Bad Responders Research Articles

Metformin Resistance Alleles in Polycystic Ovary Syndrome: Pattern and Association With Glucose Metabolism

Insulin-sensitizer treatment with metformin is common in polycystic ovary syndrome (PCOS). OCT alleles were investigated in PCOS patients to identify genetic 'bad responders' and 'nonresponders' to metformin including their possible effects on glucose metabolism without treatment. We genotyped eight SNPs in OCT1, OCT2 and ATM genes in 676 women with PCOS and 90 control women, we also measured oral glucose tolerance tests prior to treatment. Nonfunctional alleles were present in 29.8% and low-functional alleles in 57.9% of our PCOS cohort. OCT variants were significantly associated with elevated baseline and glucose-induced C-peptide levels in PCOS. Metformin bad responders or nonresponders based on OCT genotypes might be relevant in clinical practice - their modulation of metformin pharmacokinetics and pharmacodynamics and metformin-independent glucose effects remain to be elucidated.

Validation of imaging biomarker in a multicentric study to predict PFS in mRCC treated with TKI.

526 Background: Tyrosine kinase inhibitors including sunitinib are the most effective treatments of metastatic renal cell carcinoma (mRCC). A multi-centric study of 539 patients (different tumors treated anti-angiogenic treatments) evaluating dynamic contrast-enhanced ultrasound (DCE-US), showed that a decrease of AUC (Area under the curve) correlated to the blood volume at one month is predictive of response. Our first objective was to validate the correlation between this parameter and the PFS in a sub-group of mRCC treated with Sunitinib The second objective was to study the variability of AUC. Methods: Each Patient had CT-scan every 2 months in order to evaluate the Response assessment using the Response Evaluation Criteria in Solid Tumors (RECIST 1.1).DCE-US were performed at baseline and at D30. At each examination, we quantified 7 DCE-US parameters after bolus injection of contrast agent and mathematical modelization of raw linear data recorded during 3 minutes. We also estimated the variation between baseline and D30. The main endpoint was progression free survival assessed according to RECIST. We first selected the best parameters. We studied the trend between the parameter value and freedom from progression. After, the best cut-points were searched through a grid search. The best single cut-point was that with the lowest P-value for progression free survival. We performed this analysis in the sub-group of patients with mRCC treated with sunitinib. Morever, we studied the variability of AUC in 30 other patients treated with TKI . We performed in this group 2 DCE-US the same day before and after lunch. Results: A total of 81 mRCC patients treated with sunitinib were selected. All had DCE-US at baseline and one month. The median of follow-up was 18 months. For DCE-US, the decrease of 90 % of AUC at D 30 was correlated to the PFS (p =0.03). The difference of PFS between the groups defined by this cut-point was 4 months (bad responders) and 14 months (good responders). The results of variability are on-going. Conclusions: The decrease of more than 90% of AUC with DCE-US at one month is a potential predictive biomarker of response in mRCC patients treated with sunitinib. The results of variability of this parameter will be also presented. Clinical trial information: no. 912346.

Is magnetic resonance useful predicting neoadjuvant response in children with osteosarcoma?

10056 Background: Few prognostic factors had been described for pediatric high-grade osteosarcoma (HGO). Primary tumor necrosis percentage after neoadjuvant chemotherapy, analyzed by pathologists, remains as an important prognostic factor. Magnetic resonance imaging (MRI) has been postulated to predict necrosis in adults. Our aim was therefore to calculate the diagnostic utility of MRI in children. Methods: Prospective, longitudinal and clinical study was performed from September 2011 to January 2012. Twenty two patients (< 18 years), with extremity HGO and no prior chemotherapy treatment were included. All had MRI from diagnosis and just before surgery. We performed a sensibility-specificity analysis comparing percentage of necrosis by MRI vs histological Rosen grades (grades I and II were classified as bad responders; III and IV as good responders). Cut off value for MRI was 90%. Agreement between radiologists was calculated by Cohen´s Kappa coefficient. Results: Average age was 11.7 ± 3.04 (SD), 54.5% were women. All patients received neoadyuvant chemotherapy (cisplatin 120mg/m2 and adriamicin 75mg/m2). Nine patients had limb-sparing surgery, 8 limb disarticulation and 5 amputations; 72.8% were classified as bad responders by histology. MRI sensibility was 25%, specificity 86.6%, positive predictive value 50% and negative predictive value 68.4%. Kappa correlation coefficient between radiologists was 0,229. Conclusions: Diagnostic utility values in this study were lower than those reported previously. Torricelli, Dyke and Guo reported MRI sensibility of over 70% and specificity ranging from 37% to 87% (similar number of patients). Predictive values were also higher (> 80%). Differences are likely explained by the lack of correlation between radiologists. Our results support the need to establish systematized criteria to measure necrosis by MRI. In order to predict clinical response to chemotherapy in HGO, it is necessary to establish areas and cut off percentages of necrosis for appropriate clinical decision making.

OP057: HER-1 differential phosphorylation as prognostic factor and possible therapeutic target in oral squamous cell carcinoma

The EGFR (epidermal growth factor receptor) a member of the family of transmembrane protein kinase receptors known as the HER family shows a significant correlation with the presence of metastases and poorly differentiated oral cancer. Aim of the present work is to define the key-role of EGFR in oral cancer prognosis. We have analyzed the EGFR expression on 149 cases of oral squamous cell cancers (OSCC) and we have found that it was poorly expressed in normal oral epithelium, but its expression was significantly increased in OSCCs. Moreover, we have recorded that both pEGFR-Tyr 845 and pEGFRTyr 1068 were mainly distributed in high histological grading and in advanced stages. Western blotting has confirmed the total absence of EGFR phosphorylation in normal oral epithelium and the higher level of protein phosphorylation in OSCCs. The HER-1 amplification was found by fluorescence in situ hybridization (FISH) in 14% of OSCC; interestingly, EGF-R amplification was mainly observed in OSCC with higher histological grading (G2 and G3) and advanced stage (pT4) sub-groups. Kaplan-Meyer survival analysis suggested that patients with positive pEGFR-Tyr 845 tumors had a worse prognosis and were bad responders to chemotherapy. These results confirm the central role of EGF-R activation status as a prognostic biomarker in OSCC.

Transcriptional Shift Identifies a Set of Genes Driving Breast Cancer Chemoresistance

BackgroundDistant recurrences after antineoplastic treatment remain a serious problem for breast cancer clinical management, which threats patients’ life. Systemic therapy is administered to eradicate cancer cells from the organism, both at the site of the primary tumor and at any other potential location. Despite this intervention, a significant proportion of breast cancer patients relapse even many years after their primary tumor has been successfully treated according to current clinical standards, evidencing the existence of a chemoresistant cell subpopulation originating from the primary tumor.Methods/FindingsTo identify key molecules and signaling pathways which drive breast cancer chemoresistance we performed gene expression analysis before and after anthracycline and taxane-based chemotherapy and compared the results between different histopathological response groups (good-, mid- and bad-response), established according to the Miller & Payne grading system. Two cohorts of 33 and 73 breast cancer patients receiving neoadjuvant chemotherapy were recruited for whole-genome expression analysis and validation assay, respectively. Identified genes were subjected to a bioinformatic analysis in order to ascertain the molecular function of the proteins they encode and the signaling in which they participate. High throughput technologies identified 65 gene sequences which were over-expressed in all groups (P ≤ 0·05 Bonferroni test). Notably we found that, after chemotherapy, a significant proportion of these genes were over-expressed in the good responders group, making their tumors indistinguishable from those of the bad responders in their expression profile (P ≤ 0.05 Benjamini-Hochgerg`s method).ConclusionsThese data identify a set of key molecular pathways selectively up-regulated in post-chemotherapy cancer cells, which may become appropriate targets for the development of future directed therapies against breast cancer.

Tumor Regression Grades: Can They Influence Rectal Cancer Therapy Decision Tree?

Background. Evaluating impact of tumor regression grade in prognosis of patients with locally advanced rectal cancer (LARC). Materials and Methods. We identified from our colorectal cancer database 168 patients with LARC who received neoadjuvant therapy followed by complete mesorectum excision surgery between 2003 and 2011: 157 received 5-FU-based chemoradiation (CRT) and 11 short course RT. We excluded 29 patients, the remaining 139 were reassessed for disease recurrence and survival; the slides of surgical specimens were reviewed and classified according to Mandard tumor regression grades (TRG). We compared patients with good response (Mandard TRG1 or TRG2) versus patients with bad response (Mandard TRG3, TRG4, or TRG5). Outcomes evaluated were 5-year overall survival (OS), disease-free survival (DFS), local, distant and mixed recurrence. Results. Mean age was 64.2 years, and median followup was 56 months. No statistically significant survival difference was found when comparing patients with Mandard TRG1 versus Mandard TRG2 (p = .77). Mandard good responders (TRG1 + 2) have significantly better OS and DFS than Mandard bad responders (TRG3 + 4 + 5) (OS p = .013; DFS p = .007). Conclusions. Mandard good responders had a favorable prognosis. Tumor response (TRG) to neoadjuvant chemoradiation should be taken into account when defining the optimal adjuvant chemotherapy regimen for patients with LARC.

PEGFR-Tyr 845 expression as prognostic factors in oral squamous cell carcinoma

The EGFR (epidermal growth factor receptor) a member of the family of transmembrane protein kinase receptors known as the erbB family shows a significant correlation with the presence of metastases and poorly differentiated oral cancer. Aim of the present work is to define the key-role of EGFR in oral cancer prognosis. We have analyzed the EGFR expression on 149 cases of oral squamous cell cancers (OSCC) and we have found that it was poorly expressed in normal oral epithelium, but its expression was significantly increased in OSCCs. Moreover, we have recorded that both pEGFR-Tyr 845 and pEGFR-Tyr 1068 were mainly distributed in high histological grading and in advanced stages. Western blotting has confirmed the total absence of EGFR phosphorylation in normal oral epithelium and the higher level of protein phosphorylation in representative cases of OSCCs. The EGF-R amplification was found by fluorescence in situ hybridization (FISH) in 14% of OSCC; interestingly, EGF-R amplification was mainly observed in OSCC with higher histological grading (G2 and G3) and advanced stage (pT4) sub-groups. Kaplan-Meyer survival analysis suggested that patients with positive pEGFR-Tyr 845 tumors had a worse prognosis and were bad responders to chemotherapy. These results confirm the central role of EGF-R activation status as a prognostic biomarker in OSCC.

Temporal processing dysfunction in schizophrenia as measured by time interval discrimination and tempo reproduction tasks

Time perception deficiency has been implicated in schizophrenia; however the exact nature of this remains unclear. The present study was designed with the aim to delineate timing deficits in schizophrenia by examining performance of patients with schizophrenia and healthy volunteers in an interval discrimination test and their accuracy and precision in a pacing reproduction–replication test. The first task involved temporal discrimination of intervals, in which participants (60 patients with schizophrenia and 35 healthy controls) had to judge whether intervals were longer, shorter or equal than a standard interval. The second task required repetitive self-paced tapping to test accuracy and precision in the reproduction and replication of tempos. Patients were found to differ significantly from the controls in the psychoticism scale of EPQ, the proportion of correct responses in the interval discrimination test and the overall accuracy and precision in the reproduction and replication of sound sequences (p < 0.01). Within the patient group bad responders concerning the ability to discriminate time intervals were associated with increased scores in the Positive and Negative Syndrome Scale (PANSS) and in the Brief Psychiatric Rating Scale (BPRS) in comparison to good responders (p < 0.01). There were no gender effects and there were no differences between subgroups of patients taking different kinds or combinations of drugs. Analysis has shown that performance on timing tasks decreased with increasing psychopathology and therefore that timing dysfunctions are directly linked to the severity of the illness. Different temporal dysfunctions can be traced to different psychophysiological origins that can be explained using the Scalar Expectancy Theory (SET).

Early Interim PET Scans in Diffuse Large B-Cell Lymphoma: Can There Be Consensus About Standardized Reporting, and Can PET Scans Guide Therapy Choices?

The prognosis value of interim positron emission tomography (PET) remains controversial in diffuse large B-cell lymphoma (DLBCL) patients because of the absence of consensus on criteria able to early identify good and bad responders to treatment. Visual interpretation using the International Harmonization Project (IHP) criteria, primarily established for end of treatment evaluation, was related to a low positive predictive value of treatment failure. The 5-point scale (5PS) that refers the residual uptake to the liver as background tissue was shown to slightly reduce false-positive interim PET interpretations compared to IHP criteria. Semiquantification of fluorodeoxyglucose (FDG) uptake using standardized uptake value (SUV) and assessment of reduction of maximum SUV (SUVmax) between baseline and interim PET drastically improves both the interpretation accuracy and the interobserver reproducibility, and better predicts patient outcome than visual analysis. This latter approach is feasible in a multicenter setting and allows clinicians to design a risk-adapted therapeutic strategy based on early PET response assessment.

Standardized pathologic evaluation of the esophagus after neoadjuvant chemoradiation.

136 Background: The main objective of this study was to develop and validate a method to reconstruct the gross and clinical tumor volume (GTV and CTV) on the esophageal specimen in order to facilitate a good pathologic examination of the original tumor area after neo-adjuvant chemoradiation (CRT). Methods: The GTV and CTV borders of 25 patients were defined by a radiation oncologist on the planning CT in relation to 5 anatomical reference points. After CRT, the GTV and CTV borders were marked in vivo on the esophagus during surgical resection. Finally, the pathologist evaluated the presence of macroscopic and microscopic tumor in- and outside the GTV and CTV. The radiation tumor response was scored according to the standardized 5-tier Mandard classification. Radiation induced side effects were scored as well. Results: The Mandard classification could be scored on basis of the GTV alone in 68% of the cases (N=17). For the other patients (N=8), the GTV and the CTV should both be incorporated for correct evaluation of the tumor response. Five patients (20%) showed complete tumor response (Mandard 1), 68% (N=17) showed partial response (Mandard 2-3) and 12% (N=3) showed hardly any response (Mandard 4-5). In the partial responders, macroscopic tumor was found within the delineated GTV and microscopic tumor remained within the CTV both in 100% of the cases. In two patients (40%) with hardly any response, microscopic tumor was also found outside the CTV. This might be caused by tumor growth during the neo-adjuvant treatment or by geographical miss. Nine patients turned out to have positive lymph nodes. On average 18 (range 8-30) lymph nodes were evaluated per patients. Giant cell reactions, lymphocyte infiltration, and fibrosis, which indicate tumor regression were seen in the CTV and GTV, and were most pronounced in the GTV. Conclusions: This study suggested that demarcation of the GTV and CTV on the esophagus in vivo is important for standardized pathologic evaluation of the esophagus after neo-adjuvant chemoradiation. Furthermore using this method we determined microscopic tumor outside the CTV in 40% of the cases (N=2) of the bad responders (Mandard 4-5), illustrating the importance of our method in this patient category.

Erythropoietic response to erythropoiesis-stimulating agents and outcome: should we give up the haemoglobin target approach?

The hypothesis that complete anaemia correction with erythropoiesis-stimulating agents (ESA) would reduce the risk of death and cardiovascular and renal endpoints among patients with chronic kidney disease (CKD) has been largely tested in the last 15 years. Despite expectations, no study has demonstrated any clinical benefit except some improvement in quality of life by targeting at high haemoglobin (Hb) levels in comparison to partial anaemia correction. Some of these studies even found a trend towards increased risk of cardiovascular endpoints (i.e. death, stroke, acute myocardial infarction and hospitalization for congestive heart failure in different combinations) at primary or secondary analyses [1–4]. A lively debate has developed focussing on possible explanations why clinical trials have not confirmed what has been shown by observational studies: the higher the Hb the better the outcome. The Trial to Reduce Cardiovascular Events with Aranesp Therapy (TREAT) study randomized 4038 patients with Type 2 diabetes and CKD Stages III–IV to darbepoetin alfa (Hb of 13 g/dL) or to placebo (with rescue darbepoetin alfa for Hb <9.0 g/dL) [5]. Results were neutral on primary composite cardiovascular and renal outcomes, but a higher risk of stroke, death for cancer in patients with previous history of malignancies and thromboembolic events were found at secondary analyses in those randomized to complete anaemia correction. This parallelled a reduction in cardiac revascularization procedures, transfusion rates and improvement in quality of life in this group [6]. Possible implications of these findings have already been discussed [7]. In this editorial, we would like to examine carefully and comment on the recent secondary analysis of the TREAT data by Solomon et al. [8], which investigated the relationship between initial response to darbepoetin alfa, patient characteristics and outcomes. For this purpose, patients in the darbepoetin-alfa group having a Hb change <2% in the first month after the initial weight-based two doses of darbepoetin alfa were defined as having a poor response and compared with the remaining ones (upper three quartiles of Hb changes). After adjusting for outcome-related covariates, this subgroup had a higher risk of reaching the cardiovascular composite endpoint or death from any cause than the better Hb responders who had an event rate similar to the placebo group. Conversely, both subgroups had similar stroke rates that were higher than the placebo group. This post hoc analysis suggests that after just two doses of darbepoetin alfa, it is possible to classify the patients in good and bad responders and this may translate into a different outcome risk. This may be an easy method to identify high-risk patients and tailor ESA treatment accordingly, avoiding excessive doses. However, as admitted by the authors, these findings cannot identify the culprit of increased cardiovascular risk following ESA treatment. Is this a consequence of too high ESA doses especially in those who are hyporesponsive to treatment or does this merely reflect patient characteristics?

The Cyclooxygenase-1 C50T Polymorphism Is Not Associated with Aspirin Responsiveness Status in Stable Coronary Artery Disease in Tunisian Patients

In this study, we evaluate the relationships between aspirin nonresponsiveness and the cyclooxygenase-1 (Cox-1) gene C50T polymorphism in stable coronary artery disease (CAD) in Tunisian patients. One hundred twenty-five stable CAD patients were included. The Cox-1 gene C50T polymorphism was determined by the polymerase chain reaction/restriction fragment length polymorphism method. Aspirin response was evaluated by measuring the collagen epinephrine closer time and the urinary dehydro-thromboxane B2 excretion. According to the collagen epinephrine closer time values, the frequency of the -50T allele was not significantly different in bad responders when compared with good responders (36.8% vs. 15.7%; p=0.1). Similarly, the presence of the -50T mutant allele was not statistically different comparing bad and good responders according to the urinary 11-dehydro-thromboxane B2 excretion concentration (60% vs. 40%; p=0.43). Our study did not demonstrate any association between the Cox-1 gene C50T polymorphism and aspirin nonresponsiveness status in stable CAD patients.

Dynamin binding protein gene expression and memory performance in aged rats

Previous studies have shown that messenger RNA (mRNA) of the dynamin-binding protein (DNMBP), a scaffold protein regulating actin cytoskeleton and synaptic vesicle pools, is lower in neuropathologically-confirmed Alzheimer's brains. Here we investigated whether a deficit in long term memory formation during physiological aging is also associated with lower DNMBP expression. Hippocampal DNMBP mRNA was quantified by quantitative real time reverse transcriptase polymerase chain reaction (qRT-PCR) following inhibitory avoidance task in aged (26- to 27-month-old) rats that, according to memory performance, were ranked as good responders (GR) and bad responders (BR), in adult (3-month-old), late-adult (19-month-old), and aged (26–27-month-old) naive animals. We found that DNMBP mRNA levels were significantly higher in naive adults versus late adult and aged naive rats, in GR versus BR, and in pooled GR and BR versus aged-matched controls. Our data provide the first evidence that hippocampal DNMBP mRNA expression is reduced during physiological aging, and suggest that the capability to increase the expression of this mRNA may be a requirement for preserving long term memory formation during aging.

Interim FDG-PET Scan in Hodgkin's Lymphoma: Hopes and Caveats

FDG-PET has recently emerged as an important tool for the management of Hodgkins lymphoma. Although its use for initial staging and response evaluation at the end of treatment is well established, the place of interim PET for response assessment and subsequent treatment tailoring is still quite controversial. The use of interim PET after a few cycles of chemotherapy may allow treatment reduction for good responders, leading to lesser treatment toxicities as well as early treatment adaptation for bad responders with a potential higher chance for cure. Interpretation of interim PET is a rapidly moving field. Actually, visual interpretation is preferred over quantitative interpretation in this situation. The notion of minimal residual uptake emerged for faint persisting FDG uptake, but has evolved during the recent years. Guidelines using mediastinum and liver as references have been proposed at the expert meeting in Deauville 2009. Actually, several trials are ongoing both for localised and advanced disease to evaluate the FDG-PET potential for early treatment monitoring and tailoring. Until the results of these prospective randomized trials become available, treatment changes according to the interim PET results should remain inappropriate and limited to well-conducted clinical trials.

Micro‐RNA profiles in osteosarcoma as a predictive tool for ifosfamide response

Micro-RNAs (miRNA) are currently used as cancer biomarkers for hematological cancers and solid tumors. Osteosarcoma is the first primary malignant bone tumor, characterized by a complex genetic and resistance to conventional treatments. For this latter property, the median survival has not been improved since 1990 despite preoperative administration of chemotherapeutic agents. The prediction of tumor response before chemotherapy treatment would constitute a major progress for this pathology. We assessed in this study if miRNA profiling could surpass the current limitations for osteosarcoma diagnosis. We measured the miRNA expression in different osteosarcoma samples: (i) 27 osteosarcoma paraffin-embedded tumors from patients, (ii) human osteosarcoma cell lines, and (iii) tumors from a syngeneic rat osteosarcoma model, recapitulating human osteosarcoma. miRNA profiles were determined using microfluidic cards performing high-throughput TaqMan(®) -based PCR assays, called TaqMan(®) Low Density Arrays. Osteosarcoma of rat and human origins showed a miRNA signature, which could discriminate good from bad responders. In particular, we identified five discriminating miRNAs (miR-92a, miR-99b, miR-132, miR-193a-5p and miR-422a) in patient tumors, which could be easily transferable to diagnosis. These discriminating miRNAs, as well as those identified in rat, targeted the TGFβ, the Wnt and the MAP kinase pathways. These results indicate that our platform constitutes a potent diagnostic tool to predict tumor sensitivity to a drug in attempt to better adapt treatment to tumor biological specificities and also to identify new potential therapeutic strategies.

Response to Ranibizumab Therapy in Neovascular AMD – An Evaluation of Good and Bad Responders

Treatment of neovascular age-related macular degeneration (AMD) with Lucentis shows a broad spectrum regarding the course of visual acuity (VA). While some patients show a good response (increase in VA), others disclose much less promising results. A retrospective data analysis of all eyes treated for neovascular AMD at the University Hospital of Zurich, Switzerland for at least 12 months was carried out. The courses of VA between the 90th (good responders, GR) and the 10th (bad responders, BR) percentiles were compared at 3, 12 and 24 months from baseline. An analysis regarding demographic data, lesion type and size as well as injection frequency and visits was done and predictive factors for GR and BR were evaluated. Marked differences in the course of VA between GR (n = 30) and BR (n = 30) are already observed 3 months from baseline. In GR the gains in VA after 3, 12 and 24 were 15.7 +/- 9 letters ETDRS, 25.3 +/- 7 and 14.0 +/- 14. BR showed a deterioration of 8.3 +/- 11 letters ETDRS after 3, 22.1 +/- 8 after 12 and 23.6 +/- 13 after 24 months. The gender distribution was equal with a higher percentage of female patients (64 % in BR and 66 % in GR). The baseline VA was statistically significantly lower in GR (45.7 +/- 10 vs. 55.4 +/- 11, p < 0.05) than in BR. No other significant differences in baseline data were found, and no predictor for group membership could be identified. Only the course of VA in the first three months seems to be of value for an estimation of the response to treatment. In the future the response to treatment in the early phase may influence the treatment algorithm and the injection frequency.

Husserlian Self-Awareness and Selective Serotonin Reuptake Inhibitors

Abstract: The goal of the paper is to offer a model of self-awareness that fits the testimony of both good and bad responders to selective serotonin reuptake inhibitors (SSRIs), of which fluoxetine (Prozac; Lilly, Indianapolis, IN) is probably the most well known. After a review of troubling current uncertainties concerning how and for whom SSRIs are therapeutic, it is argued that SSRIs, as a rule, lessen the emotionality of SSRI subjects in favor of an increased cognitive and volitional orientation. Traditional empiricist and rationalist accounts self-awareness fail to provide models that adequately explain how such a shift from an active emotional response to an increased cognitive/volitional orientation is possible. Instead, notions of self-awareness, as understood by founding phenomenologist, Edmund Husserl, fit the testimony of SSRI subjects.

Hemoglobin and hematocrit are not such good candidates to detect autologous blood doping

In a recent manuscript published in the International Journal of Hematology, Sallet et al. [1] reported absolute norms of variation established for a maximal 15 days period to distinguish autologous blood transfusion from normal modifications, using hematocrit (Ht), hemoglobin ([Hb]) and stimulation index (Off-hr) (calculated using the following equation [Hb] – 60H reticulocytes in %) as markers. The thresholds the authors proposed are 6% for Ht, 4% for [Hb] and 20% for Off-hr. Although we grant this team for setting up new parameters to fight against blood doping, the application of the indirect detecting methodology for blood doping based on maximal variation of Ht, [Hb] and Off-hr requires additional considerations. We are concerned about the method used by the authors to establish the norms to tease out the ‘‘abnormal’’ variation. First, we are wondering how this norms based on the highest blood variations drawn from a group of 7 subjects with only 4 submitted to moderate altitude during 7 days can be statistically representative of physiological maximal variation of a ‘‘normal’’ population. Moreover, based on the literature defining good and bad responders [2], some more caution should be taken in the interpretation of the results. In addition, we believe that the conclusion of this study might lead to a type I error (i.e. false positive result) and that the norms proposed are likely ranged within the physiological values. That is why we disagree when the authors claim that ‘‘the absolute norms of variation of strictly above 6% for hematocrit; 4% for hemoglobin concentration and 20% for the stimulation index, from two blood samples spaced at an interval of maximum 15 days, were established in such a way as to detect autologous blood transfusion and distinguish an ‘abnormal’ variation’’. For the authors of the present paper [1], the purpose of using altitude exposure is to obtain a natural stimulation of the erythropoiesis allowing them to compare with the effects of the autologous transfusion. Unfortunately, as attested by their results, the hypoxic stimulus they used was rather mild and did not elicit any alteration in the measured parameters. We think that a more severe stimulus even using intermittent hypoxia would have been more appropriate first as a comparison point and second to stick with what is broadly used by the athletes. Indeed, in a study involving elite endurance runners submitted to an intermittent hypoxic training (living high–training low), we reported 5.7% increase in Ht in average after 6 days of exposure at 2,500 m during the nights (H3000) [3]. If we examine our data individually between the baseline and H3000, 2 of our subjects expressed Ht, [Hb] and OFF-hr higher than the ‘‘abnormal’’ of variation proposed by Sallet al. [1]: Ht: ?8.1%, [Hb]: ?9.7% and OFF-hr: ?20.5% V. Pialoux R. Mounier Laboratoire de Biologie des Activites Physiques et Sportives, Faculte de Medecine, 63000 Clermont-Ferrand, France

Functional imaging using DCE-US: Which parameter for the early evaluation of antiangiogenetic therapies?

3524 Background: The early evaluation of anti-angiogenic treatments is a challenge in oncology. Functional imaging methods based on the measure of tumoral vascularization have been developed using different modalities (CT, MRI, US). We analyzed the response in four studies using different targeted treatments with dynamic contrast enhanced-ultrasonography (DCE-US). Seven parameters characterizing tumoral perfusion were estimated. The objective of the study was to determine which parameter is the most appropriate to confirm earlier the efficacy of treatments. Methods: A total of 823 DCE-US were performed in 117 patients included in 4 following studies (multikinase inhibitor targeting angiogenic-receptor with a cytotoxic or thyrosine-kinase inhibitor targeted angiogenic-receptor and C-kit or monoclonal antibody anti-VEGFR). Each DCE-US was performed using contrast agent (Sonovue, Bracco) with perfusion and quantification softwares (Toshiba) from raw linear data with a high temporal resolution: 4 frames per second during 3 minutes. Seven quantitative parameters of perfusion were estimated: peak intensity (PI) and area under the total curve (AUC), area under the wash-in (AUWI), area under the wash-out (AUWO), time to peak intensity, mean transit time (MTT), wash-in slope. DCE-US were performed before treatment and after D3, D 8, 15, 21 (according each study design) and every 2 months. Patients were classified in good responders and bad responders according the response (RECIST on CT-scan) after 2 cycles or 2 months. Results: Among the 7 parameters, 2 parameters related to the blood volume studies (AUC and AUWO) were always earlier significantly modified (p = 0.04 to p = 0.004). One was never modified: MTT. For the 4 others, it's depending of each study. Conclusions: DCE-US appears as a sensitive tool to evaluate tumoral response to anti-angiogenic drugs. Functional parameters related to the blood volume are more pertinent and represent a key add value to early evaluation of these therapies studies. No significant financial relationships to disclose.

Expression of PRAME and WT1 in Multiple Myeloma Patients during High Dose Chemotherapy and Auto-SCT

High dose therapy enables further improvement in the outcome of multiple myeloma (MM) patients. However, it is still necessary to determine prognostic factors that may influence treatment results and provide additional criteria for the precise selection of treatment approaches. There are several tumor associated genes (MAGE, LAGE, GAGE, PRAME) that are over-expressed in different malignancies including MM. These genes are believed to modulate cancer properties and should be taken into account during treatment. Their significance as prognostic factors is under investigation. The aim of our study was to analyze the expression levels of PRAME and WT1 genes in MM patients during high dose chemotherapy following by auto-SCT. After having informed consent 25 primary MM patients were included into this study. The median age was 48 years (range, 31–62). All patients were treated by 3 cycles of VAD, Cyclophosphamide 6g/m2 + G-CSF to mobilize Stem cells, EDAP, melphalan 200 mg/m2 followed by auto-SCT. As second line therapy we used bortezomib+dexamethasone. Quantitative PRAME and WT1 gene expression analysis was performed by means of RQ-PCR. Results were normalized against expression of ABL gene which was used as internal control. Investigation was performed before treatment (n=25), after three VAD cycles (n=12), and before (n=5) and after auto-SCT (n=4). In primary MM patients: PRAME gene expression was found in 68% (n=17), WT1 in 24% (n=6) of patients, all of whom were PRAME-positive. Median expression levels were 0.1% (0.001–132%) for PRAME and 0.01% (0.002–0.07%) for WT1. PRAME and WT1 expression did not correlate with tumor bulk and was independent of the levels of M-protein, beta-2M and albumin. The expression of PRAME significantly decreased after 3 VAD cycles to 0.001–207% (n=8), at the moment of auto-SCT it was 0.05–6.1% (n=3) and after auto-SCT it was 0.013–4.9% (n=3). However for WT1, we observed increased of WT1 expression after 3 VAD cycles to 0.004-0.05% (n=4)and at the moment of auto-SCT it was 0.035–0.4% (n=3) and after auto-SCT – 0.019–2.03% (n=3). In the patients with high primary PRAME expression (>median expression) the frequency of CR+PR was significantly lower then in PRAME-negative primary patients and in patients with low (<median expression) primary PRAME expression (55% vs 84, p = 0.04). It was found also that WT1-positive primary patients were bad responders and they achieved only minimal response after 3 VAD cycles. It should be stressed that during treatment in a small number of initially negative PRAME and WT1 gene patients, we demonstrated detection by PCR. We detected the appearance of gene expression at low levels in 1 of 8 initially negative PRAME (6.1%) and in 5 of 19 initially negative WT1 (range of level 0.01–0.4%). The detection of gene expression did not correlate with disease status. All these patients achieve CR+ VGPR. In one of these secondary positive patients (acquired PRAME and WT1) relapse occurred.Conclusion: Expression of PRAME gene was found in 68% primary patients and the level of PRAME decreased with tumor reduction. High expression level of PRAME turned out to be a factor of unfavorable prognosis. Expression of WT1 was found in 24% of MM patients all of whom were PRAME-positive. WT1 expression increased during treatment in a small group of pts. Some initially negative pts acquired PRAME and WT1 expression during treatment, but clinical relevance of it is not clear so far.