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363 Background: Body composition has emerged as a prognostic factor in cancer patients. We investigated whether sarcopenia at the diagnosis and progressive loss of skeletal muscle were associated with survival in pancreatic cancer (PC) patients receiving palliative chemotherapy. Methods: We retrospectively reviewed PC patients receiving palliative chemotherapy between 2003 and 2010. Skeletal muscle cross-sectional area at L3 was measured by computed tomography. Sarcopenia was defined using the previously published sex-specific cutoff points for Korean people. Loss of skeletal muscle was classified by sex-specific cutoffs from ROC curve. Results: Among 484 patients, 260 (53.7%) patients were more than sixty years old and 295 (61.0%) patients were male. Overall, 187 (38.6%) patients were sarcopenic at the diagnosis (male, 2/m2; female, 2/m2). Decreased skeletal muscle index (SMI) during the chemotherapy, which was defined as a reduction by more than 0.21 for male and by more than 2.19 for female, was observed in 198 (77.3%) male patients and 61 (38.1%) female patients. Decreased body mass index (BMI) by more than 1 was observed in 149 patients (37.3%) without difference between genders. Median overall survival (OS) of whole patients was 8.4 months [95%CI 7.6-9.2]. In the multivariate analysis, sarcopenia (p=0.001), decreased SMI (p=0.003), and decreased BMI (p=0.001) was significantly poor prognostic factors for OS. When we analyzed four groups by SMI and BMI changes (maintained SMI and BMI, maintained SMI and decreased BMI, decreased SMI and maintained BMI, decreased SMI and BMI), the groups with maintained SMI had longer survival than the groups with decreased SMI regardless of BMI changes. Median OS was 11.5 months with maintained SMI and 8.1 months with decreased SMI (HR 0.534 and 1, p=0.004) in decreased BMI groups and 9.9 months with maintained SMI and 8.6 months with decreased SMI (HR 1 and 1.490, p=0.002) in maintained BMI groups, respectively. The analyses separately done by gender showed similar results. Conclusions: Decrease of SMI during chemotherapy was a significantly poor prognostic factor for survival regardless of BMI changes.

Purpose: To investigate the efficacy of gemcitabine plus UFT (uracil-tegafur) combination chemotherapy as salvage treatment in patients with metastatic or relapsed colorectal cancer (MRCRC). Methods: This single-arm phase 2 study was conducted at 3 institutions in Korea. Patients with MRCRC refractory to fluoropyrimidine, oxaliplatin and irinotecan were enrolled. Gemcitabine 800 mg/m2 was administered intravenously on days 1, 8 and 15. UFT 200 mg/m2/day was taken orally in 3 divided doses on days 1-21. Cycles were repeated every 4 weeks and tumor evaluation was carried out every 8 weeks. The primary endpoint of this study was 8-week progression-free survival (PFS) rate. Results: Forty-one patients were enrolled. Fourteen patients received gemcitabine/UFT as a third-line treatment and 37 patients as a 4th-line or later-line therapy. Toxicities were easily manageable and non-hematologic toxicities of ≥ grade 3 were rare. The most common toxicity of ≥ grade 3 was neutropenia (20.0%). One patient showed partial response (response rate, 2.4%) and 14 (34.1%) showed stable disease. The 8-week PFS rate was 42.3%. The median PFS was 1.7 months [95% confidence interval (CI), 1.6-1.8 months] and the median OS was 9.2 months (95% CI, 5.8-12.6 months). Conclusions: Overall efficacy of gemcitabine/UFT in refractory MRCRC was unsatisfactory. However, we could found a minor proportion of patients who showed prolonged tumor stabilization to gemcitabine/UFT. Further studies are warranted to identify a patient subgroup that might have benefits from gemcitabine/fluoropyrimidine therapy.

Background: Src is a nonreceptor tyrosine kinase involved in the crosstalk and mediation of many signaling pathways that promote cell proliferation, invasion and angiogenesis. Elevation of Src activity has been reported in many types of cancers including gastric cancer (GC) and biliary tract cancer (BTC). The purpose of this study is to evaluate Src as a therapeutic target and to elaborate the biomarkers of Src inhibitor in GC and BTC. Methods: Ten gastric cancer cell lines (SNU-1, 5, 16, 216, 601, 620, 638, 668, 719, NCI-N87) and 8 biliary tract cancer cell lines (SNU-245, 308, 478, 869, 1079, 1196, HuCCT1, TFK1) were used. Saracatinib and bosutinib were used as Src inhibitors. MTT assay, colony formation assay and 3D culture were done for determining growth inhibitory effect of Src inhibitors, alone or in combination with chemotherapeutic agents (5-FU, gemcitabine, cisplatin). Cell cycle analysis was done by FACS Calibur flow cytometer. Matrigel invasion assay and wound healing assay were done. The methods described by Chou and Talalay were used to determine whether a synergistic effect existed between drug combination. Tumor xenograft model was made and used for in vivo test of Src inhibitors. Results: Among 10 GC cells, SNU216 and NCI-N87 were sensitive to Src inhibitor. These sensitive cells showed high levels of pSRC(Y416) and pFAK (Y861, Y397, Y925). These 2 sensitive GC cells are both HER2 amplified cells. However, HER2-positive breast cancer (BC) cells (SKBR3, BT474, MDA-MB453) were resistant to Src inhibitor. Contrast to these resistant BC cells, SNU216 and N87 showed high expression of integrin αV, β4 and β8. Especially, in case of integrin β8, the mRNA/protein levels were highest in SNU216 and N87 among all GC cells. Src inhibitor-sensitive GC cells showed the apoptosis by Src inhibitor and synergism with 5-FU in vitro and in vivo. Among 8 BTC cells, 3 cells were sensitive to Src inhibitor (SNU308, SNU478 and HuCCT1) in terms of growth inhibition and migration/invasion inhibition. Sensitive cells showed high levels of integrin α2, α3 and β4. Src inhibitor induced G1 arrest and decreased pSrc, pFAK, and pERK in sensitive cells. Inhibition of pSrc was accompanied with increase of PTEN and decrease of pAKT. Src inhibitor showed the synergistic effects with cytotoxic chemotherapeutic agents (gemcitabine and cisplatin) in vitro and in vivo. When the Src was inhibited, pSTAT3 was increased thru increase of IL-6 in some cells. Conclusion: Taken together, Src could be a potential therapeutic target in gastric cancer and biliary tract cancer. The role of integrin as a biomarker for Src inhibitor should be further investigated. Citation Format: Ah-Rong Nam, Hyun-Jin Nam, Kyo Hwa Kang, Ji Eun Park, Tae Yong Kim, Sae-Won Han, Sang-Hyun Song, Seock-Ah Im, Tae-You Kim, Do-Youn Oh, Yung-Jue Bang. Evaluation of Src as a therapeutic target and development of biomarkers of Src inhibitor in cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 747. doi:10.1158/1538-7445.AM2014-747

Background: Src kinases have been involved in cell proliferation, invasion, and metastasis and is one of the highly expressed oncogenes in breast cancer. Therefore, it could be a reasonable target for therapeutic strategy. Although several Src inhibitors, targeting the ATP binding site, were developed, none of them have shown remarkable responses as monotherapeutic agents in breast cancer clinical trials. KX-01 (KX2-391) is a novel peptidomimetic agent, and a non ATP-competitive Src inhibitor, which has potent efficacy for inhibiting both Src as well as tubulin polymerization. Its dual inhibition effects could potentially overcome the limitations of the Src inhibitors that have been evaluated previously. Materials & Methods: We determined the anti-tumor effects of KX-01 on breast cancer cell lines using a cytotoxicity assay, cell cycle analysis, a wound healing assay and western blotting following KX-01 treatment. Results: KX-01 effectively inhibited cell growth in most breast cancer cells including ER/PR/HER2 negative breast cancer cells. Short exposure of KX-01 down-regulated the expression level of Src phosphorylation. Phosphorylated FAK, ERK, AKT and STAT3 were also down-regulated by KX-01 treatment in MDA-MB-231, MDA-MB-468 and BT-549 cells. By wound healing assay, we confirmed migration inhibitory effect of KX-01 in BT-549 cells. Increase of G2/M cell cycle arrest was observed in dose dependent manner. In addition, mitotic catastrophes were also observed in KX-01 sensitive cell lines. Consistent with induction of mitotic catastrophe, the increased levels of aneuploidy in sensitive cell lines were examined following the treatment of KX-01. Conclusion: Inhibition of cell growth and migration, as well as an induction of mitotic catastrophy, were observed in triple negative breast cancer cell lines treated with KX-01. Our data strongly supports utilizing KX-01 as a new therapeutic agents for treating triple negative breast cancer. Citation Format: Seongyeong Kim, Seock-Ah Im, Ahrum Min, Miso Lee, Heymin Jang, Kyung-Hun Lee, Hee-Jun Kim, Tae-Yong Kim, Sae-Won Han, Do-Youn Oh, Tae-You Kim, Young-Kwang Yoon, David Hangauer, Johnson Lau, Yung-Jue Bang. Antitumor effect of KX-01, a novel Src and tubulin inhibitor, in triple negative breast cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5479. doi:10.1158/1538-7445.AM2014-5479

RNA editing can increase RNA sequence variation without alteration of DNA sequence. By comparing whole genome and transcriptome sequence data of a rectal cancer, we have found a novel tumor specific RNA editing in RHOQ. The A-to-I editing results in substitution of sparagine with serine at residue 136. We observed higher level of the RHOQ RNA editing in tumor compared with normal tissue in colorectal cancer. The degree of RNA editing was associated with RhoQ activity in colorectal cancer cell lines. RhoQ N136S amino acid substitution increased RhoQ activity, actin cytoskeletal reorganization, and invasion potential. KRAS mutation further increased the invasion potential of RhoQ N136S in vitro. Among colorectal cancer patients, recurrence was more frequently observed in patients with tumors having edited RHOQ and mutant KRAS. Collectively, we show that RNA editing is another mechanism of sequence alteration contributing to colorectal cancer progression. Citation Format: Hwang-Phill Kim, Sae-Won Han, Kyung-Hun Lee, Kyu Joo Park, Jeong-Sun Seo, Jong-Il Kim, Tae-You Kim. RNA editing in rhoq promotes invasion potential in colorectal cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5189. doi:10.1158/1538-7445.AM2014-5189

4061 Background: In advanced gastric cancer (AGC), HER2 is a validated therapeutic target. However, the metabolic landscape of AGC based on the HER2 status has not been reported. Furthermore, the p...

4062 Background: Trastuzumab-based chemotherapy is considered as standard treatment for patients with HER2-positive advanced gastric cancer (AGC). However, the biomarkers to predict the sensitivity...

Novel A-to-I RNA editing in the coding sequence of RHOQ leads to an amino acid substitution that promotes invasion in colorectal cancer., RNA editing can increase RNA sequence variation without altering the DNA sequence. By comparing whole-genome and transcriptome sequence data of a rectal cancer, we found novel tumor-associated increase of RNA editing in ras homologue family member Q (RHOQ) transcripts. The adenosine-to-inosine (A-to-I) editing results in substitution of asparagine with serine at residue 136. We observed a higher level of the RHOQ RNA editing in tumor compared with normal tissue in colorectal cancer (CRC). The degree of RNA editing was associated with RhoQ protein activity in CRC cancer cell lines. RhoQ N136S amino acid substitution increased RhoQ activity, actin cytoskeletal reorganization, and invasion potential. KRAS mutation further increased the invasion potential of RhoQ N136S in vitro. Among CRC patients, recurrence was more frequently observed in patients with tumors having edited RHOQ transcripts and mutations in the KRAS gene. In summary, we show that RNA editing is another mechanism of sequence alteration that contributes to CRC progression.

252 Background: Metabolic landscape evaluated by 18F-FDG positron emission tomography (PET) in advanced biliary tract cancer (BTC) has not been studied. Furthermore, the clinical meanings of metabolic features have rarely been reported. We evaluated the metabolic features using 18F-FDG PET and its clinical significances in advanced BTC. Methods: We consecutively enrolled advanced BTC patientswho underwent PET prior to palliative chemotherapy between 2003 and 2013. We retrospectively evaluated the findings of PET such as SUVmax, lesion numbers, involved organ and pathologic findings and other clinical factors including response to chemotherapy, progression-free survival (PFS) and overall survival (OS). Results: A total of 106 patients were enrolled (intrahepatic cholangiocarcinoma (ICC):53, extrahepatic BTC :7, gallbladder cancer (GB Ca) :30, and ampulla of vater cancer (AoV Ca): 16 patients) ECOG PS 0-1 in 89 and PS 2 in11 patients. 52.8 % of patients received gemcitabine-based chemotherapy and 47.1% of patients received 5-FU-based chemotherapy. The SUVmax was median 7.8 (range 0-44). Fifty-four percent of patients showed higher SUVmax in metastatic lesion than primary site. The SUVmax was different according to primary origin (ICC: 9.10, extrahepatic BTC: 5.90, GB Ca : 9.10, AoV Ca :6.37, p=0.008) and histologic differentiation (well-differentiated: 4.95, moderately-differentiated :6.60, poorly-differentiated :14.50, p=0.004) Patients with a SUVmax of >7.5 had more poorly differentiated histology and more PET uptake-lesions (p < 0.05) than those with a SUVmax of 7.5. In multivariate analysis, SUVmax was also a significant prognostic factor for OS (p=0.012) and PFS (p=0.039). Conclusions: Metabolic landscapes of advanced BTC are different according to primary origin and histology. This metabolic feature such as SUVmax could be a potential prognostic factor for OS and PFS in advanced BTC.

351 Background: The optimal treatment strategy for locally advanced pancreatic cancer (LAPC), especially the role of chemoradiotherapy (CCRT), is still in debate. We compared the clinical outcomes of CCRT and palliative chemotherapy alone (CA) in patients with LAPC. Methods: We consecutively enrolled LAPC patients treated between 2003 and 2010. AJCC 7th edition was followed for the diagnostic criteria of LAPC. We retrospectively evaluated the clinical outcomes according to treatment groups (CCRT vs CA). Results: A total of 86 patients were enrolled. Median age was 60 years. ECOG PS was 0-1 in 77 (89.5%) and 2 in 9 (10.5%). Forty five patients (52.3%) were treated with CCRT and 41 patients (47.7%) with CA. Baseline characteristics were not significantly different between CCRT and CA group. In the CCRT group, gemcitabine (n=7, 15.6%), 5-FU (n=10, 22.2%), and capecitabine (n=28, 62.2%) were concurrently used with radiation. Radiation was delivered with 55.8Gy/ 31fraction. All of the CA group patients were treated with gemcitabine-based chemotherapy. Median progression free survival (PFS) and overall survival (OS) of whole patients were 6.9 months [95%CI 4.8-9.0] and 12.7 months [95%CI 11.6-14.3]. PFS and OS of CCRT versus CA was 8.9 months [95%CI 6.8-11.0] vs 3.7 months [95%CI 2.9-4.5] (p

Background: In biliary tract cancer (BTC), no molecular therapeutic target has been validated so far. Therefore, cytotoxic chemotherapy is the mainstay of treatment of advanced BTC and the prognosis is poor and there is a huge unmet medical need. Src is a nonreceptor tyrosine kinase involved in the crosstalk and mediation of many signaling pathways that promote cell proliferation, invasion and angiogenesis. Elevation of Src activity has been reported in many types of cancers including BTC. The purpose of this study is to evaluate of Src as a therapeutic target in BTC and to elaborate the therapeutic potential of bosutinib, an orally active small molecule c-Src/abl kinase inhibitor. Methods: Eight human biliary tract cancer cell lines (SNU-245, 308, 478, 869, 1079, 1196, HuCCT1, TFK1) were used. MTT assay, colony formation assay and 3D culture were done for determining growth inhibitory effect of bosutinib, alone or in combination with chemotherapeutic agents (gemcitabine, cisplatin). Cell cycle analysis was done by FACS Calibur flow cytometer. Matrigel invasion assay and wound healing assay were done. The methods described by Chou and Talalay were used to determine whether a synergistic effect existed between drug combination. Tumor xenograft model was made and used for in vivo test of bosutinib/gemcitabine. Results: Bosutinib inhibited the growth of BTC cells, especially SNU308, SNU478 and HuCCT1 (IC50 0.6-0.8 µmol/L). Bosutinib induced G1 arrest and decreased pSrc, pFAK, pAKT and pERK in these cells. Inhibition of pSrc was accompanied with increase of pSTAT3 in SNU308. Bosutinib inhibited the cell migration and invasion. Interestingly, bosutinib showed synergistic antiproliferative effects with gemcitabine. In the xenograft model of SNU478, bosutinib decreased tumor growth compared to control and bosutinib in combination of gemcitabine showed the synergistic antitumor effect. Conclusion: These preclinical evaluations support that Src could be a potential therapeutic target in biliary tract cancer. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):C266. Citation Format: Do-Youn Oh, Ah-Rong Nam, Tae Yong Kim, Sae-Won Han, Sang-Hyun Song, Seock-Ah Im, Tae-You Kim, Yung-Jue Bang. Evaluation of Src as a therapeutic target in biliary tract cancer. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr C266.

The therapeutic goals are palliative for metastatic breast cancer (MBC) and include prolongation of survival with good quality of life (QoL) and symptom control. The purpose of this study was to examine QoL among women with MBC treated on KCSG-BR07-02 with maintenance of paclitaxel plus gemcitabine (PG) chemotherapy after achieving disease control to initial six cycles of PG chemotherapy or observation. Patients were randomized to either maintenance chemotherapy or observation until progression. QoL was assessed using EORTC QLQ-C30 and BR-23. QoL at each cycle was compared between the two treatment arms using the 2-sample t test. Generalized estimating equation method was used to examine the overall difference between the two treatments in QoL. All reported p-values are 2 sided. There were no statistically significant differences between two arms in most of the component of the EORTC QLQ-C30 and BR-23 (p > 0.05). There was no significant difference between two treatments (p = 0.6094 for QLQ-C30, p = 0.5516 for BR23) at baseline, and there did not exist significant change over the cycle (p = 0.0914 for QLQ-C30, p = 0.7981 for BR23). There was no significant interaction effect between treatment and cycle (p = 0.5543 for QLQ-C30. p = 0.5817 for BR23). Maintenance PG chemotherapy in patients with MBC achieving disease control with an initial six cycles of PG chemotherapy resulted in better PFS and OS compared to observation without impeding QoL.

3550 Background: There have been controversies in prognostic impact of mucinous histology in colorectal cancer (CRC) and its implication in pts treated with adjuvant FOLFOX is unclear. This study aimed at elucidating the molecular characteristics and prognostic implication of mucinous histology in pts treated with adjuvant FOLFOX. Methods: Stage II and III CRC pts who received adjuvant FOLFOX were analyzed. Pts were grouped according to the mucinous content: > 50%, mucinous adenocarcinoma (MAC); < 50%, adenocarcinoma with intermediated mucinous component (AIM); and without any mucinous component, nonmucinous adenocarcinoma (NMA). Clinicopathologic features, MSI status (N = 518), CpG island methylator phenotype (CIMP) (N = 322) BRAF mutation (N = 269) and disease-free survival (DFS) were compared. Results: Among a total of 521 pts, 27 (5.2%) had MAC, 41 (7.9%) AIM, and 453 (86.9%) NMA. MAC and AIM had higher frequency of proximal location and lower angiolymphatic invasion. MAC had higher proportion of T4 tumors. AIM had higher frequency of age ≥65 years and female. In terms of molecular characteristics, MAC and AIM showed similarly higher proportion of MSI-high and CIMP-high compared to NMA. BRAF mutation also showed similar trend. In contrast to the similarities between MAC and AIM, DFS was significantly different. MAC showed significantly worse DFS compared with AIM and NMA, whereas AIM and NMA showed similar DFS. Multivariate analysis revealed MAC as an independent negative prognostic factor of DFS (adjusted HR 7.96, 95% CI 3.76-16.8). Conclusions: AIM and MAC has distinct clinico-pathologic features and molecular characteristics compared with NMA. Only MAC but not AIM has an adverse prognostic impact on stage II or III CRC treated with adjuvant FOLFOX compared with NMA. [Table: see text]

4044 Background: A causal relationship between diabetes mellitus (DM) and pancreatic cancer (PC) is well established. However, in patients with advanced PC (APC) who receive palliative chemotherapy, the impact of DM on prognosis is unclear. Methods: Between 2003 and 2010, we enrolled consecutive patients with APC, all recipients of palliative chemotherapy, with the provision that DM disease status could be properly defined. Enrollees were stratified by diagnosis, in accordance with 2010 DM criteria (AHA/ADA). DM at least 2 years' duration prior to diagnosis of APC qualified as remote-onset DM (vs recent-onset). Clinical characteristics and outcomes were then analyzed. Results: Of the 349 enrollees with APC, 183 (52.4%) had DM. In patients with DM, 160 had DM at the time APC was diagnosed (remote onset, 87; recent onset, 73); and the remaining 23 developed DM during the course of APC treatment. Ultimately, 73.2% (134/183) of DM patients received antidiabetic medication (metformin (56), sulfonylurea (62), insulin (43)). By multivariate analysis, cancer extent (HR, 1.792; p < 0.001) and weight loss during chemotherapy (HR, 1.270; p = 0.08) were associated with diminished overall survival (OS), whereas a diagnosis of DM (HR, 0.788; p = 0.05) conferred a positive effect on OS ( OS 8.4 months in DM patients vs 7.5 months in non-DM patients, p = 0.04). Among DM patients, recent-onset DM trended toward prolonged OS, compared with remote-onset/subsequent DM (9.8 months vs. 7.9 months, respectively; HR, 0.789; p = 0.142). Neither antidiabetic medication in general nor sulfonylurea or insulin specifically affected OS. However, recipients of metformin survived longer than non-recipients (HR, 0.693; 95% CI, 0.492-0.977; p = 0.036). Relative to the APC cohort overall including non-DM patients, metformin conferred better survival as well (11.0 months vs. 7.8 months, HR 0.712, p = 0.067), given similar baseline clinical characteristics. Conclusions: In patients with APC receiving palliative chemotherapy, recent-onset DM (within 2 years of APC diagnosis) and metformin treatment are positive prognosticators, associated with prolonged OS.

2565 Background: HM781-36B, a novel pan-HER tyrosine kinase inhibitor, showed potent in vitro and in vivo antitumor activities for EGFR mutant models including T790M mutation. In the previous 2-weeks on, 1-week off phase I study, the maximum tolerated dose (MTD) was determined as 24 mg/day. Phase I study with a continuous daily dosing schedule was conducted to determine the recommended dose (RD), and to assess the effect of food on pharmacokinetics (PK) in patients (pts) with advanced solid tumors. Methods: Eligible pts had advanced malignancies refractory to standard therapies. Standard 3+3 scheme was used in the dose escalation study, and additional 8 patients were enrolled to test food effects. Results: 20 pts (median age: 55 years [range32-77], M:F=13:7, ECOG PS 0/1/2/3 8/12/0/0) were enrolled (5 NSCLC, 4 colon cancer, 3 stomach cancer, 2 breast cancer, 2 rectal cancer, 2 common bile duct cancer, 1 pancreatic cancer and 1 esophageal cancer); 12 in the dose escalation and 8 in the food effect study cohort. Twelve pts were heavily pretreated(≥4 regimens). Dose limiting toxicities (DLTs) were G3 anorexia in 1 pt at 18mg/day, G3 diarrhea and anorexia in 1 pt, and drug compliance last values, whereas Cmax values were lower in fed condition than in fasted condition. Conclusions: Continuous daily dosing schedule of HM781-36B is safe and well tolerated in advanced solid tumors. It exerts anticancer activity, without being influenced by food. Updated data will be presented at the meeting. Clinical trial information: NCT01455584.

551 Background: Although young age breast cancer represents poor prognosis, no definitive explanation could have been made for the phenomenon. A tumor proliferation marker Ki67 is known to be a marker for both prognosis and prediction for chemotherapy responsiveness, and its level varies widely depending on the breast cancer subtype. This study was aimed to analyze Ki67 in relationship with age in hormone receptor positive breast cancer patients. Methods: We retrospectively reviewed 9061 consecutive cases of hormone receptor positive invasive breast cancer from data base at Seoul National University Hospital (SNUH) (between 2000 and 2012), Samsung Medical Center (SMC) (between 2004 and 2010), and National Cancer Center (NCC) (between 2001 and 2010) in Korea. Patients with estrogen receptor (ER) or progesterone receptor (PR) positive tumors were included irrespective of HER2 amplification. A multicenter data of Ki67 level identified by immunohistochemistry (IHC) and age at diagnosis were analyzed. Patients who underwent neoadjuvant systemic therapy were excluded. Results: Total 6222 cases from SNUH, 976 from SMC and 1863 from NCC were included. The three datasets were analyzed separately due to variable IHC methods in each institute. Mean ages were 49.30 years (range 20-86), 47.75 years (range 22-81) and 45.31 years (range 25-59), and mean Ki67 levels were 4.66% (range 1-100), 22.98% (range 1-97) and 14.58% (range 1-90), at SNUH, SMC, NCC respectively. Ki67 level was inversely proportional with age at diagnosis in all three datasets, and the level was significantly higher for patients

Background: Src is a nonreceptor tyrosine kinase involved in the crosstalk and mediation of many signaling pathways that promote cell proliferation, invasion and angiogenesis. Elevation of Src activity has been reported in many types of cancers including biliary tract cancer (BTC). In BTC, no molecular therapeutic target has been validated so far. Therefore, the prognosis of advanced BTC is limited and there is a huge unmet medical need in this area. The purpose of this study is to evaluate of Src as a therapeutic target in BTC and to elucidate the therapeutic potential and action mechanism of bosutinib, an orally active small molecule c-Src/Abl kinase inhibitor, alone or in combination with chemotherapy for the treatment of BTC. Methods: Experiments were conducted using 8 human biliary tract cancer cell lines (SNU-245, 308, 478, 869, 1079, 1196, HuCCT1, TFK1). MTT assay, colony formation assay and 3D culture were done for determining growth inhibitory effect of bosutinib, alone or in combination with chemotherapeutic agents (gemcitabine, cisplatin). Cell cycle distribution was analyzed by FACS Calibur flow cytometer. Matrigel invasion assay and wound healing assay were done. The methods described by Chou and Talalay were used to determine whether a synergistic effect existed between drug combination. Results: pSrc was highly expressed in most BTC cells. Bosutinib inhibited the growth of BTC cells, especially SNU308, SNU478 and HuCCT1 (IC50, 0.6-0.8 μmol/L). Bosutinib induced G1 arrest (decrease of cyclin D, cyclin E, cyclin A, cyclin B and increase of p27) in SNU308 and SNU478. It decreased pSrc, pFAK, pAKT and pERK in these cells. Bosutinib also inhibited the cell migration and invasion in parallel with decrease of snail, not only in SNU308, SNU478 cells but also in several other cells. Interestingly, bosutinib showed synergistic antiproliferative effects (Combination Index value Conclusion: Taken together, these preclinical evaluations reveal that Src could be a potential therapeutic target in biliary tract cancer and support clinical development of bosutinib alone or in combination with chemotherapy in biliary tract cancer. Citation Format: Ah-Rong Nam, Ahrum Min, Sae-Won Han, Sang-hyun Song, Do-Youn Oh, Seock-Ah Im, Tae-You Kim, Youg-Jue Bang. Evaluation of Src as a therapeutic target in biliary tract cancer using bosutinib, a c-Src/Abl kinase inhibitor. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2081. doi:10.1158/1538-7445.AM2013-2081

Background: The poly (ADP-ribose) polymerase (PARP) inhibitor, olaparib, has been found to have a therapeutic potential for treating cancers that have an impaired DNA repair ability. RAD51C has been reported to play an essential role in DNA repair mediated by homologous recombination. In addition, RAD51C is a gene that affects human cancer susceptibility, similar to BRCA1 and BRCA2. RAD51C-defective cancers can therefore be potentially treated with olaparib because DNA damage induced by olaparib cannot be effectively repaired by HR since RAD51C deficiency interferes with RAD51-mediated HR. Materials and Methods: We studied the growth inhibitory effects of olaparib on human cancer cell lines using clonogenic survival assays. Cell cycle analysis and molecular changed induced by olaparib were also performed. DNA methylation status in cancer cells and tumor tissue samples was also determined by using bisulfate sequencing. Results: Olaparib is effective to RAD51C-defective cells. Olaparib leads to the accumulation of unrepaired DSBs due to dysfunctional RAD51 foci formation along with increased caspase 3-dependent cell death and G2/M arrest in RAD51C-defective cells. Moreover, the growth of RAD51C-deficient SNU601 tumor cells in a xenograft model decreased significantly following treatment with olaparib. Furthermore, RAD51C expression was significantly decreased in cancer and the lack of RAD51C was attributed to DNA methylation and histone modification. Our findings showed that the loss of RAD51C expression due to epigenetic silencing leads to olaparib sensitivity in cancer cells. Conclusion: The loss of RAD51C expression was frequently associated with human cancers. We used a novel synthetic lethal approach using PARP inhibitors and observed that RAD51C was a participant in the DNA repair pathway. Our findings have potential clinical application for treating cancers with RAD51C deficiencies. Furthermore, we determined that RAD51C may serve as a novel biomarker for identifying olaparib-sensitive patients, thereby allowing physicians to select the most effective modalities for treating these individuals. Citation Format: Ahrum Min, Seongyeong Kim, Seock-Ah Im, Young-Kwang Yoon, Sang-Hyun Song, Hyun-Jin Nam, Hyung-Seok Hur, Kyung-Hun Lee, Sae-Won Han, Do-Youn Oh, Tae-You Kim, Mark J. O`Connor, Woo-Ho Kim, Yung-Jue Bang. Olaparib increases antitumor effects on epigenetically RAD51C-deficient human cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3442. doi:10.1158/1538-7445.AM2013-3442

Background: Amphiregulin is a ligand for the epidermal growth factor receptor (EGFR). Human epidermal growth factor receptor 2 (HER2) shares common signal pathways and forms a heterodimer with EGFR. In this study, we investigated the effect of amphiregulin on trastuzumab therapy in HER2-positive breast cancer. Methods: We analyzed serum amphiregulin levels by enzyme-linked immunosorbent assay (ELISA) from baseline serum samples obtained from HER2-positive metastatic breast cancer patients who received first-line trastuzumab plus taxane chemotherapy. In addition, in vitro experiments were performed to elucidate the biologic mechanism of clinical findings related to amphiregulin using SK-BR-3 and BT-474 cell lines. Results: Between October 2004 and July 2009, a total of 50 women with HER2-positive metastatic breast cancer were consecutively enrolled. The median age was 47 years (range, 27-72 years). Eighteen patients (36.0%) received weekly paclitaxel plus trastuzumab, 24 patients (48.0%) tri-weekly paclitaxel plus trastuzumab, and 8 patients (16.0%) tri-weekly docetaxel plus trastuzumab. Among 43 patients with measurable lesions, the response rate (RR) was 76.7%. The median follow-up duration was 29.2 months (range, 0.7-63.3 months). The median progression-free survival (PFS) was 17.6 months (95% confidence interval (CI), 13.4-21.9 months). The median overall survival (OS) was 47.0 months (95% CI, 35.3-58.6 months). The median serum amphiregulin level was 1.0 ng/mL with a maximum level of 4.4 ng/mL. Patients with high serum amphiregulin levels (≥0.5 ng/mL) had significantly shorter PFS (p=0.018) along with a tendency toward lower RR (p=0.237) and shorter OS (p=0.529) than the others. The in vitro colony forming assay demonstrated that the addition of amphiregulin resulted in increased proliferation of both SK-BR-3 and BT-474 cells. In addition, the anti-proliferative effect of trastuzumab was decreased in the presence of amphiregulin in both SK-BR-3 and BT-474 cells. The Western blot analysis showed that amphiregulin increased the phosphorylation of Akt and its downstream molecules in both SK-BR-3 and BT-474 cells. In addition, in the presence of amphiregulin, sustained phosphorylation of Akt and its downstream molecules was observed after trastuzumab treatment in both SK-BR-3 and BT-474 cells. Conclusions: High serum amphiregulin levels (≥0.5 ng/mL) predicted disease progression after first-line trastuzumab plus taxane chemotherapy in patients with HER2-positive metastatic breast cancer. Amphiregulin promoted the proliferation of HER2-positive breast cancer cells in vitro and induced trastuzumab resistance by activating PI3K/Akt pathway. Our results suggest that the measurement of serum amphiregulin levels by ELISA may provide additional information for the clinical outcome of trastuzumab-based chemotherapy in patients with HER2-positive breast cancer. Citation Format: Ji-Won Kim, Young Seok Joung, Ahrum Min, Hyun-Jin Nam, Jee Hyun Kim, Seock-Ah Im, Kyung-Hun Lee, Jin-Soo Kim, Tae-Yong Kim, Sae-Won Han, Yoon Kyung Jeon, Do-Youn Oh, Tae-You Kim, In Ae Park. Amphiregulin confers trastuzumab resistance by activating PI3K/Akt pathway in HER2-positive breast cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2386. doi:10.1158/1538-7445.AM2013-2386

Rad21 is a subunit of cohesin complex, which is well known to be involved in sister chromatid cohesion at mitotic phase. In recent studies, Rad21 has been also known to be associated with long-range chromosome interaction to dynamically regulate gene transcription. However, Very little is known about Rad21-mediated high order chromatin structure and global changes in gene transcription in human cancer. In this study, we silenced Rad21 gene using Lenti-virus knockdown system and performed transcriptome analysis to verify the changes in gene transcription by knockdown of Rad21. Among genes showing significant transcription changes by the reduction of Rad21, we chose several target genes decreasing its transcription and checked the transcriptional change with passaging over a 60 day period. Interestingly, the change in transcription level of some genes appeared by the earliest passage time point, whereas those of other genes arose during the course of long-term passage. Consistent with gene transcription changes, results from a ChIP assay demonstrated that Rad21-knockdown cells showed low level of Rad21 occupancy and active histone marks (H3K4me3 and Pol2) on the target genes, indicating that dynamic histone modifications also contribute to regulation of the target genes. To figure out whether the reduction of Rad21 binding directly affects the chromatin loops, we applied Chromosome Conformation Capture (3C) -qPCR in Rad21-knockdown cells. The results demonstrated that the decreased Rad21 expression cause overal reduction of locus interaction on the target genes, indicating that Rad21 binding was directly required for long-range interactions at the target gene regions and gene transcription regulation. Taken together, the results of our study demonstrated that long-range chromosome interaction mediated by Rad21 can dynamically regulate global gene transcription in human cancer cells, indicating that it may be important for cancer development. Citation Format: Jiyeon Yun, Sang-Hyun Song, Jee-Youn Kang, Hwang-Phill Kim, Sae-Won Han, Do-Youn Oh, Seock-Ah Im, Yung-Jue Bang, Tae-You Kim. Rad21, as a high order chromatin architecture molecule, regulates global gene transcription. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2975. doi:10.1158/1538-7445.AM2013-2975 Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.

182 Background: To evaluate the prognostic significance of phosphorylated Akt (p-Akt), phosphorylated mammalian target of rapamycin (p-mTOR), and total phosphatase and tensin homolog deleted on chromosome 10 (PTEN) expression in patients undergoing adjuvant chemoradiotherapy for proximal extrahepatic bile duct (EHBD) cancer. Methods: Sixty-three patients with proximal EHBD cancer who underwent curative resection followed by adjuvant chemoradiotherapy were enrolled into this study. Postoperative radiotherapy was delivered to tumor bed and regional lymph nodes up to a median of 40 Gy (range; 40-54). Fifty-nine patients also received fluoropyrimidine chemotherapy as a radiosensitizer. p-Akt, p-mTOR, and PTEN expression were assessed with immunohistochemical staining on the tissue microarray. Results: p-Akt, p-mTOR, and PTEN were expressed in 23 (36.5%), 17 (27.0%) and 25 patients (39.7%), respectively. p-Akt expression was associated with distant metastasis and overall survival, but not with loco-regional recurrence. The 5-yr distant metastasis-free and overall survival rates were 25.8% vs. 58.2% (p = 0.007), and 27.5% vs. 50.2% (p = 0.0167) in patients with negative and positive expression, respectively. On multivariate analysis, nodal involvement was the only significant prognosticator predicting inferior distant metastasis-free survival (p = 0.0105), while p-Akt expression had a borderline significance (p = 0.0541). As for overall survival, p-Akt expression was a marginally significant prognosticator (p= 0.0635), whereas other risk factors lost the statistical significance. Conclusions: p-Akt expression was a favorable prognostic factor for patients undergoing curative resection followed by adjuvant chemoradiotherapy for proximal EHBD cancer.

505 Background: Circumferential resection margin (CRM) and distal resection margin (DRM) have different impact on clinical outcomes after preoperative chemoradiotherapy (CRT) followed by surgery. Effect and adequate length of resection margin as well as impact of treatment response after preoperative CRT was evaluated. Methods: Total of 403 patients with locally advanced rectal cancer underwent preoperative CRT followed by total mesorectal excision between January 2004 and December 2010. After applying the criterion of margin less than 0.5 cm for CRM and/or less than 1 cm for DRM, 158 cases were included as a study cohort. All patients underwent conventionally fractionated radiation with dose over 50 Gy and concurrent chemotherapy with 5-FU or capecitabine. Median follow-up duration was 44.9 months. Results: The 5-year overall survival (OS), disease-free survival (DFS), locoregional relapse-free survival (LRFS), and distant metastasis-free survival (DMFS) were 83.3%, 75.6%, 86.3%, and 77.4% respectively. CRM of 1.5 mm and DRM of 7 mm were cutting points showing maximal difference using maximal chi-square method. In univariate analysis, the shorter CRM was significantly related with worse clinical outcomes, whereas DRM was not. In multivariate analysis, CRM of 1.5mm, ypN, and perineural invasion were prognosticators for OS, DFS, LRFS, and DMFS. CRM was not a significant prognostic factor for good responders, defined as patients with near total regression or T down-staging. However, poor responders demonstrated a significant difference according to the CRM status. Conclusions: Close CRM, defined as 1.5 mm, was a significant prognosticator, but the impact was different for treatment response. Postoperative treatment strategy may be individualized based on this finding. However, findings from this study needs to be validated with larger independent cohort. [Table: see text]

To evaluate the prognostic significance of CD24 expression in patients undergoing adjuvant chemoradiotherapy for extrahepatic bile duct (EHBD) cancer.Eighty-four patients with EHBD cancer who underwent curative resection followed by adjuvant chemoradiotherapy were enrolled in this study. Postoperative radiotherapy was delivered to the tumor bed and regional lymph nodes up to a median of 40 Gy (range: 40-56 Gy). All patients also received fluoropyrimidine chemotherapy for radiosensitization during radiotherapy. CD24 expression was assessed with immunohistochemical staining on tissue microarray. Clinicopathologic factors as well as CD24 expression were evaluated in multivariate analysis for clinical outcomes including loco-regional recurrence, distant metastasis-free and overall survival.CD24 was expressed in 36 patients (42.9%). CD24 expression was associated with distant metastasis, but not with loco-regional recurrence nor with overall survival. The 5-year distant metastasis-free survival rates were 55.1% and 29.0% in patients with negative and positive expression, respectively (P = 0.0100). On multivariate analysis incorporating N stage, histologic differentiation and CD24 expression, N stage was the only significant factor predicting distant metastasis-free survival (P = 0.0089), while CD24 expression had borderline significance (P = 0.0733). In subgroup analysis, CD24 expression was significantly associated with 5-year distant metastasis-free survival in node-positive patients (38.4% with negative expression vs 0% with positive expression, P = 0.0110), but not in node-negative patients (62.0% with negative expression vs 64.0% with positive expression, P = 0.8599).CD24 expression was a significant predictor of distant metastasis for patients undergoing curative resection followed by adjuvant chemoradiotherapy especially for node-positive EHBD cancer.

Purpose Exemestane has shown good efficacy and tolerability in postmenopausal women with hormone receptor-positive metastatic breast cancer. However, clinical outcomes in Korean patients have not yet been reported. Methods Data on 112 postmenopausal women with metastatic breast cancer were obtained retrospectively. Clinicopathological characteristics and treatment history were extracted from medical records. All patients received 25 mg exemestane daily until objective disease progression. Progression-free survival (PFS) was the primary endpoint, and secondary endpoints were overall survival (OS), objective response rate (ORR), and clinical benefit rate (CBR=complete response+partial response+stable disease for 6 months). Results The median age of the subjects was 55 years (range, 28-76 years). Exemestane treatment resulted in a median PFS of 5.7 months (95% confidence interval [CI], 4.4-7.0 months) and median OS of 21.9 months (95% CI, 13.6-30.3 months). ORR was 6.4% and CBR was 46.4% for the 110 patients with evaluable lesions. Symptomatic visceral disease was independently associated with shorter PFS (hazard ratio, 3.611; 95% CI, 1.904-6.848; p

3624 Background: CpG island methylator phenotype (CIMP) is characterized by concurrent methylation of multiple CpG islands in tumor DNA, which can inactivate tumor suppressor genes or promote carcinogenesis. The prognostic impact of CIMP on treatment outcome of colon cancer patients receiving adjuvant 5-fluorouracil/leucovorin/oxaliplatin (FOLFOX) is unclear. We investigated CIMP markers in colorectal cancer patients treated with adjuvant FOLFOX. Methods: Sporadic colorectal cancer patients treated with curative resection followed by adjuvant FOLFOX were included. DNA was extracted from formalin-fixed paraffin-embedded surgical specimen. 8 CpG island loci (CACNA1G, CRABP1, IGF2, MLH1, NEUROG1, P16, RUNX3 and SOCS1) were examined using MethyLight analysis. Disease-free survival (DFS) was evaluated according to each methylation loci. Results: A total of 322 patients were included. Methylation at 1 or more loci was observed in 150 patients (46.6%) and 6 or more loci in 15 (4.7%). During a median follow-up duration of 39.7 months, 55 recurrences were observed. Three year DFS in the patient cohort was 84%. CRABP1 (23.9%) was the most frequently methylated loci, followed by p16 (22.7%) and NEUROG1 (20.8%). Patients having methylation at NEUROG1 (3 year DFS 78% in (+) vs. 86% in (-), p = 0.014) and p16 (3 year DFS 78% in (+) vs. 86% in (-), p = 0.12) had worse DFS, whereas methylation at MLH1 had better DFS (3 year DFS 100% in (+) vs. 86% in (-), p = 0.19). In a combined analysis, patients with MLH1(-)/NEUROG1(+)/p16(+) had worst treatment outcome compared to MLH1(-)/NEUROG1(+) or p16(+), MLH1(-)/ NEUROG1(-) /p16(-), and MLH1(+) (3 year DFS 62%, 82%, 87%, and 100%, respectively; p = 0.002). In multivariate analysis, NEUROG1(+)/p16(+) was associated with significantly higher recurrence compared with other patients (adjusted hazard ratio (HR) 2.15 (95% confidence interval (CI) 1.08 - 4.27, p = 0.029). Conclusions: Methylation status of NEUROG1, p16, and MLH1 is associated with recurrence following adjuvant FOLFOX in stage II/III colorectal cancer. Further validation and translational studies to improve treatment outcome in the subset of patients are warranted in the future.

1003 Background: Chemotherapy provides a survival benefit in patients with metastatic breast cancer (MBC), but the optimal duration of chemotherapy remains controversial. Primary purpose of the study was to evaluate whether the maintenance chemotherapy with gemcitabine/paclitaxel (GP), which is one of the two regimens which showed a survival gain from a randomized trial, is superior to observation in terms of progression free survival (PFS) in responding patients with MBC after 6 cycles of GP as first-line treatment. Methods: This study is a prospective, randomized, multi-center, phase III study. Patients who achieved response (CR+PR+SD) following 6 cycles of GP chemotherapy (gemcitabine 1250 mg/m2 on day 1 and 8 plus paclitaxel 175 mg/m2 on day 1 every 3 weeks) randomized to maintenance till progression or observation arm. The trial was conducted by the Korean Cancer Study Group (KCSG). Results: Among total 324 patients enrolled between 2007 and 2010 from 10 centers, 231 responding patients to were randomly assigned to maintenance chemotherapy (n=116) or observation (n=115). Median age was 49 (range 28-76). The numbers of hormone receptor (HR)+ve and HR-ve patients were 172 (74.5%) and 59 (25.5%), respectively. The median No. of chemotherapy cycles in maintenance group was 12 (range 6-32). During median 33 months of follow-up, median PFS was superior in maintenance than in observation (12.0 vs. 8.3 months, p=0.030). Patients < age 50 years (hazard ratio 0.50, p=0.001) and HR-ve patients (hazard ratio 0.52, p=0.019) received more benefit from maintenance chemotherapy in terms of PFS. Median OS was superior in maintenance than in observation (36.8 vs 28.0 months, p=0.047). Neurotoxicity (≥ grade 2) was more common in maintenance than in observation without statistical significance (41.7% vs 33.3%, p=0.210). Serial assessment of Quality of Life (QoL) did not show any significant difference between two groups. Conclusions: Maintenance GP chemotherapy for responding patients with MBC showed clinical benefit in terms of PFS and OS without impairment of QoL.

3076 Background: HM781-36B is a pan-HER tyrosine kinase inhibitor, which showed a potent activity against the gefitinib- or erlotinib-resistant, EGFR L858R/T790M double mutant cells. A phase I study was conducted to determine the MTD, pharmacokinetics, and antitumor activity. Methods: Eligible pts had advanced malignancies refractory to standard therapies. Standard 3+3 scheme was used in the dose escalation part, and additional 12 pts were enrolled in the expansion cohort of molecular enrichment. Results: In dose-escalation part, 43 pts (median age: 55 yrs (range 25-82), M:F=25:18, ECOG PS 0/1/2/3: 23/17/2/1, median prior chemotherapy: 4) were treated. DLTs were G3 diarrheas in 5 pts, one at 12 mg, 16 mg, 24 mg, and two at 32 mg. The MTD was determined as 24mg. The most common drug-related adverse events were diarrhea, stomatitis, rash, pruritus, and anorexia. Among 41 evaluable pts, 4 pts achieved PR (1 unconfirmed, duration of response: 11.9 mo, 7.07 mo+, 4.5 mo+), and 19 pts had SD. Two of 4 PR pts were Her2-positive breast cancer pts. The median duration of treatment in pts with PR or SD was 3.87 (2.47- 15.17) months. In the dose range of 0.5 to 24 mg, it showed linear pharmacokinetics proportional to dose-escalation, relatively short half-life, and little accumulation. Additional 12 pts in the expansion cohort are under treatment at 24 mg (6 pts: EGFR-mutant NSCLC, 3 pts: Her2-positive gastric cancer, 2 pts: Her2-positive breast cancer, 1 pt: rectal cancer). Conclusions: HM781-36B was safe and well tolerable in advanced solid tumors. Preliminary evidence of anticancer activity has been observed. Updated data will be presented at the meeting.

Cancer is a result of complex genetic alterations including mutations, copy number alterations, and gene fusions. Recent advance in sequencing technology enables comprehensive genome-wide study of the complex genetic alterations. Better understanding of the genetic alteration of cancer will ultimately lead to better treatment options and improved survival of the patients. We have established a set of cancer-related genes comprised of 184 genes for targeted sequencing using next generation sequencing technology. Targeted genome sequencing of the coding regions in the gene set was performed using the Illumina Genome Analyzer IIx. We analyzed the gene set in 13 gastric cancer cell lines. Average coverage depth was 140X. A total of 237 somatic sequence variations were found in 111 genes. The frequently mutated genes were ATM in 7, MSH6 in 6, TP53 in 5, and KRAS in 4 gastic cancer cells. Using the coverage data, we could also analyze copy number variations. Copy number gain (>2.0 fold) was found in 5 genes, whereas copy number loss ( 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 5096. doi:1538-7445.AM2012-5096

Epiregulin (EREG), a ligand of the epidermal growth factor receptor (EGFR), induces cell growth by binding to the EGFR. Expression of EREG affects sensitivity to cetuximab a chimeric monoclonal antibody that inhibits the EGFR signaling pathway. The mechanism through which EREG is regulated is largely unknown, but a methyl-array study performed by our group revealed that EREG is methylated in gastric cancer cells. In the present study, we found that EREG gene expression was low in seven out of 11 gastric cancer cells and this down-regulation was mediated by aberrant CpG methylation of the EREG promoter. Treatment with 5-aza-CdR restored EREG expression and demethylated the CpG sites in the EREG promoter. Compared to DNMT1, knockdown of DNMT3b significantly increased the expression of EREG mRNA and led to the demethylation of specific CpG sites in the EREG promoter, suggesting that DNMT3b primarily regulates the CpG methylation and silencing of EREG gene. EREG methylation was observed in 30% (4/13) of human primary gastric tumor tissues. In addition to DNA methylation, results from a ChIP assay demonstrated that transcriptional levels of EREG were associated with the enrichment of active histone markers (H3K4me3 and AcH3) and of a repressive marker (H3K27me2). Treatment with 5-aza-CdR dynamically increased the low occupancy of H3K4me3 and AcH3, while decreasing the high enrichment of H3K27me2, indicating that dynamic histone modifications contribute to EREG regulation in addition to DNA methylation. Lastly, the combination of 5-aza-CdR and cetuximab exerted a synergistic anti-proliferative effect on gastric cancer cells. Taken together, the results of our study demonstrated for the first time that EREG is epigenetically silenced in gastric cancer cells by aberrant DNA methylation and histone modification. 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 4017. doi:1538-7445.AM2012-4017

Background Identification of predictive biomarkers is essential for the successful development of targeted therapy. Insulin-like growth factor 1 receptor (IGF1R) has been examined as a potential therapeutic target for various cancers. However, recent clinical trials showed that anti-IGF1R antibody and chemotherapy are not effective for treating lung cancer. Methodology/Principal Findings In order to define biomarkers for predicting successful IGF1R targeted therapy, we evaluated the anti-proliferation effect of figitumumab (CP-751,871), a humanized anti-IGF1R antibody, against nine gastric and eight hepatocellular cancer cell lines. Out of 17 cancer cell lines, figitumumab effectively inhibited the growth of three cell lines (SNU719, HepG2, and SNU368), decreased p-AKT and p-STAT3 levels, and induced G 1 arrest in a dose-dependent manner. Interestingly, these cells showed co-overexpression and altered mobility of the IGF1R and insulin receptor (IR). Immunoprecipitaion (IP) assays and ELISA confirmed the presence of IGF1R/IR heterodimeric receptors in figitumumab-sensitive cells. Treatment with figitumumab led to the dissociation of IGF1-dependent heterodimeric receptors and inhibited tumor growth with decreased levels of heterodimeric receptors in a mouse xenograft model. We next found that both IGF1R and IR were N-linked glyosylated in figitumumab-sensitive cells. In particular, mass spectrometry showed that IGF1R had N-linked glycans at N913 in three figitumumab-sensitive cell lines. We observed that an absence of N-linked glycosylation at N913 led to a lack of membranous localization of IGF1R and figitumumab insensitivity. Conclusion and Significance The data suggest that the level of N-linked glycosylated IGF1R/IR heterodimeric receptor is highly associated with sensitivity to anti-IGF1R antibody in cancer cells.

623 Background: Ethnic diversity of genetic polymorphism can result in individual differences in the efficacy and toxicity of cancer chemotherapy. Methods: A total of 292 Korean patients (183 men and 109 women) from six hospitals in Korea were prospectively enrolled. Patients had resected stage III or high-risk stage II colon cancer and were treated with 12 cycles of adjuvant oxaliplatin plus leucovorin plus 5-fluorouracil (FOLFOX) chemotherapy. 20 germline polymorphisms in 10 genes (TS, MTHFR, ERCC1, XPD, XRCC1, ABCC2, AGXT, GSTP1, GSTT1 and GSTM1) were analyzed from peripheral blood. TS genotype in 5’UTR was classified as ‘high’ (2R/3G, 3C/3G, and 3G/3G) or ‘low’ (2R/2R, 2R/3C, and 3C/3C). Results: Most patients (86.3%) received 12 complete cycles of FOLFOX chemotherapy. In contrast to previous studies in Caucasians, neutropenia (grade 3–4, 60.5%) was frequently observed in our Korean patients, whereas only 16.4% experienced grade 2 or more sensory neuropathy. Neutropenia was more frequent in MTHFR 677TT [adjusted odds ratio (OR) 2.32, 95% confidence interval (CI) 1.19–4.55] and ERCC1 19007TT (adjusted OR 4.58, 95% CI 1.20–17.40) genotypes. Patients harboring XRCC1 23885GG had lower risk of neuropathy (adjusted hazard ratio 0.56, 95% CI 0.32-0.99) and longer time to the onset of grade 2-4 neuropathy. MTHFR 677TT and XRCC1 23885GG genotype was also more prevalent in Koreans compared to Caucasians, and the difference of genotypic frequency could partly explain the ethnically different toxicity profile. After median 49.4 months of follow-up, there were 58 (19.9%) relapses and 19 (6.5%) deaths. TS ‘low’ genotype [adjusted hazard ratio (OR) 1.83, 95% CI 1.003–3.34] was significantly related to shorter disease-free survival. Overall survival was not significantly different according to the polymorphisms. Conclusions: Polymorphisms in MTHFR, XRCC1 and TS are related to toxicities and disease-free survival in patients with colon cancer. These polymorphisms may explain the ethnic difference in toxicity profile following adjuvant FOLFOX chemotherapy.

269 Background: In advanced HCC, the role of salvage chemotherapy after failure of prior chemotherapy including sorafenib has not been fully studied. The aims of this study were to evaluate the efficacy of infusional 5-fluorouracil (5-FU), doxorubicin, and mitomycin-C (iFAM) as salvage regimen in advanced HCC and to identify the prognostic factors in a salvage setting. Methods: Eligibility included the following: 1) age > 18 years; 2) Diagnosed as HCC; 3) previously treated with palliative chemotherapy; 4) ECOG PS 0-2; 5) Child-Pough Class A or B; and 6) adequate BM and renal function. iFAM consisted of 5-FU 800 mg/m2 over 10 hours on days 1-5, doxorubicin 30 mg/m2 on day 1 and mitomycin-C 8 mg/m2 on day 1, every 4 weeks. Response evaluation was done every 8 weeks with RECIST criteria. Results: A total of thirty-five patients were enrolled. The median age was 50.2 years (range: 23.3-74.7). ECOG PS was; PS 0-1 (80%) and PS 2 (20%). The etiology of HCC was; HBV (82.8%), HCV (2.9%) and others (14.3%). Liver function was; Child Pough A (91.4%) and B (8.6%). Nine patients had macrovascular invasion and all had extrahepatic metastatic lesion. Prior chemotherapies were sorafenib or 5-FU based regimens. A total of 94 cycles of iFAM were delivered (median 2: range 1-7). There were 2 PR and 6 SD, resulting in RR 5.7% (95% CI, 1.6-18.6%) and DCR 22.9% (95% CI, 12.1-39%). The PFS and OS were 2.4 months and 8.6 months, respectively. The 6- and 12-month survival rates were 67.7% and 35.3%, respectively. Grade 3/4 hematologic toxicity was neutropenia (14.3%). Frequent non-hematological toxicities were; anorexia (45.7%), alopecia (40%), stomatitis (20%), all of which were grade 1/2. In multivariate analysis, PS and serum α-FP were significantly associated with OS. Patients were classified into 3 risk groups based on the number of prognostic factors (0 vs. 1 vs. 2). This classification was well-correlated to OS (16.1 months vs. 10.5 months vs. 3.8 months, respectively; p < 0.0001). Conclusions: iFAM is an effective and tolerable treatment option in advanced HCC and could be considered as salvage regimen. PS and α-FP are significant prognostic factors in a salvage setting.

626 Background: Systemic chemotherapy is still mainstay of treatment of metastatic colorectal cancer (mCRC), while the role of palliative resection is not convincing. We intended to find out the role of palliative resection in mCRC. Methods: A total of 1,015 patients were diagnosed with mCRC at Seoul National University Hospital between 2000 and 2009. Except 169 patients who received curative metastectomy of liver and/or lung, 847 patients were retrospectively analyzed. Results: Out of 847 patients, 556 (65.6%) had metastasis at the time of diagnosis and 291 (34.4%) had recurrence after surgery. The median age was 61 (range, 16-88) and 491 (58.0%) were male. The median number of metastatic site was 1 (range, 1-6) and 738 (87.1%) had limited metastasis (number of metastatic site ≤2). The liver was most frequently involved site (451, 53.2%). Surgery was done in 527 (62.2%). One hundred three patients received resection of both primary and metastatic sites (group 1), while 347 and 78 received resection of primary (group 2) and metastatic sties (group 3), respectively. R0 resection was done in 95 patients (G1: 53, G3: 42), while R1/2 was done 431 in patients (G1: 56, G2: 336, G3:39). Of 95 patients with R0 resection, 93 (97.8%) had limited metastasis (number of metastatic organ ≤2) in the peritoneum, lymph nodes, liver, or lung. The median overall survival (OS) was 19.0 months (95% CI, 17.8-20.1) and resected patients had prolonged median OS compared with patients never resected (21.3 vs 14.1 months, p < 0.001). In multivariate analysis, R0 resection was associated with superior OS compared to R2 resection (51.3 vs 18.7 months; HR, 3.0; 95% CI, 1.86 to 4.85, P Conclusions: Palliative resection with curative intent and chemotherapy confer long-term survival on subsets of mCRC with limited metastasis.

Background This study was aimed 1) to investigate the predictive value of FDG PET/CT (fluorine-18 fluorodeoxyglucose positron emission tomography/computed tomography) for histopathologic response and 2) to explore the results of FDG PET/CT by molecular phenotypes of breast cancer patients who received neoadjuvant chemotherapy. Methods Seventy-eight stage II or III breast cancer patients who received neoadjuvant docetaxel/doxorubicin chemotherapy were enrolled in this study. FDG PET/CTs were acquired before chemotherapy and after the first cycle of chemotherapy for evaluating early metabolic response. Results The mean pre- and post-chemotherapy standard uptake value (SUV) were 7.5 and 3.9, respectively. The early metabolic response provided by FDG PET/CT after one cycle of neoadjuvant chemotherapy was correlated with the histopathologic response after completion of neoadjuvant chemotherapy (P = 0.002). Sensitivity and negative predictive value were 85.7% and 95.1%, respectively. The estrogen receptor negative phenotype had a higher pre-chemotherapy SUV (8.6 vs. 6.4, P = 0.047) and percent change in SUV (48% vs. 30%, P = 0.038). In triple negative breast cancer (TNBC), the pre-chemotherapy SUV was higher than in non-TNBC (9.8 vs. 6.4, P = 0.008). Conclusions The early metabolic response using FDG PET/CT could have a predictive value for the assessment of histopathologic non-response of stage II/III breast cancer treated with neoadjuvant chemotherapy. Our findings suggest that the initial SUV and the decline in SUV differed based on the molecular phenotype. Trial Registration ClinicalTrials.gov: NCT01396655

Background Because of the late clinical presentation of biliary tract cancer (BTC), only 10% of patients are eligible for curative surgery. Even among those patients who have undergone curative surgery, most patients develop recurrent cancer. This study is to determine the clinical role of 18F-FDG PET/CT during post-operative surveillance of suspected recurrent BTC based on symptoms, laboratory findings and contrast-enhanced CT (ceCT) findings. Methods We consecutively enrolled 50 patients with BTC who underwent curative surgery. An 18F-FDG PET/CT was obtained for assessment of recurrence based on clinical suspicion during post-operative surveillance. The final confirmation of recurrence was determined pathologically or clinically. When a pathologic confirmation was impossible or inconclusive, a clinical confirmation was used by radiologic correlation with subsequent follow-up ceCT at a minimum of 3-month intervals. Diagnostic efficacy was evaluated by comparing the results of ceCT and 18F-FDG PET/CT with the final diagnosis. Results Among the 50 patients, 34(68%) were confirmed to have a recurrence. PET/CT showed higher sensitivity (88% vs. 76%, p = 0.16) and accuracy (82% vs. 66%, p = 0.11) for recurrence compared to ceCT, even though the difference was not significant. The positive (86% vs. 74%, p = 0.72) and negative predictive values for recurrence (73% vs. 47%, p = 0.55) were not significantly different between PET/CT and ceCT. However, an additional PET/CT on ceCT significantly improved the sensitivity than did a ceCT alone (94% [32/34] for PET/CT on ceCT vs. 76% [26/34] for ceCT alone, p = 0.03) without increasing the specificity, positive predictive value, and negative predictive value. Conclusions 18F-FDG PET/CT alone is not more sensitive or specific than ceCT in the detection of recurrent BTC after curative surgery. These results do not reach statistical significance, probably due to the low number of patients. However, an additional 18F-FDG PET/CT on ceCT significantly improves the sensitivity of detecting recurrences.

Background: Src is a nonreceptor tyrosine kinase involved in the crosstalk and mediation of many signaling pathways that promote cell proliferation, adhesion, invasion, migration, metastasis, angiogenesis, and tumorigenesis. Elevation of Src activity has been reported in many types of human cancers. Therefore, Src has been promising therapeutic target for the treatment of cancers. However, the mechanism of Src inhibition in gastric cancers has not been fully understood yet. Saracatinib is an orally active small molecule c-Src/abl kinase inhibitor currently in phase II clinical trials. The purpose of this study is to evaluate the therapeutic potential of saracatinib alone or in combination with chemotherapeutic agents for the treatment of gastric cancers by testing its in vitro and in vivo efficacy. Methods: Experiments were conducted in human gastric cancer cell lines from the Korea Cell Line Bank or the American Type Culture Collection. MTT assay was used for determining growth inhibitory effect of saracatinib alone or in combination with chemotherapeutic agents (5-FU, cisplatin). Cell cycle distribution was analyzed by FACS Calibur flow cytometer and apoptotic effect was measured by annexin V assay. Western blot analysis was performed to verify molecular mechanism of this compound. The methods described by Chou and Talalay were used to determine if a synergistic effect existed. The analysis of the median effect was conducted using the Calcusyn software to determine a combination index value. (CI>1: antagonistic effect, CI=1:additive effect, CI Results: Saracatinib specifically inhibited the growth of SNU216 and NCI N87 cells among 10 gastric cancer cell lines. We found that saracatinib induces G1 arrest and apoptosis in sensitive cell lines. The apoptotic process required induction of the proapoptotic BH3-only BCL2 family member, BIM, and saracatinib led to increase in the BIM level in SNU216 and N87 cells. Knockdown of BIM using siRNA decreased the apoptotic cell numbers induced by treatment of saracatinib suggesting important role for BIM in saracatinib induced apoptosis pathway. The sensitivity to saracatinib in SNU216 and N87 cell lines was correlated with inhibiton of AKT and ERK pathways and blockade of HER family kinases. Furthermore, the combined treatment of saracatinib with 5-FU or cisplatin exerted synergistic effects in gastric cancer cell lines including NCI N87 cells. Consistent with these in vitro findings, saracatinib alone or in combination with 5-FU showed antitumor activity in a NCI-N87 xenograft model. Conclusion: Taken together, these preclinical evaluations support that Src is a potential therapeutic target in gastric cancer and clinical development of saracatinib alone or in combination with chemotherapy in gastric cancer could be considered. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3588. doi:10.1158/1538-7445.AM2011-3588

compounds with anticancer and antitumor activity. Onchidium is an airbreathing and shell-less sea slug from gastropod mollusc in the family Onchidiidae, that is distributed with a good abundance in Persian Gulf. Methods: Adult Onchidium were collected from Persian Gulf during spring of 2010. Eggs were freeze dried and used for protein extraction by homogenization in acid acetic (0.1N). After centrifugation, supernatants were freeze dried. Freeze dried extract dissolved in ammonium acetate buffer and it was subjected to gel filtration on a Sephadex G50 column (1.4×60). Purified molecules were separated by SDS-PAGE under denaturing conditions in 12.5% polyacrylamide gels which was calibrated with SDS-PAGE polypeptide high range standards. A 320 kDa protein separated and their in vitro anti-proliferative and cytotxic effects on human breast cancer cell line (T47D in RPMI-1640 culture medium) examined by MTT assay. Results: This protein showed growth inhibitory activity against T47D cells with LC50 values ranging 0.240 to 0.923 nM. This result suggests that separated 320 kDa protein has potent activity against breast cancer cell line. To understand the mechanism of their anti-cancer action, the effect of this compound on the location of cells in the cell cycle was examined using fluorescent activated cell-sorting analysis. This protein was added to T47D culture medium at varying doses, and then at 24 hr the cells were examined for cell cycle location. The addition of separated 320 kDa protein to these cells caused a dose-dependent block in the cell cycle in the G2/M phase. Conclusion: Although these primeval results showed anti-proliferative capacity of this protein, the modulation of apoptotic pathways deserves further evaluation as an anticancer strategy. Disclosure of Interest: None Declared

Background Studies about the biology, treatment pattern, and treatment outcome of metastatic/recurrent neuroendocrine tumor (NET) have been few. Methods We enrolled patients with metastatic/recurrent NET diagnosed between January 1996 and July 2007 and retrospectively analyzed. Results A total of 103 patients were evaluated. Twenty-six patients (25.2%) had pancreatic NET, 27 (26.2%) had gastrointestinal NET, 2 (1.9%) had lung NET, 28 (27.2%) had NET from other sites, and 20 (19.4%) had NET from unknown origin. The liver was the most common metastatic site (68.9%). Thirty-four patients had grade 1 disease, 1 (1.0%) had grade 2 disease, 15 (14.6%) had grade 3 disease, 9 (8.7%) had large cell disease, and 7 (6.8%) had small cell disease. Sixty-six patients received systemic treatment (interferon, somatostatin analogues or chemotherapy), 64 patients received local treatment (TACE, radiofrequency ablation, metastasectomy, etc.). Thirty-six patients received both systemic and local treatments. Median overall survival (OS) was 29.0 months (95% confidence interval, 25.0-33.0) in the103 patients. OS was significantly influenced by grade (p = .001). OS was 43.0, 23.0, and 29.0 months in patients who received local treatment only, systemic treatment only, and both treatments, respectively (p = .245). The median time-to-progression (TTP) was 6.0 months. Overall response rate was 34.0% and disease-control rate was 64.2%. TTP was influenced by the presence of liver metastasis (p = .011). Conclusions OS of metastatic/recurrent NET was different according to tumor grade. TTP was different according to metastasis site. Therefore, development of optimal treatment strategy based on the characteristics of NET is warranted.

1088 Background: Lapatinib (L) is a dual tyrosine kinase inhibitor of EGFR and HER2. Lapatinib plus capecitabine (C) is an effective treatment option in trastuzumab (T)-refractory HER2-positive (+)...