted over a 21-day period in a mouse xenograft model of HT-29 human colorectal carcinoma, following multi-dose chemotherapy treatment with irinotecan (CPT-11) or nanoliposomal irinotecan (nal-IRI, MM-398).ResultsTumors treated with 10 mg/kg nal-IRI maintained significantly lower levels of hypoxia and smaller hypoxic fractions compared to tumors that received 50 mg/kg CPT-11. Specifically, differences in FAZA uptake were detectable 9 days before any significant differences in tumor volume were observed between the treatment groups.ConclusionsThese findings highlight the potential use of FAZA-PET as an early marker of treatment response following multi-dose chemotherapy.Electronic supplementary materialThe online version of this article (doi:10.1186/s13550-015-0135-x) contains supplementary material, which is available to authorized users.

DW-MRI and (18) F-FLT PET for early assessment of response to radiation therapy associated with hypoxia-driven interventions. Preclinical studies using manipulation of oxygenation and/or dose escalation.

Early markers of treatment response may help in the management of patients by predicting the outcome of a specific therapeutic intervention. Here, we studied the potential value of diffusion-weighted MRI (DW-MRI) and (18)F-fluorothymidine ((18)F-FLT), markers of cell death and cell proliferation respectively, to predict the response to irradiation. In addition, dose escalation and/or carbogen breathing were used to modulate the response to irradiation. The studies were performed on two hypoxic rat tumor models: rhabdomyosarcoma and 9L-glioma. The rats were imaged using MRI and PET before and two days after the treatment. In both tumor models, changes in ADC (apparent diffusion coefficient) and (18)F-FLT SUV (standardized uptake value) were significantly correlated with the tumor growth delay. For both tumor models, the ADC values increased in all irradiated groups two days after the treatment while they decreased in the untreated groups. At the same time, the uptake of (18)F-FLT increased in the untreated groups and decreased in all treated groups. Yet, ADC values were not sensitive enough to predict the added value of dose escalation or carbogen breathing in either model. Change in (18)F-FLT uptake was able to predict the higher tumor response when using increased dose of irradiation, but not when using a carbogen breathing challenge. Our results also emphasize that the magnitude of change in (18)F-FLT uptake was strongly dependent on the tumor model.

Acute vascular response to cediranib treatment in human non-small-cell lung cancer xenografts with different tumour stromal architecture.

Tumours can be categorised based on their stromal architecture into tumour vessel and stromal vessel phenotypes, and the phenotypes have been suggested to define tumour response to chronic treatment with a VEGFR2 antibody. However, it is unclear whether the vascular phenotypes of tumours associate with acute vascular response to VEGFR tyrosine kinase inhibitors (TKI), or whether the early changes in vascular function are associated with subsequent changes in tumour size. This study was sought to address these questions by using xenograft models of human non-small cell lung cancer (NSCLC) representing stromal vessel phenotype (Calu-3) and tumour vessel phenotype (Calu-6), respectively. For dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI), nude mice bearing established Calu-3 or Calu-6 xenografts were treated with a potent pan-VEGFR TKI, cediranib (6 mg/kg), at 0 h and 22 h. DCE-MRI was performed 2h before the first dose and 2h after the second dose of cediranib to examine acute changes in tumour vessel perfusion. Tumours were harvested for hypoxia detection by CA9 immunohistochemistry. For tumour growth study, mice carrying established Calu-3 or Calu-6 tumours were treated with cediranib once daily for 5 days. Twenty-four hours after cediranib administration, the perfusion of Calu-3 tumours was markedly reduced, with a significant increase in hypoxia. In contrast, neither perfusion nor hypoxia was significantly affected in Calu-6 tumours. Tumour regressions were induced in Calu-3 xenografts, but not in Calu-6 xenografts, although there was a trend towards tumour growth inhibition after 5 days of cediranib treatment. These findings suggest that tumour stromal architecture may associate with acute tumour vascular response to VEGFR TKI, and this acute tumour vascular response may be a promising early predictive marker of response to VEGFR TKI in NSCLC.

Prediction of lung density changes after radiotherapy by cone beam computed tomography response markers and pre-treatment factors for non-small cell lung cancer patients

Background and purposeThis study investigates the ability of pre-treatment factors and response markers extracted from standard cone-beam computed tomography (CBCT) images to predict the lung density changes induced by radiotherapy for non-small cell lung cancer (NSCLC) patients. Methods and materialsDensity changes in follow-up computed tomography scans were evaluated for 135 NSCLC patients treated with radiotherapy. Early response markers were obtained by analysing changes in lung density in CBCT images acquired during the treatment course. The ability of pre-treatment factors and CBCT markers to predict lung density changes induced by radiotherapy was investigated. ResultsAge and CBCT markers extracted at 10th, 20th, and 30th treatment fraction significantly predicted lung density changes in a multivariable analysis, and a set of response models based on these parameters were established. The correlation coefficient for the models was 0.35, 0.35, and 0.39, when based on the markers obtained at the 10th, 20th, and 30th fraction, respectively. ConclusionsThe study indicates that younger patients without lung tissue reactions early into their treatment course may have minimal radiation induced lung density increase at follow-up. Further investigations are needed to examine the ability of the models to identify patients with low risk of symptomatic toxicity.

Usefulness of Cardiac Troponins as Markers of Early Treatment Response in Cardiac Sarcoidosis

Evaluation and treatment of cardiac sarcoidosis (CS) suffer from lack of sensitive and easily repeatable markers of disease activity. We studied measurements of high-sensitivity cardiac troponin T or troponin I (hs-cTnT/I) taken at presentation and during treatment in 62 patients with new-onset CS (48 women, mean age 49 years). Hs-cTnT was measured in 50 patients and was elevated (>13 ng/L) at presentation in 26 of them (52%). Hs-cTnI was measured in the remaining 12 patients and was elevated (>0.04 ng/mL) in 7 of them (58%). Left ventricular ejection fraction averaged 43 ± 14% in association with elevated hs-cTnT/I (n = 33) versus 53 ± 10% with normal hs-cTnT/I (n = 29; p = 0.001). Hs-cTnT/I was remeasured after 4 weeks of steroid therapy in 38 patients and was normalized in 16 of the 24 (67%) with an elevated pretreatment concentration and remained normal in the rest of the 14 patients (p <0.001). During follow-up (median, 17 months), cardiac death (n = 2), aborted sudden death (n = 5), sustained ventricular tachycardia (n = 8), or new complete atrioventricular block (n = 1) was recorded in 11 of 33 patients with elevated hs-cTnT/I versus in 5 of 29 with normal hs-cTnT/I (log-rank p = 0.068). Two-year event-free Kaplan-Meier cardiac survival estimate (95% confidence interval) was 67% (48% to 81%) with elevated hs-cTnT/I versus 93% (76% to 99%) with normal hs-cTnT/I. In CS, circulating hs-cTnT/I may help clinicians evaluate disease activity and treatment response. Their prognostic value remains tentative pending more follow-up data.

PTH-059 Faecal calprotectin identifies non responders to anti-tnfΑ therapy when measured after induction in inflammatory crohn’s disease

Introduction Poor response to anti-TNFα treatment in patients with Crohn’s disease (CD) is not infrequent. As new biologic therapies targeting other immune pathways of the intestinal inflammation are becoming available in routine clinical practice early prognostic markers of treatment failure are urgently needed. Faecal calprotectin (fcal) has been shown to be a useful prognostic marker of relapse in CD correlating well with both endoscopic and magnetic resonance imaging scores. Furthermore, a recently published study utilising gene expression profiling by micro arrays to detect prognostic markers of early response to anti-TNFα therapy identified the two genes coding for the calprotectin subunits (S100A8, S100A9) to be among the most highly expressed gene transcripts in non-responders. Method This is a retrospective observational study of a prospectively kept database testing the hypothesis that a faecal calprotectin measurement after anti-TNFα induction (infliximab: 8–12 weeks, adalimumab: 6-weeks) reflects response to treatment. CD patients, who commenced anti-TNFα therapy for active, inflammatory disease and had serial fcal measurements before and after induction were identified. Remission was assessed at 6 months and defined as the composite of Harvey Bradshaw Index (HBI Results We identified 26 patients who started either infliximab (16) or adalimumab (10) and had serial fcal at the set time points [median age: 32, disease duration: 5 (3, 8), location: 17 ileocolonic, 8 colonic and 1 ileal]. Patients on infliximab received 5mg/kg doses at 0, 2, 6 and then 8 weekly. Those on adalimumab had one dose of 160mg followed by 80 mg after two weeks and then received 40 mg doses fortnightly. At 6 months, 15/26 (58%) patients were in remission. Fcal post induction in those in remission at 6 months was lower compared to those with persistently active disease [65 (38, 118) vs. 269 (161, 542)]. Receiver operator characteristic curve analysis shows that fcal >150 has 93% sensitivity and 91% specificity in predicting non responders (likehood ratio: 10.27, area under curve: 0.98, p Conclusion Fcal measurement soon after anti-TNFα induction predicts non-response providing the opportunity to identify those patients who require further treatment tailoring early. Disclosure of interest None Declared.

Abstract P4-01-20: A novel microfluidic system for the detection, enumeration and molecular analysis of circulating tumor cells (CTCs) in metastatic breast cancer (MBC)

Abstract Background: In recent years blood testing for circulating tumor cells (CTC) has gained increasing interest in cancer research. CTC detection and enumeration can serve as a ‘liquid biopsy’ and an early marker of response to systemic therapy. Different analytical systems for CTC detection and isolation have been developed, but the CellSearch® is currently the only FDA-approved technology. We aimed to evaluate CTCs detection by a novel microfluidific technology, a size- and deformability-based capture system. This unique platform not only allows flex[not]ibility in the selection of antibody markers but also segregates the CTCs in their own chambers, thus, enabling morphological, immunological and genetic characterization of each CTC at the single cell level. Methods: We performed a prospective study to compare the detection of CTCs using the CellSearch® (Janssen Diagnostics) vs. the new microfluidic platform. Enumeration by the CellSearch® was performed according to standard protocol. For the microfluidific-device capture (De Novo Sciences, MI) peripheral blood from MBC patients was diluted 1:1 with PFA 0.8% and PBS 1%. Prior to sample loading, the microfluidic device was coated with priming buffer. After cells were fixed using 4.0% PFA and were subsequently stained with pancytokeratin, Zym 5.2 and CD45. Nuclei were coun[not]terstained with Hoechst-33342. CTCs were identified as round and bright green cells not stained with red (CK+/CD45-/DAPI+). Data were analyzed using non-parametric methods: Cohen’s kappa, Chi2 test, Spearman rank correlations and Mann-Whitney test. Associations with CTCs were calculated in two ways: CTCs as a continuous variable normalized for ml of blood and CTCs categorized as &amp;lt; 5 versus ≥ 5. Results: The two methods was concordant in 88.2% of patients with a Cohen’s kappa of 0.743 when the detection of a single CTC is considered like positive. We also found a concordance of 85% (k=0.70) when we use the CTC cut-off level of ≥5 cells per 7.5 ml of blood to identify patient with higher risk for disease progression. Thirty-one patients with MBC were tested for CTCs using both methods. CTC detection by microfluidific platform was positively associated with Her2 positive patients if we consider as categorical variable; a weak association was seen also with the CellSearch®. CTCs AssociationCTC/ml of blood ContinuousCTC categorized (≥5)DeNovomedian [min, max]p-valueNn (%)OR ( 95% CI)p-valueHer 20.5620.038No0.5 [0, 62]197 (36.8)refYes1.75 [0, 4]129 (75.0)5.14 (1.03, 25.6)CellSearch®median [min, max]p-valueNn (%)OR ( 95% CI)p-valueHer 20.7040.056No0.13 [0, 335.7]196 (41.2)refYes0.95 [0, 3.33]128 (34.7)4.33 (0.93, 20.2) Conclusions: The enumeration of CTCs showed strong prognostic significance in MBC raising interest in a more accurate molecular characterization to achieve the possibility for a dynamic molecular monitoring. Introducing novel methodologies should demonstrate comparable detection rate for epithelial cells. This new microfluidic system showed accuracy and flexibility to move to the next phase of molecular testing with the intent of assessing the value as liquid biopsy. Citation Format: Carmela Paolillo, Zhaomei Mu, Angela Toss, Priyadarshini Gogoi, Saedeh Sepehri, Yi Zhou, Kalyan Handique, Ye Zhong, Hushan Yang, Ettore Capoluongo, Massimo Cristofanilli, Paolo Fortina. A novel microfluidic system for the detection, enumeration and molecular analysis of circulating tumor cells (CTCs) in metastatic breast cancer (MBC) [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P4-01-20.

210 Contrast Enhanced Endoscopic Ultrasound of the Colon Is a Marker of Early Therapy Response Following Anti-TNF-Therapy in Patients With Acute Ulcerative Colitis

210 Contrast Enhanced Endoscopic Ultrasound of the Colon Is a Marker of Early Therapy Response Following Anti-TNF-Therapy in Patients With Acute Ulcerative Colitis

IM-03 * SERIAL METABOLISM OF DIFFUSE INTRINSIC PONTINE GLIOMA TREATED WITH PEPTIDE BASED VACCINE THERAPY

PURPOSE: Diffuse intrinsic pontine glioma (DIPG) carries the worst prognosis in pediatric neurooncology, with no improvement in outcomes for decades. The goal of this study was to examine myo-inositol (mI)/choline and other metabolites (measured by serial MRS) as predictors of overall survival in (1) a DIPG cohort treated with standard radiochemotherapy and (2) a DIPG cohort treated with radio-therapy followed by peptide-based vaccine therapy. METHODS: 50 MRS studies were performed in 11 patients with DIPG enrolled in a phase 1 clinical trial of peptide based vaccine therapy. 56 MRS studies were performed in 14 patients receiving only radio-chemotherapy. Metabolic profiles were binned into comparable time points after radiation therapy. All spectra were acquired using a single-voxel PRESS sequence (TE = 35ms, TR = 1.5s, 128 averages). Spectra were processed with fully automated LCModel (version 6.3-1c) software. Concentrations of the main metabolites (NAA, Cr, Cho, mI) and metabolite ratios relative to Cho were evaluated. Generalized estimating equations were used to evaluate longitudinal associations between metabolites and/or therapy group and overall survival. RESULTS: The 25 patients survived 6-25 months after diagnosis. Study baseline (post radiotherapy/pre-vaccine, 78-116 days post-diagnosis) mI/choline measures did not predict overall survival, but at subsequent time points higher mI/choline was associated with longer overall survival (p <0.05). A statistical interaction in the model with scans 159-250 days post-diagnosis (p < 0.001) suggested that the association with survival was much greater in the vaccine group (47 days' greater expected survival for each unit higher mI/choline ratio, after ∼2 vaccine doses) than for the non-vaccine group (13 days). DISCUSSION: mI/choline ratio may be a predictive marker for early response to peptide-based vaccine therapy of pediatric DIPG. A larger sample study is needed to further validate that MRS could be used to optimize/adjust peptide based vaccine therapy of DIPG patients.

Increased levels of choline metabolites are an early marker of docetaxel treatment response in BRCA1-mutated mouse mammary tumors: an assessment by ex vivo proton magnetic resonance spectroscopy.

BackgroundDocetaxel is one of the most frequently used drugs to treat breast cancer. However, resistance or incomplete response to docetaxel is a major challenge. The aim of this study was to utilize MR metabolomics to identify potential biomarkers of docetaxel resistance in a mouse model for BRCA1-mutated breast cancer.MethodologyHigh resolution magic angle spinning (HRMAS) 1H MR spectroscopy was performed on tissue samples obtained from docetaxel-sensitive or -resistant BRCA1-mutated mammary tumors in mice. Measurements were performed on samples obtained before treatment and at 1-2, 3-5 and 6-7 days after a 25 mg/kg dose of docetaxel. The MR spectra were analyzed by multivariate analysis, followed by analysis of the signals of individual compounds by peak fitting and integration with normalization to the integral of the creatine signal and of all signals between 2.9 and 3.6 ppm.ResultsThe HRMAS spectra revealed significant metabolic differences between sensitive and resistant tissue samples. In particular choline metabolites were higher in resistant tumors by more than 50% with respect to creatine and by more than 30% with respect to all signals between 2.9 and 3.6 ppm. Shortly after treatment (1-2 days) the normalized choline metabolite levels were significantly increased by more than 30% in the sensitive group coinciding with the time of highest apoptotic activity induced by docetaxel. Thereafter, choline metabolites in these tumors returned towards pre-treatment levels. No change in choline compounds was observed in the resistant tumors over the whole time of investigation.ConclusionsRelative tissue concentrations of choline compounds are higher in docetaxel resistant than in sensitive BRCA1-mutated mouse mammary tumors, but in the first days after docetaxel treatment only in the sensitive tumors an increase of these compounds is observed. Thus both pre- and post-treatment tissue levels of choline compounds have potential to predict response to docetaxel treatment.

AMPK-mediated energy homeostasis and associated metabolic effects on cancer cell response and resistance to cetuximab.

We previously reported that cetuximab, an EGFR-blocking antibody, inhibits cancer metabolism via downregulation of HIF-1α and reverses the Warburg effect in cancer cells. Here, we report that inhibition of HIF-1 transcriptional activity by cetuximab does not necessarily lead to successful inhibition of cell proliferation. In several head and neck squamous cell carcinoma (HNSCC) cell lines, we observed a pattern of oscillating decrease and increase of intracellular ATP level after cetuximab treatment, and the magnitude and kinetics of which varied by cell line and appeared to be linked to the extent of cellular response to cetuximab. In HNSCC cells with low basal level of AMPK activity and that responded to cetuximab-induced growth inhibition, there was a transient, LKB1-dependent activation of AMPK. In contrast, HNSCC cells that had a high basal level of AMPK activity were less sensitive to cetuximab-induced growth inhibition despite effective inhibition of EGFR downstream signaling by cetuximab. Knockdown or inhibition of AMPK markedly enhanced response to cetuximab via induction of apoptosis. These findings indicate that a transient activation of AMPK is an early metabolic marker of cellular response to cetuximab and that high and sustained AMPK activity is an important mechanism by which cancer cells survive cetuximab treatment.

A Potent Virus-Specific Antibody-Secreting Cell Response to Acute Enterovirus 71 Infection in Children.

Enterovirus 71 (EV71) remains a leading pathogen for acute infectious diseases in children, especially in Asia. The cellular basis for establishing a virus-specific antibody response to acute EV71 infections is unclear in children. We studied the magnitude of virus-specific antibody-secreting B cells (ASCs) and its relationship with serological response, clinical parameters, and virological parameters among children with laboratory-confirmed EV71 infection. A potent EV71 genogroup B- and virus-specific ASC response was detected in the first week of illness among genotype B5 EV71-infected children. The cross-reactive EV71-specific ASC response to genogroup C viral antigens composed about 10% of the response. The EV71-specific ASC response in children aged ≥3 years produced immunoglobulin G predominantly, but immunoglobulin M was predominant in younger children. Proliferation marker was expressed by the majority of circulating ASCs in the acute phase of EV71 infection. Virus-specific ASC responses significantly correlated with throat viral load, fever duration, and serological genogroup-specific neutralization titer. The presence of a virus-specific ASC response serves an early cellular marker of an EV71-specific antibody response. Further detailed study of EV71-specific ASCs at the monoclonal level is crucial to delineate the specificity and function of antibody immunity in children.

High-throughput measurements of the optical redox ratio using a commercial microplate reader.

There is a need for accurate, high-throughput, functional measures to gauge the efficacy of potential drugs in living cells. As an early marker of drug response in cells, cellular metabolism provides an attractive platform for high-throughput drug testing. Optical techniques can noninvasively monitor NADH and FAD, two autofluorescent metabolic coenzymes. The autofluorescent redox ratio, defined as the autofluorescence intensity of NADH divided by that of FAD, quantifies relative rates of cellular glycolysis and oxidative phosphorylation. However, current microscopy methods for redox ratio quantification are time-intensive and low-throughput, limiting their practicality in drug screening. Alternatively, high-throughput commercial microplate readers quickly measure fluorescence intensities for hundreds of wells. This study found that a commercial microplate reader can differentiate the receptor status of breast cancer cell lines (p < 0.05) based on redox ratio measurements without extrinsic contrast agents. Furthermore, microplate reader redox ratio measurements resolve response (p < 0.05) and lack of response (p > 0.05) in cell lines that are responsive and nonresponsive, respectively, to the breast cancer drug trastuzumab. These studies indicate that the microplate readers can be used to measure the redox ratio in a high-throughput manner and are sensitive enough to detect differences in cellular metabolism that are consistent with microscopy results.

Immune memory to Sudan virus: comparison between two separate disease outbreaks.

Recovery from ebolavirus infection in humans is associated with the development of both cell-mediated and humoral immune responses. According to recent studies, individuals that did not survive infection with ebolaviruses appear to have lacked a robust adaptive immune response and the expression of several early innate response markers. However, a comprehensive protective immune profile has yet to be described. Here, we examine cellular memory immune responses among survivors of two separate Ebolavirus outbreaks (EVDs) due to Sudan virus (SUDV) infection in Uganda—Gulu 2000–2001 and Kibaale 2012. Freshly collected blood samples were stimulated with inactivated SUDV, as well as with recombinant SUDV or Ebola virus (EBOV) GP (GP1–649). In addition, ELISA and plaque reduction neutralization assays were performed to determine anti-SUDV IgG titers and neutralization capacity. Cytokine expression was measured in whole blood cultures in response to SUDV and SUDV GP stimulation in both survivor pools, demonstrating recall responses that indicate immune memory. Cytokine responses between groups were similar but had distinct differences. Neutralizing, SUDV-specific IgG activity against irradiated SUDV and SUDV recombinant proteins were detected in both survivor cohorts. Furthermore, humoral and cell-mediated crossreactivity to EBOV and EBOV recombinant GP1–649 was observed in both cohorts. In conclusion, immune responses in both groups of survivors demonstrate persistent recognition of relevant antigens, albeit larger cohorts are required in order to reach greater statistical significance. The differing cytokine responses between Gulu and Kibaale outbreak survivors suggests that each outbreak may not yield identical memory responses and promotes the merits of studying the immune responses among outbreaks of the same virus. Finally, our demonstration of cross-reactive immune recognition suggests that there is potential for developing cross-protective vaccines for ebolaviruses.

Bone marrow minimal residual disease was an early response marker and a consistent independent predictor of survival after anti-GD2 immunotherapy.

Immunotherapy is a standard of care for children with high-risk neuroblastoma, where bone marrow (BM) is the predominant metastatic site. Early response markers of minimal residual disease (MRD) in the BM that are also predictive of survival could help individualize patient therapies. After achieving first remission (n = 163), primary refractory disease (n = 102), or second remission (n = 95), children with stage 4 neuroblastoma received anti-GD2 3F8 antibody immunotherapy. BM MRD before 3F8 treatment and after cycle 2 (postMRD) was measured using a four-marker panel (B4GALNT1, PHOX2B, CCND1, and ISL1) by quantitative reverse transcription polymerase chain reaction. Progression-free survival (PFS) and overall survival (OS) were estimated using the Kaplan-Meier method. Prognostic variables were tested in both univariable and multivariable analyses, and MRD markers were further assessed individually and in combination as binary composite (postMRD: 0 and 1) and as equal sum (postMRDSum: 0 to 4) using the Cox regression models, and their predictive accuracy was determined by the concordance index. When BM was evaluated after cycle 2, individual markers were highly predictive of PFS and OS. The prediction accuracy improved when they were combined in postMRDSum. A multivariable model taking into account all the variables significant in the univariable analyses identified postMRDSum to be independently predictive of PFS and OS. When the model for OS also included missing killer immunoglobulin-like receptor ligand, human antimouse antibody response, and the enrollment disease status, the concordance index was 0.704. BM MRD after two cycles of immunotherapy was confirmed as an early response marker and a consistent independent predictor of survival.

Effects of Diabetes Mellitus and HbA1c on Treatment Prognosis in Uncomplicated Acute Pyelonephritis

Purpose: We investigated potential difference in the clinical features of hospitalized female acute pyelonephritis (APN) patients with and without diabetes mellitus (DM). The additional purpose of the study was to determine whether HbA1c has a role in predicting clinical outcome in terms of morbidity compared with those considered to have normal glycemic control in female APN patients with DM. Materials and Methods: Among the female patients who received inpatients care due to APN from January 2011 to December 2013, 282 patients (86 with DM and 196 without DM) were selected and their medical records were analyzed. Laboratory results (white blood cell, blood urea nitrogen, serum creatinine, fasting plasma glucose, HbA1c) were dichotomized. Primary outcomes for comparison between the groups of patients with or without DM were early clinical response rate and length of in-hospital stay. Subgroup analysis was also performed in DM patients according to HbA1c status (<6.5%, ≥6.5%). Results: In multivariate logistic regression analysis, DM was not found to show independent association with poor early clinical response or longer hospital stay. HbA1c was the common risk factor of poor early clinical response (odds ratio [OR] 4.88, 95% confidence interval [CI] 1.61-14.8) and longer hospital stay (OR 4.20 95% CI 1.52-11.60) in APN patients with DM. Conclusions: Strict glucose control may be more important prognostic factor than presence or absence of DM in APN patients. As a marker of prolonged hospitalization and poor early clinical response, HbA1c can complement the present clinical and laboratory parameters used as guide in the proper treatment of DM patients with APN.

Serum total antioxidant capacity in oral carcinoma patients.

Context:Oxidative stress is a major determinant in carcinogenesis and serum total antioxidant capacity (TAC) indirectly reflects the level of oxidative stress. Although oral cancer is the third most frequent cancer in Indian population, there are no standard noninvasive marker for early detection and monitoring therapeutic response in oral cancer patients.Aims:This study was carried out to investigate implications of serum TAC in oral cancer by evaluating pre- and post-operative levels in oral cancer patients.Settings and Designs:It was a prospective single blinded study.Subjects and Methods:Serum TAC was measured using ferric-reducing antioxidant power assay. Pre- and post-operative values were estimated and compared in 30 oral cancer patients who underwent surgery.Statistical Analysis Used:Paired t-test was used to compare pre- and post-operative values.Results:Compared to the normal value, both pre- and post-operative serum TAC were significantly low in oral cancer patients. However, there was no statistically significant difference between pre- and postoperative levels.Conclusions:In this study, low mean serum TAC was detected in oral cancer patients. However, its diagnostic and prognostic significance in oral cancer needs further investigation.