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Purpose To determine the efficacy of consolidation chemotherapy (CC) with docetaxel and cisplatin (DP) after concurrent chemoradiotherapy (CCRT) with the same agents in locally advanced non–small-cell lung cancer (LA-NSCLC). Patient and Methods Patients were randomly assigned to either CCRT alone (observation arm) or CCRT followed by CC (consolidation arm). CCRT with docetaxel (20 mg/m2) and cisplatin (20 mg/m2) was administered every week for 6 weeks with a total dose of 66 Gy of thoracic radiotherapy in 33 fractions. In the consolidation arm, patients were further treated with three cycles of DP (35 mg/m2 each on days 1 and 8, every 3 weeks). The primary end point was 40% improvement in progression-free survival (PFS) compared with observation. Results From October 2005 to April 2011, 437 patients were randomly assigned. Seventeen patients did not start CCRT as a result of consent withdrawal or ineligibility reasons after random assignment, leaving 420 patients for this analysis (n = 211 for observation; n = 209 for consolidation). Patient characteristics were similar in both arms. In the consolidation arm, 143 patients (68%) received CC, of whom 88 (62%) completed three planned cycles. The median PFS was 8.1 months in the observation arm and 9.1 months in the consolidation arm (hazard ratio, 0.91; 95% CI, 0.73 to 1.12; P = .36). Median overall survival times were 20.6 and 21.8 months in the observation and consolidation arms, respectively (HR, 0.91; 95% CI, 0.72 to 1.25; P = .44). Conclusion CC with DP after CCRT with weekly DP in LA-NSCLC failed to further prolong PFS. CCRT alone should remain the standard of care.

The clinical predictors of the survival benefit of epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitor (TKI) therapy for nonesmall-cell lung cancer (NSCLC) with EGFR-activating mutations have not been well elucidated. The present study determined the clinical predictors of outcome in 224 patients with EGFRmutant NSCLC treated with EGFR-TKIs. A large tumor burden at baseline and rapid progression were predictive of inferior survival. Background: Mutations in the epidermal growth factor receptor (EGFR) have been associated with a marked therapeutic response to EGFR-tyrosine kinase inhibitors (TKIs) in patients with advanced nonesmall-cell lung cancer (NSCLC). However, the clinical predictors of the survival benefit of EGFR-TKI therapy for NSCLC with EGFR-activating mutations have not been well elucidated. Therefore, the present study evaluated the clinical predictors of survival outcome in patients with EGFR-mutant NSCLC who had been treated with EGFR-TKIs. Material and Methods: The data from 224 patients with EGFR-mutant lung adenocarcinoma treated with EGFR-TKIs were retrospectively reviewed. The treatment outcomes were evaluated according to the clinical factors, number of metastasis sites, and progression patterns. Results: The clinical factors associated with reduced progression-free survival (PFS) and overall survival (OS) on univariate analysis were Eastern Cooperative Oncology Group (ECOG) performance status (PS) � 2, intra- and extrathoracic metastasis, extrathoracic metastasis, a high number of metastatic sites, metastasis to the liver or adrenal gland at baseline, and rapid progression at the diagnosis of progressive disease (PD). On multivariate analysis, the factors that remained significantly associated with a shorter PFS were ECOG PS � 2 (odds ratio [OR], 2.189; 95% confidence interval [CI], 1.374-3.437; P < .001) and rapid tumor progression at PD (OR, 1.800; 95% CI, 1.059-3.058; P ¼ .030). Conclusion: Thus, the tumor burden, expressed as the number of metastatic sites at EGFRTKI treatment, and rapid tumor progression at PD were predictive of inferior survival in patients with lung adenocarcinoma with activating EGFR mutations.

Purpose: The increasing prevalence of distant metastases from non–small cell lung cancer (NSCLC) indicates an urgent need for novel therapeutic modalities. Brain metastasis is particularly common in NSCLC, with severe adverse effects on clinical prognosis. Although the molecular heterogeneity of NSCLC and availability of various targeted agents suggest personalized therapeutic approaches for such brain metastases, further development of appropriate preclinical models is needed to validate the strategies. Experimental Design: We established patient-derived xenografts (PDX) using NSCLC brain metastasis surgical samples and elucidated their possible preclinical and clinical implications for personalized treatment. Results: NSCLC brain metastases (n = 34) showed a significantly higher successful PDX establishment rate than primary specimens (n = 64; 74% vs. 23%). PDXs derived from NSCLC brain metastases recapitulated the pathologic, genetic, and functional properties of corresponding parental tumors. Furthermore, tumor spheres established in vitro from the xenografts under serum-free conditions maintained their in vivo brain metastatic potential. Differential phenotypic and molecular responses to 20 targeted agents could subsequently be screened in vitro using these NSCLC PDXs derived from brain metastases. Although PDX establishment from primary NSCLCs was significantly influenced by histologic subtype, clinical aggressiveness, and genetic alteration status, the brain metastases exhibited consistently adequate in vivo tumor take rate and in vitro tumor sphere formation capacity, regardless of clinical and molecular conditions. Conclusions: Therefore, PDXs from NSCLC brain metastases may better represent the heterogeneous advanced NSCLC population and could be utilized as preclinical models to meet unmet clinical needs such as drug screening for personalized treatments. Clin Cancer Res; 21(5); 1172–82. ©2014 AACR.

BACKGROUND Dermatofibroma (DF) comprises a heterogeneous group of mesenchymal tumors, with fibroblastic and histiocytic elements present in varying proportions. The cell of origin of DF has been investigated, but remains unclear. OBJECTIVE The present study attempted to investigate the expression of leukocyte-specific protein 1 (LSP1), a marker of fibrocytes, in DF. Additionally, we evaluated the effectiveness of LSP1 in the differential diagnosis of DF from dermatofibrosarcoma protuberans (DFSP). METHODS Immunohistochemical staining was performed on 20 cases of DF using antibodies against LSP1, CD68, and factor XIIIa (FXIIIa). In addition, the expression of LSP1 and FXIIIa was evaluated in 20 cases of DFSP. RESULTS Eighteen of 20 cases (90%) of DF stained positive for LSP1, with variation in the intensity of expression. CD68 was positive in 10 cases (50%), and FXIIIa was expressed in all cases of DF. There were differences between the regional expression patterns of the three markers in individual tumors. In contrast, only 2 of 20 cases of DFSP expressed LSP1, and none of DFSP cases stained positive for FXIIIa. CONCLUSION The LSP1-positive cells in DF could potentially be fibrocyte-like cells. FXIIIa and CD68 expression suggests that dermal dendritic cells and histiocytes are constituent cells of DF. It is known that fibrocytes, dermal dendritic cells and histiocytes are all derived from CD14+ monocytes. Therefore, we suggest that DF may originate from CD14+ monocytes. Additionally, the LSP1 immunohistochemical stain could be useful in distinguishing between DF and DFSP.

Aim To investigate the effects of Tubastatin A, a selective histone deacetylase-6 inhibitor, on synovial inflammation and joint destruction in a collagen antibody-induced arthritis (CAIA) mouse model. Methods Collagen antibody-induced arthritis mice were given daily intraperitoneal injections of various concentrations of Tubastatin A (0, 10, 50, 100 mg/kg). The clinical score and paw thickness were measured. Mice were sacrificed on day 15, and the expression of tumor necrosis factor (TNF)-α, interleukin (IL)-1 and IL-6 in the serum were analyzed using enyme-linked immunosorbent assay (ELISA). Two pathologists independently measured the synovitis score. Micro-computed tomography (CT) scans of the joints were performed to quantify joint destruction. The expression of IL-6 from human fibroblast-like synoviocytes (FLSs) after incubation with various doses of Tubastatin A (0, 0.75, 1.5, 3 μmol/L) was measured using ELISA. Results The clinical arthritis score was significantly attenuated and paw thickness was lower in the group treated with 100 mg/kg Tubastatin A compared with those treated with vehicle alone. The synovitis score was significantly reduced in the 100 mg/kg Tubastatin A-treated group compared with the control group. Micro-CT showed that quantitative measures of joint destruction were significantly attenuated in the 100 mg/kg Tubastatin A-treated group compared with the control. The expression of IL-6 in the sera was lower in the mice treated with Tubastatin A compared with the control. The expression of IL-6 in human FLSs decreased dose-dependently after incubation with Tubastatin A without affecting cell viability. Conclusions Tubastatin A successfully ameliorated synovial inflammation and protected against joint destruction in CAIA mice, at least in part, by modulating IL-6 expression.

// Hyo Song Kim 1,* , Seung Eun Lee 2,* , Yoon Sung Bae 3,* , Dae Joon Kim 4 , Chang-Geol Lee 5 , Jin Hur 6 , Hyunsoo Chung 7 , Jun Chul Park 7 , Da Hyun Jung 7 , Sung Kwan Shin 7 , Sang Kil Lee 7 , Yong Chan Lee 7 , Hye Ryun Kim 1 , Yong Wha Moon 1 , Joo Hang Kim 1 , Young Mog Shim 8 , Susan S. Jewell 9 , Hyunki Kim 3 , Yoon-La Choi 2 and Byoung Chul Cho 1 1 Division of Medical Oncology, Department of Internal Medicine, Yonsei University College of Medicine, Seoul, Korea 2 Departments of Pathology, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea 3 Department of Pathology, Yonsei University College of Medicine, Seoul, Korea 4 Department of Thoracic and Cardiovascular Surgery, Yonsei University College of Medicine, Seoul, Korea 5 Department of Radiation Oncology, Yonsei University College of Medicine, Seoul, Republic of Korea 6 Department of Radiology, Yonsei University College of Medicine, Seoul, Republic of Korea 7 Division of Gastroenterology, Department of Internal Medicine, Yonsei University College of Medicine, Seoul, Korea 8 Department of Thoracic Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea 9 Abbott Molecular Laboratories, Des Plaines, IL * These authors contributed equally to this work as co-first authors Correspondence: Byoung Chul Cho, email: // Yoon-La Choi, email: // Hyunki Kim, email: // Keywords : Fibroblast growth factor receptor 1, esophageal squamous cell carcinoma, gene amplification, fluorescent in situ hybridization, prognostic factor Received : September 11, 2014 Accepted : December 09, 2014 Published : December 10, 2014 Abstract To investigate the frequency and the prognostic impact of fibroblast growth factor receptor 1 ( FGFR1 ) gene amplification in 526 curatively resected esophageal squamous cell carcinoma (ESCC). Using fluorescent in situ hybridization, high amplification was defined by an FGFR1 /centromer 8 ratio is ≥ 2.0, or average number of FGFR1 signals/tumor cell nucleus ≥ 6.0, or percentage of tumor cells containing ≥ 15 FGFR1 signals or large cluster in ≥ 10%. Low amplification was defined by ≥ 5 FGFR1 signals in ≥ 50%. FGFR2 and FGFR3 mutations were assessed by direct sequencing in 388 cases and no mutation was detected. High and low amplification were detected in 8.6% and 1.1%, respectively. High FGFR1 amplification had significantly shorter disease-free survival (34.0 vs 158.5 months P =0.019) and overall survival (52.2 vs not reached P =0.022) than low/no amplification group. After adjusting for sex, smoking, stage, histology, and adjuvant treatment, high FGFR1 amplification had a greater risk of recurrence (adjusted hazard ratio [AHR], 1.6; P =0.029) and death (AHR, 1.53; P =0.050). High amplification was significantly higher in current smokers than former and never-smokers ( P trend

Angiomyolipomas (AMLs) are relatively rare hamartomatous or benign tumors that occasionally occur as part of tuberous sclerosis complex (TSC). Mutations in either of the two genes, TSC 1 and TSC 2, have been attributed to the development of TSC. Between 1994 and January 2009, 83 patients were diagnosed with AML at the Samsung Medical Center. In that group of patients, 5 (6%) had AML with TSC (AML-TSC). Mutational analysis of the TSC 2 gene was performed using 7 samples from the 5 AML-TSC patients and 14 samples from 14 patients with sporadic AML without TSC (AML-non-TSC). From this analysis, mutations in TSC genes were identified in 5 samples from the AML-TSC patients (mutation detection rate = 71%) and 3 samples from AML-non-TSC patients (mutation detection rate = 21%). In the case of AML-TSC, 6 mutations were found including 3 recurrent mutations and 3 novel mutations, while in the case of AML-non-TSC, 4 mutations were identified once, including 1 novel mutation. Also MLPA analysis of the TSC2 gene showed that TSC2 exon deletion is more frequently observed in AML-TSC patients than in AML-non-TSC patients. This is the first mutation and multiplex ligation-dependent probe amplification (MLPA) analyses of TSC 2 in Korean AMLs that focus on TSC. This study provides data that are representative of the distribution of mutations and exon deletions at TSC genes in clinically diagnosed AML-TSC cases of the Korean population.

T790M mutation is most common resistant mechanism to epidermal growth factor receptor gene (EGFR) tyrosin kinase inhibitor (TKI). Several third-generation EGFR-mutant selective TKI, such as AZD9291 (AstraZeneca), Rociletinib (Clovis), or HM61713 (Hanmi) have been developed. Acquired resistant C797S mutation was known to be one of the resistance mechanisms of AZD9291, which has not been reported for HM61713 yet. This is the first case report of C797S mutation as resistance mechanism of HM61713.

Background Anaplastic lymphoma kinase tyrosine kinase inhibitors (ALK-TKIs) are usually effective in lung adenocarcinoma patients with anaplastic lymphoma kinase (ALK) rearrangement. However, even after a good response to ALK-TKI therapy, most patients acquire resistance to these agents. Histological transformation is one of several suggested mechanisms of acquired resistance to ALK-TKIs. The clinicopathologic features of four patients with ALK-expressing adenocarcinoma and neuroendocrine features were analyzed. Methods We selected combined neuroendocrine differentiation in pulmonary adenocarcinoma cases with positive ALK immunostaining. Neuroendocrine differentiation was confirmed by CD56 immunohistochemical stain. Additional ALK fluorescence in situ hybridization (FISH) study and epidermal growth factor receptor (EGFR) mutation tests were also performed. Results All four cases were positive for ALK immunohistochemistry and no EGFR mutations were detected. Interestingly, the results of ALK FISH assays showed rearrangement in only two cases. Three cases showed combined adenocarcinoma and neuroendocrine component without history of ALK-TKI administration; one of them was treated with crizotinib and experienced partial tumor regression. The remaining case had an adenocarcinoma at initial biopsy and she showed a partial response to crizotinib, and neuroendocrine changes were visible on second biopsy. Then she was treated with ceritinib and achieved a partial response. Conclusion We suggest that ALK-rearranged adenocarcinoma with combined neuroendocrine component is responsive to ALK-TKIs. Moreover, even after neuroendocrine transformation as a result of resistance to ALK-TKIs, the tumor may have partial response to second generation ALK-TKIs., Graphical Abstract

The purpose of this study is to analyze the clinical impact of radical nephrectomy on retroperitoneal liposarcoma near the kidney. Data of patients who underwent surgery for unilateral primary retroperitoneal liposarcoma near the kidney were retrospectively collected. Patients were divided into four groups according to whether they underwent nephrectomy and combined resection of other organs. Kaplan–Meier survival analysis was used to estimate disease-free survival and overall survival. Multivariable Cox analysis was used to analyze factors related to disease-free survival and overall survival. Nephrectomy (HR = 0.260, CI = 0.078–0.873, p = 0.029) had a beneficial effect on disease-free survival, while interaction model of nephrectomy*other organ resection (HR = 4.655, CI = 1.767–12.263, p = 0.002) showed poor disease-free survival. Other organ resection was not related to disease-free survival (HR = 1.543, CI = 0.146–16.251, p = 0.718). Operation method (p = 0.007) and FNCLCC grade (p

Background Patient-derived xenograft (PDX) models are important tools in precision medicine and for the development of targeted therapies to treat cancer patients. This study aimed to evaluate our precision medicine strategy that integrates genomic profiling and preclinical drug-screening platforms, in order to personalize cancer treatments using PDX models. Methods We performed array-comparative genomic hybridization, microarray, and targeted next-generation sequencing analyses, in order to determine the oncogenic driver mutations. PDX cells were obtained from PDXs and subsequently screened in vitro with 17 targeted agents. Results PDX tumors recapitulated the histopathologic and genetic features of the patient tumors. Among the samples from lung cancer patients that were molecularly-profiled, copy number analysis identified unique focal MET amplification in one sample, 033 T, without RTK/RAS/RAF oncogene mutations. Although HER2 amplification in 033 T was not detected in the cancer panel, the selection of HER2-amplified clones was found in PDXs and PDX cells. Additionally, MET and HER2 overexpression were found in patient tumors, PDXs, and PDX cells. Crizotinib or EGFR tyrosine kinase inhibitor treatments significantly inhibited cell growth and impaired tumor sphere formation in 033 T PDX cells. Conclusions We established PDX cell models using surgical samples from lung cancer patients, and investigated their preclinical and clinical implications for personalized targeted therapy. Additionally, we suggest that MET and EGFR inhibitor-based therapy can be used to treat MET and HER2-overexpressing lung cancers, without receptor tyrosine kinase /RAS/RAF pathway alterations. Electronic supplementary material The online version of this article (doi:10.1186/s12885-017-3525-9) contains supplementary material, which is available to authorized users.

Purpose ROS1 rearrangement is a distinct molecular subset of non–small-cell lung cancer (NSCLC). We investigated the efficacy and safety of ceritinib in patients with ROS1-rearranged NSCLC. Patients and Methods We enrolled 32 patients with advanced NSCLC who tested positive for ROS1 rearrangement by fluorescent in situ hybridization. Ceritinib 750 mg was administered once daily. The primary end point was objective response rate. The secondary end points were disease control rate; duration of response; progression-free survival; overall survival; toxicity; and concordance among fluorescent in situ hybridization, immunohistochemistry, and next-generation sequencing. Results Between June 7, 2013, and February 1, 2016, 404 patients underwent ROS1 prescreening, and 32 patients with ROS1 rearrangement were enrolled. All patients except two were crizotinib-naïve. At the time of data cutoff, the median follow-up was 14.0 months, and 18 patients (56%) had discontinued treatment. Of the 32 patients enrolled, 28 were evaluable for response by independent radiologic review. Objective response rate was 62% (95% CI, 45% to 77%), with one complete response and 19 partial responses; duration of response was 21.0 months (95% CI, 17 to 25 months); and disease control rate was 81% (95% CI, 65% to 91%). The median progression-free survival was 9.3 months (95% CI, 0 to 22 months) for all patients and 19.3 months (95% CI, 1 to 37 months) for crizotinib-naïve patients. The median overall survival was 24 months (95% CI, 5 to 43 months). Of the eight patients with brain metastases, intracranial disease control was reported in five (63%; 95% CI, 31% to 86%). The most common adverse events (majority, grade 1 or 2) for all treated patients were diarrhea (78%), nausea (59%), and anorexia (56%). Conclusion Ceritinib demonstrated potent clinical activity in patients with ROS1-rearranged NSCLC who were heavily treated previously with multiple lines of chemotherapy.

Most anaplastic lymphoma kinase (ALK)-rearranged non-small cell lung cancers (NSCLCs) show good clinical response to ALK inhibitors. However, some ALK-rearranged NSCLC patients show various primary responses with unknown reasons. Previous studies focused on the clinical aspects of ALK fusions in small cohorts, or were conducted in vitro and/or in vivo to investigate the function of ALK. One of the suggested theories describes how echinoderm microtubule-associated protein-like 4 (EML4)-ALK variants play a role towards different sensitivities in ALK inhibitors. Until now, there has been no integrated comprehensive study that dissects ALK at the molecular level in a large scale. Here, we report the largest extensive molecular analysis of 158 ALK-rearranged NSCLCs and have investigated these findings in a cell line construct experiment. We discovered that NSCLCs with EML4-ALK short forms (variant 3/others) had more advanced stage and frequent metastases than cases with the long forms (variant 1/others) (p = 0.057, p0.05). In vitro experiments revealed that EML4-ALK short forms show lower sensitivity to ALK inhibitors than do long forms. Clinical analysis also showed a trend for the short forms showing worse PFS. Interestingly, we found that breakpoints of ALK are evenly distributed mainly in intron 19 and almost all of them undergo a non-homologous end-joining repair to generate ALK fusions. We also discovered four novel somatic ALK mutations in NSCLC (T1151R, R1192P, A1280V, and L1535Q) that confer primary resistance; all of them showed strong resistance to ALK inhibitors, as G1202R does. Through targeted deep sequencing, we discovered three novel ALK fusion partners (GCC2, LMO7, and PHACTR1), and different ALK fusion partners showed different intracellular localization. With our findings that the EML4-ALK variants, new ALK somatic mutations, and novel ALK-fusion partners may affect sensitivity to ALK inhibitors, we stress the importance of targeted therapy to take the ALK molecular profiling into consideration. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John WileySons, Ltd.

Dermatofibrosarcoma protuberans (DFSP) is a very rare soft tissue sarcoma, generally of low-grade malignancy. DFSP is locally aggressive with a high recurrence rate, but metastasis occurs rarely. To investigate the mechanism of metastasis in DFSP, we analyzed the whole exome sequencing data of serial tumor samples obtained from a patient who had a 10-year history of recurrent and metastatic DFSP. Tracking various genomic alterations, namely somatic mutations, copy number variations, and chromosomal rearrangements, we observed a dramatic change in tumor cell population during the occurrence of metastasis in this DFSP case. The new subclone that emerged in metastatic DFSP harbored a completely different set of somatic mutations and new focal amplifications, which had not been observed in the primary clone before metastasis. The COL1A1-PDGFB fusion, characteristic of DFSP, was found in all of the serial samples. Moreover, the break position on the fusion gene was identical in all samples. Based on these observations, we suggest a clonal evolution model to explain the mechanism underlying metastasis in DFSP and identified several candidate target genes responsible for metastatic DFSP by utilizing The Cancer Genome Atlas database. This is the first study to observe clonal evolution in metastatic DFSP and provide insight for a possible therapeutic strategy for imatinib-resistant or metastatic DFSP.

Molecular markers are helpful diagnostic tools, particularly for cytologically indeterminate thyroid nodules. Preoperative RET/PTC1 rearrangement analysis in BRAF and RAS wild-type indeterminate thyroid nodules would permit the formulation of an unambiguous surgical plan. Cycle threshold values according to the cell count for detection of the RET/PTC1 rearrangement by real-time reverse transcription-polymerase chain reaction (RT-PCR) using fresh and routine air-dried TPC1 cells were evaluated. The correlation of RET/PTC1 rearrangement between fine-needle aspiration (FNA) and paired formalin-fixed paraffin-embedded (FFPE) specimens was analyzed. RET/PTC1 rearrangements of 76 resected BRAF and RAS wild-type classical PTCs were also analyzed. Results of RT-PCR and the Nanostring were compared. When 100 fresh and air-dried TPC1 cells were used, expression of RET/PTC1 rearrangement was detectable after 35 and 33 PCR cycles, respectively. The results of RET/PTC1 rearrangement in 10 FNA and paired FFPE papillary thyroid carcinoma (PTC) specimens showed complete correlation. Twenty-nine (38.2%) of 76 BRAF and RAS wild-type classical PTCs had RET/PTC1 rearrangement. Comparison of RET/PTC1 rearrangement analysis between RT-PCR and the Nanostring showed moderate agreement with a κ value of 0.56 (p = 0.002). The RET/PTC1 rearrangement analysis by RT-PCR using routine air-dried FNA specimen was confirmed to be technically applicable. A significant proportion (38.2%) of the BRAF and RAS wild-type PTCs harbored RET/PTC1 rearrangements.

To make an optimal treatment decision for early stage breast cancer, it is important to identify risk of recurrence. Here, we developed and validated a new prognostic model for predicting the risk of distant metastasis in patients with pN0-N1, hormone receptor-positive, HER2-negative (HR+/HER2−) breast cancer treated with hormone therapy alone. RNA was extracted from formalin-fixed, paraffin-embedded tumor tissues and gene expression was measured by quantitative real-time reverse transcription-PCR. The relative expression of six novel prognostic genes was combined with two clinical variables (nodal status and tumor size) to calculate a risk score (BCT score). In the validation cohort treated with hormone therapy alone, the 10 year rate of distant metastasis in the high-risk group (26.3%) according to BCT score was significantly higher than that in the low-risk group (3.8%) (P

The BRAFMutations of the BRAF and RAS genes and rearrangement of the RET/PTC1, NTRK1, and ALK genes using 769 preoperative fine-needle aspiration specimens and resected PTCs were analyzed.Molecular alterations were found in 687 (89.3%) of 769 PTCs. BRAFGenetic alterations in PTC vary among its different histologic variants and seem to be different in each ethnic group.

BACKGROUND: Programmed death ligand 1 (PD-L1) expression may predict response to anti-programmed death 1 (anti-PD-1) or anti-PD-L1 treatment. There is limited information on changes in PD-L1 expression over time in patients with non-small cell lung cancer (NSCLC).PATIENTS AND METHODS: Eligible patients with NSCLC who received surgery or underwent biopsy at Samsung Medical Center, Seoul, Republic of Korea, and Aarhus University Hospital, Aarhus, Denmark, between February 2004 and April 2012 were included. PD-L1 expression in paired tumor tissue samples from the same patients at different dates and lesions was measured using a laboratory-developed prototype immunohistochemistry assay (22C3 antibody). PD-L1 positivity was defined as tumor cell membrane positivity in ≥ 1% of tumor cells (proportion score). Concordance of PD-L1 expression was analyzed by treating proportion score as categoric or continuous variables.RESULTS: Ninety-one patients were included in the analysis. The median interval between the 2 tumor collection dates was 20 months, with 91% of paired samples collected > 3 months apart. The concordance rate for PD-L1 classification between paired samples was 67% (95% confidence interval, 57%-77%). When treating the immunohistochemistry proportional score as a continuous variable, a significant correlation of PD-L1 expression was observed between the paired samples (Pearson correlation coefficient, 0.61; P < .001).CONCLUSION: There are good correlations of PD-L1 expression from paired NSCLC samples. For patients whose PD-L1 status is negative, it may be valuable to obtain additional tissue samples for retesting PD-L1 expression when anti-PD-1 immunotherapy is considered.

Objectives Lung adenocarcinoma (LUAD) is a common subtype of non-small cell lung cancer prevalent in Asia. There is a dearth of understanding regarding the transcriptome landscape of LUAD without primary known driver mutations. In this study, LUAD samples without well-known driver mutations occurring in EGFR, KRAS, ALK, ROS1 or RET (quintuple-negative) were used for transcriptome study with a focus on long noncoding RNAs (lncRNAs), alternative splicing and gene fusions. Materials and methods 24 pairs of LUAD and adjacent normal samples and 13 tumor-only samples derived from 37 quintuple-negative patients were used. Differentially expressed lncRNA transcripts were detected by paired t-test and were validated by qPCR. Functions of lncRNAs were predicted by co-expressed mRNAs. Aberrant splicing events in LUAD were identified using MISO. In addition, gene fusions were screened by SOAPfuse. Results and conclusion In total, 90 and 153 up- or down-regulated lncRNA transcripts were detected in LUAD samples in comparison with the adjacent normal samples. The most significantly differentially expressed lncRNA transcript was ENST00000598996.1 (FENDRR) down-regulated in LUAD. By lncRNA-mRNA co-expression analysis, functions of 14 lncRNAs were predicted. The predicted functions included vasculature development, immune response, cell cycle and respiratory gaseous exchange. Furthermore, six co-expressed pairs of lncRNAs and their nearby protein coding genes were identified as associated with lung development. This study also identified two highly recurrent (22 in 24) differential exon skipping events occurring in MYH14 and ESYT2 with exon including isoforms of both genes up-regulated in isoform percentage in LUAD samples. On the other hand, two out of 24 LUAD samples possessed the driver mutation exon 14 skipping of MET. The transcriptional alterations of LUAD samples without well-known driver mutations identified in the study can be used as references for future research. The translational values of these transcriptional changes are also worthy of further investigation.

Girdin is an actin-binding Akt substrate that is an integral component of the PI3K/Akt signalling pathway. However, the clinicopathological significance of Girdin expression in breast cancer has not been clarified. The purpose of this study was to characterise the clinicopathological implication of Girdin expression in breast cancer. Immunohistochemistry-based protein expression analyses of 892 human breast cancer tissues showed that Girdin was expressed in 289 (32.4%) cases. Girdin expression was significantly associated with larger tumour size, frequent lymph node invasion, advanced cancer stage, and expression of oestrogen and progesterone receptors. Patients who had breast cancer with Girdin expression experienced significantly poorer overall survival (OS) (p=0.021) and disease-free survival (DFS) (p=0.002) than those without Girdin expression. In subtype analyses, Girdin expression was significantly correlated with poorer OS and DFS in HER2 subtype (p=0.004 and p=0.034, respectively). In triple negative breast cancer (TNBC) subtype, Girdin expression was significantly correlated with poorer DFS (p=0.035), and there was a trend toward poorer OS (p=0.060) in TNBC patients with Girdin expression. Multivariate analysis revealed that Girdin expression was an independent prognostic factor for OS (p=0.022) and DFS (p=0.030) in patients with breast cancer. In HER2 subtype under multivariate analysis, Girdin expression retained its role as an independent prognostic predictor for worse OS (p=0.023), and there was a trend toward poorer DFS (p=0.086) in patients with HER2 subtype expressing Girdin. Girdin expression may serve as a useful prognostic factor for invasive breast cancer, especially for the HER2 subtype.

Background Kirsten rat sarcoma 2 viral oncogene homolog (KRAS) mutation in pulmonary adenocarcinoma is clinically important due to its association with resistance to EGFR inhibitors and poor prognosis. To our knowledge, there has not been a comparative study focusing on cytological nuclear features of pulmonary adenocarcinoma harboring KRAS mutation (KRAS-AD). Hence, we compared the cytomorphology of metastatic KRAS-AD and EGFR-positive adenocarcinoma (EGFR-AD) in aspiration specimens from lymph nodes. Methods Forty lymph node aspiration specimens from forty KRAS-AD patients were collected at Samsung Medical Center (Seoul, Korea) from 2009 to 2013. As a control group, 40 EBUS-FNA lymph node specimens from 20 EGFR-AD patients were collected. EGFR-AD specimens were evaluated at Samsung Medical Center (Seoul, Korea) from 2012 to 2013. All 80 specimens were histologically confirmed to metastatic adenocarcinoma. Two pathologists performed a blinded review of all specimens. Results Compared with EGFR-AD, KRAS-AD exhibited more severe nuclear pleomorphism (P

Pazopanib is an orally bioavailable, ATP-competitive, multitargeted tyrosine kinase inhibitor mainly targeting VEGFR2 and PDGFR tyrosine kinases, but the biologic sequences of pazopanib activities beyond antiangiogenesis are poorly defined. We used a panel of 38 gastric cancer cell lines to test the efficacy of pazopanib. In a growth inhibition assay, genomic changes indicated that pazopanib had differential effects on cell growth. Treatment of the KATO-III, OCUM-2M, SNU-16, and HSC-39 gastric cancer cell lines harboring FGFR2 amplification with pazopanib resulted in marked decreases of cell survival with IC50 in ranges of 0.1 to 2.0 μmol/L, whereas the same treatment of those cell lines without FGFR2 amplification had no growth-inhibitory effects. In the ectopic FGFR2-expressing model, treatment with the indicated concentrations of pazopanib significantly inhibited cell growth and colony formation by FGFR2-expressing NIH 3T3 cells with wild-type (WT) FGFR2 and mutant FGFR2 (S252W). Pazopanib also selectively suppressed constitutive FGFR2 signaling and phosphorylation of downstream effectors. In cell-cycle analysis, FGFR2-amplified cells underwent cell-cycle arrest at the G1–S phase after pazopanib treatment, whereas there were no significant effects on cell-cycle progression in cells without FGFR2 amplification treated with pazopanib. In addition, pazopanib increased a substantial fraction of sub-G1 only in FGFR2-amplified cells. These findings show that the activation of FGFR2 signaling by amplification may be a critical mediator of cell proliferation in a small subset of gastric cancer patients and that pazopanib may provide genotype-correlated clinical benefits beyond the setting of highly vascular tumors. Mol Cancer Ther; 13(11); 2527–36. ©2014 AACR.

Synchronous primary lung cancers are relatively rare. The accurate diagnosis remains challenging, despite of the routine use of bronchoscopy and computed tomography (CT) of the chest. Herein we report a case of synchronous triple primary cancers of the right lung in a 72-year-old male patient in whom each tumor presented distinct CT imaging findings.

// Ju-Ae Hwang 1,5,* , Heung-Mo Yang 1,* , Doo-Pyo Hong 1 , Sung-Yeon Joo 1,2 , Yoon-La Choi 4 , Joo-Hung Park 5 , Alexander J. Lazar 6 , Raphael E. Pollock 7 , Dina Lev 6 and Sung Joo Kim 1,3,8 1 Transplantation Research Center, Samsung Biomedical Research Institute, Seoul, Republic of Korea 2 Samsung Advanced Institute for Health Sciences & Technology, Graduate School, Department of Health Sciences & Technology, Sungkyunkwan University 3 Department of Surgery, Samsung Medical Center, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Seoul, Republic of Korea 4 Department of Pathology, Samsung Medical Center, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Seoul, Republic of Korea 5 Department of Biology, Changwon National University, Changwon, Kyungnam, Republic of Korea 6 Department of Cancer Biology, University of Texas MD Anderson Cancer Center, Houston, TX, USA 7 Division of Surgical Oncology, James Comprehensive Cancer Center, Ohio State University, Columbus, OH, USA 8 Sarcoma Research Center, Samsung Medical Center, Seoul, Republic of Korea * These authors are contributed equally to this work Correspondence: Sung Joo Kim, email: // Dina-Lev , email: // Keywords : Liposarcoma, 2-DE, Gankyrin, Predictive factor, Tumorigenesis Received : May 25, 2014 Accepted : August 20, 2014 Published : August 21, 2014 Abstract Liposarcoma is one of the most common histologic types of soft tissue sarcoma and is frequently an aggressive cancer with poor outcome. Hence, alternative approaches other than surgical excision are necessary to improve treatment of well-differentiated/dedifferentiated liposarcoma (WDLPS/DDLPS). For this reason, we performed a two-dimensional gel electrophoresis (2-DE) and matrix-assisted laser desorption/ionization-time of flight mass spectrometry/mass spectrometry (MALDI-TOF/MS) analysis to identify new factors for WDLPS and DDLPS. Among the selected candidate proteins, gankyrin, known to be an oncoprotein, showed a significantly high level of expression pattern and inversely low expression of p53/p21 in WDLPS and DDLPS tissues, suggesting possible utility as a new predictive factor. Moreover, inhibition of gankyrin not only led to reduction of in vitro cell growth ability including cell proliferation, colony-formation, and migration, but also in vivo DDLPS cell tumorigenesis, perhaps via downregulation of the p53 tumor suppressor gene and its p21 target and also reduction of AKT/mTOR signal activation. This study identifies gankyrin, for the first time, as new potential predictive and oncogenic factor of WDLPS and DDLPS, suggesting the potential for service as a future LPS therapeutic approach.

Background Despite significant improvements in colon cancer outcomes over the past few decades, preclinical development of more effective therapeutic strategies is still limited by the availability of clinically relevant animal models. To meet those clinical unmet needs, we generated a well-characterized in vivo preclinical platform for colorectal cancer using fresh surgical samples. Methods Primary and metastatic colorectal tumor tissues (1–2 mm 3 ) that originate from surgery were implanted into the subcutaneous space of nude mice and serially passaged in vivo . Mutation status, hematoxylin and eosin staining, short tandem repeat profiling, and array comparative genomic hybridization were used to validate the similarity of molecular characteristics between the patient tumors and tumors obtained from xenografts. Results From surgical specimens of 143 patients, 97 xenograft models were obtained in immunodeficient mice (establish rate = 67%). Thirty-nine xenograft models were serially expanded further in mice with a mean time to reach a size of 1000–1500 mm 3 of 90 ± 20 d. Histologic and immunohistochemical analyses revealed a high degree of pathologic similarity including histologic architecture and expression of CEA, CK7, and CD20 between the patient and xenograft tumors. Molecular analysis showed that genetic mutations, genomic alterations, and gene expression patterns of each patient tumor were also well conserved in the corresponding xenograft tumor. Conclusions Xenograft animal models derived from fresh surgical sample maintained the key characteristic features of the original tumors, suggesting that this in vivo platform can be useful for preclinical development of novel therapeutic approaches to colorectal cancers.

Introduction: Anaplastic lymphoma kinase ( ALK ) fusion is the most common mechanism for overexpression and activation in non–small-cell lung carcinoma. Several fusion partners of ALK have been reported, including echinoderm microtubule-associated protein-like 4, TRK-fused gene, kinesin family member 5B , kinesin light chain 1 (KLC1), protein tyrosine phosphatase and nonreceptor type 3 , and huntingtin interacting protein 1 ( HIP1 ). Methods and Results: A 60-year-old Korean man had a lung mass which was a poorly differentiated adenocarcinoma with ALK overexpression. By using an Anchored Multiplex polymerase chain reaction assay and sequencing, we found that tumor had a novel translocated promoter region ( TPR)-ALK fusion. The fusion transcript was generated from an intact, in-frame fusion of TPR exon 15 and ALK exon 20 (t(1;2)(q31.1;p23)). The TPR-ALK fusion encodes a predicted protein of 1192 amino acids with a coiled-coil domain encoded by the 5'-2 nd of the TPR and juxtamembrane and kinase domains encoded by the 3'-end of the ALK . Conclusions: The novel fusion gene and its protein TRP-ALK, harboring coiled-coil and kinase domains, could possess transforming potential and responses to treatment with ALK inhibitors. This case is the first report of TPR-ALK fusion transcript in clinical tumor samples and could provide a novel diagnostic and therapeutic candidate target for patients with cancer, including non–small-cell lung carcinoma.

Approximately 7% of non-small cell lung carcinomas (NSCLCs) harbor oncogenic fusions involving ALK, ROS1, and RET. Although tumors harboring ALK fusions are highly sensitive to crizotinib, emerging preclinical and clinical data demonstrate that patients with ROS1 or RET fusions may also benefit from inhibitors targeting these kinases. Using a transcript-based method, we designed a combination of 3' overexpression and fusion-specific detection strategies to detect ALK, ROS1 and RET fusion transcripts in NSCLC tumors. We validated the assay in 295 NSCLC specimens and showed that the assay is highly sensitive and specific. ALK results were 100% concordant with fluorescence in situ hybridization (FISH) (n = 52) and 97.8% concordant with IHC (n = 179) [sensitivity, 96.8% (95% CI 91.0%-98.9%); specificity, 98.8% (95% CI 93.6%-99.8%)]. For ROS1 and RET, we also observed 100% concordance with FISH (n = 46 and n = 15, respectively). We identified seven ROS1 and 14 RET fusion-positive tumors and confirmed the fusion status by RT-PCR and FISH. One RET fusion involved a novel partner, cutlike homeobox 1 gene (CUX1), yielding an in-frame CUX1-RET fusion. ROS1 and RET fusions were significantly enriched in tumors without KRAS/EGFR/ALK alterations. ALK/ROS1/RET/EGFR/KRAS alterations were mutually exclusive. As a single-tube assay, this test shows promise as a more practical and cost-effective screening modality for detecting rare but targetable fusions in NSCLC.

Objectives Anaplastic lymphoma kinase (ALK) rearrangement is a validated predictive marker to define patients with non-small cell lung cancer (NSCLC) who can benefit from selective ALK inhibitors. Therefore, accurate assessment of its prevalence and clinical characteristics is increasingly important in the treatment of NSCLC. Also, this ALK rearrangement was previously reported to be more common in patients with no smoking history or those with adenocarcinoma. Patients and methods Never-smokers with completely resected pulmonary adenocarcinoma were screened for ALK rearrangements using Nanostring's gene expression platform. Clinicopathologic data, such as information about epidermal growth factor receptor (EGFR) and KRAS mutation status were retrospectively reviewed. Results Of 231 tumors screened, 20 (9%) had an ALK rearrangement and all were confirmed to be positive with immunohistochemical and fluorescent in situ hybridization analysis. Of the tumors with available data on the EGFR/KRAS mutation status, EGFR and KRAS mutation rates were 64% (69/108) and 5% (5/102), respectively. Amongst the tumors that were free of EGFR and KRAS mutations, the proportion of ALK rearrangements reached up to 33%. At the time of data cut-off, total of 68 tumors were recurred. Although the recurrence rate was similar between the ALK-positive and negative groups (30% vs. 29%), there was a tendency for ALK-positive tumors to recur more frequently in the pleural space (15% vs. 5%). The five-year disease-free survival (61%) and overall survival rates (79%) in the ALK-positive group were similar to those in the ALK-negative group (51% and 83%, respectively). Even after excluding two patients treated with crizotinib after disease recurrence, overall survival was similar between the two groups. Conclusion In an NSCLC subpopulation based on smoking history, histology, and EGFR and KRAS mutation status, the prevalence of ALK rearrangements is considerably high. However, ALK rearrangement status itself has no prognostic relevance in patients with completely resected NSCLC.

Purpose Lung squamous cell carcinoma (SCC) is the second most prevalent type of lung cancer. Currently, no targeted therapeutics are approved for treatment of this cancer, largely because of a lack of systematic understanding of the molecular pathogenesis of the disease. To identify therapeutic targets and perform comparative analyses of lung SCC, we probed somatic genome alterations of lung SCC by using samples from Korean patients. Patients and Methods We performed whole-exome sequencing of DNA from 104 lung SCC samples from Korean patients and matched normal DNA. In addition, copy-number analysis and transcriptome analysis were conducted for a subset of these samples. Clinical association with cancer-specific somatic alterations was investigated. Results This cancer cohort is characterized by a high mutational burden with an average of 261 somatic exonic mutations per tumor and a mutational spectrum showing a signature of exposure to cigarette smoke. Seven genes demonstrated statistical enrichment for mutation: TP53, RB1, PTEN, NFE2L2, KEAP1, MLL2, and PIK3CA). Comparative analysis between Korean and North American lung SCC samples demonstrated a similar spectrum of alterations in these two populations in contrast to the differences seen in lung adenocarcinoma. We also uncovered recurrent occurrence of therapeutically actionable FGFR3-TACC3 fusion in lung SCC. Conclusion These findings provide new steps toward the identification of genomic target candidates for precision medicine in lung SCC, a disease with significant unmet medical needs.

Triple-negative breast cancer is characterized by the absence of estrogen and progesterone receptors and human epidermal growth factor receptor 2, and is associated with a poorer outcome than other subtypes of breast cancer. Moreover, there are no accurate prognostic genes or effective therapeutic targets, thereby necessitating continued intensive investigation. This study analyzed the genetic mutation landscape in 70 patients with triple-negative breast cancer by targeted exome sequencing of tumor and matched normal samples. Sequencing showed that more than 50% of these patients had deleterious mutations and homozygous deletions of DNA repair genes, such as ATM, BRCA1, BRCA2, WRN, and CHEK2. These findings suggested that a large number of patients with triple-negative breast cancer have impaired DNA repair function and that therefore a poly ADP-ribose polymerase inhibitor may be an effective drug in the treatment of this disease. Notably, homozygous deletion of three genes, EPHA5, MITF, and ACSL3, was significantly associated with an increased risk of recurrence or distant metastasis in adjuvant chemotherapy-treated patients.

Purpose Data are limited on programmed death ligand 1 (PD-L1) expression in epidermal growth factor receptor (EGFR)-mutant non-small cell lung cancer (NSCLC). Materials and Methods We retrospectively evaluated the relationship between PD-L1 expression and recurrence-free survival (RFS) and overall survival in 319 patients with EGFR-mutant NSCLC who were treated at Samsung Medical Center from 2006 to 2014. Membranous PD-L1 expression on tumor cells was measured using the PD-L1 IHC 22C3 pharmDx antibody and reported as tumor proportion score (TPS). Kaplan-Meier methods, log-rank test, and Cox proportional hazards models were used for survival analysis. Results All patients had ≥1 EGFR mutation—54% in exon 19 and 39% in exon 21. Overall, 51% of patients had PD-L1–positive tumors. The prevalence of PD-L1 positivity was higher among patients with stages II-IV versus stage I disease (64% vs. 44%) and among patients with other EGFR mutations (75%) than with L858R mutation (39%) or exon 19 deletion (52%). PD-L1 positivity was associated with shorter RFS, with an adjusted hazard ratio of 1.52 (95% confidence interval [CI], 0.81 to 2.84; median, 18 months) for the PD-L1 TPS ≥ 50% group, 1.51 (95% CI, 1.02 to 2.21; median, 31 months) for the PD-L1 TPS 1%-49% group, and 1.51 (95% CI, 1.05 to 2.18) for the combined PD-L1–positive groups (TPS ≥ 1%) compared with the PD-L1–negative group (median, 35 months). Conclusion PD-L1 expression is associated with disease stage and type of EGFR mutation. PD-L1 positivity might be associated with worse RFS among patients with surgically treated EGFR-mutant NSCLC.

Background: ROS1 rearrangement accounts for 1%–2% of non-small cell lung cancer (NSCLC) with a remarkable response to crizotinib. Although ROS1-rearranged tumors are known to have characteristic histologic features, only a few studies have investigated the histologic features of advanced-stage ROS1-rearranged tumors. Methods: We analyzed the histopathologic features of ROS1-rearranged tumors in advanced-stage NSCLC patients and assessed the ROS1 immunohistochemistry (IHC) staining patterns of ROS1-rearranged cases. Results: A total of 37 ROS1 fluorescence in situ hybridization (FISH)-positive cases and 64 ROS1 FISH-negative cases were analyzed, and all tumors were EGFR-, ALK-, and RET-negative. Solid pattern, round nuclei with macronucleoli, solid signet-ring cells, extracellular mucin, and a close relation with adjacent bronchioles were significantly associated with ROS1 rearrangement, and the solid signet-ring cell feature was exclusively identified in ROS1-rearranged tumors. ROS1 IHC showed a 97.3% sensitivity when weak to strong protein expression was considered positive. Conclusions: Our findings highlight distinct histologic features of ROS1-rearranged tumors, including their nuclear features. A thorough understanding of ROS1 rearrangement-related histologic features would be helpful to identify ROS1-rearranged tumors in advanced-stage NSCLC.

The molecular bases underlying neoadjuvant chemotherapy (NAC) response are poorly understood. To elucidate the effects of NAC on breast tumor biology and its association with clinical outcome, we have conducted WES and RNA-Seq profiling of a longitudinal breast cancer (BC) cohort consisting of 146 cases (281 tumors, 109 pairs), including 55 (38%) that achieved pathologic complete responses (pCR) and 91 (62%) that harbored residual diseases at time of surgery. Tumor biopsies were collected for each patient at three time points - pre-treatment, three weeks after the first cycle of anthracycline and cyclophosphamide (AC) and at the time of surgery, after 3 more cycles of AC followed by 4 cycles of taxane. In addition to somatic mutations and copy number alterations, we also derived a comprehensive set of genomic and molecular features for each tumor including chromosomal instability, loss-of-heterozygosity, mutation burden, mutation signatures and expression signatures for oncogenic signaling pathways and immune cell subsets. Virtual microdissection analysis inferred 14 factors that represent distinct tissue compartment including a tumor infiltrating lymphocyte (TIL) factor and revealed that initial NAC treatment increased stromal and adjacent normal tissue fractions while reducing tumor cellularity. NAC also induced dynamic changes in immune gene expressions over time, a pattern that was validated through detecting and quantifying the density of TILs from H&E images. To investigate NAC induced changes in tumor intrinsic biology we classified tumors into five oncogenic cellular states on a reference Onco-GPS map defined by transcriptional signatures from breast cancer cell lines. We observed that, as a result of NAC treatment, tumors often change from one oncogenic state to another, transiently upregulating an EMT program that appears to mediate drug resistance and increase the likelihood of residual disease. Multiple regression and multivariate analyses were performed to identify predictive biomarkers of pCR status while adjusting for BC subtypes and tumor purity. We found that ER+ subtype and estrogen response signature but not Ki-67 were independently associated with NAC response. Within TNBC, the immunomodulatory subtype was enriched in responders while the basal-like subtype had the poorest response. Pretreatment TIL and changes in TIL level over time were independently associated with NAC response, implicating anti-tumor immunity in mediating the efficacy of chemotherapies. Through multi-omics characterization of longitudinally paired tumor biopsies, we have revealed dynamic changes in the tumor molecular states in BC patients undergoing NAC treatment, identified molecular markers of treatment outcome and derived insights into the mechanism of action as well as resistance to an important class of therapy. Citation Format: Samir Lal, Ying Ding, Jeong Eon Lee, Soo-Hyeon Lee, Se Kyung Lee, Jae-Yong Nam, Jong Han Yu, Yoon-la Choi, Seok Won Kim, Seok Jin Nam, Ji-Yeon Kim, Sripad Ram, Eric Powell, Keith A. Ching, Pablo Tamayo, William Kim, Huwate Yeerna, Soo Youn Cho, Vinicius Bonato, Shibing Deng, Jinho Kim, Hyuntae Shin, Woong-Yang Park, Paul A. Rejto, Jadwiga Bienkowska, Yeon-Hee Park, Zhengyan Kan. Multi-omics profiling of breast cancers during neoadjuvant chemotherapy identified distinct molecular changes and biomarkers associated with clinical response [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1684.

9094 Background: In the era of immunotherapy, immune checkpoint inhibitor (ICI) has changed the treatment paradigm in metastatic non-small cell lung cancer (NSCLC). Along with clinical trials, there is an ongoing investigation to discover the predictive biomarker of ICI which so far has unsatisfactory reliability. As an effort to enhance the predictive value of ICI treatment, we applied deep learning and developed artificial intelligent (AI) score (range from 0 to 1) to analyze the specific context of immune-tumor microenvironment (TME) extracted by scanned images from H&E slides. Methods: As a ground work, deep learning-based H&E image analyzer, Lunit SCOPE, has been trained with H&E images (n = 1824) from ICI naive NSCLC samples. For the calculation of AI score, training was conducted using responder/non-responder labeled ICI treated samples from the exploratory cohort. The ICI responder was defined as the patient with a best overall response of partial or complete response and stable disease for more than 6 months. The positivity of PD-L1 immunohistochemistry (IHC) was assessed manually by pathologists. Results: The exploratory cohort is composed of NSCLC patients treated with ICI (n = 189) in Samsung Medical Center, and response to ICI was observed in 72 (38.1%) patients. Median follow-up duration was 6.8 months (6.6~8.2). Samples with PD-L1 IHC positive, defined by ≥ 1%, was observed in 138 (73.0%) patients. AI score was significant higher in the responder group (median: 0.391 vs 0.205, P = 6.14e-5), and the patients with AI score above the cut-off (0.337) showed a better response to ICI (odds ratio [OR] 3.47 P = 7.34e-5) which is higher than patients with PD-L1 ≥ 1% (OR 1.92, P = 0.069). High AI score group (n = 83) showed significantly favorable PFS compared to low AI score group (n = 106, median PFS: 5.1m vs 1.9m, hazard ratio [HR] 0.51, P = 9.6e-5) and this outcome was independent with PD-L1 status (P = 6.0e-5). In subgroup analysis, PFS of PD-L1 high / AI score high group (n = 63) had longer median PFS (6.7m) compared to both PD-L1 high / AI score low group (n = 70, 4.0m, P = 0.001) and PD-L1 low/AI score low group (n = 35, 1.9m, P = 4.0e-6). Tumor infiltrating lymphocyte (TIL) density of cancer epithelium was significantly correlated with AI score (Pearson’s r = 0.310, P = 1.43e-5), which suggests that AI score may partly reflect TME represented by TIL. Conclusions: The AI score by machine-learned information, extracted from H&E images without additional IHC stain, could predict responsiveness and PFS of ICI treatment independent of PD-L1 IHC positivity.

MET, a cell surface receptor for hepatocyte growth factor, is involved in the development of triple-negative/basal-like breast cancer (TNBC/BLBC). However, its utility as a therapeutic target in this subtype of breast cancer is poorly understood. To evaluate MET fully as a potential therapeutic target for TNBC/BLBC, we investigated the relationship between MET expression and clinical outcomes of patients with breast cancer and the functional effect of MET inhibition. Using automated immunohistochemistry (Ventana), we analyzed MET expression in 924 breast cancer patients with relevant clinicopathologic parameters. BLBC showed the strongest relationship with MET expression (57.5%, p

Objectives Although T790M mutation is considered to be the major mechanism of acquired resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) in patients with non-small cell lung cancer (NSCLC), its clinical implication remains undetermined. Methods Post-progression tumor specimens were prospectively collected for T790M mutation analysis in NSCLC patients with acquired resistance to initial EGFR TKIs. Clinical features were compared between patients with and without T790M. Results Out of 70 cases, 36 (51%) were identified to have T790M mutation in the rebiopsy specimen. There was no difference in the pattern of disease progression, progression-free survival for initial TKIs (12.8 and 11.3 months), post-progression survival (14.7 and 14.1 months), or overall survival (43.5 and 36.8 months) in patients with and without T790M. In total, 34 patients received afatinib after post-progression biopsy as a subsequent treatment, and the response rate was 18%. The median progression-free survival for afatinib was 3.7 months for the entire group, and 3.2 and 4.6 months for the groups with and without T790M, respectively (P = 0.33). Conclusions The identification of T790M as acquired resistance mechanism was clinically feasible. Although T790M had no prognostic or predictive role in the present study, further research is necessary to identify patients with T790M-mutant tumors who might benefit from newly developed T790M-specific TKIs.

Epithelioid hemangioendothelioma (EHE) is a rare vascular neoplasm that has the ability to recur locally and metastasize. Thus, it is important to distinguish this tumor from other epithelioid vascular neoplasms. A 47-year-old man presented to our hospital with a pelvic mass with severe ischialgia and weight loss. Surgical resection was performed, and the mass was found to have dark red multiloculated cysts with hemorrhage and calcification. The histopathologic examination showed a central sclerotic, hypocellular zone and a peripheral cellular zone. Only the peripheral portion of the wall revealed nested tumor cells in light blue myxoid stroma. These tumors are typically composed of short strands or cords of bland epithelioid cells with occasional intracytoplasmic lumens embedded in a myxohyalinized stroma. The tumor cells were positive for CD31 and CD34 and negative for factor VIII-related antigen, CK (AE1/AE3) and S-100. The tumor nuclei showed distinct break-apart signals with individual green and/or red signals, indicating the presence of CAMTA1 rearrangement. In this study, we report a case of EHE that was difficult to diagnose based on histology alone. Therefore, we also performed fluorescence in situ hybridization, and found that the tumor harbored a CAMTA1 gene rearrangement, which confirmed the diagnosis.

Melanoma is the most aggressive skin cancer and the number of cases worldwide has doubled in the past 20 years.1 In particular, metastatic malignant melanoma is largely refractory to existing therapies and has a poor prognosis with a median survival rate of 6 months and a 5-year survival rate of less than 5%.2 The management of melanoma is rapidly evolving due to an improved understanding of its molecular biology and the development of effective, personalized, and targeted therapy strategies. Of particular importance in melanoma is the mitogen-activated protein kinase (MAPK) pathway, which normally regulates cell growth, proliferation, and differentiation.3 Aberrant activation of the MAPK pathway is present in over 80% of primary melanomas,4 and mutations in proteins along the RAS-RAF-MEK-ERK pathway are thought to be mutually exclusive. Among them, the most commonly mutated component of this pathway is BRAF.1 Constitutively activating mutations in the BRAF oncogene have been reported in 33% to 47% of primary melanomas and 41% to 55% of metastatic melanomas.5 Since its discovery in 2002, the BRAF V600E mutant kinase has been considered a promising therapeutic target for malignant melanoma. Recently, metastatic melanoma patients with the BRAF V600E mutation in a phase 3 randomized clinical trial on BRAF inhibitor (vemurafenib) showed significantly prolonged survival with a 63% relative reduction in the risk of death and a 74% relative reduction in the risk of tumor progression.6,7 Therefore, it is of great importance to accurately select patients with BRAF mutations who are candidates for BRAF inhibitors. Furthermore, in preclinical studies with mutant-selective BRAF inhibitors, melanoma patients with wild-type BRAF showed no response and paradoxically increased tumor growth.8-10 These findings once again highlighted the importance of accurate BRAF detection in the management of malignant melanoma. Given the importance of BRAF mutation detection in this highly aggressive tumor, companion diagnostic testing using the cobas 4800 BRAF V600 mutation test, a real-time polymerase chain reaction (PCR) test, was approved by the Food and Drug Administration (FDA) in 2011. There is limited data on the direct comparison of BRAF detection approaches such as PCR-based methods and sequencing methods.11-13 A more recently developed method, cobas, was reported to have a lower failure rate and more sensitivity for detecting V600E mutations than Sanger direct sequencing.12,13 However, all previous studies have only compared the analytic performance of the cobas test to Sanger sequencing. We performed direct comparison of the cobas BRAF V600 mutation test with dual-priming oligonucleotide-polymearse chain reaction (DPO-PCR) and direct sequencing in Korea, where the incidences of mucosal and acral melanomas are higher compared to melanomas that develop on sun-exposed skin for the first time.

Background Progression of hepatocellular carcinoma (HCC) often leads to vascular invasion and intrahepatic metastasis, which correlate with recurrence after surgical treatment and poor prognosis. The molecular prognostic model that could be applied to the HCC patient population in general is needed for effectively predicting disease-free survival (DFS).

Ovarian clear cell adenocarcinoma (Ov-CCA) is a distinctive subtype of ovarian epithelial carcinoma. In this study, we performed array comparative genomic hybridization (aCGH) and paired gene expression microarray of 19 fresh-frozen samples and conducted integrative analysis. For the copy number alterations, significantly amplified regions (false discovery rate [FDR] q0.05) were 1q21.3 and 8q24.3, and significantly deleted regions were 3p21.31, 4q12, 5q13.2, 5q23.2, 5q31.1, 7p22.1, 7q11.23, 8p12, 9p22.1, 11p15.1, 12p13.31, 15q11.2, 15q21.2, 18p11.31, and 22q11.21 using the Genomic Identification of Significant Targets in Cancer (GISTIC) analysis. Integrative analysis revealed 94 genes demonstrating frequent copy number alterations (25% of samples) that correlated with gene expression (FDR0.05). These genes were mainly located on 8p11.21, 8p21.2-p21.3, 8q22.1, 8q24.3, 17q23.2-q23.3, 19p13.3, and 19p13.11. Among the regions, 8q24.3 was found to contain the most genes (30 of 94 genes) including PTK2. The 8q24.3 region was indicated as the most significant region, as supported by copy number, GISTIC, and integrative analysis. Pathway analysis using differentially expressed genes on 8q24.3 revealed several major nodes, including PTK2. In conclusion, we identified a set of 94 candidate genes with frequent copy number alterations that correlated with gene expression. Specific chromosomal alterations, such as the 8q24.3 gain containing PTK2, could be a therapeutic target in a subset of Ov-CCAs.

Summary Background and objective An animal model for bile duct stenosis using intraductal thermal injury has not yet been established. The aims of the current study were to develop biliary stenosis in a swine model by inducing intraductal thermal injury using a heat probe or radiofrequency ablation electrode and to investigate an effective and safe energy dose. Methods Intraluminal thermal injury was applied to the common bile duct with a heat probe in three swines and a radiofrequency ablation electrode in the other three swines by either endoscopic retrograde cholangiography or open laparotomy. Cholangiography and histologic evaluation of common bile duct were taken 2 weeks after thermal injury. Results Thermal injury with a heat probe at 25 J for 40 seconds produced a stricture in all three animals. Application of a radiofrequency ablation electrode produced a stricture in two of three animals. An energy dose of 40 W at 80 °C for 30 seconds produced biliary stenosis without any complications initially and 2 weeks after thermal injury. Conclusions The application of a heat probe and a radiofrequency ablation electrode for intraductal thermal injury resulted in a reproducible animal model of biliary stenosis.

SummaryFrequent discrepancies between preclinical and clinical results of anticancer agents demand a reliable translational platform that can precisely recapitulate the biology of human cancers. Another critical unmet need is the ability to predict therapeutic responses for individual patients. Toward this goal, we have established a library of orthotopic glioblastoma (GBM) xenograft models using surgical samples of GBM patients. These patient-specific GBM xenograft tumors recapitulate histopathological properties and maintain genomic characteristics of parental GBMs in situ. Furthermore, in vivo irradiation, chemotherapy, and targeted therapy of these xenograft tumors mimic the treatment response of parental GBMs. We also found that establishment of orthotopic xenograft models portends poor prognosis of GBM patients and identified the gene signatures and pathways signatures associated with the clinical aggressiveness of GBMs. Together, the patient-specific orthotopic GBM xenograft library represent the preclinically and clinically valuable “patient tumor’s phenocopy” that represents molecular and functional heterogeneity of GBMs.

Purpose More accurate prediction of patient outcome based on molecular subtype is required to identify patients who will benefit from specific treatments. Methods We selected novel 16 candidate prognostic genes, including 10 proliferation-related genes (p-genes) and 6 immune response-related genes (i-genes), from the gene list identified in our previous study. We then analyzed the association between their expression, measured by quantitative real-time reverse transcription-PCR in formalin-fixed, paraffin-embedded tissues, and clinical outcome in 819 breast cancer patients according to molecular subtype. Results The prognostic significance of clinical and gene variables varied according to the molecular subtype. Univariate analysis showed that positive lymph node status was significantly correlated with the increased risk of distant metastasis in all subtypes except the hormone receptor-negative, HER2-positive (HR−/HER2+) subtype. Most p-genes were significantly associated with poor prognosis in patients with the HR+/HER2− subtype, whereas i-genes correlated with a favorable outcome in patients with HR−/HER2+ breast cancer. In HR−/HER2+ breast cancer, four genes (three i-genes BTN3A2, CD2, and TRBC1 and the p-gene MMP11) were significantly associated with distant metastasis-free survival (DMFS). A new prognostic model for HR−/HER2+ breast cancer based on the expression of MMP11 and CD2 was developed and the DMFS for patients in the high-risk group according to our model was significantly lower than that for those in the low-risk group. Multivariate analyses revealed that our risk score is an independent prognostic factor for DMFS. Moreover, C-index showed that our risk score has a superior prognostic performance to traditional clinicopathological factors. Conclusions Our new prognostic model for HR−/HER2+ breast cancer provides more accurate information on the risk of distant metastasis than traditional clinical prognostic factors and may be used to identify patients with a good prognosis in this aggressive subtype of breast cancer. Electronic supplementary material The online version of this article (doi:10.1007/s10549-017-4234-4) contains supplementary material, which is available to authorized users.

Objective A mutational analysis of tumor tissue samples is an important part of advanced lung cancer treatment strategies. This study evaluated the efficacy of a triple gene analysis using samples obtained via endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA). Methods Either metastatic lymph nodes or primary lung mass samples obtained by EBUS-TBNA were collected between May 2011 and May 2013. We consecutively analyzed epidermal growth factor receptor (EGFR), V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS), and anaplastic lymphoma kinase (ALK) fusion genes using remnant tissue samples. Results A total of 109 patients were diagnosed with non-small cell lung cancer (NSCLC). Of these, 70% were adenocarcinoma, 27% squamous cell carcinoma with NSCLC, and 3% were related to other types of lung cancer. EGFR mutations were detected in 23 cases (21.1%), KRAS mutations in 13 cases (11.9%), and ALK fusion genes in 5 cases (4.9%). The ALK fusion genes could not be analyzed in four cases because of insufficient tissue samples remaining after routine histochemistry and an EGFR/KRAS mutation analysis. We found that small biopsy samples from EBUS-TBNA were adequate for performing a triple gene analysis in 97 patients (96%). ALK fusion protein immunohistochemistry (IHC) was 100% consistent with fluorescence in situ hybridization (FISH). Conclusion Small samples obtained by EBUS-TBNA were found to be sufficient for performing a triple gene analysis following routine histology and IHC. ALK IHC showed a very good concordance with FISH for detecting ALK fusion genes.

Introduction Response to mesenchymal-epithelial transition (MET) inhibitors in NSCLC with mesenchymal-epithelial transition gene ( MET ) exon 14 skipping ( MET ex14) has fueled molecular screening efforts and the search for optimal therapies. However, further work is needed to refine the clinicopathologic and prognostic implications of MET ex14 skipping. Methods Among 795 East Asian patients who underwent a surgical procedure for NSCLC, we screened 45 patients with quintuple-negative ( EGFR -negative /KRAS -negative / anaplastic lymphoma kinase gene [ ALK ]-negative /ROS1 -negative / ret proto-oncogene [ RET ]-negative) lung adenocarcinomas by using reverse-transcriptase polymerase chain reaction and found 17 patients (37.8%) with MET ex14 skipping. We also investigated the effect of small interfering RNA (siRNA) targeting skipping junction in cells with MET ex14 skipping. Results The median age of the 17 patients was 73 years. The acinar subtype was predominant (52.9%), followed by the solid subtype (35.3%). MET immunohistochemistry demonstrated 100% sensitivity and 70.4% specificity. Multivariate analyses showed that patients with MET ex14 skipping had a higher recurrence rate than those with ALK fusion (versus MET ex14 skipping) (hazard ratio = 0.283, 95% confidence interval: 0.119 – 0.670) in stage I to IIIA disease; however, the differences in overall survival were not significant after adjustment for pathologic stage ( p = 0.669). Meanwhile, siRNA decreased MET-driven signaling pathways in Hs746T cells, and combined treatment with siRNA and crizotinib inhibited cell proliferation in crizotinib-resistant H596 cells. Conclusions The prevalence of MET ex14 skipping was quite high in East Asian patients without other driver mutations in lung adenocarcinomas. MET ex14 skipping was associated with old age, the acinar or solid histologic subtype, and high MET immunohistochemical expression. The prognosis of patients with MET ex14 skipping was similar to that of patients with major driver mutations. siRNA targeting the junction of MET ex14 skipping could inhibit MET-driven signaling pathways in cells with MET ex14 skipping.

Aim To compare the abilities of conventional magnetic resonance imaging (MRI) and apparent diffusion coefficient (ADC) in differentiating between benign and malignant soft-tissue tumours (STT). Material and methods A total of 123 patients with STT who underwent 3 T MRI, including diffusion-weighted imaging (DWI), were retrospectively analysed using variate conventional MRI parameters, ADC mean and ADC min . Results For the all-STT group, the correlation between the malignant STT conventional MRI parameters, except deep compartment involvement, compared to those of benign STT were statistically significant with univariate analysis. Maximum diameter of the tumour ( p =0.001; odds ratio [OR], 8.97) and ADC mean ( p =0.020; OR, 4.30) were independent factors with multivariate analysis. For the non-myxoid non-haemosiderin STT group, signal heterogeneity on axial T1-weighted imaging (T1WI; p =0.017), ADC mean , and ADC min ( p =0.001, p =0.001), showed significant differences with univariate analysis between malignancy and benignity. Signal heterogeneity in axial T1WI ( p =0.025; OR, 12.64) and ADC mean ( p =0.004; OR, 33.15) were independent factors with multivariate analysis. Conclusion ADC values as well as conventional MRI parameters were useful in differentiating between benign and malignant STT. The ADC mean was the most powerful diagnostic parameter in non-myxoid non-haemosiderin STT.