BACKGROUND

1. NRP1 Glut-1 VEGFA HIF1a HIF2a LDHA VEGFR1 VEGFR2 VEGFR3 Serum LDH -0.049 0.061 0.351 0.025 0.093 0.280 0.501 0.158 0.054 p-value 0.781 0.725 0.036 0.883 0.588 0.098 0.004 0.379 0.786 Glut-1 -0.144 1.000 0.447 0.005 -0.039 0.440 0.008 -0.248 -0.199 p-value 0.409 0.006 0.975 0.823 0.007 0.967 0.164 0.309 VEGFA -0.079 0.447 1.000 0.159 0.170 0.568 0.515 0.206 0.195 p-value 0.651 0.006 0.354 0.322 0.0001 0.003 0.251 0.320 HIF1a 0.414 0.005 0.159 1.000 0.571 0.411 0.168 0.216 -0.010 p-value 0.013 0.975 0.354 0.0001 0.013 0.366 0.227 0.959 LDHA -0.005 0.440 0.568 0.411 0.338 1.000 0.400 0.016 -0.120 p-value 0.976 0.007 0.0001 0.013 0.044 0.026 0.930 0.543 Abstract # Tumor VEGFA gene expression is associated with serum lactate dehydrogenase (LDH) levels and intratumoral mRNA expression of genes involved in glycolysis in patients with metastatic colorectal cancer (mCRC) 3530 Mizutomo Azuma1, Michael M. Shi2, Christian J. Jacques2, Carl Barrett2, Kathleen D. Danenberg3, Syma Iqbal1, Anthony El-Khoueiry1, Dongyun Yang4, Wu Zhang1 and Heinz-Josef Lenz1 1 Division of Medical Oncology, University of Southern California/Norris Comprehensive Cancer Center, Los Angeles, CA, USA, 2 Novartis Pharmaceuticals Corporation, East Hanover, NJ, USA3 Response Genetics Inc. Los Angeles, CA, USA, 4 Department of Preventive Medicine, University of Southern California/Norris Comprehensive Cancer Center, Los Angeles, CA, USA BACKGROUND RESULTS Table 2. Serum LDH showed a significant positive correlation with intratumoral gene expression of VEGFA (P=0.036, r=0.35) and VEGFR1 (p= 0.006, r=0.45). Intratumoral gene expression of LDHA showed a significant positive correlation with Glut-1 (P=0.007, r=0.44), VEGFA (P<0.001, r=0.57), HIF1a (P=0.013, r=0.41 ), HIF2a (P=0.044, r=0.34) and VEGFR1 (p=0.026, r=0.40).But weak positive correlation with serum LDH levels (P=0.098, r=0.28). (Table.1) It is well known that angiogenesis and glycolysis are regulated by hypoxic conditions. Recent clinical trials (CONFIRM1 and CONFIRM2) have shown that patients with mCRC with high serum LDH benefited from PTK787/ZK 222584 (PTK/ZK), which blocks all the known VEGF receptor (VEGFR) tyrosine kinase. We tested the hypothesis that patients with high serum LDH have increased intratumoral expression of genes involved with hypoxia [hypoxia inducible factor (HIF1a and 2a)], glycolysis [lactate dehydrogenase A (LDHA) and glucose transporter 1 (Glut-1)], and angiogenesis [vascular endothelial growth factor A (VEGFA), VEGFR1-3 and neuropilin 1 (NRP1)] in patients with mCRC. Gene expression levels relative to the internal reference geneβ-actin of the analyzed genes. Table 1. Spearman Rank Correlation Analysis of Associations Between Serum LDH levels and Gene Expression Values PATIENTS AND METHOD 78 formalin fixed paraffin embedded (FFPE) tumor samples were collected from 36 patients (20 males, 16 females) with a median age of 59 years (range 29-84) with metastatic CRC who underwent first line therapy (not from CONFIRM trials). Intratumoral mRNA expression were analyzed for genes involved with hypoxia (HIF1a and 2a), glycolysis (LDHA and Glut-1), and angiogenesis (VEGFA, VEGFR1-3 and NRP1). Tumor gene expression was correlated with serum LDH levels from the same group of patients. FFPE tissues were dissected using laser-captured microdissection and analyzed for gene expression using a quantitative real-time RT-PCR method. Gene expression values (relative mRNA levels) are expressed as ratios between the target gene and internal reference gene (b-actin). CONCLUSIONS Our results demonstrate that intratumoral gene expression of LDHA, HIF1a, HIF2a, Glut-1, NRP1, VEGFA and VEGFR1 are significantly correlated. Patients with high serum LDH have increased intratumoral gene expression of VEGFA and VEGFR1. These results support the hypothesis that serum LDH levels may serve as a surrogate marker for activation of the HIF related genes in the tumor. These observations may explain, at least partially, the clinical benefit of PTK/ZK in metastatic colorectal cancer patients with high serum LDH levels. Abbreviations: LDH; lactate dehydrogenase, HIF;hypoxia inducible factor, Glut-1; glucose transporter 1, VEGF(R); vascular endothelial growth factor (receptor), NRP; neuropilin