bluuebunny/crossref_metadata_2025 huggingface dataset:
Response to the anti-EGFR antibody panitumumab combined with standard neoadjuvant chemotherapy in triple-negative breast cancer (TNBC): The immune and IGFR pathways.
Authors: Penault-Llorca Frederique , Dauplat Marie-Melanie , Abrial Catherine , Weber Beatrice E. , Mouret-Reynier Marie-Ange , Gligorov Joseph , Tredan Olivier , Privat Maud , Uhrhammer Nancy , Desrichard Alexis , Bidet Yannick , Radosevic-Robin Nina , Cayre Anne , Aubé Cécile , Romero Pierre Charles , Kwiatkowski Fabrice , Chalabi Nassera , Bignon Yves-Jean , Chollet Philippe , Nabholtz Jean-Marc
Published: March 2020

1058

Background: EGFR overexpression is one of the hallmarks of the “basal-like” TNBC definition by immunohistochemistry (IHC). In a phase II neoadjuvant clinical trial targeting EGFR in TNBC, we investigated various biomarkers to better identify an EGFR-sensitive population for potential further regimen development. Methods: Sixty patients (pts) with stage II-IIIA TNBC were prospectively included. Systemic treatment (ST) consisted of the anti-EGFR antibody panitumumab combined with FEC 100, followed by 4 cycles of docetaxel. All pts underwent surgery after ST completion. Patient characteristics: median tumor size: 40 mm (20-120); invasive ductal carcinoma: 96.7%; SBR grade III: 71.7%; complete pathological response (pCR) rate: 55.3% and 46.8% (according to Sataloff’s and Chevallier’s classifications, respectively). Paraffin-embedded and frozen tumor samples were collected before and after ST for biologic studies. Germinal BRCA1 mutations, and EGFR, KRAS, BRAF and PI3KCA somatic mutations were analyzed by NGS. EGFR, IGF-1R, MET, cytokeratins 5/6 and 8/18, PTEN, P-cadherin, ALDH1, Ki-67, p53, tumoral FOXP3 expression and the number of FOXP3+ or CD8+ tumor-infiltrating lymphocytes (TILs) were evaluated by IHC. Results: High CD8+ TILs was response-predictive (p=3.4.10-6). Tumor FOXP3 expression and high FOXP3 TILs tended to be predictive. High IGF-1R expressors responded better than low expressors (p = 0.028). Comparison of EGFR, IGF-1R and Her3 in biopsies versus surgical samples showed reduced EGFR levels in non-responders (p = 0.037), while Her3 (p = 0.049) and IGF-1R (p = 0.08) increased. Sequencing revealed BRCA1 mutations in 10% of pts. No difference of response (pCR) was observed between mutated patients and others. Somatic mutations of PI3K were observed in 6 pts. No mutations were observed in BRAF, KRAS, or EGFR. Conclusions: The CD8+ TIL count seems to predict the response to panitumumab. Tumor FOXP3 expression and high FOXP3 TILs also tended to be predictive. Tumor levels of IGF-1R seem to play a determinant role in TNBC response to anti-EGFR antibodies, in concordance with our observations in a head-and-neck cancer cohort. Clinical trial information: NCT00933517.
A prospective multicenter phase II study of sunitinib in patients with advanced aggressive fibromatosis.
Authors: Jo Jae-Cheol , Hong Yong Sang , Kim Kyu-Pyo , Lee Jae-Lyun , Lee Jeeyun , Park Young Suk , Kim Sun Young , Ryu Jin-Sook , Lee Jong-Seok , Kim Tae Won
Published: March 2020

10589

Background: There have been reports on responses and prolonged disease stabilizations with imatinib which may be associated with PDGFR-β pathway and KIT mutation in the treatment of aggressive fibromatosis (AF). Sunitinib has not only PDGFRs, KIT, and FLT3 inhibiting activity, but also VEGFRs blockade as antiangiogenesis. The aim of this study is to evaluate the efficacy and safety of sunitinib for patients with advanced AF. Methods: Nineteen patients with pathologically proven AF were accrued between Jun 2008 and Mar 2012. Sunitinib was administered with 37.5 mg/day for 4 weeks without break, comprising one cycle. Primary endpoint was response rate. Results: Ten (53%) patients were female and the median age was 30 years (range, 22-67). Most of the primary sites were intra-abdominal (12, 63.2%), and AF associated with familial adenomatous polyposis was observed in 10 (52.6%). With a median 6 cycles per patients (range, 1-47 cycles), 5 (26.3%) achieved a partial response and 8 (42.1%) had stable disease; the overall response rate was 26.3% (95% CI, 6.3-45.7) in intention-to-treat analysis. With median follow-up time of 20.3 months (range, 1.8-50.7), the 2-year progression-free survival and overall survival were 74.7% and 94.4%, respectively. Grade 3 or 4 adverse events of sunitinib with frequency > 5% of patients included neutropenia (33.3%), diarrhea (5.3%), and hand-foot syndrome (5.3%). In 3 patients out of 12 patients with mesenteric AF, mesenteric mass bleeding (n=1), bowel perforation (n=1), and bowel fistula (n=1) with tumor mass necrosis, were observed during an early period of sunitinib. Conclusions: Sunitinib showed potential antitumor activity and was relatively tolerated in patients with AF, especially non-mesenteric AF. Further investigation of sunitinib treatment is necessary in patients with AF.
N0937(Alliance): Final clinical results and correlative data from the phase II trial of cisplatin (C) and the novel agent brostallicin (B) in patients with metastatic triple-negative breast cancer (mTNBC).
Authors: Moreno-Aspitia Alvaro , Liu Heshan , Hillman David W. , Rowland Kendrith M. , Geiger Xochiquetzal , Stella Philip J. , Kourlas Peter , Gross Howard M. , Karlin Nina J. , Bury Martin J. , Soori Gamini S. , Nassar Aziza , Perez Edith A. ,   Alliance for Clinical Trials in Oncology
Published: March 2020

1059

Background: Relapsed TNBC is characterized by its poor prognosis. B is a synthetic DNA minor groove binding non-cross resistant agent that is fully active against DNA-mismatch repair deficient tumor cells. C increases GSH/GST in which enhances the efficacy of B. Methods: Phase II study in pts with mTNBC. C Day 1 followed by B Day 2, repeated every 21 days. Aim: Efficacy of C + B in mTNBC. Primary endpoint: PFS at 3 mo. Secondary endpoints include ORR, duration of response (DOR), 6-mo PFS, OS and AE profile. Tertiary endpoints include assessment of baseline GSH levels, prevalence of BCRA1 mutation and pERK, and correlations with outcome. Results: Closed 3/2012. 48 pts accrued (1 ineligible pt). Median f/u 11.4 mo (6.6-14.8); 2 pts still on treatment. 47 pts evaluable for the PFS endpoint and AEs. 49% received therapy as 3rd-5th line. Median number of cycles = 4 (range 1-15). There are currently 9 confirmed responses (8 PR and 1 CR); DOR = 2.6-14.5 mo. The 3-mo PFS = 51%; 6-mo PFS = 28%; the median TTP is 3.2 mo. AE data: 75% G3/4 hematologic AE: febrile neutropenia 19%. Non-hematologic AEs included G3 (47%) and G4 (15%): G3 fatigue 17%; and no G5 non-heme AE. No correlation of GSH/GST levels with outcome was found. Among 35 pts with evaluable data, no association between BRCA1 expression and outcomes was found. pERK expression was negatively associated with 3-mo PFS, but not 6-mo PFS. Cytoplasmic pERK expression was negatively associated with tumor response (p=0.03). No significant effect of BRCA-1 or pERK expression was found on OS. Conclusions: This trial showed B + C is an active regimen, especially in heavily pretreated mTNBC pts. Correlative data demonstrated a negative association between pERK expression and outcomes. A randomized phase II trial is in development. Clinical trial information: NCT01091454.
Efficacy and safety of metronomic capecitabine and gefitinib in heavily pretreated metastatic triple-negative breast cancer.
Authors: Ranade Anantbhushan , Govind Babu Kanaka , Parikh Purvish M. , Singh Jk , Singh Manisha , Bhattacharyya Gouri Shankar
Published: March 2020

1071

Background: Metronomic chemotherapy regimens have shown efficacy in patients with metastatic breast cancer by antiangiogenic mechanisms. When used metronomically the toxicity profile of capecitabine is low. Triple negative breast cancer is a common problem in India and developing countries. Approximately 30% of triple negative breast cancer express EGFR and its mutation. Methods: Since October 2003 to December 2011 we objectively tested response rates, clinical benefit, and safety of gefitinib and capecitabine administered with a metronomic schedule of 500 mg thrice daily in heavily pretreated metastatic breast cancer patients with gefitinib 250 mg once daily. 300 patients were screened for EGFR expression. Among 85 enrolled patients with EGFR positivity, 76 were evaluable. ECOG performance status (PS) was 0-2, median age 52 years (range 36-65), bone plus visceral metastasis in 40% of patients. Rest had only visceral metastasis. All the patients were pretreated with anthracyclines and taxanes. The combination was administered for a median duration of 32 weeks (range 12-166). Results: We observed 18 partial responses (PR: 24%), 42 (55%) stable disease (SD). Median time to progression was 53 weeks, (95% CI, range 12-166 weeks). Safety of metronomic capecitabine with gefitinib was excellent. Neither grade 2-4 haematological or clinical side effects were recorded. Only 12 patients experienced grade I (WHO) hand-foot syndrome. Conclusions: Treatment with metronomic capecitabine and gefitinib was effective and minimally toxic in heavily pretreated breast cancer patients.
Etirinotecan pegol (EP) target-specific pharmacodynamic (PD) biomarkers measured in circulating tumor cells (CTCs) from patients in the phase III BEACON study in patients with metastatic breast cancer (mBC).
Authors: Hoch Ute , Fry Dennis G. , Chia Yen Lin , Caygill Katie , Hannah Alison L. , Perez Edith A. , Cortez Javier , Awada Ahmad , O'Shaughnessy Joyce , Twelves Christopher , Rugo Hope S. , Im Seock-Ah , Xu Binghe , Anderes Kenna Lynn , Davis Darren W.
Published: March 2020

1087

Background: EP is a unique topoisomerase 1 inhibitor that provides continuous exposure to SN38. EP demonstrated a 29% overall response rate in patients with mBC, leading to a phase III global BEACON study in patients with mBC. CTCs in patient blood samples provide a minimally invasive approach to detect PD markers of drug activity. We developed quantitative multiplex immunofluorescent assays to measure target-specific PD biomarkers for EP in CTCs isolated pre- and post-treatment. Methods: Assays for Top1, Top2, g-H2Ax, Rad51, ABCG2, and Ki-67 were developed using control (0.1% DMSO) and drug-treated (SN38, 10 uM) tumor cell lines (HCT116, MCF7, A549, SKBr3) and PBMCs from healthy donors. The optimal antibody for each biomarker was then multiplexed in a panel with antibodies against cytokeratin, CD45 and DAPI for phenotypic identification of CTCs. For analysis of BEACON pts, serial 7.5 mL whole blood samples were drawn and shipped ambient to Apocell (Houston, TX) for further processing. PBMCs were separated and CTCs were isolated using ApoStream technology. CTCs were stained for PD markers and analyzed using an iCys laser scanning cytometer equipped with image analysis software. Results: Antibodies bound to tumor cells showed staining confined to the nucleus (Top1, Top2, g-H2AX, Ki-67) or membrane (ABCG2), exhibited defined peak separation from their isotype controls, and signal strength correlated with cellular expression of high and low levels of markers. To date, ~ 80% of BEACON pts consent to participate in the CTC sub-study. As of 30 Oct 2012, data from 167 pre-dose blood samples from BEACON pts were available. 99% of blood samples were successfully processed. 93% had detectable CTCs, yielding a median of 216 CTCs (range: 7.5-14816). Staining was positive for Top1, Top2, g-H2Ax, Rad51, ABCG2, and Ki-67 in 82%, 89%, 16%, 53%, 31%, 52% of samples, respectively. Conclusions: EP target-specific pharmacodynamic biomarkers can be reliably measured in CTCs isolated from patients participating in BEACON. Sample collection and analysis of pre- and post-treatment samples is ongoing. Clinical trial information: NCT01492101.
Integrating molecular profiling into cancer treatment decision making: Experience with over 35,000 cases.
Authors: Gatalica Zoran , Millis Sherri , Chen Sting , Basu Gargi Dan , Wen Wenhsiang , Paul Les , Bender Ryan P. , Von Hoff Daniel D.
Published: March 2020

11001

Background: Molecular profiling of both common and rare cancer types provides for the identification of actionable targets for chemotherapy with many unexpected associations. Methods: Caris Life Sciences database of >35,000 profiled cancers was reviewed for well-established driver gene mutations and copy number alterations, and protein expression patterns that are relevant for selection of targeted therapy. Based on the published literature, these tumor characteristics were then associated with potential benefit or no benefit to the specific therapeutic agents. All relevant published studies were evaluated using the USPSTF grading scheme for study design and validity. Assay methodologies included sequencing (Sanger, pyrosequencing), PCR, FISH, CISH, and immunohistochemistry. Results: All common malignancies (10 most common cancer types in men and women) and 10 rare cancer types were well represented (minimum of 100 cases in each individual cancer type). Well established driver mutations and protein expression in common cancers were all identified with expected frequencies (e.g. HER2 amplification in breast, PIK3CA mutations in ER+ breast cancer, EGFR mutations in NSCLC, etc.). Importantly, unexpected new and potentially actionable targets were identified in common (e.g., 6.7% HER2 amplification in NSCLC, 1.6% KRAS mutation in prostatic adenocarcinoma) and rare cancers (e.g., 8.3% ALK alteration in soft tissue sarcomas, 10.5% c-MET and 26.4% EGFR gene amplification in melanomas, 16.3% KRAS mutation in cholangiocarcinomas, 10% AR expression in STS), as well in cancers of unknown primary site (approximately 4% of all tested cases). Conclusions: This review of the large referral cancer profiling database provided an unparalleled insight in the distribution of common and rare genetic and protein alterations with direct and potential treatment implications. Numerous targets were discovered that had a potential to be treated by the conventional chemotherapy as well as targeted therapy not usually considered for the cancer type. Comparison between an individual patient tumor profile and database for the matched cancer type provides additional level of support for targeted treatment choices.
The genetic landscape of clinical resistance to RAF inhibition in melanoma.
Authors: Van Allen Eliezer Mendel , Wagle Nikhil , Carter Scott L. , Sucker Antje , Farlow Deborah Norman , Hodis Eran , Taylor-Weiner Amaro , Berking Carola , Egberts Friederike , Hassel Jessica Cecile , Gogas Helen , Gutzmer Ralf , Goldinger Simone M. , Loquai Carmen , Ugurel Selma , Zimmer Lisa , Gabriel Stacey B. , Getz Gad , Garraway Levi A. , Schadendorf Dirk ,   DeCOG
Published: March 2020

11009

Background: Although single-agent RAF inhibition has proved effective in metastatic BRAFV600-mutant melanoma, most patients relapse and some are intrinsically resistant. While several genetic resistance effectors have been identified, a comprehensive assessment of the genetic resistance spectrum in a large patient cohort may further inform resistance patterns and treatment strategies. Methods: Pre-treatment and post-relapse biopsies were obtained from BRAFV600melanoma patients treated with vemurafenib or dabrafenib. Whole exome sequencing of tumor and normal samples was performed to identify exome-wide mutations, insertion/deletions, and chromosomal copy number alterations. Since somatic allelic fraction comparisons across treatment time points must account for variable stromal admixture between biopsies, novel algorithms were applied to impute purity and tumor cell ploidy, thus determining the “cancer cell fraction” (CCF) of each alteration. Those with enriched CCF in post-relapse samples were nominated as potential resistance effectors. Results: 30 patients (towards a goal of 77) were sequenced to a mean depth of 145-fold coverage. In acquired resistance patients (on drug ≥ 12 weeks, n=18), NRAS Q61R (28%) and BRAF amplifications (6%) were observed. The remaining patients had multiple clonally enriched alterations in the post-relapse tumors that may drive resistance, including MEK1 mutations at residues previously noted in an in vitro mutagenesis screen for vemurafenib resistance. In the intrinsic resistance cohort (on drug < 12 weeks, n=12), multiple putative resistance mutations not previously observed in the setting of acquired resistance were identified. Conclusions: In addition to reported genetic resistance mechanisms, we identified new MEK1 mutations in resistant samples. For the majority of patients (67%) without events in these genes, computational algorithms identified new candidate drivers of RAF inhibitor resistance. Alterations that predict intrinsic or acquired resistance may also inform new approaches to melanoma therapy. Future studies incorporating longitudinal biopsies and genetic profiling will underpin a robust assessment of clinical resistance in melanoma and other cancers.
Fusion transcript discovery in formalin-fixed paraffin-embedded human breast cancer tissues and its relation to tumor progression.
Authors: Ma Yan , Ambannavar Ranjana , Stephans James , Jeong Jennie , Dei Rossi Andrew , Morlan John , Liu Mei-Lan , Levy Samuel , Baker Joffre , Sinicropi Dominick , Qu Kunbin
Published: March 2020

11018

Background: While several recently discovered gene fusions already play an important role in personalized cancer treatment, many cancer gene fusions remain to be discovered. Next generation sequencing has enabled identification of many rare gene fusion events in fresh or frozen solid tumors. There is a need to detect gene fusions in transcriptomes of formalin-fixed paraffin-embedded (FFPE) tumor tissue, for which there is long-term clinical outcome data. We therefore sought to develop bioinformatics methods to detect fusion transcripts in FFPE tissue and to characterize their association with clinical outcomes. Methods: RNA sequencing libraries were created and sequenced from tumor biopsy tissues (Plos One 2012 7(7): e40092) of two ER+ breast cancer cohorts consisting of 136 and 77 patients, for which clinical outcomes were available. The fusion junctions were nominated by the RNA-seq aligner GSNAP and further filtered to consider discontinuous expression patterns at exon/intron levels. Results: A total of 108 candidate fusion transcripts were detected and RT-PCR assays confirmed 89% of the top ranking fusion transcript candidates. The majority (82%) of identified fusion gene partners are listed in the COSMIC database of known cancer sequence variations. Of note, several patients expressed multiple fusion transcripts that are significantly associated with tumor progression (P<0.001), including genes associated with cell proliferation and cellular metabolism. Furthermore, these patients also harbored inter-chromosomal gene fusions. It is noteworthy that several gene fusions were present in multiple patients. In one of these recurrent fusions the estrogen receptor gene acts as the fusion pair donor. Conclusions: Novel bioinformatics approaches developed here demonstrate the ability to detect fusion transcripts as biomarkers from archival FFPE tissues that associate with breast cancer progression. Some gene fusions were common in multiple patients and deserved further study.
ROR1 mRNA expression in EGFR-mutant non-small-cell lung cancer (NSCLC) patients (p) with the T790M mutation: A potential therapeutic target.
Authors: Karachaliou Niki , Drozdowskyj Ana , Costa Carlota , Molina-Vila Miguel Angel , Gimenez Capitan Ana , Vergnenegre Alain , Massuti Bartomeu , Moran Teresa , Majem Margarita , Felip Enriqueta , Carcereny Costa Enric , Garcia-Campelo M. Rosario , Viteri Ramirez Santiago , Heidecke Cordula Nicole , Estrada-Tejedor Roger , Teixido Jordi , Bivona Trever Grant , Giannikopoulos Petros , Ono Mayumi , Rosell Rafael ,   Spanish Lung Cancer Group
Published: March 2020

11027

Background: Progression-free survival (PFS) is short in NSCLC driven by EGFR mutations treated with erlotinib alone, due to crosstalk with other signaling pathways that can cause secondary dependency. ROR1 knockdown inhibited the growth of NCI-H1975 cells (with EGFR L858R and T790M mutations). A pro-survival function for ROR1/MEK/ERK signaling has been demonstrated, with cooperation with AKT. In a subset of 95 p in the EURTAC trial (clinicaltrials.gov NCT00446225), 65% had pre-treatment T790M mutations. We have assessed ROR1 expression in 45 of these 95 p. Methods: The T790M mutation was determined by Taqman with a PNA to inhibit amplification of the wild-type (wt) allele. Tumor samples were run in octuplicates; this method can detect 1 mutated allele among 10,000 wt alleles. ROR1 mRNA expression was examined by quantitative RT-PCR and categorized by terciles. p were classified as having low/intermediate or high ROR1 expression. The impact of ROR1 expression on outcome was examined in all 45 p and in a subset of 15 p with concomitant T790M mutations. Results: Median age 65; 68.9% female; 57.8% never-smokers; 95.6% ECOG PS <2; 91.1% adenocarcinoma; 68.9% exon 19 deletion. No differences in baseline characteristics were observed according to ROR1 expression levels. 24 p (53.3%) were treated with erlotinib and 21 p (46.7%) with chemotherapy. 10 (41.7%) erlotinib-treated p and 6 (28.6%) chemotherapy-treated p had ROR1 mRNA levels in the top tercile. Among erlotinib-treated p, response rate was 40% for p with high ROR1 levels vs 71.4% for p with low/intermediate levels (P=0.0918). Among chemotherapy-treated p, only p with low ROR1 levels responded (6.7%). PFS was 11.8 months (m) for erlotinib-treated p with low/intermediate ROR1 levels vs 5.8 m for p with high levels. PFS for chemotherapy-treated p was 5.6 and 9 m, respectively (P=0.033). Among 15 erlotinib-treated p with concomitant T790M mutations, PFS was10.8 m for p with low/intermediate ROR1 levels vs 2.7 for p with high levels (P=0.0174). Conclusions: HighROR1 expression significantly limits PFS in p with T790M mutations. ROR1-directed therapies can enhance the efficacy of erlotinib in EGFR-mutant NSCLC p overexpressing ROR1. Clinical trial information: NCT00446225.
Low cytokeratin- and low EpCAM-expressing circulating tumor cells in pancreatic cancer.
Authors: Adams Daniel , Tsai Susan , Makarova Olga V. , Zhu Peixuan , Li Shuhong , Amstutz Platte T. , Tang Cha-Mei
Published: March 2020

11046

Background: To date, detecting circulating tumor cells (CTCs) in the peripheral blood of pancreatic patients using standard immuno-capture techniques has met with limited success. As pancreatic cancer is prone to metastasize at distant sites, and therefore should have high numbers of CTCs, it is possible that immuno-capture methods are not suitable for this disease. Using a microfiltration approach, we show that CTCs are present in the peripheral blood in over 75% of pancreatic cancer patients, and that two distinct subtypes can be identified. Methods: Pancreatic patient samples were provided by Medical College of Wisconsin, Milwaukee, WI. CellSieve microfilters, with precision 7 micron diameter pores distributed in uniform arrays were employed. 7.5 mL of whole blood was diluted in pre-fixation solution and filtered through CellSieve microfilters. CTCs collected by this size exclusion technique were fixed, permeabilized, and stained with DAPI, an antibody cocktail against cytokeratin 8, 18 and 19 (FITC), EpCAM (PE), and CD45 (Cy5). CTCs, defined as cytokeratin positive and CD45 negative, were found in two distinct subtypes. One subtype had the “classic” characteristics of a CTC, with high EpCAM and cytokeratin expression, identifiable cytokeratin filamentation, and a cancer-like nuclear structure. The second subtype is indicative of a CTC undergoing epithelial-mesenchymal transition (EMT), with low or no EpCAM, weak cytokeratin expression, and a smooth oval nuclear structure. Results: The “classic” CTCs were found in ~20% (n=40) of patient samples. The EMT-like CTCs were found in ~75% of the same patient cohort. Neither cell was present in any healthy subjects (n=30). EMT-like CTCs consistently lacked EpCAM expression and commonly presented as multi-cell clusters, or microemboli, in ~40% of the cases. Conclusions: We show that two distinct CTC subtypes circulate in the blood of most pancreatic patients. The low expression of cytokeratin and EpCAM of the EMT-like subtype implies that immuno-capture based CTC isolation methods have limited utility for pancreatic cancer. Further, this subtype provides a useful strategy for tracking pancreatic CTCs over the course of treatment.
The prognostic impact from the difference of surgical procedure after ipsilateral breast cancer recurrence.
Authors: Yoshida Atsushi , Arima Nobuyuki , Nakatsukasa Katsuhiko , Okumura Yasuhiro , Iwase Takuji , Shien Tadahiko , Masuda Norikazu , Tanaka Satoru , Tanabe Masahiko , Tanaka Takehiro , Komoike Yoshifumi , Taguchi Tetsuya , Nishimura Reiki , Inaji Hideo , Yamauchi Hideko , Ishitobi Makoto
Published: March 2020

1106

Background: Mastectomy is the current standard of surgical procedure for ipsilateral breast cancer recurrence (IBTR). However, there is little evidence about a prognostic impact of surgical procedure after IBTR, due to small number of incidence of IBTR. Methods: A total of 271 consecutive patients who had histologically confirmed IBTR without distant metastases and underwent definitive surgery for the IBTR between 1989 and 2008 were included from eight institutions as a scientific research from the Japanese Breast Cancer Society. Distant Disease free survival (DDFS) rates were calculated by the Kaplan-Meier method. Univariate analysis was performed using the log rank test and multivariate analysis by Cox's proportional hazard model. Results: Of the 271 patients, 149 patients (55%) underwent lumpectomy and 122 patients (45%) underwent mastectomy after IBTR. One hundred thirty four patients (49%) did not receive radiation therapy after initial lumpectomy (52% in lumpectomy patients and 47% in mastectomy patients). The median follow-up period from definitive surgery for IBTR was 55 months. Seventy-six patients (28.2%) relapsed after IBTR (23.7% of lumpectomy patients, 33.6% of mastectomy patients). In comparing groups by type of surgery, patient and tumor demographics in each group were significantly less positive HER2 status and smaller tumor size of IBTR, and more receiving adjuvant endocrine therapy and less adjuvant chemotherapy for IBTR in lumpectomy patients compared with mastectomy patients. On univariate analysis, lumpectomy group was significantly longer DDFS than mastectomy group (p=0.012), but this difference was not appeared on multivariate analysis (p=0.35). Furthermore, only using small IBTR (<2cm) population, this difference was not appeared (p=0.71). On multivariate analysis, the time intervals from initial surgery to IBTR (<5years) (HR, 1.93; 95%CI, 1.01-3.67; p=0.047) and lymphovascular invasion of the IBTR (HR, 2.31; 95%CI, 1.18-4.52; p=0.015) were independent predictive factor for poor DDFS. Conclusions: Our study suggests that the type of surgical procedure after IBTR does not affect DDFS. Further analyses are needed. (UMIN-CTR number UMIN000008136).
Frequency of MET amplification determined by comprehensive next-generation sequencing (NGS) in multiple solid tumors and implications for use of MET inhibitors.
Authors: Palma Norma Alonzo , Palmer Gary A. , Ali Siraj M. , Stephens Philip J. , Ross Jeffrey S. , Miller Vincent A. , Lipson Doron
Published: March 2020

11068

Background: MET expression has been shown to be prognostic and possibly predictive in several tumor types and both small molecule inhibitors of MET kinase as well as monoclonal antibodies are under study as therapies for this subset of patients. However, the methods to assess MET overexpression are not well standardized. In the clinic, NGS is an advanced diagnostic method for identifying known and unknown targeted-therapy options. We therefore sought to explore the ability of NGS to detect high level MET amplifications in a general oncology population. Methods: We review here the results from analysis of 2,221 FFPE tissue samples across a range of tumor types by an NGS assay in a CLIA-certified laboratory (Foundation Medicine). We specifically report base pair substitutions, small insertions/homozygous deletions (indels), high level (> 6 copies/nucleus) amplification (amp) and select rearrangements in 189 genes, including MET. Results: MET was altered in 28/2,223 (1.2%) of patient samples. MET alterations were limited to amplifications (median 15X, range 6X-40X); no base pair substitutions, indels, or rearrangements were found. MET amp was present in 3/48 (6.2%) primary bone sarcomas, 1/20 (5.0%) myogenic sarcomas, 2/58 (3.4%) gastric, 1/38 (2.6%) kidney, 2/78 (2.5%) hepatocellular, 1/41 (2.4%) uterine, 4/188 (2.1%) unknown primary, 2/99 (2.0%) ovarian, 8/386 (2.0%) lung, 3/157 (1.9%) colorectal, and 1/109 (0.9%) pancreatic carcinomas. Conclusions: Use of NGS to characterize MET alterations in a 2,000+ patient population provides evidence for the role of NGS in identifying the future use of MET-targeted therapy in a variety of common and uncommon solid tumors. Clinical follow-up of the subset of cases treated with a MET inhibitor is ongoing.
Prognostic value of microRNAs (miRs) expression in stage I-IIIA lung adenocarcinoma (AC).
Authors: Skrzypski Marcin Tomasz , Goryca Krzysztof , Czapiewski Piotr , Jassem Ewa , Biernat Wojciech , Pawlowski Ryszard , Rzyman Witold , Jassem Jacek
Published: March 2020

11069

Background: About 50% of NSCLC patients (pts) will develop distant metastases following pulmonary resection. Currently, apart from clinical stage at diagnosis, there are no reliable factors to select high-risk pts for adjuvant chemotherapy. We previously demonstrated prognostic value of 22 miRs in frozen samples of early squamous cell lung carcinoma, and the feasibility of their assessment in formalin fixed paraffin embedded (FFPE) samples (Skrzypski et. al. J Clin Oncol 2010;28;15s). In this study, we investigated the expression of 350 microRNAs in operable AC pts. Methods: FFPE tumor samples were obtained from 80 stage I-IIIA pts who underwent curative pulmonary resection, 48% of whom subsequently developed distant metastases. Median follow-up of pts who were disease recurrence-free was 5.8 years (range, 4.0-9.1 years). RNA was isolated from tumor tissue with RecoverAll kit (Ambion). Expression of 350 miRs was analyzed by qRT-PCR (Appliedbiosystems). Raw data were normalized vs. the geometric mean of U6, RNU48 and RNU44 expression. Relative miRs expression was correlated with distant metastases-free survival (MFS) and the mean expression was compared between the groups with and without relapse. Results: Expression of 41 miRs correlated with MFS in Cox regression analysis and 21 of these showed different level in pts with and without relapse in the t-Student test (both at p<0.05 level). The top prognostic miRs included miR-99a* (p=0.006), miR-1255B (p=0.005), miR-1233 (p=0.013) and miR-1303 (p=0.03); all previosly shown to be prognostic in AC or NSCLC. We also found 14 new microRNAs (patent pending) potentially prognostic for relapse in AC. Conclusions: Expression of selected miRs may aid in prediction of distant relapse in AC pts undergoing pulmonary resection.
Clinical and analytical performance of non-small cell lung cancer biomarkers.
Authors: Anderson Steven M. , Cai Li , Tepperberg James H. , Faruki Hawazin
Published: March 2020

11070

Background: A variety of biomarkers are currently used to help guide treatment decisions for patients with non-small cell lung cancer (NSCLC). These include mutation analysis for the EGFR and KRAS genes, along with gene rearrangement analysis for the ALK and ROS1 loci. In this study we have evaluated the clinical and analytical performance features of these assays in a series of formalin-fixed paraffin-embedded (FFPE) tissue samples. Methods: FFPE samples submitted for analysis of the EGFR, KRAS, ALK and ROS1 genes were evaluated using molecular and FISH assays. EGFR mutation analysis was performed using Sanger nucleic acid sequencing methods for exons 18-21. KRAS mutations were detected using allele specific PCR or pyrosequencing methods. Rearrangements involving the ALK gene were detected using break-apart FISH probes (Abbott Molecular). Genetic alterations involving the ROS1 gene were determined using FISH probes (Kreatech Diagnostics). Over 6,200 test results for these 4 markers are included in this analysis. Results: Mutations in the EGFR gene were detected in 10.1% of samples evaluated (n=3,872). A slightly higher percentage of samples from female patients (13%) had a detectable mutation compared to samples from males (7%) (chi-square p<0.0001). Deletions in exon 19 (51%) were the most common alterations detected, followed by point mutations in exon 21 (35%). KRAS mutations were detected in approximately 22% of specimens. ALK gene rearrangements were observed in 3.1% of samples (n=1,524). Specimens from individuals <50y of age were more likely to provide a positive result (11%) compared to samples from individuals >50y of age (2.5%) (chi-square p<0.0001). Gene amplification for the ALK gene was a common finding in the NSCLC samples evaluated. ROS1 alterations were detected in 2.8% of the specimens. In this cohort, no specimens were positive for both an EGFR mutation and an ALK gene rearrangement. Conclusions: Biomarker testing is well established in clinical practice for NSCLC, with results from the tests used to guide important therapy decisions. Assays for biomarkers such as EGFR, KRAS, ALK and ROS1 are robust, allowing for the analysis of multiple analytes in FFPE samples, even when the amount of tissue may be limiting.
Prognostic value of waterfall plots with the addition of nontarget lesion data.
Authors: Mietlowski William Leonard , Stein Andrew Marc , Bao Weichao , Waltzman Roger J. , Wood Patricia Ann
Published: March 2020

11082

Background: In patients with solid tumors, the use of a waterfall plot displaying the best percentage change in sum of the longest diameters of target lesions per patient is a common way to depict anti-tumor activity for cytostatic agents (Booth 2008, Dhani 2009). This representation assumes that the best percentage change represents the maximum anti-tumor activity for each patient. Information about new lesions and/or changes in non-target lesions is not incorporated in the waterfall plot; yet these additional events were found to be significant prognostic factors for overall survival adjusting for change in the target lesions (Litiere 2012, Suzuki 2012). Methods: We analyzed two phase III lung cancer trials of 1st and 2nd line combination chemotherapy ± ASA404 (ATTRACT-1,n=1299, ATTRACT-2, n= 920). For patients whose best response in the target lesions was shrinkage, we calculated how often this best response was synchronously accompanied by non-target disease progression. Results: See Table. Conclusions: There can be substantial tumor shrinkage in target lesions synchronously with progressive disease outside the target lesions. Therefore, graphical displays of anti-tumor activity should consider incorporating new and non-target lesion information, as well as target lesion tumor burden. We propose an extended waterfall plot presenting a more complete assessment of anti-tumor activity by incorporating non-target lesion information. We illustrate its utility in an additional data set, the RECORD-1 phase III renal cell cancer trial. Clinical trial information: NCT00662597. [Table: see text]
Application of next-generation sequencing (NGS) for evaluation of advanced epithelial cancers: A single institution experience.
Authors: Boland Patrick McKay , Skarbnik Alan P. , Cristofanilli Massimo , Alpaugh R. Katherine , Olszanski Anthony J.
Published: March 2020

11092

Background: The use of molecular targeted therapeutic agents may require the application of sophisticated diagnostic technologies for patients’ selection. Next generation DNA sequencing (NGS) has the ability to identify genetic alterations (GA) including mutations, copy number alterations, insertions/deletions, and rearrangements in tumor specimens. We sought to evaluate patients with advanced and refractory epithelial tumors to detect potentially actionable molecular abnormalities. Therapeutic intervention driven by GA findings was determined solely by the patient’s treating physician. Methods: Tumor samples from 77 patients ≥ 18 years old with any solid malignancy were sequenced. NGS of 186 genes was performed by FoundationOne through funding from Foundation Medicine using archival or newly acquired tumor tissue. Results: Seventy-four patients had specimens with adequate material for DNA extraction and analysis. Characteristics: 74% female, median age 55 years (19-82). Tumor sites included inflammatory breast (50%), colon (12%), unknown primary (5%) and other (33%). At least one genetic alteration was seen in 71 (96%) patients. The most common GA included mutations in 65 (60%) samples revealing TP53 (32%), KRAS (10%), PIK3CA (8%), and APC (6%) and amplifications in 38 (35%) samples which included MYC (18%), MCL1 (14%), CCND1 (12%), and ERBB2 (7%). Copy number loss (4%), fusion (1%) and deletions (2%) were also discovered. An actionable GA was seen in 46 of 74 patients successfully tested (62%), with 54% of GAs being amplifications and 43% mutations. Patients had a median of 3 GA (range 0-7). One patient with anal cancer had a concomitant PIK3CA mutation and amplification. NGS in association with immunohistochemistry helped identify site of origin for one patient with an occult primary. Conclusions: NGS identified GAs in the majority of patients with advanced epithelial cancers, including actionable abnormalities in a large fraction of this heterogeneous population. NGS shows promise in the diagnostic evaluation of advanced malignancies. Future studies should include the potential prognostic implications of genomic-driven personalized therapy.
MRI apparent diffusion coefficient in a murine orthotopic glioblastoma model as a clinically translatable early readout of efficacy for AMG 595, an antibody drug conjugate targeting EGFRvIII.
Authors: Yerby Brittany , Estrada Juan , Silva Matthew D. , Hamblett Kevin J. , Coberly Suzanne K. , Hill John S. , Ungersma Sharon E. , Beltran Pedro J.
Published: March 2020

11095

Background: Epidermal growth factor receptor variant III (EGFRvIII) is a constitutively active mutant of EGFR present in thirty to fifty percent of glioblastoma (GBM) patients. AMG 595, currently in Phase I trials, is composed of a fully human anti-EGFRvIII-specific antibody conjugated to the maytansinoid DM1 via a non-cleavable linker. The MRI apparent diffusion coefficient (MRI ADC) has been shown to correlate with tissue cellularity, and changes in MRI ADC can be an early indicator of therapeutic efficacy. The aim of this work is to evaluate MRI ADC as a clinically translatable early readout of tissue changes due to AMG 595 treatment in a murine orthotopic GBM model. Methods: D317 human GBM cells were intracranially injected into in female CB17 SCID mice at Day 0. Mice were randomized at Day 7, using tumor volumes calculated by MRI, and were treated with vehicle, 6.5, 11, or 22 mg/kg AMG 595 i.v. twice per week, or temozolomide 10 mg/kg p.o. daily five days per week (N=8/group). MRI was repeated at days 14 and 21. Tumor volumes were manually traced on multi-slice T2-weighted RARE images covering the entire tumor volume. The mean MRI apparent diffusion coefficients for each tumor in the vehicle and 22 mg/kg AMG 595-treated groups were calculated from diffusion-weighted spin echo images (b = 100-1200 s/mm2). Results: A dose-dependent effect of AMG 595 on tumor volume was observed at Day 21; growth was inhibited in both the temozolomide and AMG 595-treated groups (22 and 11 mg/kg) relative to vehicle (p<0.0001). At Day 14, this significant treatment effect on tumor volume was not yet detectable. However, mean MRI ADC values were already significantly higher in the AMG 595 (22 mg/kg) treated group than in the vehicle group (23% higher at Day 14, p<0.01 vs vehicle; 32% higher at Day 21, p<0.0001 vs vehicle). The increase in MRI ADC in the AMG 595-treated group preceded observable tumor growth inhibition in the AMG 595-treated animals. Conclusions: Increases in tumor MRI ADC in response to AMG 595 treatment precede measurable inhibition of tumor growth, supporting the use of MRI ADC as a clinically relevant early biomarker for therapeutic efficacy.
Cross-species synthetic lethal interaction screening as a strategy for the identification of novel therapeutic targets in cancer.
Authors: Shen John P. , Srivas Rohith , Bojorquez-Gomez Ana , Licon Katherine , Li Jian Feng , Sobol Robert W. , Ideker Trey
Published: March 2020

11105

Background: Mutation, deletion, or epigenetic silencing of tumor suppressor genes is a near universal feature of malignant cells. However, therapeutic strategies for restoring the function of mutated or deleted genes have proven difficult. Synthetic lethality, an event in which the simultaneous perturbation of two genes results in cellular death, has been proposed as a method to selectively target cancer cells. Identifying and pharmacologically inhibiting proteins encoded by genes that are synthetic lethal with known tumor suppressor mutations should result in selective toxicity to tumor cells. Methods: To identify candidate target proteins we measured all pair-wise genetic interactions between all known orthologs of human tumor suppressor genes (162 genes) and all orthologs of druggable human proteins (~400 genes) in the model organism S. Cerevisiae. Analysis of the data uncovered 2,087 distinct synthetic lethal interactions between a tumor suppressor and druggable gene. A computational algorithm was then developed to identify those interactions which were likely to be conserved in humans based on conservation of the synthetic lethal relationship in the distant fission yeast S. pombe. Results: Our bioinformatic analysis suggested a high probability of conservation of the synthetic lethal interactions between the yeast RAD51 (ortholog of BRCA1) and RAD57 (ortholog of XRCC3) with HDA1 (a histone deacetylase; HDAC). We confirmed this by treating LN428 cells with stable lentiviral knockdown of BRCA1 or XRCC3 with the HDAC inhibitors vorinostat (SAHA) and entinostat (MS-275). Both the BRCA1 and XRCC3 knockdown cell lines were significantly more sensitive to HDAC inhibition relative to wild-type (non-silencing lentiviral control) cell line (Table). Conclusions: These results demonstrate that high-throughput approaches for screening synthetic lethal interactions in model organisms such as S. cerevisiae and S. pombecan serve as a valuable resource in helping to identify novel therapeutic targets in human cancer. [Table: see text]
Highly active combination of BRD4 antagonist and histone deacetylase inhibitor against human AML.
Authors: Bhalla Kapil N. , Fiskus Warren , Peth Karissa
Published: March 2020

11107

Background: The bromodomain-containing protein 4 (BRD4) is a member of the BET (bromodomain and extraterminal) protein family, which regulate the assembly of chromatin complexes and transcriptional regulators at the target gene promoters, e.g., of Myc and BCL2. JQ1 competitively binds and inhibits the bromodomains of the BET proteins, including BRD4. Treatment with JQ1 attenuates Myc and BCL2 levels and promotes growth arrest, differentiation and apoptosis of AML cells. Methods: Here, we determined the growth inhibitory, differentiation and apoptotic effects of JQ1 (100 to 500 nM) alone and in combination with low concentrations of the deacetylase inhibitor panobinostat (PS) (5 to 20 nM) (gift from Novartis) in cultured (OCI-AML3, MV4-11 and MOLM13) and primary AML blast progenitor cells. We also determined the in vivo anti-AML activity of JQ1 (50 mg/kg/day IP) and/or PS (5 mg/kg/MWF IP) for 3 weeks against the OCI-AML3 xenograft in the NOD/SCID mice. Results: Treatment with JQ1 decreased the binding of BRD4 at the chromatin associated with Myc and BCL2 promoters (by ChIP-qPCR) and attenuated the mRNA and protein levels of Myc, BCL2 and cyclin D1, while increasing the levels of p21, CEBPα, BIM and cleaved PARP in the cultured and primary AML blasts. This was associated with an increase in the % of G1 phase cells, attenuation of the colony growth, and induction of apoptosis of the cultured (OCI-AML3 >MV4-11 >MOLM13) and primary AML cells. JQ1 treatment also dose-dependently inhibited the viability of CD34+ primary AML progenitor cells (n=10). Gene expression microarray analysis revealed a JQ1 treatment-associated signature of the most up- and down-regulated mRNAs that correlated with its anti-AML effects. Co-treatment with JQ1 and PS synergistically induced apoptosis of AML cells (by isobologram analysis; CI < 1.0). As compared to the mice treated with vehicle alone, or with each agent alone, co-treatment with JQ1 and PS, without inducing toxicity, significantly improved the survival of the OCI-AML3 engrafted NOD/SCID mice (p < 0.01). Conclusions: These pre-clinical findings warrant further in vivo evaluation of the efficacy of the combination of a BRD4 antagonist and PS, and its predictive biomarkers, against human AML.
NGS-based targeted RNA sequencing for expression profiling and relative quantitation of specific gene isoforms and fusions in tumor-specific panels.
Authors: Vansant Gordon , Landers Mark , Vo Lien , Oades Kahuku , Kim Hyunsoo , Lee Jerry , Meredith Rhonda , Lee Byung-In , Monforte Joseph
Published: March 2020

11108

Background: Gene expression signatures have become a useful tool for the identification of tumor subtypes and response to specific therapies. Expression of tumor, metastatic and macrophage specific transcripts utilizing alternative promoters and transcriptional start sites can further characterize these tumors . NGS is a powerful tool for gene expression analysis, however larger sample input requirements (>100ng) and excessive sequencing depth requirements (30-40M tags/sample) to detect the expression of rare isoforms or fusions in tumor samples are prohibitive for clinical assay development. We describe the development of a targeted RNA sequencing assay for the relative quantitation of specific gene expression signatures, known splice variants and gene fusions from less than 100 ng of starting material in a single tube universal amplification format. Methods: Primers for 52 genes, isoforms and gene fusion products were designed using the universal amplification strategy. 10 ng of RNA from 5 matched tumor/adjacent normal breast cancer tumor pairs were assayed. Libraries were prepared for sequencing by emPCR and sequenced on Ion Torrent PGM. Data were aligned via TMAP. Relative expression was determined vs. housekeeping genes or wild type transcripts. Results: All gene targets were detected at significant levels in at least one tumor sample. Robust expression profiling (5 log dynamic range) was obtained from FFPE macrodissected tumor and normal samples with as little as 200K reads/sample. Immune specific transcripts demonstrated differential expression (CCL3, AIF, FCGR3A and CSF1) across patients and matched pairs as well as an upregulation of CXCL12, indicative of tumor associated macrophages. Conclusions: Targeted RNAseq demonstrates detection and quantitation of relative expression levels of not only tumor subclass specific gene expression signatures, but immune cell specific transcripts from 10ng of FFPE derived total RNA derived from macrodissected tumor samples. The lower input requirements, quicker turnaround time and incredible sensitivity of targeted RNAseq make this assay a useful tool for clinical assay development.