bluuebunny/crossref_metadata_2025 huggingface dataset:
An analysis of ERBB2 alterations (amplifications and mutations) found by next-generation sequencing (NGS) in 2000+ consecutive solid tumor (ST) patients.
Authors: Palmer Gary A. , Wang Kai , Miller Vincent A. , Yelensky Roman , Stephens Phil , Ross Jeffrey S. , Lipson Doron
Published: March 2020

11000

Background: Testing for ERBB2 amplification by FISH and IHC is routine in breast and gastro-esophageal cancer. There are 3 approved and multiple targeted therapies in clinical trials that rely on the results of these tests. ERBB2 amplification and activation by mutation/fusion has also been described in a wide variety of other ST. As these alterations are not routinely tested for but may predict response to anti-ERBB2 agents we sought to determine their frequency in an unselected cohort of specimens from advanced ST patients. Methods: We reviewed genomic profiles from the first 2,223 formalin-fixed, paraffin-embedded specimens received and analyzed by our CLIA-certified lab (Foundation Medicine) with our NGS platform. 3,230 exons in 182 cancer-related genes and 14 genes frequently rearranged were assayed for base pair substitutions, small insertions/deletions (indels), amplifications, and rearrangements. Results: 110/2,223 (4.9%) specimens had 116 ERBB2 alterations: 67 (58%) amplifications, 29 (25%) substitutions, 16 (14%) indels, 2 (2%) splice site variants and 2 (2%) translocations, including a potential fusion. Six samples (5%) had multiple alterations, and two had both ERBB2 substitution and amplification. 14 ST types had evidence of ERBB2 alterations including 29% of esophageal, 20% of uterine, 14% of breast, and 12% of stomach carcinomas. 6% of all lung cancer samples had ERBB2 alterations. Amplifications predominated, but lung specimens had predominantly indels. Durable responses exist to anti-ERBB2 agents in STs with activating ERBB2 mutations. Conclusions: Use of a broad NGS panel identifies an unprecedented number of actionable genomic changes including a significant rate of ERBB2 alterations across 14 different solid tumor types. The discovery of unanticipated ERBB2 amplifications and activating mutations in a wide variety of ST highlights the need to study a broad range of genes at a high level of sensitivity and specificity when searching for novel targets of therapy. Widespread use of this approach could provide more treatment options and enable more rapid accrual to ongoing and planned trials of agents targeting pathways under study.
Amplification of the MET receptor to drive resistance to anti-EGFR therapies in colorectal cancer.
Authors: Bardelli Alberto , Corso Simona , Bertotti Andrea , Hobor Sebastijan , Siravegna Giulia , Sartore-Bianchi Andrea , Migliardi Giorgia , Galimi Francesco , Lauricella Calogero , Zanon Carlo , Amatu Alessio , Gambacorta Marcello , Diaz Luis A. , Velculescu Victor E. , Sausen Mark , Comoglio Paolo M. , Trusolino Livio , Di Nicolantonio Federica , Giordano Silvia , Siena Salvatore
Published: March 2020

11005

Background: The anti EGFR monoclonal antibodies cetuximab and panitumumab are used to treat metastatic colorectal cancer patients but their clinical efficacy is limited by the development of acquired resistance. We recently reported that secondary KRAS mutations are responsible for acquired resistance in approximately 50% of the patients who initially respond to cetuximab or panitumumab (Misale et al., Nature 2012; Diaz et al., Nature 2012). Here we studied the molecular bases of relapse in CRC patients who do not develop KRAS mutations during the course of anti-EGFR therapy. Methods: Next generation sequencing was applied to tumor biopsies to identify genetic alterations associated with relapse to cetuximab and panitumumab in mCRC patients. Detection and quantitation of genetic alterations in circulating tumor DNA was used to monitor the occurrence of KRAS mutations and MET amplification in blood samples. Results: Molecular analyses of tumor biopsies from patients who did not develop KRAS mutations during anti-EGFR therapy revealed high level of amplification of the MET proto-oncogene in 3/5 cases. Quantitative PCR, FISH and IHC analysis confirmed high level of MET amplification in the post-therapy samples but not in the matched pre-treatment tissues. We developed a PCR based assay to detect the presence of the MET amplicon in circulating, cell-free, DNA. We found that MET amplification could be detected in the blood as early as 3 months after initiation of anti EGFR therapy. To functionally evaluate the role of MET amplification on resistance to anti EGFR antibody therapies we exploited patient-derived CRC xenografts (‘xenopatients). We found that (2/2) xenopatients established from MET amplified tumors were completely refractory to cetuximab but showed sensitivity to the Met inhibitor crizotinib. Conclusions: Amplification of the MET proto-oncogene is responsible for acquired acquired resistance to anti-EGFR antibody therapy in a subset of CRCs. The emergence of MET amplification in circulating, cell-free, DNA may be used to select patients most likely to benefit from anti MET therapies.
Early PET/CT scan compared with RECIST to predict long-term outcome of patients with liver metastases from colorectal cancer treated with preoperative chemotherapy plus bevacizumab.
Authors: Piccirillo Maria Carmela , Lastoria Secondo , Nasti Guglielmo , Caraco Corradina , Aloj Luigi , Arrichiello Cecilia , Ottaiano Alessandro , Izzo Francesco , De Lutio Elisabetta , Albino Vittorio , Romano Carmen , Palaia Raffaele , Daniele Gennaro , Di Maio Massimo , Giordano Pasqualina , Signoriello Simona , Delrio Paolo , Iaffaioli Rosario Vincenz , Romano Giovanni , Perrone Francesco
Published: March 2020

11008

Background: Early changes in tumor metabolism measured with positron-emission-tomography/computerized tomography (PET/CT) could predict the long-term efficacy of treatment better than dimensional RECIST response. Methods: We performed PET/CT before and after 1 cycle of treatment in patients with resectable liver metastases from colorectal cancer, within a phase II trial of preoperative FOLFIRI plus bevacizumab. For each lesion, the maximum SUV (SUVmax) and the total lesion glycolisis (TLG) were determined. For both, based on previous studies, a ≤-50% change from baseline was used as threshold for significant response. Metabolic response was categorized no/yes by using three different methods that enter into the calculation (i) the largest observed value (highest SUVmax/TLG), or (ii) the sum of all the observed values (Total SUVmax/TLG), or (iii) each observed values (SUVmax/TLG-by-lesion). Standard RECIST response was assessed after 3 months of treatment. The association between metabolic and RECIST response was tested with the Mc Nemar’s test and their agreement was expressed as Kappa statistics; the ability to predict progression-free (PFS) and overall (OS) survival was tested with Log-rank test and a multivariable Cox model. Results: 33 patients were analyzed. After treatment, there was a notable decrease of all PET/CT parameters, with a median change of -33.9% for the highest SUVmax, -61.5% for the highest TLG, -34.9% for the total SUVmax, and -65.5% for the total TLG. The association of SUV-based metabolic response (but not the TLG-based) with RECIST was statistically significant. However, the agreement between RECIST and PET/CT responses was consistently small. PFS and OS were significantly longer among PET/CT responding patients, whichever the measure used. On the contrary, no significant outcome difference was evident according to RECIST response. Conclusions: Early PET/CT response was significantly predictive of long-term outcomes during preoperative treatment of patients with liver metastases from colorectal cancer and its predictive ability was higher than that of RECIST response. Clinical trial information: 2006-006572-38.
Olaparib monotherapy in patients with advanced cancer and a germ-line BRCA1/2 mutation: An open-label phase II study.
Authors: Kaufman Bella , Shapira-Frommer Ronnie , Schmutzler Rita K. , Audeh M. William , Friedlander Michael , Balmaña Judith , Mitchell Gillian , Fried Georgeta , Bowen Karin , Fielding Anitra , Domchek Susan M.
Published: March 2020

11024

Background: The oral PARP inhibitor olaparib has shown antitumor activity as monotherapy in patients (pts) with breast and ovarian cancer with gBRCA1/2 mutations. This multicenter non-comparative study evaluated whether tumors in gBRCA1/2 mutation carriers are responsive to olaparib regardless of tumor type (NCT01078662). Methods: Heavily pretreated pts with advanced cancer refractory to standard therapy (98% of breast cancer pts had ≥3 lines of prior chemotherapy for metastatic disease) and with a gBRCA1/2 mutation, received olaparib 400 mg bid (capsule) until disease progression. Primary objective: tumor response by RECIST 1.1. Secondary objectives: PFS, OS and safety. Results: 298 pts received treatment and were evaluable. Enrollment is complete, 33 pts remain on study. Median duration of treatment in this heavily pretreated population was 5.5 months (range <1–28.5 months). Most common AEs (generally grade 1/2) were fatigue (59%), nausea (59%) and vomiting (37%). Grade ≥3 AEs were reported for 162 pts (54%); most common was anemia (17%). 11 pts (4%) had AEs that led to treatment discontinuation. Conclusions: The observedtumor response rates indicate antitumor activity of olaparib monotherapy in gBRCA mutated pts with advanced cancer refractory to standard therapy. A clinical benefit was seen in prostate and pancreatic cancer and activity in ovarian and breast cancer was confirmed. Prolonged responses to olaparib across all tumor types support the hypothesis that therapy directed against a genetically-defined target has activity regardless of anatomic organ of origin. Olaparib was generally well tolerated with toxicities consistent with prior studies. Clinical trial information: NCT01078662. [Table: see text]
Components of homologous recombination and translesion synthesis (TLS) in pemetrexed/cisplatin-treated non-small-cell lung cancer (NSCLC) patients (p).
Authors: Lopez-Vivanco Guillermo , Marti Thomas , Kotov Ilya N. , Chaib Imane , Ponce-Aix Santiago , García Campelo Rosario , Sanchez Jose Miguel , Artal Angel , Bover Isabel , Taron Miquel , Sanchez-Ronco Maria , Rolfo Christian Diego , Massuti Bartomeu , Stahel Rolf A. , Rosell Rafael ,   Spanish Lung Cancer Group
Published: March 2020

11028

Background: REV3, the catalytic subunit of the TLS polymerase ξ, can continue replication past DNA adducts. Depletion of REV3 sensitizes A549 lung cancer cells to cisplatin. REV3 expression is part of a gene signature that predicted pemetrexed sensitivity in 17 NSCLC cell lines. Homologous recombination and TLS pathways have non-redundant functions in response to cisplatin. We hypothesized that low REV3 mRNA expression – alone or in combination with low expression levels of genes involved in homologous recombination – could correlate with better outcome to cisplatin/pemetrexed in NSCLC. Methods: REV3, BRCA1, RAP80, TS and AEG1 mRNA was examined by quantitative RT-PCR and categorized by terciles. Expression of each gene was correlated with outcome in 47 cisplatin/pemetrexed-treated NSCLC p. Results: 63.8% male; 47% smokers; 80.9% ECOG PS 1; 80.8% adenocarcinoma. Overall response rate was 51%, with no differences according to expression levels of any of the genes. Progression-free survival (PFS) for p with low, intermediate and high BRCA1 levels was 13.4, 5.5 and 3.9 months (m), respectively (P=0.005). Similar differences in PFS were observed according to TS (P=0.003) and AEG1 (P<0.001) expression. Hazard ratio (HR) for PFS for p with high BRCA1 levels was 4 (P=0.002). Overall survival (OS) for p with low, intermediate and high BRCA1 levels was 29.7, 7.4 and 6.3 m, respectively (P=0.05). Similar differences in OS were observed according to TS (P=0.005) and AEG1 (P=0.001) expression.HR for OS for p with high BRCA1 levels was 3.6 (P=0.004). There were no differences in PFS or OS according to REV3 or RAP80 levels. However, the joint effect of BRCA1 and REV3 was significant for predictive modeling. PFS for p with low, intermediate and high levels of both genes was 14.9, 7.2 and 2.8 m, respectively (P=0.001). OS for p with low, intermediate and high levels of both genes was 29.7, 7.8 and 6.3 m, respectively (P=0.04). Conclusions: Low BRCA1 expression predicts longer PFS and OS in pemetrexed/cisplatin-treated NSCLC p. Low TS and AEG1 levels have similar predictive value. Analysis of these genes could be useful for customizing pemetrexed/platinum chemotherapy.
Differential luminal breast cancer (LBC) reprogramming in response to BIBF1120 (BIBF) or bevacizumab (B).
Authors: Quintela-Fandino Miguel , Navarro Paloma , Mondejar Tamara , Sanchez Jesus , Garcia Elena , Gomez Gonzalo , Gonzalez-Pisano David , Colomer Ramon
Published: March 2020

11033

Background: FVB MMTV PyMT is a great mouse model for studying LBC due to its stochastic nature, immune system preservation, natural growth kinetics and ER/HER2 status. We approached the transcriptomic (T), proteomic (P), phospho-proteomic (P-P) and metabonomic (M) layers of regulation of the tumor phenotype attempting to unravel mechanisms of resistance to antiangiogenic drugs (ADs) and find new targets. Methods: Treatments started when tumors measured 0,1cm3; with the VEGFR/PDGFR inhibitor BIBF 85mg/kg/day; a murine analog of B 10mg/kg/biweekly; or vehicle (V). Tumors were harvested at time 0, week 1 (T1), or at sacrifice (Tend, 1cm3). Cd31 and pimomidazole measured vascularization and O2. T; P/P-P; and M layers were interrogated with sure print 3G arrays; titanium separation - HPLC fractionation - orbitrap spectrometer run; and GC/MS and LC/MS. Gene ontology pathways were discovered with GSEAS; kinase networks by consensus domain analysis; metabolic pathways rearrangement with in-house built bioinformatic tools. Pairwise comparisons included >15 mice/condition. All results: p or false-discovery ratio<0,05. Results: Total mice in study:2,327. BIBF-treated showed: early (T1 vs. 0) ↓glycolysis (55%) and ↑pyruvate diversion to Krebs mediated by ↓800% phosphorylation of PDH, ↑ ketones uptake and degradation (220%), mediated by 2700% ↑AMPK activity; late (Tend) 900% ↑ lipid uptake and aerobic degradation mediated by >800% ↑ ERK/B-adrenergic/PKA and PPARg (1600%) signaling. All changes supported by GSEAS. Adding blockade of AMPK, ERK, B-adrenergic or PPARg (ShRNAs or compounds) to BIBF yielded at least 50% growth delay (Anova). Tend B-treated tumors showed 210% ↑degradation of tryptophan to immunosuppressive (IS) kynurenine, and >200% ↑ levels of IS/vasodilator prostaglandins. Celecoxib added to B delayed growth 90%. B and V induced similar ↑of hypoxic/necrotic areas (T0-1-end: 5%-15%-52%, Poisson), no change in CD31. BIBF avoided necrosis/hypoxia despite >30% ↓ in CD31. Conclusions: Our strategy identified differential mechanisms of action of ADs in LBC. BIBF induces a switch from Warburg to aerobic metabolism; B reprograms the immune response. Regulators of these processes constitute targets.
Targeted next-generation sequencing (NGS) of circulating tumor cells (CTCs) in hormone-sensitive prostate cancer (HSPC).
Authors: Liu Stephen V. , Dempsey Paul W. , Strauss William , Xu Yucheng , Xu Tong , Pinski Jacek K. , Dorff Tanya B. , Quinn David I. , Triche Timothy J. , Winer-Jones Jessamine , De Fusco Andre , McMurdie Janine , Goldkorn Amir
Published: March 2020

11040

Background: Recently a succession of new hormonal therapies has emerged, highlighting the need for biomarkers to guide the management of HSPC. Biomarker development in HSPC has been hampered by the absence of primary tumor tissue in men who undergo radiation or present with metastatic disease. CTCs can address this challenge by providing real-time cancer tissue for biomarker analysis in HSPC. To test this approach we conducted a pilot of CTC capture and targeted NGS in HSPC. Methods: Under IRB approval, blood samples from patients with HSPC were labeled with EpCAM ferrofluid and placed into the LiquidBiopsy platform (Cynvenio Biosystems, Inc.), an immunoaffinity-based microfluidic device tailored to query genomic events. CTCs were identified by CK, CD45 and DAPI expression. A matched WBC pellet served as a control representing germline sequence. Amplicon libraries were generated using Life Technologies AmpliSeq 2.0 and sequenced on an Ion Torrent platform. Somatic single nucleotide variants (SNV) present in CTCs but not in WBC were identified. Results: CTCs were detected in all 8 patients with HSPC (CTC median 64.5, range 17-217). Germline variants were consistently detected in matched CTC and WBC samples. Significant SNVs (occurring in > 1% of DNA in a sample) were found in 4 of 8 CTC samples (range 1-5 SNVs/sample, frequency 1.2%-11.9% with 620X-14,422X coverage). Notably, 3 patients had biochemical recurrence only (no clinical metastases) yet still yielded CTCs associated with SNVs in KIT, APC, RET, SMAD4 and PTEN. One patient who had untreated metastatic disease had the highest number of CTCs which harbored 4 SNVs. Conclusions: This pilot demonstrates the feasibility of using CTCs as real-time disease relevant substrate for NGS to identify personalized genomic targets in HSPC. A high number of CTCs were detectable in all patients and CTC germline variants correlated with matched WBC controls. Encouragingly, even with a relatively narrow, primary tumor-derived AmpliSeq platform, cancer relevant SNVs were detected in half of the patients including those with only biochemical recurrence, making targeted NGS of CTCs a promising approach for biomarker discovery and validation in HSPC.
Persistence of HER2 overexpression on circulating tumor cells in patients after systemic treatment for HER2-positive breast cancer: Follow-up results of the German Success B trial.
Authors: Neugebauer Julia Katharina , Rack Brigitte Kathrin , Jaeger Bernadette Anna Sophia , Andergassen Ulrich , Pestka Aurelia , Fasching Peter A. , Friedl Thomas W. P. , Mouarrawy Doraid , Clemens Michael R. , Ziemendorff Gabriele , Heinrich Georg , von Abel Ekkehard , Schneeweiss Andreas , Beckmann Matthias W. , Pantel Klaus , Janni Wolfgang
Published: March 2020

11043

Background: The discordance between HER2-expression on circulating tumor cells (CTC) in peripheral blood and the primary tumor has already been shown by our study group for early breast cancer patients with HER2-positive tumors. Here, we compare the results to CTC prevalence and HER2-status of CTC after adjuvant chemotherapy. Methods: The SUCCESS B trial compares FEC-Docetaxel vs. FEC-Docetaxel-Gemcitabine and HER2-targeted therapy as adjuvant treatment for patients with early, HER2-positive, node positive or high risk node negative primary breast cancer. We prospectively analyzed 23ml peripheral blood before and after chemotherapy. CTC and HER2-status were assessed with the CellSearchSystem (Veridex, USA). After immunomagnetic enrichment with an anti-Epcam-antibody, cells were labeled with anti-CK 8/18/19, anti-CD45 antibodies as well as a fluorescein conjugate antibody for HER2-phenotyping. Cutoff for CTC positivity was ≥ 1 CTC. HER-positivity of CTC was assigned if at least one CTC showed strong HER2 staining (3+). Results: CTCs and their HER2-status both before and after chemotherapy were available for 392 patients. In 179 (45.7%) patients no CTC were detected before and after chemotherapy. CTC status changed from positive before to negative after chemotherapy in 104 (26.5%) patients and from negative before to positive after chemotherapy in 69 (17.6%) patients, while 40 (10.2%) patients had a consistently positive CTC status. Patients were significantly more likely to change their CTC status from positive to negative than from negative to positive (p = 0.01). Of the 40 patients with CTC both before and after chemotherapy, 14 (35%) patients had HER2-positive CTC before and after therapy, and 9 (22%) patients had HER2-negative CTC at both time points. 7 (18%) patients had HER2-positive CTC before but not after chemotherapy, while 10 (25%) patients showed the reverse pattern (p = 0.63). Conclusions: Cytotoxic treatment does not seem to influence the HER2-status on CTC. Follow-up data within the Success B trial will analyze the relevance of the HER2-expression of CTC to predict the efficacy of targeted treatment.
Characterization and identification of specific EGFR mutations in circulating tumor cells (CTCs) isolated from non-small cell lung cancer patients using antibody independent method, ApoStream.
Authors: Tran Hai T. , Melnikova Vladislava O. , Tsao Anne S. , Fossella Frank V. , Johnson Faye M. , Papadimitrakopoulou Vassiliki , Garza Miguel , Neal Chris , Hasegawa Dave , Kruempel Amy , Wu Grant , Richardson Katherine , Lewis Marcia E. , Legendre Benjamin Jr , Anderes Kenna Lynn , Davis Darren W. , Heymach John
Published: March 2020

11044

Background: A variety of methods for isolation of CTCs of epithelial origin are available; most employ antibodies to epithelial cell adhesion molecule (EpCAM). Using classic phenotypic definition, a CTC is nucleated, cytokeratin CK(+), CD45(-) cell. However, some CTCs may elude capture as they originate from primary tumor cells which have undergone epithelial-mesenchymal transition (EMT). We report here the use of ApoStream, a novel dielectrophoresis field-flow-assisted, antibody-free method to isolate CTCs from blood. Methods: Blood was collected from consented NSCLC patients and processed using ApoStrea. For CTC enumeration comparison, CellSearch FDA-approved kit was used. Isolated cells were evaluated with multiplexed immunofluorescent assay and laser scanning cytometry analysis were applied to identify multiple combinations of positive and/or negative staining for CK/CD45/DAPI and EpCam. To determine specific EGFR mutations from captured CTCs, samples were analyzed using Improved and Complete Enrichment with CO-amplification at Lower Denaturation temperature (ICE COLD-PCR). Results: Blood samples from 32 NSCLC patients and 3 healthy volunteers were processed. ApoStream isolated 0 to 65 CK(+)/CD45(-) CTCs(n=32) and CellSearch isolated 0 to 13 EpCAM(+)/CK(+)/CD45(-) CTCs(n=7). Additionally, ApoStream™ recovered 37-3536 CK(-)/CD45(-) and 4-10702 CK(+)/CD45(+) cells. EpCAM expression was detected in 7-100% of CK(+)/CD45(-) and 0-5% of CK(-)/CD45(-) cells, and 18-100% of CK(+)/CD45(+) cells. EGFR mutations [exon 19 deletion and exon 21 L858R] were determined and found to be concordant when compared to tumor tissue analysis by Sanger sequencing. Conclusions: The ApoStream platform enriched EpCAM(+) and EpCAM(-) CTCs from the blood of NSCLC patients demonstrating utility in recovering cancer cells with multiple phenotypes. From recovered CTCs, detection of EGFR mutations was possible indicating the clinical relevance and potential utility of CTCs as an alternative to tissue biopsy. Complete mutation analysis will be presented.
Prognostic significance of ATP-binding cassette (ABC) and solute carrier (SLC) transporters in pancreatic ductal adenocarcinoma (PDAC).
Authors: Mohelnikova-Duchonova Beatrice , Brynychova Veronika , Oliverius Martin , Hlavsa Jan , Honsova Eva , Mazanec Jan , Melichar Bohuslav , Kala Zdenek , Soucek Pavel
Published: March 2020

11051

Background: Resistance against anticancer drugs significantly limits the clinical use and efficacy. One of the most important mechanisms of the multidrug resistance is low accumulation of the drug in cancer cells caused by an increased efflux (mediated mainly by ABCs) or by a decreased uptake (mediated by some members of SLCs). The aim of the present study was to investigate prognostic importance of 13 anticancer drug-relevant SLCs and all 49 human ABCs in PDAC patients in association with clinical and pathological characteristics and the tumor KRAS mutation status. Methods: Tumors and adjacent non-neoplastic pancreatic tissues were obtained from 50 patients with histologically verified PDAC. The transcript profile of ABCs and SLCs was assessed using quantitative real-time PCR. KRAS mutations in exon 2 were assessed by high-resolution melting analysis and sequencing. Associations of transcript levels with clinical parameters were assessed by non-parametric Mann-Whitney tests. The Kaplan-Meier method with the log-rank test and Cox regression were used for analysis of overall survival (OS). Results: Most transporters investigated were deregulated in PDAC and 14 ABCs or SLCs were associated with clinical characteristics. Up-regulation of 7 drug resistance-associated ABC efflux transporters and down-regulation of 5 drug uptake-relevant SLC transporters was observed in tumors vs. non-neoplastic tissues. Moreover, expression levels of SLC28A1 and SLC22A1 were associated with OS of all patients (P=0.001). High transcript levels of SLC29A3 and SLC22A3 significantly predicted longer OS in chemotherapy-treated patients (P=0.004 and P=0.038, respectively). No association of ABC and SLC expression with KRAS mutation status was observed. Conclusions: Up-regulation of numerous drug resistance-associated ABCs and down-regulation of SLCs implicated in drug uptake may explain the generally very poor response of PDAC to cytotoxic agents. This study also identified some ABCs and SLCs as potential prognostic and predictive factors in PDAC. Supported by the Czech Science Foundation grant P301/12/1734 and CZ.1.05/2.1.00/03.0076 European Regional Development Fund.
Genomic profiling of breast cancer in African-American women.
Authors: Nunes Raquel , Wray Lynette , Mete Mihriye , Godwin Thomas , Herbolsheimer Pia Maarit , Smith Karen L. , Bijelic Lana , Boisvert Marc E. , Swain Sandra M.
Published: March 2020

11054

Background: Molecular profiling of breast cancer (BC) identifies intrinsic subtypes with distinct gene expression and clinical characteristics. In the US, BC is less frequent in African-American females (AAF); however mortality is higher, particularly among younger women. Unfavorable subtypes of BC seem to be more frequent in premenopausal AAF. Methods: Tumor gene expression in AAF presenting with early stage or locally advanced BC was performed using the Symphony platform on fresh and paraffin-embedded tissue (Agendia inc), a microarray-based method which classifies tumors according to prognosis (MammaPrint, MP), molecular subtype (BluePrint, BP) and estrogen receptor (ER), progesterone receptor (PR) and Human Epidermal Growth Factor Receptor 2 (HER2) mRNA levels (TargetPrint, TP). Genomic information is correlated with clinical and pathologic characteristics and Oncotype DX recurrence score (RS) when available. We plan to enroll 100 patients. Results: Results available in 46 patients. Median age 62 years (range 24-100), 20 stage I, 15 stage II, 11 stage III disease. There was no significant association between MP risk and stage, but MP risk was significantly associated with grade 3 disease (p=.006). 9 cancers were triple negative by IHC; using BP, 8 of these were Basal-type and 1 HER2-type. Basal-type was the most common subtype in patients ≤ 40 years old (p < .001). In the 6 cases ER positive by IHC but negative by TP, 3 were Basal-type and 3 were HER2-type.14 patients had Oncotype RS results available: 2 were High Risk by Oncotype and MP; 3 had intermediate RS, 2 of which were High Risk by MP; 9 had a low RS, 4 of which were High Risk by MP. Conclusions: African-American women with stage I to III BC often present with High-Risk disease irrespective of stage. BP classified all young patients (≤40) as Basal-type. Molecular subtyping confirmed the biologic heterogeneity in triple negative and hormone positive tumors. Oncotype RS and MP offered different prognostic information. Follow up will be needed to determine correlation with outcome. Funding: MP, BP and TP test provided by Agendia. Biostatistical support by GHUTTCS-CTSA. [Table: see text]
Induction of an aggressive phenotype and poor survival in early-stage lung adenocarcinoma and epigenetic inactivation of microRNA-34b/c.
Authors: Nadal Ernest , Gallegos Marc , Chen Guoan , Palmero Ramon , Capella Gabriel , Cardenal Felipe , Beer David G.
Published: March 2020

11056

Background: The microRNA-34b/c (miR-34b/c) is a transcriptional target of TP53 that is frequently mutated in primary lung adenocarcinoma (AC). We investigated the clinical implications of miR-34b/c methylation in early stage lung AC patients and the functional role of miR-34b/c re-expression in lung AC cell lines. Methods: DNA methylation of miR-34b/c promoter was assessed by real-time PCR temperature dissociation in 15 lung AC cell lines and 140 early stage lung AC resected at the Bellvitge Hospital (Barcelona) and at the University of Michigan (Ann Arbor). Patient characteristics: 65% males, 88% smokers, 68.5% stage I and 31.5% stage II. Expression of miR-34b/c was determined by TaqMan RT-PCR. Two lung AC cell lines (NCI-H1838 and SK-LU-1) were transfected with an expression vector containing both miRs and the effects upon cell proliferation, migration and invasion were determined. Results: MiR-34b/c was methylated in 40% of cell lines and in 46% of primary tumors with significant association with higher tumor stage (P=0.033), recurrence (P=0.017) and death (P=0.005). In the training set (n=58), patients with higher levels of miR-34b/c methylation had significantly shorter median DFS (28.5 months) compared to low to medium levels (not reached, log-rank P=0.016). In the test set (n=82), higher levels of miR-34b/c methylation were also associated to shorter median DFS (19 months) compared to patients with low to medium levels (not reached, log-rank P=0.005). MiR-34b/c methylation remained an independent prognostic marker for DFS after adjusting by age, gender and stage. Tumors harboring TP53 mutations and miR-34b/c methylation expressed significantly lower levels of miR-34b/c (P≤0.001). Stable cells expressing miR-34b/c had lower proliferation rate relative to cells transfected with empty vector (P≤0.001). Expressing miR-34b/c in SK-LU-1 cells reduced migration and invasion ability. Conclusions: Epigenetic inactivation of miR-34b/c by DNA methylation is an independent prognostic marker in early stage lung AC. Ectopic expression of miR-34b/c generated a less aggressive phenotype in lung AC cell lines suggesting a potential for therapeutic targeting.
Integrated genomic analysis of EGFR-mutant non-small cell lung cancer immediately following erlotinib initiation in patients.
Authors: Bivona Trever Grant , Giannikopoulos Petros , Costa Carlota , Karachaliou Niki , Viteri Santiago , Garcia-Campelo M. Rosario , St. John John , Uzilov Andrew V. , Wellde Anne S. , Polkinghorn William Reilly , Majem Margarita , Felip Enriqueta , Carcereny Enric , Heidecke Cordula Nicole , Massuti Bartomeu , Wellde George W. , Weissman Jonathan S. , Rosell Rafael ,   Cancer Therapeutics Innovation Group
Published: March 2020

11067

Background: Major obstacles limiting the clinical success of EGFR TKI therapy in non-small cell lung cancer (NSCLC) patients are heterogeneity and variability in the initial anti-tumor response to treatment. The underlying molecular basis for this heterogeneity has not been explored in patients immediately after initiation of therapy. Methods: We conducted CT-guided core needle biopsies immediately prior to erlotinib treatment initiation and at 6 days post erlotinib initiation in a patient with histologically confirmed NSCLC harboring an established activating mutation in EGFR. DNA from the paired frozen biopsies and matched normal tissue was analyzed by whole exome sequencing and RNA from the biopsies was analyzed by whole transcriptome sequencing. High-resolution CT images were obtained at the time of each biopsy to compare the degree of molecular and radiographic response observed. Results: Two established activating somatic mutations were identified in EGFR (p.G719A and p. R776H). Selective depletion of each EGFR mutant allele, but not the EGFR wild type allele, was observed upon erlotinib treatment. Gene expression analysis of the paired transcriptomes revealed that erlotinib treatment resulted in significant upregulation of proapoptotic genes including BAD, BAX, BID, CASP3 and growth inhibitory genes including CDKN1A, GADD45B, GADD45G and downregulation of growth-promoting genes including CCNB1 and CCND3. Several unexpected and novel molecular biomarkers were identified by transcriptome analysis and the complete dataset will be presented. High-resolution CT scans revealed no interval radiographic changes in the target lesion and no clinical complications were encountered. Conclusions: This study is the first reported integrated genomic analysis of EGFR-mutant NSCLC immediately following EGFR TKI initiation. We documented the feasibility, safety and utility of this strategy to establish initial drug efficacy at the molecular level prior to any radiographic evidence of response. Additional, serial integrated genomic analysis is ongoing in the index patient and others on therapy to enhance the management of NSCLC patients on targeted therapy.
Noninvasive measurement of prostate-specific membrane antigen (PSMA) expression with radiolabeled J591 imaging: A prognostic tool for metastatic castration-resistant prostate cancer (CRPC).
Authors: Tagawa Scott T. , Akhtar Naveed Hassan , Christos Paul J. , Osborne Joseph , Vallabhajosula Shankar , Freedy Shoaib , Goldsmith Stanley J. , Nanus David M. , Bander Neil Harrison
Published: March 2020

11081

Background: PSMA is nearly universally expressed by PC, upregulated with increased grade and castration-resistance. Evidence points towards PSMA expression as a downstream cellular biomarker of androgen receptor (AR) activity and non-invasive measurement of PSMA expression has recently been demonstrated to be a novel biomarker of AR activity. Methods: Planar gamma camera images following radiolabeled J591 (111In-J591 or 177Lu-J591) were semi-quantitatively scored using 2 methods by 2 independent radiologists blinded to outcome.A 5-point visual score (VS) of 0 - 4+ was assigned. Tissue Targeting Index (TTI), a novel metric designed to semi-quantitatively score images was calculated using the ratio of lesion count density (corrected for background) to whole body count density, with maximum (TTImax) and mean (TTIave) scores recorded. Follow up tabulating subsequent therapies and overall survival (OS) was recorded and imaging scores were associated with OS using Cox regression analysis. Results: 130 men with metastatic CRPC underwent radiolabeled J591 imaging. 86.2% had bone metastases, 51.5% lymph node, 16.9% lung, 9.2% liver. 87.7% had accurate targeting of known sites of disease by planar imaging. CALGB (Halabi) nomogram scores were prognostic for the population. As continuous variables, TTImax (p=0.013) and TTIave (p=0.002) were associated with worse survival. VS demonstrated a trend for worse survival (p=0.09). In multivariate analysis, TTI maintained independent prognostic value when controlling for Halabi score: TTImax HR 1.05 [95% CI 1.01, 1.10; p=0.02], TTIaveHR 1.09 [1.03, 1.16; p=0.004]. Conclusions: Level of PSMA expression measured by planar gamma-camera imaging following radiolabeled J591 is associated with OS in men with metastatic CRPC. High PSMA expression may indicate more aggressive tumor biology with increased AR pathway dysfunction. Improvements in quantitative molecular imaging techniques such as PSMA PET/CT with 89Zr-J591 may prove to be a valuable prognostic and predictive biomarker, particularly in the setting of AR- and PSMA-targeted therapy. Clinical trial information: NCT00195039, NCT00538668.
Developing a molecular imaging agent for Met using onartuzumab (MetMAb).
Authors: Wright Cara Elizabeth , Jagoda Elaine , Cecchi Fabiola , Bhadrasetty Veerendra , Histed Stephanie , Williams Mark , Kramer-Marek Gabriela , Mena Esther , Rosenblum Lauren , Lang Lixin , Szajek Lawrence , Paik Chang , Choyke Peter L. , Marik Jan , Tinianow Jeffrey , Merchant Mark , Bottaro Donald P.
Published: March 2020

11083

Background: Developing an imaging agent to assess Met expression would aid in diagnosis and monitoring tumor response to Met-targeted therapies. Onartuzumab (MetMAb), a Met selective humanized one-armed monoclonal antibody, has been studied in Phase I-II clinical trials in which it was generally well tolerated and has shown the most benefit in patients with MET positive tumors. Methods: Studies to assess Met-binding were executed using the human gastric carcinoma cell line MKN-45 which exhibits a high level of Met expression. Murine PET studies and biodistribution assays were performed using MKN-45 xenografts. Results: Plasma shed Met concentration is directly related to total tumor burden (p < 0.001). The absence of a positive correlation between shed Met and %ID in blood indicates that binding of tracer to shed Met present in plasma is unlikely. There are positive correlations between tumor mass, Met abundance, and phosphoMet content and uptake of 89Zr-df-onartuzumab in MKN-45 mouse xenografts. Lastly, tumor mass, Met, pMet and 89Zr-df-onartuzumab uptake were all significantly decreased by drug treatment. Conclusions: MKN-45 tumor uptake of 89Zr-df-onartuzumab correlated significantly with tumor mass and Met abundance. Blood tracer uptake did not positively correlate with the presence of plasma shed Met. The amounts of Met, pMet, as well as 89Zr-df-onartuzumab image intensity correlated significantly with tumor size (all Spearman p values < 0.001).Tumor volumes and Met content were significantly decreased in TKI treated versus control mice, which correlated with imaging results.89Zr-df-onartuzumab has potential utility for imaging Met to identify patients for treatment with Met-targeted therapeutics and to identify the emergence of Met-driven acquired resistance to other molecularly targeted cancer therapies.
BRAF mutation testing with a novel, rapid, fully automated molecular diagnostics prototype platform.
Authors: Huang Helen J. , Devogelaere Benoit , Falchook Gerald Steven , Fu Siqing , Angelo Laura S. , Hong David S. , Piha-Paul Sarina Anne , Naing Aung , Holley Veronica R. , Tsimberidou Apostolia Maria , Stepanek Vanda M. T. , Kim Kevin , Subbiah Vivek , Wheler Jennifer J. , Zinner Ralph , Wolff Robert A. , Sablon Erwin , Maertens Geert , Kurzrock Razelle , Janku Filip
Published: March 2020

11086

Background: Mutations in the BRAF gene provide actionable targets for cancer therapy in melanoma and other tumor types. Novel, fast, and accurate diagnostic systems are needed for further implementation of personalized therapy. Methods: The molecular diagnostics (MDx) prototype platform (Biocartis, Mechelen, Belgium) is a fully integrated real-time PCR-based system with high sensitivity (1% mutant in wild-type [wt] background) and fast turnaround time (< 90 minutes), which requires no sample preparation and <2 min hands-on time. Archival formalin-fixed paraffin-embedded tumor samples (1 to 5 shavings of 10 µm) from patients (pts) with advanced cancers previously tested for V600 BRAF mutations in a CLIA-certified Molecular Diagnostic Laboratory (PCR-based sequencing or Sequenom MassARRAY) were tested for BRAF V600 mutations using the MDx prototype platform. Concordance between methods and treatment outcomes with BRAF/MEK inhibitors were analyzed. Results: Forty-seven pts (melanoma, n=26; colorectal, n=8; papillary thyroid, n=3; other cancers, n=10) with available tissue and CLIA laboratory BRAF results were selected (BRAF V600 mutant, n=37; BRAF V600 wt, n=10). Of the 40 pts for whom the same tissue block was used for MDx and CLIA, BRAF status was concordant in 38 (95%; kappa 0.87; 95% CI 0.69-1.05) of them. BRAF status by MDx was discordant with CLIA in 3 of 47 cases (mutant by CLIA, but not MDx); one discrepant case contained a different mutation subtype (resp. V600E vs. V600K/R), and in another case different tissue blocks were used for MDx vs. CLIA testing. Of 34 pts with BRAF mutations detected by MDx, 28 were treated on protocols (on the basis of the CLIA results) with BRAF/MEK inhibitors and 8 (29%) had a partial (n=7) or complete response (n=1). Of interest, 1 pt with prostate cancer (V600E by CLIA, wt by MDx) received a BRAF/MEK inhibitor and did not respond. Detailed patient characteristics, mutation types and discrepancy analysis will be presented. Conclusions: The BRAF V600 mutation MDx prototype assay is a fast (turn-around time about 1.5 hours) and simple (<2 minutes hands-on time) test to determine BRAF mutation status with 95% concordance with CLIA laboratory if identical tissue blocks are used.
Molecular subtyping to predict better clinical and pathologic tumor response in operable early-stage breast cancer treated with docetaxel-capecitabine with or without trastuzumab.
Authors: Gluck Stefan , Royce Melanie , Stork-Sloots Lisette , De Snoo Femke
Published: March 2020

11090

Background: Classification into molecular subtypes is important for the selection of therapy for patients with early breast cancer. Here we determine rates of pathological complete response (pCR) in early stage breast cancer to neoadjuvant capecitabine plus docetaxel, +/- trastuzumab, and investigate MammaPrint together with the molecular subtyping profile BluePrint as markers of pathological response in comparison to other biomarkers. Methods: This analysis was carried out on data from 122 patients enrolled in a multicenter study (XeNA) of neoadjuvant therapy for four 21-day cycles with capecitabine 825 mg/m2 plus docetaxel 75 mg/m2if HER2-, and a standard trastuzumab dose if HER2+ (Glück , BCRT 2011). Clinical and pathological features, TP53 mutation analysis and PAM50 results were collected through GEO at NCBI (GSE22358). MammaPrint and BluePrint outcomes were determined from the available gene expression data and resulted in 4 distinct molecular groups: Luminal A (MammaPrint Low Risk/Luminal-type), Luminal B (MammaPrint High Risk/ Luminal-type), Basal-type and HER2-type. Results: In patients who completed 4 cycles of chemotherapy and surgery the overall pCR rate was 16%. Stratified by BluePrint pCR was observed in 1/15 (7%) of the Luminal A and 2/44 (5%) of Luminal B, in 10/22 (45%) of the HER2-type and in 7/41 (17%) of the Basal-type. The response rate among TP53 mutated patients was 6/61 (26%), which was significantly higher than TP53 wild-type patients (3/54 4%; p=0.012). Concordance of BluePrint/MammaPrint with PAM50 molecular subtyping was 61%. Conclusions: Molecular Subtyping with BluePrint and MammaPrint can identify better outcomes of patients in the neo-adjuvant setting. Patients with Luminal A breast cancer have a good baseline prognosis with excellent survival and may not benefit from chemotherapy (Glück, SABCS 2013). MammaPrint and BluePrint provide predictive information for patients treated with treated with docetaxel-capecitabine +/- trastuzumab.
Utilizing a collaborative working model to optimize molecular analysis of solid tumors in the Cancer Research UK's Stratified Medicine Programme.
Authors: Walker Ian , MacDonald Fiona , Stuart Helen , Butler Rachel , Ellwood-Thompson Rhianedd , Brown Celia , Eden James , Gonzalez de Castro David , Thompson Lisa , Lillis Sue , Mair Debbie , Cloke Vicky , Rehal Pauline , Bell Jenny , Smith Matthew , O'Sullivan Brendan , Taniere Philippe , Griffiths Michael , Johnson Peter W. M. ,   Stratified Medicine Programme Team
Published: March 2020

11094

Background: The Stratified Medicine Programme is demonstrating large scale molecular testing of solid tumours in the UK using a range of technologies. The collaborative model has allowed three laboratories to share and compare data on mutation frequencies including mutation exclusivity, test turnaround times and failure rates, to inform a future routine service for clinical care. Methods: Phase One is a two-year pilot study of molecular analysis of surplus diagnostic FFPE tumour tissue obtained from patients with cancer of the breast, colorectum, lung, ovary, prostate or malignant melanoma. Samples are tested for specified genes of clinical and research interest (for example KRAS, BRAF, NRAS, PIK3CA, TP53, PTEN, TMPRSS2-ERG, EGFR, EML4-ALK and KIT). The labs have developed and validated protocols with comparable sensitivity for the simultaneous molecular analysis of multiple genes. Results: By 31 December 2012, 4,734 sets of molecular results were completed with 60% of tumour-site specific reports issued within the target 15 days (from sample receipt). Failure rates vary with both sample quality and the type of analysis performed. The Table illustrates the link between turnaround times and failure rates, showing that repeat testing for specimens which initially fail may reduce overall failure rates but consequently increase average turnaround times. Conclusions: We will report comparative data across the three testing labs and identify multiple factors that affect mutation detection rates, failure rates, turnaround times and reporting procedures. The Stratified Medicine Programme acknowledges funding from Cancer Research UK, AstraZeneca, and Pfizer. [Table: see text]
Liquid biopsy-based assays to monitor residual disease in cancer.
Authors: Mei Gangwu , Sebisanovic Dragan , Mir Alain , Gulzar Zulfiqar , Brooks James D. , Jeffrey Stefanie S. , Talasaz AmirAli
Published: March 2020

11096

Background: One of the main challenges in cancer management is monitoring the non-organ-confined disease post metastasis. Recently analysis of circulating tumor nucleic acids has generated a new paradigm for monitoring the progression and molecular pathology of the residual disease through a non-invasive test. Methods: We have developed a new sequencing workflow, which increases the sensitivity and specificity of detecting and quantifying rare tumor-derived nucleic acids among healthy fragments by at least ten-fold. Unlike conventional sequencing library preparation protocols, the majority of extracted circulating DNA fragments are hybridized to sequencing flow cells with minimal modifications. The sequencing data are processed using our proprietary bioinformatics pipelines to search for rare genetic abnormalities within the heterogeneous circulating fragments. Results: To study the performance of our technology, we first evaluated its sensitivity in analytical samples. We spiked varying amounts of LNCaP cancer cell line gDNA into a background of normal gDNA and were able to successfully detect somatic mutations down to 0.1% sensitivity. Subsequently, we investigated the correlation of circulating DNA and tumor gDNA in human xenograft models in mice. In seven different mice models of two different human tumors, we found very strong correlation between somatic mutations detected in all pairs of tumor gDNA and mouse blood cfDNA. After preclinical studies, we initiated a pilot study on human samples across different cancer types. We found more than 90% correlation between tumor mutations in stage IIIb CRC cancer patients and the mutations detected in their circulation (n=7). In a pilot study in prostate cancer patients (n=3), we detected chromosomal abnormalities in multi locations in circulating DNA genome in both advanced and post-relapse stages of the disease. Conclusions: The present work indicates the potential of using circulating nucleic acids to facilitate the integration of real-time molecular pathology into routine cancer care. Our assay will be used to identify residual disease after neoadjuvant chemotherapy and/or surgery, but may also identify a subset of patients who may benefit from alternative therapies.