Stratification Of Patients For Therapy Research Articles

Prediction of anxious depression using multimodal neuroimaging and machine learning

Anxious depression is a common subtype of major depressive disorder (MDD) associated with adverse outcomes and severely impaired social function. It is important to clarify the underlying neurobiology of anxious depression to refine the diagnosis and stratify patients for therapy. Here we explored associations between anxiety and brain structure/function in MDD patients. A total of 260 MDD patients and 127 healthy controls underwent three-dimensional T1-weighted structural scanning and resting-state functional magnetic resonance imaging. Demographic data were collected from all participants. Differences in gray matter volume (GMV), (fractional) amplitude of low-frequency fluctuation ((f)ALFF), regional homogeneity (ReHo), and seed point-based functional connectivity were compared between anxious MDD patients, non-anxious MDD patients, and healthy controls. A random forest model was used to predict anxiety in MDD patients using neuroimaging features. Anxious MDD patients showed significant differences in GMV in the left middle temporal gyrus and ReHo in the right superior parietal gyrus and the left precuneus than HCs. Compared with non-anxious MDD patients, patients with anxious MDD showed significantly different GMV in the left inferior temporal gyrus, left superior temporal gyrus, left superior frontal gyrus (orbital part), and left dorsolateral superior frontal gyrus; fALFF in the left middle temporal gyrus; ReHo in the inferior temporal gyrus and the superior frontal gyrus (orbital part); and functional connectivity between the left superior temporal gyrus(temporal pole) and left medial superior frontal gyrus. A diagnostic predictive random forest model built using imaging features and validated by 10-fold cross-validation distinguished anxious from non-anxious MDD with an AUC of 0.802. Patients with anxious depression exhibit dysregulation of brain regions associated with emotion regulation, cognition, and decision-making, and our diagnostic model paves the way for more accurate, objective clinical diagnosis of anxious depression.

Imaging evaluation of thymic tumors.

An integral part of managing patients with thymoma and thymic carcinoma is imaging. At diagnosis and staging, imaging helps demonstrate the extent of local invasion and distant metastases which allows the proper stratification of patients for therapy. For decades, the predominant staging system for thymic tumors was the Masaoka-Koga staging system. More recently, however, the International Association for the Study of Lung Cancer, the International Thymic Malignancies Interest Group (ITMIG), the European Society of Thoracic Surgeons, the Chinese Alliance for Research on Thymomas, and the Japanese Association of Research on Thymus partnered together to develop a tumor-node-metastasis (TNM) staging system specifically for thymic tumors based on a retrospective database of nearly 10,000 patients. The TNM 8th edition defines specific criteria for thymic tumors. Imaging also serves to assess treatment response and detect recurrent disease after various treatment modalities. The Response Evaluation Criteria in Solid Tumors (RECIST) version 1.1 is currently used to assess response to treatment. ITMIG recommends certain modifications to RECIST version 1.1, however, in thymic tumors due to unique patterns of spread. While there is often overlap, computed tomography (CT), magnetic resonance imaging (MRI), and positron emission tomography/computed tomography (PET/CT) characteristics can help differentiate thymoma and thymic carcinoma, with newer CT and MRI techniques under evaluation showing encouraging potential.

The value of prognostic and predictive parameters in early-stage lung adenocarcinomas: A comparison between biopsies and resections

IntroductionSince lung adenocarcinoma (LUAD) biopsies are usually small, it is questionable if their prognostic and predictive information is comparable to what is offered by large resection specimens. This study compares LUAD biopsies and resection specimens for their ability to provide prognostic and predictive parameters. MethodsWe selected 187 biopsy specimens with stage I and II LUAD. In 123 cases, subsequent resection specimens were also available. All specimens were evaluated for growth pattern, nuclear grade, fibrosis, inflammation, and genomic alterations. Findings were compared using non-parametric testing for categorical variables. Model performance was assessed using the area under the curve for both biopsies and resection specimens, and overall (OS) and disease-free survival (DFS) was calculated. ResultsThe overall growth pattern concordance between biopsies and resections was 73.9%. The dominant growth pattern correlated with OS and DFS in resected adenocarcinomas and for high-grade growth pattern in biopsies. Multivariate analysis of biopsy specimens revealed that T2-tumors, N1-status, KRAS mutations and a lack of other driver mutations were associated with poorer survival. Model performance using clinical, histological and genetic data from biopsy specimens for predicting OS and DSF demonstrated an AUC of 0.72 and 0.69, respectively. ConclusionsOur data demonstrated the prognostic relevance of a high-grade growth pattern in biopsy specimens of LUAD. Combining clinical, histological and genetic information in one model demonstrated a suboptimal performance for DFS prediction and good performance for OS prediction. However, for daily practice, more robust (bio)markers are required to predict prognosis and stratify patients for therapy and follow-up.

Understand SLE heterogeneity in the era of omics, big data, and artificial intelligence

AbstractSystemic lupus erythematosus (SLE) is a systemic autoimmune disease characterized by extraordinary heterogeneity, due to the complex pathogenesis and diverse manifestations. Stratification of patients for therapy and prognosis represents a major challenge to manage SLE. Conventional biomarkers for disease diagnosis and activity assessment provide very limited insight into immunological pathogenesis and therapeutic response rates. The advancement of “omics” technologies including genomics, transcriptomics, proteomics, and metabolomics has constituted an unprecedented opportunity to characterize the immunopathological landscape in individual patients with SLE. Indeed, genomic studies reveal a subset of SLE patients carrying one or more functional single nucleotide polymorphisms (SNPs) underlying immune dysregulation while transcriptomic studies have revealed subgroups in SLE patients showing distinct signatures for Type I interferon (TI‐IFN) pathway activation or aberrant differentiation of B cells into plasma cells. This review will summarize results from the latest studies using omics technology to understand SLE heterogeneity. In addition, we propose that the application of artificial intelligence, such as by machine learning‐based nonlinear dimensionality reduction method uniform manifold approximation and projection (UMAP) can further strengthen the analysis of omics big data. The combination of new technology and novel analysis pipeline can lead to breakthroughs in stratifying SLE patients for a better monitoring of disease activity and more precise design of treatment regime, not only for conventional immunosuppression but also novel immunotherapies targeting B‐cell activating factor (BAFF), TI‐IFN, and interleukin 2 (IL‐2).

Biomarkers for Antiplatelet Therapies in Acute Ischemic Stroke: A Clinical Review.

Stroke is one of the world's leading causes of disability and death. Antiplatelet agents are administered to acute ischemic stroke patients as secondary prevention. Clopidogrel involves biotransformation by cytochrome P450 (CYP) enzymes into an active metabolite, and single nucleotide polymorphisms (SNPs) can influence the efficacy of this biotransformation. Despite the therapeutic advantages of aspirin, there is significant inter-individual heterogeneity in response to this antiplatelet drug. In this clinical review, the recent advances in the biomarkers of antiplatelet agents in acute ischemic stroke are discussed. The studies reviewed herein highlight the clinical relevance of antiplatelet resistance, pharmacotherapy of antiplatelet agents predicting drug response, strategies for identifying aspirin resistance, pharmacogenetic variants of antiplatelet agents, miRNAs, and extracellular vesicles (EVs) as biomarkers toward the personalized approach in the management of acute ischemic stroke. The precise pathways contributing to antiplatelet resistance are not very well known but are presumably multi-factorial. It is essential to understand the clinical relevance of clopidogrel and aspirin-related single nucleotide polymorphism (SNPs) as potential predictive and prognostic biomarkers. Prasugrel is a next-generation antiplatelet agent that prevents ADP-platelet activation by binding irreversibly to P2Y12 receptor. There are sporadic reports of prasugrel resistance and polymorphisms in the Platelet endothelial aggregation receptor-1 (PEAR1) that may contribute to a change in the pharmacodynamics response. Ticagrelor, a direct-acting P2Y12-receptor antagonist, is easily absorbed and partly metabolized to major AR-C124910XX metabolite (ARC). Ticagrelor's primary active metabolite, ARC124910XX (ARC), is formed via the most abundant hepatic cytochrome P450 (CYP) enzyme, CYP3A4, and CYP3A5. The integration of specific biomarkers, genotype as well as phenotype-related data in antiplatelet therapy stratification in patients with acute ischemic stroke will be of great clinical significance and could be used as a guiding tool for more effective, personalized therapy.

Abstract B08: Tumor-educated platelets for early prostate cancer diagnosis, and therapy stratification in patients with metastasized castration resistant prostate cancer

Abstract One in every ten men diagnosed with prostate cancer will have metastases at some point, causing the high mortality of prostate cancer. Only earlier diagnosis has the chance to save those lives. However, correct therapy stratification may prolong the life of the patients without severely reducing the quality of life, thereby possibly turning a deadly disease into a chronic disease. For this to happen, we need predictive biomarkers that can be employed to longitudinally follow the disease in order to identify vulnerabilities and to monitor acquired resistance for personalized medicine. Using the platelet-enriched fraction of a blood sample, we here show that we can use the ability of platelets to sequester material during circulation to provide information derived from the tumor. This information could be used for early detection of prostate cancer. By using the platelet test in combination with PSA measurements, we may be able to rule out a majority of the false negatives associated with the PSA test. Furthermore, the platelet platform can be used in synergy with other biomarker sources such as circulating tumor DNA or protein biomarkers, providing the possibility to predict and monitor outcome of abiraterone therapy, and providing alternatives to the nonfunctional therapy by guiding therapy switching, in metastatic castration-resistant patients. Citation Format: Lee-Ann Tjon-Kon-Fat, Linda Köhn, Myron Best, Marie Lundholm, Thomas Wurdinger, Anders Widmark, Camilla Thellenberg-Karlsson, Pernilla Wikström, Jonas Nilsson. Tumor-educated platelets for early prostate cancer diagnosis, and therapy stratification in patients with metastasized castration resistant prostate cancer [abstract]. In: Proceedings of the AACR Special Conference on Advances in Liquid Biopsies; Jan 13-16, 2020; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(11_Suppl):Abstract nr B08.

Regulatory T cells and cytotoxic T cells close to the epithelial–stromal interface are associated with a favorable prognosis

ABSTRACT Cytotoxic T cells and regulatory T cells play a crucial role in the outcome of cancer patients. Besides the density of these cells, it was shown recently that the spatial distribution is equally important. Here, we specifically analyzed the spatial distribution of these T cell subtypes at the epithelial–stromal interface in a rectal cancer cohort and its relevance for prognosis. We studied a cohort of 191 patients with advanced rectal cancer treated by radiochemotherapy (RCT). Tissue microarrays were immunohistochemical double-stained by FoxP3+ and CD+. Cell densities were analyzed in the stromal and epithelial compartment. Additionally, an image analysis software calculated the distances of lymphocytes to the epithelial–stromal interface (ESI). CD8+ and FoxP3+ cell counts decreased clearly after RCT with the decrease of FoxP3+ being more pronounced than of CD8+ cells. In the invasive front, short distances of the ESI to CD8+ and to FoxP3+ cells were associated with improved overall survival. Cell counts in the stromal compartment had no influence on prognosis. No correlation between stromal and epithelial lymphocyte densities was observed. The distance of epithelial–stromal interface to CD8+ and FoxP3+ cells was more accurate in predicting prognosis in the stromal compartment of rectal cancer patients than mere cell counts and could thereby be means of better stratifying patients for therapy. This observation will have to be validated in future prospective studies with regard to other tumor entities and its implications for the responsiveness of tumors to new therapeutic modalities.

Highlights of This Issue

While trastuzumab-based chemotherapy has shown remarkable clinical benefits for HER2-positive breast cancer patients, a subset of patients (30%–40%) shows little or no effect. This highlights an important clinical need for biomarkers in addition to HER2 for better stratification of patients for therapy. In this multicenter retrospective study by Kang and colleagues, patients with HER2-positive breast cancer that also overexpress WBP2 had better pathologic complete response to trastuzumab-based neoadjuvant therapy. The potential to predict which patients would attain successful downstaging of their tumors from neoadjuvant therapy could help guide surgical decisions; for example, breast-conserving surgery versus mastectomy, consequently improving patients' outcome.Extending previous studies in more restricted populations, Nassar and colleagues examined 472 patients across the spectrum of urothelial carcinoma from nonmuscle-invasive bladder cancer (NMIBC) to upper-tract urothelial carcinoma (UTUC) for mutations in 237 cancer genes. Low-grade NMIBC has a distinct mutation spectrum from invasive disease, suggesting that it is truly a separate pathologic entity unrelated to invasive disease in most instances. Other key findings include enrichment of KDM6A mutations in women with low-grade NMIBC compared to men and identification of a subtype of high-grade UTUC with mutations in FGFR3, EP300, and PIK3CA associated with poorer survival.Drug-resistance is a major obstacle for the effective treatment of patients with high-grade serous ovarian cancer (HGSOC), often resulting in the selection of an ineffective treatment course. Using PET imaging with the radiotracer (4S)-4-(3-[18F]fluoropropyl)-L-glutamate ([18F]FSPG), Greenwood and colleagues showed decreased radiotracer uptake in chemotherapy-resistant A2780 tumors compared with parental drug-sensitive tumors, demonstrating the ability of [18F]FSPG to detect upregulated antioxidant pathways present in drug-resistant cancer. [18F]FSPG may, therefore, enable the identification of HGSOC patients who are refractory to standard-of-care, allowing the transferal of drug-resistant patients to alternative therapies and thereby improving outcomes in this disease.To understand the immunological nature of glioblastoma, Park and colleagues examined the expression of immune checkpoint inhibitory receptors in tumor-infiltrating CD8+ T cells from patients with primary glioblastoma and their differentiation status. In addition, they investigated the reinvigoration ability of tumor-infiltrating CD8+ T cells by ex vivo treatment with anti-PD-1 and anti-CTLA-4. They demonstrated that the differentiation status of tumor-infiltrating CD8+ T cells determines the reinvigoration ability. This study provides a rationale for developing optimal strategies for immune checkpoint inhibitor treatment in patients with glioblastoma.

P53 Immunoexpression in Laryngeal Squamous Cell Carcinoma

ABSTRACT: p53 is a marker described in the premalignant lesions with a high risk of malignant transformation for laryngeal cancer. It is a tumor suppressor gene that during the cancer gains also oncogenic activity. We aimed to study the p53 immunoexpression in 38 cases of laryngeal squamous cell carcinomas and the relation with the clinicopathological aspects. We obtained variable p53 expression regarding the differentiation degree and tumor stage. The higher p53 immunotaining values were observed in high grade and advanced stages lesions. P53 may be useful in identifying aggresive laryngeal squamous carcinomas, a useful aspect for better stratification of patients for therapy.

Reduced IL-2R signaling in T1D subjects is pervasive in the memory CD4 T effector compartment and is mediated through intrinsic mechanisms

Abstract The IL-2/IL-2R pathway is strongly implicated in type 1 diabetes (T1D). Yet, identifying the functional significance and underlying mechanisms of altered IL-2R signaling in T1D has been challenging due to variance in the subjects, cell types and the assays utilized. We performed a large, well-matched, cross-sectional study of control and T1D subjects (n>75 subjects per group), with an optimized IL-2/pSTAT5 assay (internal control CoV <7%). We found reduced responses to IL-2 in memory CD4 Treg and Teff cell populations of T1D subjects relative to controls. Known genetic risk alleles accounted for some, but not all, of the reduction in pSTAT5. To determine additional factors underlying reduced IL-2R signaling, we interrogated CD4 Teff cells of control and T1D subjects held constant for risk alleles associated with IL-2R signaling. IL-2 signaling in T1D memory CD4 Teff cells remained significantly lower than controls after in vitro activation and proliferation, suggesting a role for intrinsic factors. These intrinsic factors were not caused by divergence to MAPK or Akt signaling pathways, as measured by western blot. Instead, differences in basal activation state prior to in vitro activation correlated with IL-2 response, as measured by targeted flow and qPCR of activation and negative regulation. Thus, multiple factors including genetics, disease and activation state contribute to natural variation in IL-2 responsiveness, requiring precise assays and well-defined cohorts to reach solid conclusions. These findings offer a better understanding of the mechanisms contributing to reduced IL-2R signaling in T1D and have implications for development of more targeted IL-2-based therapies and stratification of patients for therapy.

Abstract P6-14-03: Genome wide association study (GWAS) to identify variants conferring ramucirumab-associated hypertension in the ROSE/TRIO-012 breast cancer trial

Abstract Background: In a candidate single nucleotide polymorphism (SNP) association study, we previously identified VEGFR-1 and VEGFR-2 SNPs strongly associated with treatment emergent hypertension (HT) in the ramucirumab (RAM) and docetaxel (Doc) arm in the ROSE/TRIO-012 study, a double-blinded multinational phase III trial that randomized 1,144 patients with advanced breast cancer to receive first-line Doc in combination with RAM or placebo (Mackey et al, JCO Jan 10, 2015:141-148; Mackey et al, JCO, Volume 33, Issue 15_suppl, May 20, 2015: 547). However, candidate SNP studies limit the number of genes for interrogation and a more comprehensive genome wide search may identify critical variants associated with the phenotype of HT. Preliminary analysis indicated that patients experiencing HT with RAM showed better overall survival (Mackey JR, et al (2015). Reply to H. Lee, et al. JCO; in press). These observations provide the potential to identify those patients with genetic variants for predisposition to RAM-associated HT to inform therapeutic decisions. Methods: Genotyping of samples is underway using Affymetrix SNP 6.0 arrays. Genotype data will be filtered for deviations from Hardy Weinberg Equilibrium and minor allele frequency of >0.05. Study subjects (n=792) provided ethics-committee approved prospective consent for this genetic study of whom 478 subjects were allocated RAM + Doc arm. Toxicity grades 0-1 (n= 394 controls; low toxicity) vs. grade >2 (n= 84 cases, high toxicity) is our binary outcome. Dominant genotypic model is assumed. Chi-square test, FDR and/or 10000 permutation tests will be employed (Golden Helix-SVS v8.3) and p<0.05 considered statistically significant. Population stratification will be identified (EIGENSTRAT) and association statistics will be corrected using Eigenvectors along with age as covariates. Fine mapping of loci showing significant associations will be attempted using imputation tools. Results and conclusions: We expect up to 700,000 SNPs to be retained after filtering based on our previous breast cancer GWAS analyses (Damaraju et al. Cancer Research (suppl); Vol 70 (24), page 258s, 2010 and Sehrawat et al Hum Genet. 2011 Oct;130(4):529-37) and 30,000 SNPs to show significance at a nominal p-value (0.05); these will be analysed for regions of high linkage disequilibrium to narrow down potential loci showing association with HT to serve as candidate markers in further independent validation studies. Cumulative dose to adverse events will be considered in the analysis. Identified loci will be interrogated for potential genes in the flanking regions with biological relevance based on pathway analysis. Identified variants from candidate SNP and GWAS may allow developing predictive tools to enable stratification of patients for therapies. The analysis is expected to be completed by mid- November, 2015. Citation Format: Mackey JR, Lipatov O, Martín M, Webster M, Hegg R, Verma S, Ramos-Vázquez M, Fresco R, Thireau F, Houé V, Press MF, Kumaran M, Damaraju S. Genome wide association study (GWAS) to identify variants conferring ramucirumab-associated hypertension in the ROSE/TRIO-012 breast cancer trial. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P6-14-03.

Synthesis, physicochemical and biological evaluation of technetium-99m labeled lapatinib as a novel potential tumor imaging agent of Her-2 positive breast cancer

Tumors that are Her-2-positive tend to grow and spread more quickly than other types of breast cancer. Overexpression of Her-2 can be a predictive biomarker for stratification of patients for therapy with Herceptin (containing humanized IgG1 monoclonal antibody trastuzumab) or Tykerb (containing lapatinib di-p-toluenesulfonate) drug. Usually, Her-2 status is determined by immunohistochemical (IHC) as well as fluorescent or chromogenic in situ hybridisation (FISH or CISH) analysis of biopsy material. The objective of the present work was to standardize the conjugation of anti-cancer drug lapatinib (which recognizes selectively the Her-2 extracellular domain) with technetium-99m complex, of type ‘4+1’, to obtain 99mTc(NS3)(CN-lapatinib) conjugate for use as in vivo tracer of the Her-2 expression in breast cancer. The conjugate 99mTc(NS3)(CN-lapatinib) was formed with high yield, high radiochemical purity and specific activity within the range 25–30 GBq/μmol. The biological in vitro and in vivo studies of the conjugate showed its high affinity to Her-2 receptor (Kd = 3.5 ± 0.4 nM, Ki = 2.9 ± 0.5 nM, Bmax = 2.4 ± 0.3 nM, approximate number of 2.4 × 106 binding sites per cell, IC50 = 41.2 ± 0.4 nM) and also pointed out to the clearance through the hepatic and renal route in comparable degree. Basing on these results one can conclude that 99mTc(NS3)(CN-lapatinib) conjugate could be a promising radiopharmaceutical for in vivo diagnosis of the Her-2 status in breast with impact on treatment planning.

New Oral Anticoagulants Increase Risk for Gastrointestinal Bleeding: A Systematic Review and Meta-analysis

A new generation of oral anticoagulants (nOAC), which includes thrombin and factor Xa inhibitors, has been shown to be effective, but little is known about whether these drugs increase patients' risk for gastrointestinal bleeding (GIB). Patients who require OAC therapy frequently have significant comorbidities and may also take aspirin and/or thienopyridines. We performed a systematic review and meta-analysis of the risk of GIB and clinically relevant bleeding in patients taking nOAC. We queried MEDLINE, EMbase, and the Cochrane library (through July 2012) without language restrictions. We analyzed data from 43 randomized controlled trials (151,578 patients) that compared nOAC (regardless of indication) with standard care for risk of bleeding (19 trials on GIB). Odds ratios (ORs) were estimated using a random-effects model. Heterogeneity was assessed with the Cochran Q test and the Higgins I(2) test. The overall OR for GIB among patients taking nOAC was 1.45 (95% confidence interval [CI], 1.07-1.97), but there was substantial heterogeneity among studies (I2,61%). Subgroup analyses showed that the OR for atrial fibrillation was 1.21 (95% CI, 0.91-1.61), for thromboprophylaxis after orthopedic surgery the OR was 0.78 (95% CI, 0.31-1.96), for treatment of venous thrombosis the OR was 1.59 (95% CI, 1.03-2.44), and for acute coronary syndrome the OR was 5.21 (95% CI, 2.58-10.53). Among the drugs studied, the OR for apixaban was 1.23 (95% CI, 0.56-2.73), the OR for dabigatran was 1.58 (95% CI, 1.29-1.93), the OR for edoxaban was 0.31 (95% CI, 0.01-7.69), and the OR for rivaroxaban was 1.48 (95% CI, 1.21-1.82). The overall OR for clinically relevant bleeding in patients taking nOAC was 1.16 (95% CI, 1.00-1.34), with similar trends among subgroups. Studies on treatment of venous thrombosis or acute coronary syndrome have shown that patients treated with nOAC have an increased risk of GIB, compared with those who receive standard care. Better reporting of GIB events in future trials could allow stratification of patients for therapy with gastroprotective agents.

Evaluation of thyroid nodules – combined use of 99mTc‐methylisobutylnitrile scintigraphy and aspiration cytology to assess risk of malignancy and stratify patients for surgical or nonsurgical therapy – a retrospective cohort study

Thyroid nodules are a common clinical problem, and differentiation between benign and malignant nodules is essential. The aim of this study was to evaluate an approach for cold thyroid nodules including (99m)Tc-methylisobutylnitrile (MIBI) scintigraphy to assess risk of malignancy and stratify patients for therapy. Retrospective cohort study; 391 patients with at least one cold thyroid nodule were consecutively admitted (between 1 January 2004 and 31 December 2006) and recommended for surgical or nonsurgical therapy. Thyroid ultrasonography, (99m)Tc-pertechnetate scintigraphy, laboratory tests, fine needle aspiration cytology (FNAC) and MIBI scintigraphy. 57·3% (224/391) had one cold nodule, 17·9% (70/391) had several cold nodules, and 24·8% (97/391) had both cold and hot nodules. MIBI scintigraphy was classified into 'positive' (16·1%, 63/391), 'weakly positive' (19·2%, 75/391) or 'negative' (64·7%, 253/391). FNAC was classified into benign (87·9%, 247/281), nondiagnostic (6·8%, 19/281) or suspicious/malignant (5·3%, 15/281). 127 patients received surgery, revealing malignancy in 13·3% (17/127), predominantly papillary (64·7%, 11/17) and follicular carcinoma (23·5%, 4/17). MIBI scintigraphy was 'positive' (64·7%, 11/17) or 'weakly positive' (23·5%, 4/17) in most patients with malignant findings. FNAC was unavailable in 23·5% (4/17) with malignancy, positive in 38·5% (5/13) and negative in 61·5% (8/13). Among patients undergoing surgery, sensitivity, specificity, negative and positive predictive values for MIBI scintigraphy were 88·2%, 35·5%, 95·1% and 17·4%, for FNAC 38·5%, 90·6%, 90·6% and 38·5%, respectively, and for the combination (MIBI scan + FNAC) 92·3%, 30·6%, 96·3% and 16·9%. Benign MIBI-positive nodules were predominantly follicular adenomas (68%, 33/48). Evaluation of cold thyroid nodules by MIBI scintigraphy aids therapeutic decisions: MIBI-negative findings support nonsurgical management in about two-thirds of patients, while MIBI-positive findings have an increased risk of malignancy, supporting surgical therapy. However, the positive predictive value was low, which requires further research.

Whole Genome Copy Number Variation Analysis of Chronic Lymphocytic Leukemia (CLL) Cells From Early-Intermediate Stage, High Risk CLL Patients Prior to First Treatment Reveals New Loss of Heterozygosity and Duplication Events in the CLL Genome.

Abstract 1265Poster Board I-287 IntroductionFluorescence in situ hybridization (FISH), in combination with other markers, is used as a prognostic tool for CLL patients at diagnosis. The presence or absence of trisomy 12 and deletions at 13q, 11q, and 17p helps to predict disease progression and to stratify patients for therapeutic decisions. We hypothesized that whole genome single nucleotide polymorphism (SNP)-based copy number variation (CNV) analysis would capture all of the information in current CLL FISH panels and would reveal new CNV features in the CLL genome. Patients and MethodsNineteen early-intermediate clinical stage, untreated CLL patients aged 29 to 77 were determined to be at high risk for disease progression by FISH, IgVH mutation status, ZAP-70, and CD38 prognostic markers. CLL cells and normal cells were separated by magnetic bead selection from patient peripheral blood samples with absolute lymphocyte counts that ranged from 7.4 to 162 × 109/L. CNV analysis was performed on purified genomic DNA from the CLL cells and from normal cells for each patient in order to distinguish acquired CNVs in malignant cells from polymorphic CNVs in the human genome. We used the Illumina human660w-quad beadchip, a SNP-based microarray for whole-genome genotyping and CNV analysis that contains more than 550,000 tag SNPs and approximately 100,000 additional markers that target regions of common CNV. CNV data was analyzed using CNV partition (Illumina Genome Studio software) and PennCNV. Results100% concordance is found between del(13q), del(11q), and del(17p) FISH abnormalities and loss of heterozygosity (LOH) at 13q, 11q, and 17p by CNV analysis. All three patients with trisomy 12 by FISH show copy number(CN)=3 of chromosome 12 by CNV analysis. Of 15 patients with del(13q) by FISH, 12 out of 15 have regions of hemizygous deletion on 13q that vary from ∼830 Kb to ∼38 Mb. The smallest region of LOH is located within 13q14.3. Three out of 15 patients show homozygous deletion within 13q14.3. One of these 3 patients has copy-neutral LOH of the entire 13q arm with an embedded 835 Kb segment of homozygous deletion at 13q14.3. Two patients have large discontinuous segments of LOH on 13q, indicating complex interstitial deletion events. Two out of 5 patients with del(13q) as a sole FISH abnormality show additional CNV events in the CLL genome. One of these patients has copy neutral LOH at 2q33.1-telomere(tel). One other patient with sole del(13q) FISH shows LOH events at 10q23.31-23.33 and at 15q15.1. Five out of six patients with del(11q) by FISH have either 13q LOH (n=4) or chromosome 12 CN=3 (n=1) without any other CNV events detected in the CLL genome. One patient with trisomy 12 and del(11q) by FISH has three additional acquired CNV abnormalities in the CLL genome: LOH at 7p15.2-tel, LOH at 11p13, and CN=3 at 3q24-tel. In contrast to patients with del(11q), del(13q), and trisomy 12, patients with del(17p) by FISH have numerous acquired CNV abnormalities in the CLL genome. These include LOH events at 1p34.3-p34.2, 2q34-q36.3, 3p21.31-tel, 4p13, 4p15.1-tel, 15q11.2-q14 and 15q14-q15.3, 16p13.3-tel, 16p13.11, 16p13.2, 18p11.21-tel, 20p11.21-tel, and 20q13.2-q13.31. CN=3 at 2p12-tel is detected in 2 out of 5 patients with 17p hemizygous deletion. One out of 5 patients with 17p hemizygous deletion shows CN=3 at 10q22.2-tel. One other patient also with 17p hemizygous deletion shows CN=3 at 22q12.2-tel. ConclusionsWhole genome CNV analysis by SNP-based microarrays greatly expands our ability to detect acquired genomic events in CLL cells. These events include hemizygous deletion, homozygous deletion, copy-neutral LOH, and CN=3 duplication. Detection of copy-neutral LOH is not possible by FISH or array comparative genomic hybridization technology. The current study reveals a high number of acquired CNV events in earlier stage, untreated CLL patients with 17p hemizygous deletion. This observation, indicative of genomic instability, is consistent with the known poor prognosis of del(17p) patients. The new somatic CNV abnormalities detected in CLL cells may help to discover additional genes or signaling pathways involved in CLL initiation and progression. In addition, the new CNV markers may be used in larger clinical studies to improve CLL prognosis and patient stratification for therapy. DisclosuresShanafelt:Genentech: Research Funding; Hospira: Membership on an entity's Board of Directors or advisory committees, Research Funding; Polyphenon E International: Research Funding; Celgene: Research Funding; Cephalon: Research Funding; Bayer Health Care Pharmaceuticals: Research Funding. Kay:Genentech, Celgene, Hospira, Polyphenon Pharma, Sanofi-Aventis: Research Funding; Biogenc-Idec, Celgene, Genentech, genmab: Membership on an entity's Board of Directors or advisory committees. Zent:Genentech, Bayer, Genzyme, Novartis: Research Funding.

BCL2 gene aberration as an IPI-independent marker for poor outcome in non-germinal-centre diffuse large B cell lymphoma

Aim:Diffuse large B cell lymphoma (DLBCL) is the most common lymphoid malignancy in the western hemisphere, and is characterised by a highly variable outcome that impedes individual risk assessment. Lacking...

Molecular abnormalities in Ewing’s sarcoma

Ewing’s sarcoma is one of the few solid tumors for which the underlying molecular genetic abnormality has been described: rearrangement of the EWS gene on chromosome 22q12 with an ETS gene family member. These translocations define the Ewing’s sarcoma family of tumors (ESFT) and provide a valuable tool for their accurate and unequivocal diagnosis. They also represent ideal targets for the development of tumor-specific therapeutics. Although secondary abnormalities occur in over 80% of primary ESFT the clinical utility of these is currently unclear. However, abnormalities in genes that regulate the G1/S checkpoint are frequently described and may be important in predicting outcome and response. Increased understanding of the molecular events that arise in ESFT and their role in the development and maintenance of the malignant phenotype will inform the improved stratification of patients for therapy and identify targets and pathways for the design of more effective cancer therapeutics.

Minimal disease monitoring by QRT–PCR: guidelines for identification and systematic validation of molecular markers prior to evaluation in prospective clinical trials

Real-time RT-PCR (QRT-PCR) is a sensitive method for the detection of minimal disease (MD) and may improve monitoring of disease status and stratification of patients for therapy. Where tumour-specific mRNAs have not been identified, the selection of which target(s) is(are) optimal for the detection of MD remains a challenge. This reflects the heterogeneity of tumour cells, the stability of mRNAs and low-level of transcription in cells of the normal haemopoietic compartments. The aim of this study was to establish for the first time guidelines for the systematic prioritization of potential markers of MD detected by QRT-PCR prior to evaluation in multicentre prospective clinical outcome studies. We combined microarray analysis, ESTs gene expression profiles, improved probe-sets sequence annotation, and previously described standard operating procedures for QRT-PCR analysis to identify and prioritize potential markers of MD. Using this methodology, we identified 49 potential markers of MD in neuroblastoma (NB), of which 11 were associated with neuronal function. We found that, in addition to TH, Phox2B and DCX mRNA may be useful targets for the detection of MD in children with NB. This same strategy could be exploited to select MD markers of other solid tumours from the large number of potential targets identified by microarray gene expression profiles.

Five-year follow-up in patients after therapy stratification based on intracoronary pressure measurement

Invasive coronary angiography (ICA) alone fails to accurately select patients with intermediate stenoses who should be treated by percutaneous coronary intervention (PCI). Previous studies have demonstrated the usefulness of fractional flow reserve (FFR) for identifying patients in whom deferring an intended PCI would be more beneficial than performing the planned procedure. The long-term safety of FFR-based therapy stratification, however, remains unknown. Therefore, the aim of this study was to retrospectively evaluate the long-term safety of an FFR-based therapy stratification in patients with intermediate coronary lesions detected by ICA. We included 56 patients presenting with a 50% to 75% angiographic stenosis by visual assessment on ICA, with negative, inconclusive, or no stress test, and in whom the intended PCI was deferred based on the result of the FFR measurement (ie, FFR > or = 0.75). The occurrence of major adverse cardiac events during a 5-year follow-up period was recorded. A complete 5-year follow-up was available in all patients. Mean follow-up was 1868 +/- 380 days. During follow-up, 16 events (1 cardiac death, 4 noncardiac deaths, 11 revascularization procedures) occurred. The combined rate of cardiac death and nonfatal myocardial infarction was 1.8% over 5 years. Nine PCI procedures (5 target vessel, 4 nontarget vessel) were performed during follow-up based on objective signs of ischemia. The angina status was not different between inclusion and the 5-year follow-up. Deferring PCI in patients without critical reduction in FFR may be a safe option during long-term follow-up.

Long-term safety of therapy stratification in patients with intermediate coronary lesions based on intracoronary pressure measurements

Long-term safety of therapy stratification in patients with intermediate coronary lesions based on intracoronary pressure measurements