Markers For Prediction Of Outcome Research Articles

Ratio of intratumoral macrophage phenotypes is a prognostic factor in epithelioid malignant pleural mesothelioma.

HypothesisThe tumor micro-environment and especially the different macrophage phenotypes appear to be of great influence on the behavior of multiple tumor types. M1 skewed macrophages possess anti-tumoral capacities, while the M2 polarized macrophages have pro-tumoral capacities. We analyzed if the macrophage count and the M2 to total macrophage ratio is a discriminative marker for outcome after surgery in malignant pleural mesothelioma (MPM) and studied the prognostic value of these immunological cells.Methods8 MPM patients who received induction chemotherapy and surgical treatment were matched on age, sex, tumor histology, TNM stage and EORTC score with 8 patients who received chemotherapy only. CD8 positive T-cells and the total macrophage count, using the CD68 pan-macrophage marker, and CD163 positive M2 macrophage count were determined in tumor specimens prior to treatment.ResultsThe number of CD68 and CD163 cells was comparable between the surgery and the non-surgery group, and was not related to overall survival (OS) in both the surgery and non-surgery group. However, the CD163/CD68 ratio did correlate with OS in both in the total patient group (Pearson r −0.72, p<0.05). No correlation between the number of CD8 cells and prognosis was found.ConclusionsThe total number of macrophages in tumor tissue did not correlate with OS in both groups, however, the CD163/CD68 ratio correlates with OS in the total patient group. Our data revealed that the CD163/CD68 ratio is a potential prognostic marker in epithelioid mesothelioma patients independent of treatment but cannot be used as a predictive marker for outcome after surgery.

P04.11 * GENETIC POLYMORPHISMS OF EGF 5'-UTR IN PATIENTS WITH GLIOMA: A POSSIBLE PREDICTIVE MARKER OF OUTCOME

BACKGROUND: Epidermal growth factor (EGF) plays an important role in carcinogenesis. An adenine (A) to guanine (G) single nucleotide polymorphism at position 61 in the 5'-untranslated region (5'-UTR) of the EGF gene has been found to be associated with levels of EGF production and contribute to the risk of glioma. However, published data are contradictory. EGF +61G/A polymorphism may contribute to the risk of glioma in different ethnic group. Patients with glioma and GG genotype have been reported to have a risk of poorer outcome than patients with AA genotype. Purpose of this study is to investigate the potential role of this polymorphism in cancer progression and its role as predictive marker of outcome in glioma caucasian patients. METHODS: The significant SNP rs4444903, EGF 61A/G, was analyzed in glioma patients and was determined by means of Polymerase Chain Reaction and Direct Sequencing method from blood samples. Association of this genetic polymorphism with clinical and pathological data of patients was evaluated. RESULTS: We investigated EGF +61G/A polymorphism in 28 glioma patients. EGF +61G allele has been found in 68% of glioma patients (22% G/G genotype and 46% A/G genotype). In astrocytomas, EGF +61G allele represents a 83% frequency; in glioblastomas and in oligodendrogliomas, EGF +61G allele frequency represents respectively 73% and 54%. In WHO IV gliomas, the EGF +61G allele represents a 72% frequency, (27% G/G and 45% A/G ), in WHO III gliomas a 81% frequency (54% G/G and 27% A/G ) and in WHO II gliomas a 33% frequency (80% A/G ). Median PFS of glioblastoma patients was 9 months. 79% of glioblastoma patients with a relapsing disease showed the G/G and A/G genotype. No difference was detected in the others histotypes. CONCLUSIONS: Our data confirm previous studies which reported G allele as a risk factor for glioma in Caucasian. G/A and G/G genotypes seem to be more rappresentative in high grade gliomas . Despite limited number of patients, our study supports the predictive role of EGF 61 A/G polymorphism in GBM. Additional large studies are warranted to confirm the role of EGF polymorphism as indipendent prognostic factor in glioma.

Acquired hypothyroidism as a predictive marker of outcome in patients with metastatic renal cell carcinoma (mRCC) treated with tyrosine-kinase inhibitors (TKIs): A literature-based meta-analysis.

Acquired hypothyroidism as a predictive marker of outcome in patients with metastatic renal cell carcinoma (mRCC) treated with tyrosine-kinase inhibitors (TKIs): A literature-based meta-analysis.

The active form of MMP-3 is a marker of synovial inflammation and cartilage turnover in inflammatory joint diseases

BackgroundMatrix metalloproteinase-3 (MMP-3) plays an important role in the pathology of rheumatoid arthritis (RA) and ankylosing spondylitis (AS). Measurement of active MMP-3 in clinical samples could provide information about progression of rheumatoid diseases, and potentially response to treatment. Hence, we aimed to develop a sensitive assay specifically measuring the active form of MMP-3 (act-MMP-3) both in ex vivo models and in human sera.MethodsA monoclonal antibody against the first 6 amino acids of act-MMP-3 was developed, and the specificity was carefully tested by comparing total and active MMP-3. A technically robust act-MMP-3 ELISA was produced. For biological validation, human synovial membrane and human cartilage explant (HEX) culture models were measured and compared by ELISA and immunoblots. For clinical relevance, the serum levels of act-MMP-3 in AS and RA patients before and after anti-TNF-α treatment were evaluated.ResultsA highly specific and technically robust ELISA detecting act-MMP-3 in serum was developed. The lower limit of detection was 33.7 pg/mL. The dilution and spiking recovery of human serum was within 100 ± 20%. The average intra- and inter-assay variations were 3.1% and 13.5% respectively.High levels of act-MMP-3 expression were observed in human synovial membrane culture and oncostatin M and TNF-α stimulated human cartilage. In a cross-sectional study of both AS and RA patients, serum act-MMP-3 level was correlated with C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR). In addition, in patients receiving anti-TNF-α treatment, the serum level of act-MMP-3 was significantly reduced compared to baseline level reflecting the anti-inflammatory effects of the treatment.ConclusionWe have successfully developed an assay measuring act-MMP-3 in human serum showing correlation to inflammatory markers. Further studies are required to clarify, whether act-MMP-3 can serve as a predictive marker for outcome in chronic rheumatoid disorders.

Metabolic tumour burden assessed by 18F-FDG PET/CT associated with serum CA19-9 predicts pancreatic cancer outcome after resection

Tumour burden is one of the most important prognosticators for pancreatic ductal adenocarcinoma (PDAC). The aim of this study was to investigate the predictive significance of metabolic tumour burden measured by (18)F-FDG PET/CT in patients with resectable PDAC. Included in the study were 122 PDAC patients who received preoperative (18)F-FDG PET/CT examination and radical pancreatectomy. Metabolic tumour burden in terms of metabolic tumour volume (MTV) and total lesion glycolysis (TLG), pathological tumour burden (tumour size), serum tumour burden (baseline serum CA19-9 level), and metabolic activity (maximum standard uptake value, SUVmax) were determined, and compared for their performance in predicting overall survival (OS) and recurrence-free survival (RFS). MTV and TLG were significantly associated with baseline serum CA19-9 level (P = 0.001 for MTV, P < 0.001 for TLG) and tumour size (P < 0.001 for MTV, P = 0.001 for TLG). Multivariate analysis showed that MTV, TLG and baseline serum CA19-9 level as either categorical or continuous variables, but not tumour size or SUVmax, were independent risk predictors for both OS and RFS. Time-dependent receiving operating characteristics analysis further indicated that better predictive performances for OS and RFS were achieved by MTV and TLG compared to baseline serum CA19-9 level, SUVmax and tumour size (P < 0.001 for all). MTV and TLG showed strong consistency with baseline serum CA19-9 level in better predicting OS and RFS, and might serve as surrogate markers for prediction of outcome in patients with resectable PDAC.

Acquired hypothyroidism as a predictive marker of outcome in patients with metastatic renal cell carcinoma (mRCC) treated with tyrosine-kinase inhibitors (TKIs): A literature-based meta-analysis.

500 Background: The development of hypothyroidism in patients with mRCC during treatment with the tyrosine kinase inhibitors (TKIs) sunitinib and sorafenib is a well-established side effect. The potential role of hypothyroidism as predictive marker of outcome has been studied with conflicting results. The aim of the present meta-analysis was to assess the predictive value of hypothyroidism for both progression free (PFS) and overall survival (OS) in patients with mRCC treated with TKI. Methods: We searched PubMed as well as the electronic abstract databases of the major international congresses’ proceedings to identify all eligible studies which reported a correlation of the development of hypothyroidism during treatment with TKI and outcome in patients with mRCC. Hazard ratios (HRs) with 95% confidence intervals (CIs) for PFS and OS were obtained from these publications and pooled in a meta-analysis by using random-effects or fixed-effects models based on the heterogeneity of the included studies. Results: Eleven studies with a total of 500 patients fulfilled the inclusion criteria. We found no statistical significant difference in PFS between patients who developed hypothyroidism and patients without hypothyroidism during sunitinib therapy (HR: 0.82, 95% CI: 0.59-1.13, p-value = 0.22) (6 studies; 250 patients). For studies that included patients treated with either sunitinib or sorafenib, the development of hypothyroidism was found to be predictive for longer progression free survival (HR: 0.59, 95% CI: 0.42-0.84, p-value = 0.003) (3 studies; 205 patients). The HR for OS was 0.52 (95% CI: 0.31-0.87, p-value = 0.01) for patients who developed hypothyroidism during sunitinib therapy compared to patients without hypothyroidism (4 studies; 147 patients). Conclusions: The development of hypothyroidism during TKIs therapy is not clearly shown to be predictive in patients with mRCC. The observed advantage in overall survival for patients with treatment related hypothyroidism should be interpreted with caution due to the potential imbalance of treatments received after first line treatment in each group.

Genetic polymorphisms of EGF 5'-UTR in patients with glioma: A possible predictive marker of outcome.

e13009 Background: Epidermal growth factor (EGF) plays an important role in carcinogenesis. An adenine (A) to guanine (G) single nucleotide polymorphism at position 61 in the 5'-untranslated region (5'-UTR) of the EGF gene has been found to be associated with levels of EGF production and contribute to the risk of glioma. However, published data are contradictory. EGF +61G/A polymorphism may contribute to the risk of glioma in different ethnic group. Patients with glioma and GG genotype have been reported to have a risk of poorer otucome than patients with AA genotype. Purpose of this study is to investigate the potential role of this polymorphism in cancer progression and its role as predictive marker of outcome in glioma caucasian patients. Methods: EGF 61A/G polymorphism (rs4444903) was analyzed in glioma patients and was determined by means of Polymerase Chain Reaction and Direct Sequencing method (GenBank reference sequence-accession no. AC005509) from blood samples. Association of this genetic polymorphism with clinical and pathological data of patients was evaluated. Results: We investigated EGF +61G/A polymorphism in 24 glioma patients . EGF +61G allele has been found in 67% of glioma patients (25% G/G genotype and 42% A/G genotype). In astrocytomas, EGF +61G allele represents a 83% frequency; in glioblastomas and in oligodendrogliomas, EGF +61G allele frequency represents respectively 71% and 50%. In WHO IV gliomas, the EGF +61G allele represents a 63% frequency, (27% G/G and 36% A/G ), in WHO III gliomas a 77% frequency (33% G/G and 44% A/G ) and in WHO II gliomas a 50% frequency (100% A/G ). Median PFS of glioblastoma patients was 9 months. 83% of glioblastoma patients with a relapsing disease showed the G/G and A/G genotype. No difference was detected in the others histotypes. Conclusions: Our data conferm previous studies which reported G allele as a risk factor for glioma in Caucasian. G/A and G/G genotypes seem to be more rappresentative in high grade gliomas . Despite limited number of patients, our study supports the predictive role of EGF 61 A/G polymorphism in GBM. Additional large studies are warranted to confirm the role of EGF polymorphism as indipendent prognostic factor in glioma.

MR Imaging Assessment of Tumor Perfusion and 3D Segmented Volume at Baseline, during Treatment, and at Tumor Progression in Children with Newly Diagnosed Diffuse Intrinsic Pontine Glioma

DIPG is among the most devastating brain tumors in children, necessitating the development of novel treatment strategies and advanced imaging markers such as perfusion to adequately monitor clinical trials. This study investigated tumor perfusion and 3D segmented tumor volume as predictive markers for outcome in children with newly diagnosed DIPG. Imaging data were assessed at baseline, during, and after RT, and every other month thereafter until tumor progression for 35 patients (ages 2-16 years) with newly diagnosed DIPG enrolled in the phase I clinical study, NCT00472017. Patients were treated with conformal RT and vandetanib, a vascular endothelial growth factor receptor 2 inhibitor. Tumor perfusion increased and tumor volume decreased during combined RT and vandetanib therapy. These changes slowly diminished in follow-up scans until tumor progression. However, increased tumor perfusion and decreased tumor volume during combined therapy were associated with longer PFS. Apart from a longer OS for patients who showed elevated tumor perfusion after RT, there was no association for tumor volume and other perfusion variables with OS. Our results suggest that tumor perfusion may be a useful predictive marker for the assessment of treatment response and tumor progression in children with DIPG treated with both RT and vandetanib. The assessment of tumor perfusion yields valuable information about tumor microvascular status and its response to therapy, which may help better understand the biology of DIPGs and monitor novel treatment strategies in future clinical trials.

Impact of KRAS mutational status on clinical outcomes in patients receiving capecitabine based chemotherapy.

436 Background: Mutations affecting the KRAS gene are established predictive markers of outcome with anti–epithelial growth factor receptor antibodies in metastatic colorectal cancer (mCRC). The relevance of these markers for chemotherapy has not been established. This analysis was performed to assess the predictive impact of KRAS mutation status in patients receiving chemotherapy. Methods: KRAS mutational status was available for 223 patients treated for mCRC. Predictive analysis of mutational status by type of fluoropyrmidine the 1st-line regimen contained (either capecitabine [C] based chemotherapy or infusional 5Fluorouracil [I-5Fu]) for clinical outcomes: progression-free survival (PFS), time to chemotherapy resistance (TTCR) and overall survival (OS). Results: KRAS mutations were observed in 43.5% of the patients. 165 patients received I-5Fu, 44 patients received C. KRAS mutation status (wild type [WT] v mutated [MT]) had no prognostic impact for OS (hazard ratio [HR], 0.81; CI, 0.69 to 1.1 p=0.17) for PFS (hazard ratio [HR], 0.87; CI, 0.66 to 1.14 p=0.3) and TTCR (hazard ratio [HR], 0.85; CI, 0.65to 1.12 p=0.26). C based 1st-line chemotherapy vs. I- 5FU based was predictive of PFS (hazard ratio [HR], 0.52; CI, 0.37 to 0.74 p=0.0003) and TTCR (hazard ratio [HR], 0.54; CI, 0.38 to 0.75p=0.0005) and not of OS (hazard ratio [HR], 0.74; CI, 0.52 to 1.1 p=0.1). KRAS mutational status had predictive impact in patients receiving C based 1st-line chemotherapy on OS (hazard ratio [HR], 0.47; CI, 0.23 to 0.948 p&lt;0.0001) TTCR (hazard ratio [HR], 0.49; CI, 0.25 to 0.97 p=0.0398) and was not predictive of PFS (hazard ratio [HR], 0.78; CI, 0.4 to 1.53 p=0.47) Conclusions: KRAS gene mutation status was predicitve for OS and for TTCR in patients who received C based 1st-line chemotherapy.

Reduced TET2 expression in Patients with Myelodysplastic Syndromes and Acute Myeloid Leukemia.

AbstractAbstract 2822 Introduction:Acquired mutations in TET2 and DNMT3A have been found in myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML), and may predict a worse survival in these diseases. TET2 mutations are considered to be a loss-of-function mutation and results in decreased 5-hydroxymethylcitosine (5-hmc) levels. In normal CD34+ cells, TET2 silencing skews progenitor differentiation towards the granulomonocytic lineage at the expense of lymphoid and erythroid lineages. Dnmt3a participates in the epigenetic silencing of hematopoietic stem cell regulatory genes, enabling efficient differentiation. Here, we attempted to evaluate the expression of TET2 and DNMT3A in total bone marrow cells from normal donors, patients with MDS and AML, and in CD34+ cells from MDS and normal controls during erythroid differentiation. Materials and Methods:The study included normal donors (n = 21), patients with MDS (n = 43) and AML (n = 42) at diagnosis. All normal donors and patients provided informed written consent and the study was approved by the ethics committee of the Institution. MDS patients were stratified into low and high-risk according to WHO classification (RCUD/RCMD/RARS=31 and RAEB1/RAEB2=12). TET2 and DNMT3A mRNA expression was assessed by quantitative PCR. CD34+ cells from normal donors (n = 9) and low-risk MDS patients (n = 7) were submitted to erythroid differentiation. Cells were collected and submitted to immunophenotyping for GPA and CD71 (days 6 and 12) and q-PCR for TET2 and DNMT3A expression (days 6, 8 and 12). Results of gene expression in normal donors and patients are presented as median, minimum-maximum, and were compared using Mann-Whitney test. Student t test was used for comparison of gene expression during CD34+ erythroid diferentiation. Overall survival was defined from the time of sampling to the date of death or last seen. Univariate analysis for overall survival was conducted with the Cox proportional hazards model. Results:TET2 expression was significantly reduced in both AML (0.62; 0.01–32.69) and MDS (1.46; 0.17–21.30) compared to normal donors (2.72; 0.43–31.49); P<0.0001 and P=0.01, respectively. TET2 expression was also significantly reduced in AML compared to MDS (P=0.0007). MDS patients were stratified into low and high-risk disease, and we still observed a significant reduction in TET2 expression in high-risk (0.73, 0.17–7.25) when compared to low-risk (1.58; 0.48–21.30; P=0.02) patients, but no difference was noted between normal donors vs. low-risk MDS, and high-risk MDS vs. AML. In MDS cohort, the median overall survival was 14 months (range 1–83), increased TET2 expression was associated with a longer survival (HR, 0.44; 95% CI, 0.21–0.91, P=0.03), and, as expected, WHO high-risk disease was associated with a shorter survival (HR, 10.16; 95% CI, 3.06–33.72, P<0.001), even though the confidence interval (CI) was large. TET2 expression did not impact survival in our cohort of AML patients. The erythroid differentiation was effective in cells from normal donors and MDS patients, as demonstrated by the flow cytometry analyses of GPA and CD71. TET2 expression was significantly increased on day 12 of erythroid differentiation, P<0.05. On the other hand, DNMT3A expression was similar between normal donors (0.74; 0.22–1.53), MDS (0.78; 0.26–3.46) and AML (0.95, 0.15–6.46), and during erythroid differentiation, with no impact on survival. Conclusion:These data suggest that decreased TET2 expression may participate in leukemogenesis, and supports the participation of TET2 in the erythroid differentiation of MDS. DNMT3A was not differentially expressed in AML and MDS, indicating that the presence of mutations in this gene may be the predominant mechanism of changes in protein function. We thus suggest that decreased TET2 expression may explain the reduced levels of 5-hmc found in TET2 wild type patients, and may become a predictive marker for outcomes in MDS and other myeloid diseases. Further studies would be necessary to better elucidate the clinical relevance and biologic significance of our findings, and whether the decreased TET2 expression results in hypermethylation in these diseases. Disclosures:Maciejewski:NIH: Research Funding; Aplastic Anemia&MDS International Foundation: Research Funding.

Impact of the adaptor protein GIPC1/Synectin on radioresistance and survival after irradiation of prostate cancer

Studies have shown that GIPC1/Synectin is an essential adaptor protein of receptors that play an important role in cancer progression and therapy resistance. This is the first study to explore the role of GIPC1/Synectin in radioresistance of prostate cancer and as a possible predictive marker for outcome of primary radiation therapy. The effect of RNA interference-mediated GIPC1/Synectin depletion on clonogenic cell survival after irradiation with 0, 2, 4, or 6 Gy was assayed in two different GIPC1/Synectin-expressing human prostate cancer cell lines. The clinical outcome data of 358 men who underwent radiotherapy of prostate cancer with a curative intention were analyzed retrospectively. Uni- and multivariate analysis was performed of prostate-specific antigen recurrence-free survival and overall survival in correlation with protein expression in pretreatment biopsy specimens. Protein expression was evaluated by standard immunohistochemistry methods. In cell culture experiments, no change was detected in radiosensitivity after depletion of GIPC1/Synectin in GIPC1/Synectin-expressing prostate cancer cell lines. Furthermore, there was no correlation between GIPC1/Synectin expression in human pretreatment biopsy samples and overall or biochemical recurrence-free survival after radiotherapy in a retrospective analysis of the study cohort. Our results do not show a predictive or prognostic function of GIPC1/Synectin expression for the outcome of radiotherapy in prostate cancer. Furthermore, our in vitro results do not support a role of GIPC1 in the cellular radiation response. However, the role of GIPC1 in the progression of prostate cancer and its precursors should be subject to further research.

P-0270 Impact of Kras and Braf Gene Mutation Status on Outcomes of Chemotherapy in Combination with Bevacizumab in Metastatic Colorectal Cancer

ABSTRACT Introduction Mutations affecting KRAS are established predictive markers of outcome with anti-epithelial growth factor receptor antibodies, in metastatic colorectal cancer (mCRC). Several ongoing trials are investigated the predictive role of these and novel targeted agents in patients (pts) with BRAF mutation. The relevance of these biomarkers for anti-vascular endothelial growth factor therapy is still unclear. We aimed to evaluate the possible role of KRAS and BRAF status on therapeutic outcome and to assess the clinical response in different mCRC subgroup. Methods This was a retrospective study enrolling 67 pts treated with first-line FOLFIRI or FOLFOX chemotherapy plus bevacizumab. Tissue samples were analyzed for DNA sequencing to identify KRAS and BRAF mutations. Before therapy and after 3 months of treatment pts were investigated with CT scan. Mutation status was correlated with efficacy outcomes such as objective response rate (RR), mean progression-free survival (mPFS) and mean overall survival (mOS). Results KRAS and BRAF genes mutations were observed in 34.3% and 13.4% respectively. RR was available in all pts while mPFS and mOS in 63 pts. RR obtained in KRAS wild-type (wt) group was 54.5% vs 11.5% in KRAS mutated (mut) group, mPFS was 11.7 months vs 9.9 months respectively and mOS was 27.6 months vs 27.3 months. In BRAF wt group RR was 46.5% vs 22.2% in BRAF mut group. mPFS obtained in BRAF wt group was 11.4 months vs 7 months in BRAF mut group while mOS was 28.2 months vs 19.6 months. By analyzing RR, mPFS and mOS in the following subpopulations (40 pts KRAS wt/BRAF wt; 4 pts KRAS wt/BRAF mut; 18 pts KRAS mut/BRAF wt; 5 pts KRAS mut/BRAF mut) gave these results: in first subpopulation RR was 57.5%, mPFS 19.3 months and mOS 28.7 months; in second group RR 25%, mPFS 8 months, mOS 20.5 months; in KRAS mut/BRAF wt RR 22.2%, mPFS 10 months, mOS 27.5 months, finally in KRAS mut/BRAF mut RR 20%, mPFS 9.6 months, mOS 33 months. Also we evaluated the difference in RR, mPFS and mOS between KRAS wt/BRAF wt group and pts with a single or both oncogenes mutation. RR in pts with KRAS and BRAF wt was 57.5% vs 22.2% in pts with KRAS and/or BRAF mut; mPFS in the two groups was 19.3 months vs 7.9 months respectively and mOS was 28.7 months vs 21.9 months. Conclusion KRAS and BRAF oncogenes mutations were not clearly predictive of anti-vascular endothelial growth factor therapy outcome. Clinical response, mPFS and mOS in KRAS and BRAF groups were different with advantage in KRAS and BRAF wt groups. Also RR, mPFS and mOS tended to be higher in KRAS wt/BRAF wt group, while in pts with only one or both oncogenes mutation these values were lower.

Abstract 3455: p53 status determines the radio-sensitizing effect of an Akt kinase inhibitor in malignant gliomas

Abstract Purpose: It has been reported that PI3K-Akt pathway, which plays a role in cell survival and apoptosis, is deregulated in many cancer types. In gliomas, amplification and constitutive activation of Akt, which can result from amplifications in TKIs including EGFR, IGFR1α, PDGFRα, etc. and mutations in PI3K, PTEN, etc. are thought to play a role in radiation resistance. Some have tried to modulate the PI3K/Akt pathway by targeting molecules upstream of Akt such as Ras and PI3K, while others have used inhibitors targeting molecules downstream of Akt, such as mTOR. These inhibitors have been shown to reduce cancer cell growth in vitro and in vivo, but have had limited success in clinical settings as single agents. In our study, we have investigated the use of a phosphatidylinositol ether lipid analogue (PIA) that directly inhibits Akt activation as a radiosensitizer for GBM. Materials and Methods: We utilized multiple established (U87, LN18, LN229) and primary (VC3, MGH8, OSU61) cell lines with varying molecular and genetic signatures to determine radiosensitizing effects of PIA. We performed Annexin V, clonogenic survival, and COMET assays on glioma cells treated with PIA, radiation (RT), and a combination of PIA and RT. Western blot was performed using these cell lines to examine the changes in expression of PI3K-Akt pathway, apoptotic, and pro-survival proteins following treatment. Furthermore, we sequenced tumor suppressor genes p53 and PTEN in all cell lines used in this study. Gene silencing of p53 was done in U87 cells via a lentiviral transfection of shRNA. Results and Conclusions: PIA decreases the activation of Akt to a considerable extent (∼80%), induces apoptosis, and reduces clonogenicity of all cell lines tested. A subset of cell lines did show enhanced radiosensitization when PIA is combined with RT. Further detailed investigation of differential radiosensitizing effects led us to discover the link between p53 status, Akt kinase inhibition, and radiosensitization. GBMs with intact p53 had no radiosensitizing effect after Akt kinase inhibition, whereas radiosensitization was observed if either p53 is null or mutated. This observation was further validated using isogenic U87 p53 knockdown cells. Surprisingly, MGMT status, a predictive marker of outcome in GBM, did not determine radiosensitizing effects of PIA in the GBM cells tested. Through this preclinical study, we have identified which subset of GBM patients will derive benefit from Akt kinase inhibition and the investigations conducted justify future clinical trials and lays ground work for phase I study. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3455. doi:1538-7445.AM2012-3455

Overexpression of wild-type or mutants forms of CEBPA alter normal human hematopoiesis

C/EBPα (CEBPA) is mutated in approximately 8 % of AML in both familial and sporadic AML and, with FLT3 and NPM1, has received most attention as a predictive marker of outcome in patients with normal karyotype disease. Mutations clustering to either the N- or C-terminal (N-and C-ter) portions of the protein have different consequences on the protein function. In familial cases the N-ter form is inherited with patients exhibiting long latency period before the onset of overt disease, typically with the acquisition of a C-ter mutation. Despite the essential insights murine models provide the functional consequences of wild-type C/EBPα in human hematopoiesis and how different mutations are involved in AML development have received less attention. Our data underline the critical role of C/EBPα in human hematopoiesis and demonstrate that C/EBPα mutations (alone or in combination) are insufficient to convert normal human hematopoietic stem/progenitors (HSC/HPCs) into leukemic initiating cells, although individually each altered normal hematopoiesis. It provides the first insight into the effects of N- and C-terminal mutations acting alone and to the combined effects of N/C double mutants. Our results mimicked closely what happens in CEBPA mutated patients.

Pretransplant serum ferritin level may be a predictive marker for outcomes in patients having undergone allogeneic hematopoietic stem cell transplantation

Iron overload increases the risk of infections, veno-occlusive disease and hepatic dysfunction in post-transplant period. Our objective was to investigate the association of pre-transplant ferritin levels with complications and survival after allogeneic hematopoietic stem cell transplantation (alloHSCT).We retrospectively analysed 84 patients' data who had undergone allogeneic HSCT into two groups: patients with a serum ferritin level ≥ 1000 ng/ml, and patients with <1000 ng/ml at the time of HSCT.Cox-regression analysis showed that pre-transplant serum ferritin levels were significantly higher in patients who had at least one infectious event compared with those who had no any infectious event in the post-transplant 100 days (p<0.023). Overall survival (OS) and disease-free survival (DFS) rates were significantly higher in patients with a time-to-tx interval 12 months (p=0.002 and p=0.008 respectively). A higher risk of death was observed in high-ferritin group (hazard ratio=2.27, CI:1.01-5.09, p=0.023 for OS and hazard ratio=2.49, CI:1.12-5.53 p=0.039 for DFS). No significant effect on OS and DFS among groups was observed for variables conditioning regimen, gender and diagnosis. Acute GVHD was more common in patients with a ferritin level ≥ 1000 ng /mL, but this was not statistically significant (p>0.05). There was no statistical significance in both groups (ferritin ≥ 1000 ng /mL and ferritin <1000 ng/mL) for relapse rates (p>0.05). Platelet and neutrophil engaftment day was not found statistically significant compared with both groups (p=0.273 and p=0.882, respectively). Pre-transplant ferritin levels may predict poor outcomes in patients who had undergone allogeneic hematopoietic stem cell transplantation.

Low ANXA10 expression is associated with disease aggressiveness in bladder cancer

Background:Markers for outcome prediction in bladder cancer are urgently needed. We have previously identified a molecular signature for predicting progression in non-muscle-invasive bladder cancer. ANXA10 was one of the markers included in the signature and we now validated the prognostic relevance of ANXA10 at the protein level.Methods:We investigated ANXA10 expression by immunohistochemistry using a tissue microarray with 249 Ta and T1 urothelial carcinomas. The expression of ANXA10 was also investigated in an additional set of 97 more advanced tumours. The functional role of ANXA10 in cell lines was investigated by siRNA-mediated ANXA10 knockdown using wound-healing assays, proliferation assays, and ingenuity pathway analysis.Results:Low expression of ANXA10 correlated with shorter progression-free survival in patients with stage Ta and T1 tumours (P<0.00001). Furthermore, patients with more advanced tumours and low ANXA10 expression had an unfavourable prognosis (P<0.00001). We found that ANXA10 siRNA transfected cells grew significantly faster compared with control siRNA transfected cells. Furthermore, a wound-healing assay showed that ANXA10 siRNA transfected cells spread along wound edges faster than control transfected cells.Conclusion:We conclude that ANXA10 may be a clinical relevant marker for predicting outcome in both early and advanced stages of bladder cancer.

Can Response to Induction Chemotherapy Be a Predictive Marker for Ultimate Outcome in Hypopharyngeal Cancer?

Induction chemotherapy (ICT) may reduce rates of distant metastases and enhance organ preservation and survival rates in patients with hypopharyngeal cancer. The authors compared survival data in patients who underwent chemoradiotherapy or surgery after ICT and investigated whether response to ICT is a predictive marker for outcome in patients with hypopharyngeal cancer. Historical cohort study. Tertiary-care hospital. The authors enrolled 97 patients (89 men, 8 women; mean age 61.2 years; range, 29-80 years) with previously untreated hypopharyngeal cancer who underwent ICT between January 1997 and December 2006 at Asan Medical Center. Disease-free survival (DFS), overall survival (OS), and laryngectomy-free survival (LFS) were analyzed. At a mean follow-up of 38.0 months, the 3-year DFS and OS for all patients were 48.3% and 49.2%, respectively. The 3-year LFS rate of patients who underwent nonsurgical therapy (n = 85) after ICT was 48.0%. Only response to ICT was associated with DFS (P = .047), OS (P = .003), and LFS (P = .009) in multivariate analysis. When the authors compared survival data in patients who underwent surgical and nonsurgical treatments after ICT, they found that there was no statistical difference in terms of the 3-year DFS in partial response (42.9% vs 50.5%, P = .77) and nonresponse groups (50.0% vs 0%, P = .43) between the 2 treatment types. ICT permits assessment of tumor responsiveness and alters subsequent therapy accordingly. Response to ICT may be useful in the prediction of ultimate outcomes and organ conservation in patients with hypopharyngeal cancer.

Response to the comment of Brusselaers et al. questioning the clinical benefit of circulating free DNA/neutrophil extracellular traps (cfDNA/NETs) as a laboratory marker for outcome prediction after severe burn injuries.

Response to the comment of Brusselaers et al. questioning the clinical benefit of circulating free DNA/neutrophil extracellular traps (cfDNA/NETs) as a laboratory marker for outcome prediction after severe burn injuries.

Use of KRAS mutation status to predict clinical outcomes in patients with metastatic colorectal cancer (mCRC) treated with the VEGF signaling inhibitor cediranib.

3613^ Background: The value of KRAS mutations as predictive markers of clinical benefit of anti-EGFR antibody therapy in patients with mCRC has been shown in randomized trials. Less is known of the influence of KRASmutations in patients with mCRC treated with VEGF signaling inhibitors and literature on the prognostic value of KRAS mutations in this setting is inconsistent. The Phase III HORIZON II (HZII) study evaluated the efficacy of cediranib 20 mg + FOLFOX (FF) or XELOX (X) vs placebo + FF or X in patients with mCRC; cediranib 20 mg met the co-primary endpoint of progression-free survival (PFS; HR=0.84) but not overall survival (OS; HR=0.94). A retrospective analysis of KRAS mutation status and correlation with PFS and OS in patients was performed to evaluate its prognostic and predictive value. Methods: Analysis of KRAS codon 12 and 13 mutations was performed at a central laboratory using a commercially available, allele-specific, ARMS-based polymerase chain reaction assay. Results: KRAS mutation status was determined in 489/860 patients (cediranib 20 mg arm, n=285/502; placebo arm, n=204/358); of these 489 patients, 42.3% had mutant KRAS and 57.7% were wild-type. Baseline characteristics were similar in the KRAS mutant, KRAS wild-type and KRAS status unknown groups. There was a trend towards improved PFS and OS in the KRAS wild-type subgroup compared with the KRAS mutant subgroup, which was independent of treatment arm (PFS: HR=0.85, 95% CI: 0.70,1.03, median 8.5 vs 8.3 months; OS: HR=0.71, 95% CI: 0.57,0.88, median 20.9 vs 16.9 months). There were no statistically significant differences in treatment effect between KRAS subgroups for the cediranib vs placebo arm (PFS: wild-type HR=0.78, mutant HR=0.82; OS: wild-type HR=0.92; mutant HR=1.01). Conclusions: This is the first reported analysis of KRAS mutation status in a randomized Phase III clinical study with a VEGFR-TKI. KRAS mutation status did not influence the PFS or OS treatment effect and therefore cannot be used as a predictive marker for outcome. These data show that KRAS codon 12 or 13 mutations have prognostic value in mCRC independent of treatment.

Gene polymorphisms in cyclophosphamide metabolism pathway,treatment-related toxicity, and disease-free survival in SWOG 8897 clinical trial for breast cancer.

There are no established genetic markers for prediction of outcomes after cyclophosphamide (CP)-containing adjuvant therapy for breast cancer. In an ancillary study to a SWOG (Southwest Oncology Group) trial (S8897), we investigated functional polymorphisms in 4 genes in CP pharmacokinetic pathways in relation to hematologic toxicity and disease-free survival (DFS). Germline DNA was available from 458 women who were at high risk of relapse and was randomized to CAF (CP, intravenous doxorubicin, and 5-fluorouracil) versus CMF (CP, intravenous methotrexate, and 5-fluorouracil) ± tamoxifen, and from 874 women who had a presumed favorable prognosis and did not receive adjuvant therapy. Odds ratios for grade 3 and 4 hematologic toxicity in the treated group and hazard ratios for DFS associated with selected functional polymorphisms in CYP2B6CYP3A4GSTA1 and GSTP1 were estimated by logistic regression and Cox proportional hazard regression. Compared with women with AA genotypes, those with at least 1 GSTP1 variant G allele had reduced risk of grade 3 and 4 neutropenia [odds ratios (OR) = 0.63, 95% CI = 0.41-0.97] and leucopenia (OR = 0.59, 95% CI = 0.39-0.89). No other associations between single nucleotide polymorphisms and toxicity or survival were found in the treated or untreated group. Known genetic variants in genes involved in CP pharmacokinetics may not have major effects on DFS in breast cancer patients. The lower risk of developing high-grade hematologic toxicity among women with variant GSTP1 alleles suggests that genetic markers in combination with clinical factors may be useful in defining a subgroup of women who are less susceptible to adverse hematologic toxicities with CP-containing therapies.