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Purpose Isoform 2 of tight junction protein claudin-18 (CLDN18.2) is a potential target for gastric cancer treatment. A treatment targeting CLDN18.2 has shown promising results in gastric cancer. We investigated the clinical significance of CLDN18.2 and other cell-adherens junction molecules (Rho GTPase-activating protein [RhoGAP] and E-cadherin) in metastatic diffuse-type gastric cancer (mDGC). Materials and Methods We evaluated CLDN18.2, RhoGAP, and E-cadherin expression using two-plex immunofluorescence and quantitative data analysis of H-scores of 77 consecutive mDGC patients who received first-line platinum-based chemotherapy between March 2015 and February 2017. Results CLDN18.2 and E-cadherin expression was significantly lower in patients with peritoneal metastasis (PM) than those without PM at the time of diagnosis (P=0.010 and 0.013, respectively), whereas it was significantly higher in patients who never developed PM from diagnosis to death than in those who did (P=0.001 and 0.003, respectively). Meanwhile, CLDN18.2 and E-cadherin expression levels were significantly higher in patients with bone metastasis than in those without bone metastasis (P=0.010 and 0.001, respectively). Moreover, we identified a positive correlation between the expression of CLDN18.2 and E-cadherin (P

Background/Aims: The outcome of local treatment for advanced non-small cell lung cancer (NSCLC) remains poor, with therapies such as induction chemother apy (IC) yielding conflicting results. This study aimed to assess the clinicopath ologic and prognostic significance of the excision repair cross-complementation group 1 (ERCC1), beclin-1, and glucose-regulated protein of molecular mass 78 (GRP78) in patients with locally advanced NSCLC receiving docetaxel-platinum IC, along with efficacy and safety. Methods: This is a retrospective observational cohort study. We reviewed medical records of 31 NSCLC patients receiving docetaxel-platinum IC, and conducted im- munohistochemical staining of ERCC1, beclin-1, and GRP78. Results: Response rate was 67.8% with 10.7 months of median relapse-free surviv al (RFS) and 23.1 months of median overall survival (OS), and no treatment-related death was reported. High expression of ERCC1, beclin-1, and GRP78 was identi fied in 67.7%, 87.1%, and 67.7%, respectively. Expression of ERCC1 and GRP78 did not reveal statistical significance in survival, whereas high beclin-1 expression re vealed longer OS (7.6 months vs. 23.2 months; log-rank p = 0.024). In multivariate analysis, histologic differentiation (hazard ratio [HR], 3.48; p < 0.001), stage (HR, 8.5; p = 0.024), and adjuvant treatment (HR, 16.1; p = 0.001) were related to RFS, and in OS, stage (HR, 5.4; p = 0.037), adjuvant treatment (HR, 8.6; p = 0.004), and beclin-1 expression (HR, 8.2; p = 0.011) were identified as significant prognostic factors. Conclusions: Our findings suggest that high beclin-1 expression predicts longer survival in locally advanced NSCLC and docetaxel-platinum IC is a treatment op tion that deserves consideration.

Background Immune checkpoint blockades (ICBs) are characterized by a durable clinical response and better tolerability in patients with a variety of advanced solid tumors. However, we not infrequently encounter patients with hyperprogressive disease (HPD) exhibiting paradoxically accelerated tumor growth with poor clinical outcomes. This study aimed to investigate implications of clinical factors and immune cell composition on different tumor responses to immunotherapy in patients with non-small cell lung cancer (NSCLC). Methods This study evaluated 231 NSCLC patients receiving ICBs between January 2014 and May 2018. HPD was defined as a > 2-fold tumor growth kinetics ratio during ICB therapy and time-to-treatment failure of ≤2 months. We analyzed clinical data, imaging studies, periodic serologic indexes, and immune cell compositions in tumors and stromata using multiplex immunohistochemistry. Results Of 231 NSCLC patients, PR/CR and SD were observed in 50 (21.6%) and 79 (34.2%) patients, respectively and 26 (11.3%) patients met the criteria for HPD. Median overall survival in poor response groups (HPD and non-HPD PD) was extremely shorter than disease-controlled group (SD and PR/CR) (5.5 and 6.1 months vs. 16.2 and 18.3 months, respectively, P = 0.000). In multivariate analysis, HPD were significantly associated with heavy smoker (p = 0.0072), PD-L1 expression ≤1% (p = 0.0355), and number of metastatic site ≥3 (p = 0.0297). Among the serologic indexes including NLR, PLR, CAR, and LDH, only CAR had constantly significant correlations with HPD at the beginning of prior treatment and immunotherapy, and at the 1st tumor assessment. The number of CD4+ effector T cells and CD8+ cytotoxic T cells, and CD8+/PD-1+ tumor-infiltrating lymphocytes (TIL) tended to be smaller, especially in stromata of HPD group. More M2-type macrophages expressing CD14, CD68 and CD163 in the stromal area and markedly fewer CD56+ NK cells in the intratumoral area were observed in HPD group. Conclusions Our study suggests that not only clinical factors including heavy smoker, very low PD-L1 expression, multiple metastasis, and CAR index, but also fewer CD8+/PD-1+ TIL and more M2 macrophages in the tumor microenvironment are significantly associated with the occurrence of HPD in the patients with advanced/metastatic NSCLC receiving immunotherapy.

BackgroundHypoxic tumors are known to be highly resistant to radiotherapy and cause poor prognosis in non-small cell lung cancer (NSCLC) patients. CKD-516, a novel vascular disrupting agent (VDA), mainly affects blood vessels in the central area of the tumor and blocks tubulin polymerization, thereby destroying the aberrant tumor vasculature with a rapid decrease in blood, resulting in rapid tumor cell death. Therefore, we evaluated the anti-tumor efficacy of CKD-516 in combination with irradiation (IR) and examined tumor necrosis, delayed tumor growth, and expression of proteins involved in hypoxia and angiogenesis in this study.MethodsA xenograft mouse model of lung squamous cell carcinoma was established, and the tumor was exposed to IR 5 days per week. CKD-516 was administered with two treatment schedules (day 1 or days 1 and 5) 1 h after IR. After treatment, tumor tissues were stained with hematoxylin and eosin, and pimonidazole. HIF-1α, Glut-1, VEGF, CD31, and Ki-67 expression levels were evaluated using immunohistochemical staining.ResultsShort-term treatment with IR alone and CKD-516 + IR (d1) significantly reduced tumor volume (p = 0.006 andp = 0.048, respectively). Treatment with CKD-516 + IR (d1 and d1, 5) resulted in a marked reduction in the number of blood vessels (p p = 0.02) and decreased HIF-1α, Glut-1, VEGF, and Ki-67 expression. Long-term administration of CKD-516 + IR reduced tumor volume and delayed tumor growth. This combination also greatly reduced the number of blood vessels (p = 0.0006) and significantly enhanced tumor necrosis (p = 0.004). CKD-516 + IR significantly increased HIF-1α expression (p = 0.0047), but significantly reduced VEGF expression (p = 0.0046).ConclusionsTaken together, our data show that when used in combination, CKD-516 and IR can significantly enhance anti-tumor efficacy compared to monotherapy in lung cancer xenograft mice.

Background Immune checkpoint blockers (ICBs) are characterized by a durable clinical response and better tolerability in patients with a variety of advanced solid tumors. However, we not infrequently encounter patients with hyperprogressive disease (HPD) exhibiting paradoxically accelerated tumor growth with poor outcomes. This study aimed to determine differences in factors among patients with non-small cell lung cancer (NSCLC) displaying different tumor responses to immunotherapy. Methods This study evaluated 231 NSCLC patients receiving ICBs between January 2014 and May 2018. HPD was defined as a > 2-fold tumor growth kinetics ratio during ICB therapy and time-to-treatment failure of ≤ 2 months. We analyzed clinical data, imaging studies, periodic serologic indexes, and immune cell compositions in tumors and stromata using multiplex immunohistochemistry. Results Twenty-six patients (11.3%) met the HPD criteria. HPD was more frequent in patients with oncogenic driver mutations, ≥ 3 metastatic sites, ≥ 4 prior systemic treatments, and low PD-L1 expression (

Background Isoform 2 of the tight junction protein claudin-18 (CLDN18.2) is a potential target of gastric cancer treatment. A treatment targeting CLDN18.2 has shown promising results in gastric cancer. We investigated the clinical significance of CLDN18.2 and other cell-adherens junction molecules (Rho GTPase-activating protein [RhoGAP] and E-cadherin) in metastatic diffuse-type gastric cancer (mDGC).Methods We evaluated CLDN18.2, RhoGAP, and E-cadherin expression by performing two-plex immunofluorescence and quantitative data analysis of H-scores of 77 consecutive mDGC patients who received first-line, platinum-based chemotherapy between March 2015 and February 2017.Results CLDN18.2 and E-cadherin expression was significantly reduced in patients with peritoneal metastasis (PM) compared with those without PM at the time of diagnosis (P=0.010 and 0.013, respectively), while it was significantly higher in patients who never exhibited PM from diagnosis to death than in those who did (p=0.001 and 0.003, respectively). Meanwhile, CLDN18.2 and E-cadherin were expressed at significantly higher levels in patients with bone metastasis than in those without it (p=0.010 and 0.001, respectively). We identified a positive correlation between the expression of CLDN18.2 and E-cadherin (p < 0.001), RhoGAP and CLDN18.2 (p=0.004), and RhoGAP and E-cadherin (p=0.001). CDLN18.2, RhoGAP, and E-cadherin expression was not associated with chemotherapy response and survival.Conclusions CLDN18.2 expression was reduced in PM but significantly intact in bone metastasis. Furthermore, a positive correlation was identified between the expression of CLDN18.2 and other adherens junction molecules, which has clinical implications for mDGC and PM pathogenesis.

Epidermal growth factor receptor (EGFR) mutation testing is essential for individualized treatment using tyrosine kinase inhibitors. We evaluated two EGFR mutation tests, cobas v2 and PANAMutyper, for detection of EGFR activating mutations Ex19del, L858R, and T790M in tumor tissue and plasma from 244 non-small cell lung cancer (NSCLC) patients. The Kappa coefficient (95% CI) between the tests was 0.82 (0.74−0.92) in tumor samples (suggesting almost perfect agreement) and 0.69 (0.54−0.84) in plasma (suggesting substantial agreement). In plasma samples, both tests showed low to moderate sensitivity depending on disease stage but high diagnostic precision (86%−100%) in all disease stages (sensitivity: percentage of mutations in tumors that are also detected in plasma; precision: percentage of mutations in plasma which are also detected in tumors). Among the 244 patients, those previously diagnosed as T790M carriers who received osimertinib treatment showed dramatically better clinical outcomes than T790M carriers without osimertinib treatment. Taken together, our study supports interchangeable use of cobas v2 and PANAMutyper in tumor and plasma EGFR testing. Both tests have high diagnostic precision in plasma but are particularly valuable in late-stage disease. Our clinical data in T790M carriers strongly support the clinical benefits of osimertinib treatment guided by both EGFR mutation tests.

Background We screened resected tumor tissues from patients with lung cancer for EGFR mutations, ALK rearrangements, and rearranged during transfection (RET) gene variants (including RET rearrangements and the Kinesin Family Member 5B (KIF5B)-RET fusion gene) using various methods including reverse transcription polymerase chain reaction (RT-PCR), transcript assays, fluorescence in situ hybridization (FISH), and immunohistochemistry (IHC). We also examined the protein expression of associated downstream signaling molecules to assess the effect of these variants on patient outcome. Method We constructed a tissue microarray (TMA) comprising 581 resected tumor tissues from patients with lung adenocarcinoma and analyzed the microarray by both FISH (using RET break-apart and KIF5B-RET SY translocation probes) and a commercial RET transcript assay. We evaluated the expression of RET and RET-related signaling molecules, including p-AKT and p-ERK, by TMA -based IHC staining. Results Among the 581 specimens, 51 (8.8%) specimens harbored RET rearrangements, including 12 cases (2.1%) carrying a KIF5B-RET fusion gene. Surprisingly, RET expression was lower in KIF5B-RET fusion gene-positive than in RET wild-type specimens. We detected activating EGFR mutations in 11 (21.6%) of the 51 RET variant-positive specimens. Among the KIF5B-RET fusion gene-positive specimens, p-ERK expression was significantly lower in the EGFR mutation subgroup showing RET expression than in the EGFR mutation subgroup that did not express RET. Similarly, the RET rearrangement group showed significant variation in the expression level of p-AKT (P = 0.028) and p-ERK, whose expression remarkably increased in specimens not expressing RET. The expression of p-ERK markedly increased in the RET rearrangement group regardless of RET expression. Conclusion This result suggests that a combination of RET and ERK inhibitors may be an effective treatment strategy for lung adenocarcinoma patients harboring RET variants.

Background: To evaluate the anti-tumor efficacy of CKD-516 in combined with irradiation (IR) and examine tumor necrosis, delayed tumor growth, and expression of molecules involved in hypoxia and angiogenesis. Methods : A xenograft mouse model of lung cancer was established. The tumor was exposed to irradiation (IR) for 5 days per week. CKD-516 was administered with ​​two treatment schedules (day 1 or days 1 and 5) at one hour after IR. After the administration, tumor tissues were stained with hematoxylin and eosin and pimonidazole. HIF1 , Glut-1, VEGF, CD31 and Ki-67 expression were evaluated by Immunohistochemical staining. Results: With short-term administration, IR and CKD-516+IR (d1) significantly reduced tumor size ( p = 0.0062 and p = 0.0051, respectively). CKD-516+IR groups were remarkably reduced blood vessels ( p < 0.005). In particular, CKD-516+IR (d1) resulted in the most extensive tumor necrosis, which was significantly increased with large hypoxic area ( p = 0.02) and decreased HIF1 , Glut-1, VEGF, and Ki-67 expressions. Long-term administration of CKD-516+IR reduced tumor size and delayed tumor growth. This combination also greatly reduced the number of blood vessels ( p = 0.0006) and significantly enhanced tumor necrosis ( p = 0.004). CKD-516+IR notably increased HIF1 expression ( p = 0.0047), but significantly diminished VEGF expression ( p = 0.0046). Conclusion: Taken together, our results demonstrate that CKD-516 in combination with IR can significantly enhance the anti-tumor efficacy compared to CKD-516 or IR alone in lung cancer xenograft mice. Keywords: Irradiation, Tumor necrosis, Hypoxia, Squamous cell carcinoma of lung, Xenograft mice

Background Atypical hemolytic uremic syndrome (aHUS) involves dysregulation of the complement system, but whether this also occurs in thrombotic thrombocytopenic purpura (TTP) remains unclear. Although these conditions are difficult to differentiate clinically, TTP can be distinguished by low (

Simple Summary In this study, Sel-CapTM, a next-generation sequencing (NGS)-based genotyping platform, showed high sensitivity for detection of epidermal growth factor receptor (EGFR) gene mutations in plasma samples collected from 185 patients with non-small cell lung cancer (NSCLC). In the early-stage NSCLC, Sel-Cap liquid biopsy was able to detect more than half the EGFR mutations, which were detected in tumor tissue (sensitivity: 50% and 78% for Ex19del and L858R respectively, with tumor results as the references), while the conventional NGS could not detect any. Sel-Cap liquid biopsy was particularly sensitive for resistant mutation T790M (sensitivity: 88%). In addition, we conducted a retrospective study to monitor T790M using Sel-Cap in 34 patients who progressed on first-line tyrosine kinase inhibitors (EGFR-TKIs). The study suggested that the first appearance of T790M in plasma, ranging from at treatment baseline to over three years post-EGFR-TKI initiation, may be useful for prediction of disease progression (around 5 months in advance). Abstract Sel-CapTM, a digital enrichment next-generation sequencing (NGS)-based cancer panel, was assessed for detection of epidermal growth factor receptor (EGFR) gene mutations in plasma for non-small cell lung cancer (NSCLC), and for application in monitoring EGFR resistance mutation T790M in plasma following first-line EGFR-tyrosine kinase inhibitor (EGFR-TKI) treatment. Using Sel-Cap, we genotyped plasma samples collected from 185 patients for mutations Ex19del, L858R, and T790M, and compared results to those of PNAclampTM tumor biopsy (reference method, a peptide nucleic acid-mediated polymerase chain reaction clamping) and two other NGS liquid biopsies. Over two-thirds of activating mutations (Ex19del and L858R), previously confirmed by PNAclamp, were detected by Sel-Cap, which is 4–5 times more sensitive than NGS liquid biopsy. Sel-Cap showed particularly high sensitivity for T790M (88%) and for early-stage plasma samples. The relationship between initial T790M detection in plasma and progression-free survival (PFS) following first-line EGFR-TKIs was evaluated in 34 patients. Patients with T790M detected at treatment initiation (±3 months) had significantly shorter PFS than patients where T790M was first detected >3 months post treatment initiation (median PFS: 5.9 vs. 26.5 months; p < 0.0001). However, time from T790M detection to disease progression was not significantly different between the two groups (median around 5 months). In conclusion, Sel-Cap is a highly sensitive platform for EGFR mutations in plasma, and the timing of the first appearance of T790M in plasma, determined via highly sensitive liquid biopsies, may be useful for prediction of disease progression of NSCLC, around 5 months in advance.

Background: c-MET is co-expressed with EGFR in approximately 70% of EGFR mutant-positive tumors. In addition, c-Met amplification was found in 10-20% of lung cancer patients with acquired tolerance to EGFR TKI, and c-Met overexpression was found in 14-69% of patients receiving EGFR TKI. CKD-702, a double antibody, is designed to simultaneously block both EGFR and c-MET expression. Purpose: We evaluated the anti-tumor efficacy of CKD-702 with PDX model derived from lung adenocarcinoma patients. Methods: We constructed the PDX model by transplanting surgical tissue from lung adenocarcinoma patient into NOD/SCID mice after the approval of research protocol in Seoul St. Mary's Hospital IRB. We confirmed the co-existence of activating EGFR mutation and c-MET amplification with PNAclamp and FISH methods. We also checked overexpression of EGFR and c-MET protein by immunohistochemistry. Results: The characteristics of tumor tissue transplanted for PDX model were: 1) Korean, lung cancer with EGFR TKI chemotherapy failure confirmed c-MET amplification (> 5 copies) and overexpression (IHC score 3+), 19del Positive EGFR mutation, 2) European lung adenocarcinoma, c-MET amplification (> 2.5 copy) and overexpression (IHC score 3+) were confirmed, and L858R EGFR mutation positive tumor tumor confirmed. The anti-cancer efficacy of CKD-702 was reduced by more than 80% compared to the tumor size of the control group regardless of the concentration. In the CKD-702 20mg/kg group, there was no significant difference in the change in the overall tumor size until the time of disappearing or ending during drug administration. The %TGI was 93.6 and 97.5 according to the concentration of CKD-702, and the % TGD was 118.5 and 311.8, respectively. CKD-702 has a significantly higher tumor growth inhibitory effect regardless of the concentration, but the tumor growth retardation effect is 2.6 times different depending on the concentration. % TGI of CKD-702 20mg / kg group was similar to Hu8c4 group or Hu8c4 + Vectibix group. However, % TGD was 1.4 times higher than Hu8c4 group efficacy and 3.1 times higher than Hu8c4 + Vectibix group% TGD. Even after the completion of drug administration, tumor still remained at 47 days and 67 days in the CKD-702 10 mg/kg and 20 mg/kg groups, respectively. Although c-MET protein expression decreased during the administration of CKD-702, it was confirmed that c-MET protein expression increased again in tumor tissue at the time of tumor growth. Although the tumor size was rather large (605.6 ~ 816.2mm3), CKD-702 showed twice the tumor growth inhibition effect (% TGI: 32.8 vs. 68.1) and the tumor growth delay effect was 4.4 times higher depending on the drug concentration (% TGI: 2.8 vs 12.3). Conclusion: Our data showed that CKD-702 have effective anti-tumor activity in EGFR TKI-resistant PDX model harboring both activating EGFR mutation and c-MET overexpression. Citation Format: Jung-Young Shin, Min-Young Kim, Mi-Ran Lee, Jeong-Oh Kim, Eun-Ju Jeon, Ji Hye Choi, Soon-Ki Hong, Kyu-Jin Park, Jin Hyoung Kang. Anti-tumor efficacy of CKD-702 in EGFR TKI-resistant patient-derived xenograft model [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3046.

To better understand the immune profile within the tumor microenvironment (TME) and the potential immune-evasion capacity that may arise from administration of immune checkpoint inhibitors, we investigated immunological phenotypes and their correlations with genomic alterations in order to provide predictive/prognostic implications in lung adenocarcinoma (LAC).Surgical sections with corresponding peripheral blood mononuclear cells were collected from 76 patients with pathologically confirmed Ia-IIIa. The histological contents expressing immune checkpoint molecules in the tumor nest and stromal area were imaged and quantified using a multi-spectral imaging system. Distribution of tumor mutation burden (TMB) and non-synonymous somatic mutations were analyzed in whole exome sequencing (WES) data.Total 76 patients consisted of EGFR-mutant (n=37) and EGFR-wild type (n=39) tumors. As is well known, the number of tumor-infiltrating cytotoxic T cells (174.7 ± 24.4 cells/mm2) and TMB (3.8 ± 0.4) in EGFR-mutant tumors was significantly lower than those of EGFR wild type tumors (394.2 ± 82.3 cells/mm2 and 6.7 ± 1.2) (p=0.0146 and p=0.0254, respectively), elucidating EGFR mutation-mediated immune evasion. The subjects carrying EGFR wild-type were classified into two subgroups by the density of tumor-infiltrating cytotoxic T cells, defined as the high TIL (n=20) and low TIL (n=19). There was great difference of mean density of cytotoxic T cells between low TIL group (129.6 ± 16.8 cells/mm2) and high TIL group (672.8±143.5 cells/mm2). Tumor-infiltrating T cells were granzyme B-positive, suggesting most cells were tumor-specific. Interestingly, no significant difference in TMB was found between the two subgroups (7.5 ± 1.9 in low TIL group vs. 5.8±1.3 in high TIL group; p=0.4642). Tumor proportion score (TPS) of PD-L1 was remarkably higher in high TIL group than low TIL group (29.7 ± 6.5% vs. 13.9 ± 2.8%, p=0.0008, respectively), but no significant differences of tumoral CD73 and IDO between the two groups (p=0.643 and p=0.923, respectively). Even there were no significant differences in counts of tumor-infiltrating macrophages in all types of macrophage, significantly higher counts of PD-L1+ M1-type macrophages (CD68+ CD163low) in the stromal area were discovered in the high TIL group compared to those of the low TIL group (396.2 ± 93.6 vs. 139.8 ± 30.1, p=0.0114, respectively), indicating that most of macrophages resided in the stromal area. Interestingly, the low TIL group exhibited higher counts of CD73+ M2-type macrophages (CD68+ CD163high) in the stromal area than the high TIL group (644.5 ± 110.9 vs. 387.9 ± 73.2, p=0.0639, respectively). Cell count ratio (M1/M2) of macrophages was significantly higher in the high TIL group (p=0.0042), suggesting that the TME with a smaller number of TILs tends to contain more M2-type macrophages. The tumor recurrence was more frequent in the low TIL group compared to the high TIL group with no statistical significance (p=0.127). In surgically resected lung adenocarcinoma patients, the subgroup with a low density of tumor-infiltrating cytotoxic T cells had a large number of CD73+ M2-type macrophages, which may contribute to poor prognosis. Acknowledgement: This study was sponsored by Ono Pharma Co., Ltd Citation Format: Jae-Hwan Kim, Tae-Min Kim, Sung-Gu Her, Kyoung-Ho Pyo, Jeong- Oh Kim, Beung-Chul Ahn, Byoung Chul Cho, Jin Hyoung Kang. Comprehensive analysis of immune phenotypes identifies CD73+ CD163high macrophages as a prognostic biomarker in lung adenocarcinoma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4981.

Numerous clustering schemes have been proposed to increase energy efficiency in wireless sensor networks. Clustering schemes consist of a hierarchical structure in the sensor network to aggregate and transmit data. However, existing clustering schemes are not suitable for use in wireless multimedia sensor networks because they consume a large quantity of energy and have extremely short lifetime. To address this problem, we propose the Energy-Aware Cooperative Communication (EACC) method which is a novel cooperative clustering method that systematically adapts to various types of multimedia data including images and video. An evaluation of its performance shows that the proposed method is up to 2.5 times more energy-efficient than the existing clustering schemes.

Purpose Dose-limiting toxicities of docetaxel are widely considered to be neutropenia, anemia, skin toxicity, and nausea. One of the factors that limit the use of docetaxel is its unpredictability of inter-individual variation in toxicity. Materials and methods In order to identify the genetic factors that affect the risk of docetaxel-induced toxicities, we recruited patients who received docetaxel chemotherapy. We genotyped 92 patients with single-nucleotide polymorphisms (SNPs) in 5 genes: CYP3A4 (CYP3A4(*)1B, CYP3A4(*)18, and CYP3A4(*)3), CYP3A5 (CYP3A5(*)2 and CYP3A5(*)3), ABCB1 (C1236T, G2677G/T, and C3435T), SLCO1B3 (rs11045585), and ABCC2 (rs12762549). Results Out of 92 patients, 70 had grade 3 or 4 neutropenia; 4 had grade 1 or 2; and 18 had no toxicity (76.1%, 4.3%, and 19.6%, respectively). The findings of the SNP analysis showed that patients with TT genotype of ABCB1 3435C>T polymorphism showed significantly higher risk of neutropenia and anemia (p=0.029 and p=0.044, respectively). There were significant associations between docetaxel-induced leucopenia and 2677G/T of ABCB1 and rs12762549 of ABCC2 (p=0.025 and p=0.028, respectively). In a multivariate analysis, we observed that patients carrying 2677G>T in ABCB1might be associated with higher risk of chemo-resistance when treated with docetaxel (odds ratio [OR], 6.48; confidence interval, 1.92 to 21.94; p=0.003). In a subgroup analysis of non-small cell lung cancer patients, a significant association of tumor response with G2677T/A (OR, 4.54) in ABCB1 and SLCO1B3 (OR, 9.44) was observed. Conclusion Our data suggest that ABCB1 (2677G/T) and SLCO1B3 (rs11055585) might be major genetic predictors of docetaxel-related toxicities in patients receiving docetaxel chemotherapy.

Epilepsy is the most prevalent chronic neurological disorder and can be controlled by antiepileptic drugs (AEDs) in up to 70% of patients. We performed an association study between adverse drug reactions and the genetic polymorphisms of CYP2C9, CYP2C19, ABCB1, and SCN1A. The clinical data of 83 epilepsy patients who had received AEDs containing carbamazepine (CBZ) were collected. We extracted genomic DNA from peripheral blood and then genotyped CYP2C9 (CYP2C9*2, CYP2C9*3), CYP2C19 (CYP2C19*2, CYP2C19*3), ABCB1 (C3435T), and SCN1A (IVS5N+5 G>A) using direct sequencing. The allele frequencies of CYP2C9*3, CYP2C19*2, CYP2C19*3, ABCB1 (3435C>T), and SCN1A (IVS5N+5 G>A) were 0.93, 0.72, 0.91, 0.61, and 0.55, respectively. Statistically significant differences were indicated from the data obtained. Patients with SCN1A genotype CC or CT were compared with patients with SCN1A genotype TT while using more than 500mg of carbamazepine. We have associated functional polymorphisms with the dose used in regular clinical practice for Korean epilepsy patients who had received antiepileptic drugs (AEDs) containing carbamazepine. For AEDs, we found that one of the SCN1A genotypes is associated with a 500 mg dose. There was no association found with CNS ADR caused by AEDs.

Background: Mechanisms of acquired resistance for EGFR tyrosine kinase inhibitor (TKI) in non-small cell lung cancer (NSCLC) has been studied broadly, and T790M mutation, bypass signaling, and phenotype change have been suggested. Questions remain in terms of the selection of sub-clones according to the sites during EGFR TKI therapy, especially in brain due to blood-brain barrier. Three metastatic NSCLC patients harboring sensitive EGFR mutation were treated with EGFT TKI with good response but became refractory and symptomatic brain oligo-metastasis occurred. Metastectomy was done and next-generation sequencing (NGS) was done with the brain tissue and blood. Methods: Fresh brain metastasis tissue was obtained during surgery and frozen in liquid nitrogen. Blood sampling was done within a week of the surgery and plasma was used for NGS. Amplicon sequencing (Thunderbolt) was performed. Library preparation and quality control sequencing amplicon libraries were prepared using ThunderBolts Cancer Panel, according to the manufacturer's instructions. The ThunderBolts Cancer Panel uses 230 amplicons to interrogate mutations and hotspots in 50 oncogenes, tumor suppressors, and drug resistance markers. Results: All 3 brain tissues had the same sensitive EGFR mutation (exon 19 deletion) as initial presentation, with PDGFRA, APC and TP53 mutations, while in the plasma, PDGFRA and APC mutations were identified in the 3 patients. From case 1, PDGFRA, APC, MLH1, and EGFR mutations were identified in the plasma, and additional PIK3CA, KDR, TP53, ERBB2, and GNA11 mutations were found in the brain. From case 2, PDGFRA, APC, EGFR, MET, PTEN, HRAS, and TP53 mutations were found in the plasma, and in the brain, PIK3CA, KDR, and ERBB2 mutations were identified in addition. From case 3, PDGFRA, APC, HRAS, and TP53 mutations in the plasma, and additional EGFR mutation in the brain. Conclusions: Our data identified several additional mutations in brain metastasis compared to the plasma from patients with acquired EGFR TKI resistance, other than T790M mutation. Every mutation in the blood sample was identified in the brain tissue, while additional mutations were found in the brain tissue. Citation Format: Hee Yeon Lee, Jeong-Oh Kim, Jin-Hyoung Kang. Comparison of next-generation sequencing results between brain metastasis and blood in NSCLC patients with acquired EGFR-TKI resistance other than T790M mutation [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr A131.

Purpose Matrix metalloproteinase (MMP) and tissue inhibitor of matrix metalloproteinase (TIMP) play an important role in tumor invasion and progression. The aim of this study was to evaluate the prognostic role of MMP-1 and TIMP-1 in non-small cell lung cancer (NSCLC). To find out a potential serum biomarker, tissue and serum levels were investigated together. Patients and methods For 85 surgically resected NSCLC patients who had pre-operative serum samplings, MMP-1 and TIMP-1 expression were investigated using immunohistochemistry in tumor tissue and ELISA in serum. Tumor cells and surrounding stromal cells were assessed separately. Results Higher expression of MMP-1 in tumor cells comparing to stromal cells was related to male gender (P = 0.006), ever smoker (P = 0.004), and pooly differentiated tumor (P = 0.043). For TIMP-1, adenocarcinoma showed higher tumor cell expression, while squamous cell carcinoma showed higher stromal expression (P = 0.007). Patients with high carcinoembryonic antigen (CEA) level, the presence of vascular invasion, recurrence or death showed higher serum MMP-1 level. There was no correlation between the tissue and serum levels of MMP-1 and TIMP-1. A tumor/stroma TIMP-1 intensity ratio ≥1 was strongly associated with early recurrence in multivariate analysis (hazard ratio = 280.55, 95% confidence intervals; 11.12–7080.45; P = 0.001). High serum MMP-1 (≥3,500 pg/ml) showed a trend for short overall survival (P = 0.080). When serum MMP-1 was combined with CEA level or presence of vascular invasion, its prognostic implication was statistically significant (P = 0.045 and P = 0.015, respectively). Conclusion The tumor/stroma TIMP-1 intensity ratio in tissue is useful to predict tumor recurrence. Serum MMP-1 level showed a possibility as a prognostic biomarker.

e20610 Background: RET rearrangements have been identified in 1-2% of lung adenocarcinomas. The most common fusion is the KIF5B-RET, the function and roles of the RET fusion oncogene, and its downstream signaling molecules remain unclear. Methods: We constructed a tissue microarray (TMA) comprising 581 resected tumor tissues from lung adenocarcinoma patients and investigated them using FISH with RET break-apart and KIF5B-RET SY translocation probes. NanoString’s nCounter technology was used to assay RETtranscripts. We evaluated the protein expressions of RET and RET-related signaling molecules, including p-AKT and p-ERK, using TMA-based IHC staining. Results: Using FISH, we identified 51 cases (8.8%) of RET variants and 10 cases (1.7%) of KIF5B-RET fusion genes among the 581 cases. RET protein expression was lower in the group harboring KIF5B-RET fusion gene than that in the group harboring a wild type RET gene. We found the activating EGFR mutations in 11 (21.6%) cases of 51 RET variants. For the group with KIF5B-RET fusion gene, the expression of p-ERK was significantly lower in EGFR mutation subgroup with presence of RET protein compared to EGFR mutation subgroup with absence of RET protein. For the group with RET rearrangement, there were significant differences in the expression level of p-AKT (P = 0.028) and, p-ERK protein expression was remarkably increased, especially in cases with no RET protein expression. Conclusions: Taken together, the expression of p-ERK protein was meaningfully increased in the RET variants group regardless of RET protein expression. This result suggests that RET inhibitors combined with ERK inhibitors may be an effective treatment strategy for lung adenocarcinoma patients harboring the RET variants.

e20017Background: Activating or resistant somatic mutations (19 Del, L858R, T790M) occurring in the specific sites of EGFR gene are the powerful predictive biomarker for EGFR tyrosine kinase inhibi...

Studies on non-small cell lung cancer (NSCLC) patients with KRAS or BRAF mutations are urgently needed to improve clinical outcomes. We evaluated the cytotoxicities of paclitaxel and sorafenib alone and in combination in NSCLC cell lines with KRAS or BRAF mutations and investigated the mechanism of the interaction between the drugs. We found synergistic antitumor efficacy with paclitaxel followed by sorafenib in in vitro and in vivo models of NSCLC. And, we determined that downregulation of the phosphorylated ERK and Rb, and Mcl-1 plays a critical role in the synergistic activity of the drugs. Further clinical trials are needed to verify the antitumor efficacy of this combination.

Warfarin is a commonly prescribed anticoagulant drug for the prevention of thromboembolic disorders. We investigated the contribution of genetic variations of four genes and clinical factors to warfarin dose requirement and provided a warfarin-dosing algorithm based on genetic and clinical variables in Korean patients. We recruited 564 Korean patients on stable anticoagulation. Single nucleotide polymorphisms (SNPs) for the VKORC1, CYP2C9, CYP4F2 and GGCX were analyzed. Using multiple regression analysis, we developed a model to predict the warfarin requirement. The SNPs of VKORC1, CYP2C9, CYP4F2 and GGCX showed significant correlation with warfarin dose. Patients with the 3730AA genotype received significantly higher doses of warfarin than those with the 3730GG (P=0.0001). For CYP2C9, the highest maintenance dose was observed in the patients with wild-type genotype compared with the variant allele carriers (P

Introduction: The interaction between tumor cells and tumor micro-environment (TME) plays important role in tumor initiation, progression, treatment resistance, and prognosis in solid tumors including non-small cell lung cancer (NSCLC). The aim of this study is to evaluate the prognostic role of TME-related molecules in NSCLC patients who received surgical resection. To find out a possibility of a potential serum biomarker, protein expression in tissue from surgical specimen and pre-operative serum were investigated together. Methods: For 85 NSCLC patients who underwent surgical resection from August 2006 to September 2010 and had pre-operative serum samplings, we screened the serum levels of 60 TME-related molecules with angiogenesis array kit for 8 patients who had recurrences and 4 patients who did not. For matrix metalloproteinase (MMP)-1 and tissue inhibitor of matrix metalloproteinase (TIMP)-1 in which significant difference was observed between recurrence and recurrence-free groups, protein expressions were investigated using ELISA in serum and immunohistochemistry in tumor tissue. Results: High serum MMP-1 was related to carcinoembryonic antigen (CEA) level≥5 at diagnosis (P = 0.034), higher frequency of the presence of vascular invasion (P = 0.041). High expression of MMP-1 in tumor cells (intensity score≥2) was related to tumor differentiation (P = 0.057), whereas high MMP-1 expression in stroma was significantly associated with lymph node metastasis and TNM stage (P = 0.021 and P = 0.022, respectively). In the case of TIMP-1, expression in tumor cells was higher for adenocarcinoma but higher in stromal cells for squamous cell carcinoma (P = 0.046). There was no correlation between the tissue and serum levels of MMP-1 and TIMP-1. High serum MMP-1 showed a trend for short overall survival (P = 0.080). When serum MMP-1 was combined with CEA level or presence of vascular invasion, the prognostic implication for overall survival was statistically significant. Neither tumor nor stromal positivity for MMP-1 and TIMP-1 was associated with survival outcome; however, a tumor/stroma TIMP-1 intensity ratio≥1 was strongly associated with early recurrence in multivariate analysis (HR = 13.1, P = 0.008). Conclusion: In surgically resected NSCLC patients, serum MMP-1 showed a possibility as a prognostic biomarker. In addition, the tumor/stroma TIMP-1 ratio (≥1) in tissue may be a useful to predict tumor recurrence. Citation Format: Ho Jung An, Jin-Hyoung Kang, Yoon-Jin Lee, Soon Uk Hong, Kyo Young Lee, Jeong-Oh Kim. The clinical implication of MMP-1 and TIMP-1 on prognosis in patients with non-small cell lung cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1547. doi:10.1158/1538-7445.AM2015-1547

7523 Background: To assess the role of mutant EGFR and C-MET in survival benefit obtained from platinum-based adjuvant chemotherapy (PBAC) among resected lung adenocarcinoma (RLADC) patients. Metho...

1540 Background: Activating EGFR mutations are strongly predictive of EGFR tyrosine kinase inhibitor (TKI) activity in NSCLCs, however, some patients having wild type EGFRdo not infrequently show clinically favorable outcome to EGFR TKIs. The key proapoptotic proteins have been reported to be involved in TKI-induced cell death. The up-regulation of BIM, a pro-apoptotic protein of BCL2 family, is required for TKIs to induce apoptosis in cancers. Methods: For 150 advanced or metastatic NSCLC pts. receiving gefitinib or erlotinib, we assayed mutation status of EGFR in paraffin embedded tumor tissue using PNA clamping (PANAGENE, Korea) and direct sequencing. Six different genetic variants of EGFR (-191C>A, -216 G>T, CA repeat number, R497K, 2607G>A, and D994D) and BIMdeletion in genomic DNA extracted from peripheral blood were analyzed. Results: M:F ratio was 72:78 and median age was 63 (35~82). Histological subtypes are as follows: 131 adenocarcinoma (87.3%), 16 squamous cell ca. (10.7%) and 3 large cell ca. (1.3%). Objective tumor response (CR+PR) was observed in 60 pts. (40.0%) and SD in 44 pts. (29.3%). Median PFS and OS were 4.3 (0.4~67.4) and 18.6 (0.9~78.1) months, respectively. Of 129 pts. in whom EGFR mutation could be analyzed, 41 pts. (31.8%) had activating EGFR mutation. In univariate analysis, only gender was associated with objective response to EGFR TKI in activating EGFR mutant group (P=0.048). In wild type EGFR group (n=88), R497K (P=0.04) and BIM (P=0.04) as well as gender (P=0.05) were significantly correlated with objective response. In addition, multivariate analysis demonstrated that gender, R497K and BIM were associated with objective response to EGFR TKIs in wild type EGFR group with statistical significance (P=0.023, 0.027 and 0.018, repectively). Neither six EGFR polymorphisms nor BIM was correlated with PFS and OS in activating EGFR mutant group. Meanwhile, in wild type EGFR group, R497K was significantly associated with OS (P=0.028). Conclusions: Taken together, our data suggest that R497K of EGFR and BIM deletion polymorphisms are useful pharmacogenetic biomarkers to predict objective tumor response for EGFR TKIs in advanced NSCLC pts. harboing wild type EGFR.

Background: This study is desiged to evaluate biomarkers that are correlated with the efficacy of pemetrexed in patients with non-small-cell lung cancer (NSCLC). Methods: Patients with stage III or IV NSCLC who received pemetrexed monotherapy after failure of one or more chemotherapy regimen between January 2003 and August 2011 were included. Polymorphism of reduced folate carrier gene (SLC19A, rs1051298), 5,10-methylenetetrahydroflate reductase (MTHFR, rs1801133) and thymidylate synthase (TS, rs45445694, rs16430) gene were analyzed by PCR and direct sequencing method using peripheral blood. Results: Total 123 patients were included. The median age was 61 years (range 35-82). Fifty eight (47.2%) patients were female. Predominant histology was adenocarcinoma (92.7%). Metastatic diseases were 114 (80.5%) patients. Patients received one or more lines of systemic therapy before pemetrexed therapy, one line in 53 (43.1%) patients, two in 51(41.5%), three in 14 (11.02%), and four in 4 (3.3%). Median 4 cycles of chemotherapy were given (range 1-44). Response rate was 22.7% (CR, n=2, PR, n=26) and stable disease was detected in 54 patients (43.9%). Median follow up duration was 28.7months (range 4.07-84.33months). Median progression free survival (PFS) was 4.2 months (95% CI: 2.9-5.6) and median overall survival (OS) was 18.9 months (95% CI: 13.3-24.6). Clinical factors like, sex, histology of tumor (adenocarcinoma vs, nonadenocarcinoma), differentiation, smoking status and EGFR mutation status (active mutation vs. other mutation vs. wild type) did not have any significant difference on response and PFS. Only polymorphisms of TS promoter enhancer region (TSER) were related with PFS. 2R/2R repeat in the TSER (2R/2R vs 2R/3R, 3R/3R 1.0m vs 4.3m, p Conclusions: This study suggests that polymorphisms of TSER associated PFS in pemetrexed treatment. Further prospective studies will be needed to validate these polymorphism as a useful biomarker of pemetrexed treatment in NSCLC patients. Citation Format: Hiun S. Chae, Yoon Ho Ko, Eun Kyoung Jeon, Sook Hee Hong, Jeong-Oh Kim, Seung Joon Kim, Kyo Young Lee, Young Kyoon Kim, HoonKyo Kim, Seok young Park, In Sook Woo, Jin Hyoung Kang. Predictive markers in patients with non-small cell lung cancer with pemetrexed treatment. [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 2372. doi:10.1158/1538-7445.AM2013-2372

e18050 Background: The EGFR mutation on particular exons has been known as a strong predictive biomarker to EGFR TKIs (tyrosine kinase inhibitors) in NSCLC pts. However, some pts having wild type EGFR do not infrequently show clinically favorable outcome to EGFR TKIs. Accordingly, we hypothesized that clinical outcome and skin toxicity to EGFR TKIs might be related to specific single nucleotide polymorphisms regulating the expression of EGFR gene as well as EGFR sensitive mutation. Methods: In 211 advanced or metastatic NSCLC pts receiving gefitinib or erlotinib, we assayed mutation status of EGFR in paraffin embedded tumor tissue using PNA clamping method and direct sequencing. Six different SNPs in genomic DNA extracted from peripheral blood were analyzed; promoter 191C>A, 216 G>T, CA repeat number, exon 13 R497K, exon 20 2607G>A, and exon 25 D994D. Results: M:F ratio was 106:105 and mean age was 63.2 (35.0-82.0). Histological subtypes are as follows: 175 adenocarcinoma, 32 squamous cell ca. and 3 large cell ca. Objective response (CR+PR) was observed in 78 pts (36.9%) and SD in 69 pts (32.7%). Median PFS and OS were 8.7 ± 11.3 and 15.9 ± 14.8 months, respectively. Of 167 pts in whom EGFR mutation was analyzed, 68 pts (40.7%) had EGFR harboring sensitive mutation. In R497K, wild type (RR) was 35 (21.0%) and K alleles (RK + KK) were 132 (79.0%). In D994D, wild type (GG) was 76 (45.5%) and heterozygote and homozygote variants (GA+AA) were 91(54.5%). Statistically significant differences of PFS and OS were observed between wild and hetero-/homozygote variants of R497K (11.9 m vs. 36.2 m, p=0.037 and 23.7 m vs. 39.1 m, p=0.054, respectively) in the pts harboring mutant type EGFR. Meanwhile, in the pts harboring wild type EGFR, GA+ AA genotype of D994D demonstrated much longer PFS and OS compared with GG genotype (PFS: 18.5 vs. 3.7 m, p=0.013 and 37.6 vs. 15.2 m, p=0.035, respectively). In addition, skin rash showed statistically significant association with R497K polymorphism (P=0.031). Conclusions: Our data suggest that two germline genetic variations of EGFR gene, R497K and D994D, be useful pharmacogenetic biomarker to predict longer PFS and OS for EGFR TKIs in advanced NSCLC pts harboring mutant type and wild type EGFR, respectively.

64 Background: Metastasis of cancer cells is associated with numerous activating molecules, including vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and tumor necrosis factor (TNF)-α. Excess VEGF production induces hematogenous metastasis in gastric cancer (GC) cells. To understand the anti-metastatic effect of bevacizumab, an anti-VEGF monoclonal antibody developed for selective inhibition of tumor angiogenesis, we investigated VEGF-induced expression of key adhesion molecules in human umbilical vein endothelial cells (HUVECs) and the inhibitory effect of bevacizumab on the adhesion of GC cells expressing high levels of VEGF. Methods: We measured the soluble VEGF (sVEGF) produced by 7 GC cells by ELISA. We evaluated intercellular adhesion molecule (ICAM)-1, vascular cell adhesion molecule (VCAM)-1, and E-selectin expression in HUVECs pretreated with SNU-1-conditioned medium (SNU-1 CM) or recombinant VEGF (rhVEGF) at different time points by western blotting. Results: We identified that SNU-1 cells secreted the highest sVEGF concentration in 7 GC cells. After rhVEGF pretreatment, ICAM-1 and E-selectin expression were highest at 4–6 h and 2 h, respectively, but VCAM-1 expression was unchanged. Bevacizumab cotreatment markedly decreased VCAM-1 and E-selectin expression to basal levels, whereas ICAM-1 expression was unaffected. The SNU-1 adherent cell percentage decreased following bevacizumab cotreatment of SNU-1 CM- or rhVEGF-pretreated HUVECs. Investigation of SNU-1-cell invasiveness through activated HUVECs revealed less than 10 invasive adherent cells, whereas no cells were found after bevacizumab cotreatment. Conclusions: Our preclinical data suggests that bevacizumab, might contribute to anti-metastasis by inhibiting SNU-1 cell adhesion to HUVECs by reducing E-selectin and VCAM-1 expression.

11077 Background: The aromatase inhibitors cause bone loss by estrogen depletion. Zoledronic acid (ZA) can prevent bone mineral density (BMD) loss associated with the use of aromatase inhibitors. Accordingly interest has arisen in measuring surrogate markers of bone resorption to monitor the response of treatment of BMD loss in place of radiologic assessment. This study was designed to determine whether ZA would prevent bone loss that is known to occur with letrozole and identified surrogate markers of bone resorption in an animal model. Methods: In ovariectomized or sham-operated rat, we administrated ZA and letrozole to 5 different groups including: a sham operation control group (OC), a group in which an ovariectomy was performed followed by saline administration (OS), an ovariectomy with ZA treatment group (OZ), an ovariectomy with letrozole treatment group (OL) and an; ovariectomy with ZA and letrozole combined treatment group (OZL). The levels of serum osteocalcin, serum bone alkaline phosphatase (BALP), serum calcium and urine N-telopeptide (NTX) and BMD were estimated and compared at the same periods for each group. The distinct microscopic findings of proximal tibia at week sixteen were also compared. Results: Significantly intense increment of BDM in the OZ group and the OZL group and lower increment in the OL group were valued in correlation to the OS group, 31.3%, 35.0%, 10.5% and 13.0% respectively. Serum osteocalcin levels and urine calcium levels were close to each group. Serum calcium levels and BALP levels were relatively lowering in the OZ group and the OZL group than other group. Meaningfully lower levels of urine NTX were measured in the OZ group and the OZL group, 6.6% and 16.5% respectively. In the OL group levels of urine NTX were rise of 55.9%. Less cancellous bone loss and increase bone trabeculae were noted in the microscopic findings of tibia. Conclusions: The combination treatment with ZA and letrozole are effective in the inhibition of bone resorption and in the maintenance of BMD. Measurement of serum osteocalcin, urine NTX, and BMD, levels are recommended as surrogate markers for determining the response for the treatment of bone loss. No significant financial relationships to disclose.