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Objective Targeted next generation sequencing (NGS) was evaluated for its ability to identify unanticipated targetable genomic alterations (GA) for patients with relapsed ovarian epithelial carcinoma (OC). Methods DNA sequencing was performed for 3320 exons of 182 cancer-related genes and 37 introns of 14 genes frequently rearranged in cancer on indexed, adaptor ligated, hybridization-captured libraries using DNA isolated from FFPE sections from 48 histologically verified relapsed OC specimens. The original primary tumor was sequenced in 26 (54%) of the cases and recurrent/metastatic tumor site biopsies were sequenced in 22 (46%) of the cases. Actionability was defined as: GA that predict sensitivity or resistance to approved or standard therapies or are inclusion or exclusion criteria for specific experimental therapies in NCI registered clinical trials. Results There were 38 (80%) serous, 5 (10%) endometrioid, 3 (6%) clear cell, 1 mucinous (2%) and 1 (2%) undifferentiated carcinomas. 141 GA were identified with an average of 2.9 GA (range 0–8) per tumor, of which 67 were actionable for an average of 1.4 actionable GA per patient (range 0–5). 33/48 (69%) of OC patient samples harbored at least one actionable GA. Most common GA were TP53 (79%); MYC (25%); BRCA1 / 2 (23%); KRAS (16.6%) and NF1 (14.5%). One tumor featured an ERBB2 point mutation. One of 3 (33%) of clear cell tumors featured cMET amplification validated by both FISH and IHC. Conclusions NGS assessment of therapy resistant OC identifies an unexpectedly high frequency of GA that could influence targeted therapy selection for the disease.

The purpose of this study is to identify prognostic markers and treatment targets using a clinically certified sequencing panel in multiple myeloma. We performed targeted sequencing of 578 individuals with plasma cell neoplasms using the FoundationOne Heme panel and identified clinically relevant abnormalities and novel prognostic markers. Mutational burden was associated with maf and proliferation gene expression groups, and a high-mutational burden was associated with a poor prognosis. We identified homozygous deletions that were present in multiple myeloma within key genes, including CDKN2C, RB1, TRAF3, BIRC3 and TP53, and that bi-allelic inactivation was significantly enriched at relapse. Alterations in CDKN2C, TP53, RB1 and the t(4;14) were associated with poor prognosis. Alterations in RB1 were predominantly homozygous deletions and were associated with relapse and a poor prognosis which was independent of other genetic markers, including t(4;14), after multivariate analysis. Bi-allelic inactivation of key tumor suppressor genes in myeloma was enriched at relapse, especially in RB1, CDKN2C and TP53 where they have prognostic significance.

Background: As more therapies targeting genomic alterations become available, next-generation sequencing (NGS) is increasingly performed in tumor types where mutational status may drive treatment choice. In addition to its ability to identify base substitutions, insertions and deletions across entire exons, NGS can detect relevant copy number changes such as amplification of HER2 in breast tumors. However, for NGS to be clinically applicable, it must reliably analyze FFPE tumor samples and show concordance with the best current diagnostic methods. Methods: To confirm a clinical role for NGS in detecting copy number alterations, we identified 35 FFPE invasive breast carcinomas previously tested for HER2 status by FISH, including 15 HER2 positives (≥7 copies) and 20 HER2 negatives (900X uniquely-mapping reads was obtained. Sequence data were analyzed for HER2 copy number (blinded to FISH results) based on a statistical model using allele frequencies and coverage depth of HER2 exons versus a process-matched normal control, classifying cases as HER2 positive (≥6 average copies), HER2 negative ( Results: High concordance was noted between HER2 copy number status determined by FISH and NGS: 30 of the 35 samples were classified as positive or negative by NGS, 1 was classified as intermediate and 4 as unknown due to low purity. Using FISH as a gold standard, NGS HER2 calls demonstrated an accuracy of 97% (29/30, 95% CI 83–99%), 93% sensitivity (13/14, 95% CI 69–99%) and 100% specificity (16/16, 95% CI 81–100%). One discordant case was noted (FISH positive, NGS negative). Furthermore, NGS revealed 70 additional alterations (38 base substitutions, 10 insertions/deletions, 22 copy number alterations) in 23 cancer genes (an average of 2.0 alterations per sample). Genomic alterations that predict sensitivity or resistance to approved or experimental targeted therapies and thus plausibly guide treatment decisions were found in 69% of patients. These include PIK3CA (16 cases, PI3 kinase/mTOR inhibitors), PTEN (3 cases, PI3K/AKT/mTOR inhibitors), KRAS (1 case, resistance to cetuximab and panitumumab), and NF1 (1 case, mTOR/MAPK inhibitors) plus amplifications of CCND1 (4 cases, CDK4 inhibitors), FGFR1 (3 cases, FGF inhibitors) and MCL1 (3 cases, BCL-2 inhibitors, resistance to anti-tubulin therapies). Four cases included co-amplification of RARA with HER2. Conclusions: We conclude that HER2 status can be reliably determined by NGS on FFPE breast cancers and that NGS uncovers additional actionable genomic alterations that could impact disease management in a high proportion of patients. Further evaluation of NGS as a guide to therapy in breast cancer is warranted. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr PD02-07.

Introduction L’association d’inhibiteurs de BRAF et de MEK a recemment obtenu l’AMM dans le traitement du melanome mute V600, avec des taux de reponse avoisinant les 70 %. Des reponses aux therapies ciblees ont aussi ete rapportees pour des mutations autres que BRAF V600. Ainsi, une detection sensible, fiable et non exhaustive des alterations de BRAF est indispensable pour pouvoir offrir aux patients le plus de chances d’avoir acces a une therapie ciblee correspondant au profil genetique de leur melanome. Materiel et methodes Etude multicentrique retrospective de 385 cas consecutifs de melanomes pour lesquels une mutation ou un rearrangement de BRAF a ete identifie par technique NGS (sequencage de nouvelle generation) a partir de capture par hybridation. Resultats Les resultats d’analyse de BRAF, faite avant NGS a partir de capture par hybridation, etaient disponibles pour 79 cas (21 %), par PCR (n = 30), Sanger (n = 13), immuno-histochimie (n = 10), NGS sans capture par hybridation (n = 9), ou autre methode non precisee (n = 17). Parmi les cas mutes BRAF V600, 11/57 (19 %) avec donnees disponibles etaient negatifs par la technique anterieure, dont 2/11 (18 %) avec confirmation qu’il s’agissait du meme prelevement analyse. Parmi les cas avec mutations de BRAF V600 finalement trouvee, il n’y avait pas de difference significative en termes de frequence allelique (mediane 35 % vs. 40 %, p = 0,25) ou de pourcentage de cellules tumorales (mediane 50 % dans les deux groupes, p = 0,97) entre les prelevements ayant un test auparavant positif vs auparavant negatif pour BRAF. Des resultats auparavant negatifs pour BRAF etaient aussi trouves dans 16/20 (80 %) des cas avec mutation non-V600, parmi lesquels deux presentaient de multiples alterations de BRAF [K601E (4), D594A/G/N (4), S467L (2), L584F (2), G464 V, G466 V, G469 V, E586 K, N581I, L597Q, A589_T599insT]. 2/2 (100 %) des cas avec fusion activatrice de BRAF identifiee par capture par hybridation avaient un test auparavant negatif. Les reponses cliniques de certains patients seront presentees. Discussion Bien que la recherche de mutation du gene BRAF soit desormais pratique courante dans les centres suivant des patients atteints de melanome metastatique, il existe une grande diversite de methodes approuvees. Dans cette etude, le NGS a partir de capture par hybridation a permis d’identifier des mutations et fusions activatrices variees, dont la mutation BRAF V600E, dans une proportion de cas pour lesquels la methode de reference locale avait rendu un resultat negatif pour BRAF. En raison du benefice clinique demontre des therapies ciblees en cas de melanome avec alteration de BRAF, le NGS a partir de capture par hybridation devrait pouvoir etre propose a tout patient atteint de melanome metastatique, en cas de test initial negatif.

An inspiring if flawed documentary, Made in L.A. follows three Los Angeles sweatshop workers as their lawsuit against a low-price retail label and the community campaign to support them developed from 2001 through 2004. Enthusiastically received by critics and activists in 2007 when it was broadcast on PBS’s P.O.V. series, the film received even more praise in 2008 when it won an Emmy for “Outstanding Continuing Coverage of a News Story- Long Form.”

As the nation debates once again proposals to guarantee health insurance for everyone, a new biography of Frances Perkins reminds us of just how long the argument has been going on. In 1933, when Franklin Roosevelt offered Perkins the job of secretary of labor, she proposed an agenda that was to become the backbone of the New Deal: unemployment insurance, hours and wages legislation, abolition of child labor, social security, and national health insurance. Famously, all but the latter were to be won and even more. Now, even as the American labor movement leads the uphill battle for some form of public health insurance, it is fighting for its own life. The Magna Carta of labor—the Wagner Act of 1935 that Perkins did not particularly support—has run its course, and the unions want a successor law to clear the field for them.

Background Squamous cell cancers of the anal canal (ASCC) are increasing in frequency and lack effective therapies for advanced disease. Although an association with human papillomavirus (HPV) has been established, little is known about the molecular characterization of ASCC. A comprehensive genomic analysis of ASCC was undertaken to identify novel genomic alterations (GAs) that will inform therapeutic choices for patients with advanced disease. Patients and methods Hybrid-capture-based next-generation sequencing of exons from 236 cancer-related genes and intronic regions from 19 genes commonly rearranged in cancer was performed on 70 patients with ASCC. HPV status was assessed by aligning tumor sequencing reads to HPV viral genomes. GAs were identified using an established algorithm and correlated with HPV status. Results Sixty-one samples (87%) were HPV-positive. A mean of 3.5 GAs per sample was identified. Recurrent alterations in phosphoinositol-3-kinase pathway (PI3K/AKT/mTOR) genes including amplifications and homozygous deletions were present in 63% of cases. Clinically relevant GAs in genes involved in DNA repair, chromatin remodeling, or receptor tyrosine kinase signaling were observed in 30% of cases. Loss-of-function mutations in TP53 and CDKN2A were significantly enhanced in HPV-negative cases (P Conclusions This is the first comprehensive genomic analysis of ASCC, and the results suggest new therapeutic approaches. Differing genomic profiles between HPV-associated and HPV-negative ASCC warrants further investigation and may require novel therapeutic and preventive strategies.

BACKGROUND: Primary CNS tumors, which include both low-grade and high-grade gliomas, are the most common pediatric solid malignancies. We queried whether CGP could uncover a significant frequency of CRGA that could inform treatment decisions and improve clinical outcome for these aggressive tumors. METHODS: DNA was extracted from 75 pediatric glioma formalin-fixed paraffin-embedded clinical specimens. Hybridization captured libraries of 236 (FoundationOne, n = 43) or 405 (FoundationOne Heme, n = 32) genes, plus select introns frequently rearranged in cancer were sequenced to high (>500x), uniform coverage. All classes of genomic alterations including base substitutions, small insertions and deletions, rearrangements, and copy number alterations, were evaluated and reported. CRGA were defined as GA associated with FDA approved therapies or targeted therapies in mechanism-driven clinical trials. RESULTS: The median age of the patients was 9 years (range 0 to 18 years). There were 41 male (57%) and 34 female (43%) patients. The study included glioblastomas (n = 30, 42%), astrocytomas not otherwise specified (NOS) (n = 17, 24%), gliomas NOS (n = 10, 14%), pilocytic astrocytomas and anaplastic astrocytomas (9 each, 13%). 89% of cases harbored at least one CRGA. Pediatric gliomas frequently harbored short variants or copy number alterations in TP53 (39%); BRAF (15%) for which 82% were the V600E base substitution; CDKN2A/B (15%), NF1 (14%), PIK3CA (14%), ATRX (13%) and EGFR (11%). Ten cases (14%) harbored a BRAF fusion, including 44% of pilocytic astrocytomas. BRAF fusions included the known KIAA1549-BRAF and the novel CCDC6-BRAF and BCAS1-BRAF. ATG7-RAF1, ALK-PPCB1 and QK1-RAF1 rearrangements were also identified. Outcomes from cases where targeted therapy was utilized will be presented. CONCLUSIONS: CRGA are frequently identified in pediatric glial tumors of both low-grade and high-grade histology and can be identified by CGP in the course of clinical care. CGP has the potential to identify opportunities for targeted therapies or enrollment in clinical trials for these patients.

Objective: To determine whether loss of melastatin (MLSN) is a universal phenomenon in American Joint Committee on Cancer (AJCC) stage I and II melanoma patients who experienced recurrence. Material and methods: Paraffin blocks of primary melanomas (PMs) were retrieved from 30 patients who had a negative sentinel lymph node biopsy and developed recurrent melanoma (AJCC stage I and II). Chromogenic in situ hybridization (CISH) methods were utilized to evaluate the expression of MLSN mRNA. These results were correlated with clinicopathologic data. Results: Variable, heterogeneous expression of MLSN mRNA was identified in normal, in situ and invasive melanocytes within and between cases. For the invasive PM component, 24 (80%) had focal, regional or complete loss of MLSN mRNA. The remaining 20% had either regional or total partial downregulation of MLSN mRNA. Intact MLSN mRNA expression was present regionally in 14/30 (47%), with mean relative tumor area of 38%, range 5–85%. Increasing loss of MLSN mRNA significantly correlated with increasing tumor depth and microsatellites (r = 0.1/0.4, p = 0.04). However, thin, AJCC T stage 1a PM had higher relative mean loss than intermediate AJCC T stage 2a/2b/3a thickness PM (65% vs. 34%/48%/25%). Increasing loss of MLSN mRNA significantly impacted on disease free survival (DFS) by multivariate analysis (58 vs. 0% 2 years DFS, ≤ 75 vs. >75% mRNA loss, p = 0.02). Decreased overall survival significantly correlated with increasing age and vascular invasion on multivariate analysis. Conclusion: Extensive loss of MLSN in PM correlated with aggressive metastatic melanoma. Ancillary testing for MLSN mRNA expression by CISH could offer a means to more accurately identify AJCC stage I and II patients at risk for metastatic disease, who could benefit from adjuvant therapy.

Eighty-nine patients with chronic actinic dermatitis (CAD) were retrospectively studied: 69 (78%) male and 20 (22%) female, with mean ages of 66 and 64 years, respectively; nine (10%) were dark skinned. Eight (9%) were abnormally sensitive to UVB wavelengths alone, 74 (83%) to UVB and UVA, and seven (8%) to UVB, UVA and visible radiation. Eighty-six patients were patch tested to an extended standard European series of contact allergens, including in 80 cases a 0.1% mix of three sesquiterpene lactones, and photopatch tested to a standard photopatch series. Sixty-four of these 86 patients (74%) had positive patch or photopatch tests; 36% (29 of 80) were sensitive to the sesquiterpene lactone mix, 21% (18 of 86) to fragrance compounds, 20% (17 of 86) to colophony, and 14% (12 of 86) to rubber chemicals. Ten (12%) had positive photopatch tests; five (6%) to musk ambrette, six (7%) to sunscreens and one to both. Fourteen of the eighty-nine patients with CAD (16%) had preceding endogenous eczema. In 18 of 86 patients (21%), CAD occurred alone, with neither detectable contact nor photocontact allergy, nor a preceding history of endogenous eczema. This study confirms the association between Compositae (sesquiterpene lactone) dermatitis and CAD.

This thanksgiving 2006 will witness the anniversary almost to the day ninety-seven years ago, when the immigrant shopgirls of New York's shirtwaist factories called a general strike. This past April, and then on May Day, immigrant workers, 120 years after the first strike movement for the eight-hour day (May Day, 1886), asserted their claim to just treatment. The fall and rise of sweatshop labor in New York and elsewhere in the global garment industry is a story with lessons for our time, for the struggles over the sweatshop issue today echo the issues and contentions of a century ago. Then, as now, there was tension between workers and their middle-class allies.

BACKGROUND: FGFR family members are infrequently mutated but are frequently overexpressed in breast cancer and often accompanied by increased, or altered, expression of FGF ligands. In this retrospective study, we reviewed a large series of FGFR altered breast cancer cases that received comprehensive genomic profiling (CGP) in the course of clinical care. MATERIAL AND METHODS: CGP was performed on hybridization-captures, adaptor ligation-based libraries using DNA extracted from 40 μm formalin-fixed paraffin-embedded (FFPE) section cut at 10 μm performed in a CLIA-certified lab (Foundation Medicine, Inc.). The pathologic diagnosis of each case was confirmed on routine hematoxylin and eosin-stained slides, and all samples forwarded for DNA extraction contained a minimum of 20% of DNA derived from tumor cells. The FoundationOne test sequences the full coding regions of up to 315 cancer-related genes, and up to 28 genes that are frequently altered in cancer to detect all classes of genomic alterations including base substitutions, indels, copy-number alterations (CNA), and fusions/rearrangements. The average depth of coverage is greater than 600X. The genomic profiles of 2,617 patients with diverse advanced malignancies who were evaluated at Cancer Treatment Centers of America between 12/24/12 and 03/11/15 were reviewed. 176 FGFR alterations (7.8%) were detected, of which 76 (43.5%) were found in breast cancer cases out of 434 (16.5%). The study was carried out in accordance with WIRB Institutional Review Board. RESULTS: A total of 76 female breast cancer patients, having a median age 50 (range, 28-69), with FGFR alterations were reviewed. All patients had metastatic/relapsed advanced breast cancer. 54 patients were Estrogen Receptor-positive (70%), and 15 were HER2+ (20%). 6 patients had gBRCA deleterious mutations. 84% of the samples (n=67) tissue block were analyzed, and the anatomic sites represented by the samples were 24 breast primary tumor (31%), 15 liver (19%), 10 lymph nodes (13%), and other sites (37%). The median number of chemotherapies cycles was 4 (range, 1-12), and the median time to metastasis was 31 months (range, 0-175). At the time of this report, 31 patients (40%) were deceased. 79 FGFR gene alterations were identified in 76 patients, including FGFR1 (65), FGFR2 (6), FGFR3 (2), and FGFR4 (4), with all but 7 of these being amplifications. The most co-existent altered gene was TP53 (66%), and other altered genes included PIK3CA (37%), MYC (28%), FGF3/4/19 (17%), CCND1 (17%), and CCNE1 (16%). The subset of co-amplified FGF3/4/19 and FGFR amplified patients were all (7) ER+ except for 1 patient. CONCLUSIONS: FGFR genomic alterations in breast cancer patients are predominantly amplifications and are most commonly observed in ER+ patients. Further review of treatment history will be performed to evaluate the hypothesis that alterations of FGFR is a modifier of response to endocrine therapy, and co-amplified FGF3/4/19 and FGFR breast cancer cases may be a distinct clinic-pathologic entity. Any patients in this series initiated on anti-FGFR targeted therapy will also be reported. Citation Format: Alvarez RH, Thomas JW, Kramer K, Niu J, Ahn E, McKnight JE, Dhillon N, Pabbathi H, Johnson AT, Wang K, Ross JS, Miller VA, Stephens PJ, Daneker GW, Ali S, Markman M. Clinicopathologic characterization and comprehensive genomic profiling (CGP) of advanced breast cancer patients with fibroblast growth factor receptor (FGFR) alterations. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P6-07-06.

Multiple myeloma is commonly associated with genomic alterations that result in hyperactivation of the mitogen-activated protein kinase pathway. BRAF (v-raf murine sarcoma viral oncogene homolog B) V600E activating mutations have been observed in 4% of multiple myeloma cases. Patients with BRAF V600Eemutated myeloma may have an unusually aggressive clinical course associated with prominent extramedullary disease and a short duration of response to standard therapies. Vemurafenib, a BRAF V600Eespecific inhibitor and an FDA approved agent for treatment of melanoma, also has clinical activity in BRAF V600E mutationepositive multiple myeloma.

To determine if carcinogenic events in vulvar skin precede the onset of morphologic atypia, the authors investigated for derangements in DNA content, cell proliferation, and cell death in vulvar carcinomas and surrounding skin in 140 samples of tumor and surrounding skin collected from 35 consecutive vulvectomy specimen for squamous cell carcinoma (SCC) or vulvar intraepithelial neoplasia (VIN) 3. Vulvar non-cancer excisions were used as controls. Investigations consisted of histologic classification and measurement of 9 variables--epidermal thickness (acanthosis and rete ridge length), immunolabeling index (LI) for 3 proteins (p53 protein, Ki-67, and mdm-2), pattern of p53 expression (dispersed vs. compact), DNA content index, and presence of aneuploidy by image analysis and apoptotic rate by Apotag labeling. Significant positive correlations were found for all nine variables studied versus increasing histologic severity in two proposed histologic stepwise models of vulvar carcinogenesis (lichen sclerosus (LS) and VIN 3 undifferentiated associated SCC groups). High p53 LI (>25) and the compact pattern of p53 expression (suspected oncoprotein) significantly correlated with LS and its associated vulvar samples compared with samples not associated with LS (P < or = 0.001). Furthermore, p53 LI, mdm-2 LI, and pattern of p53 expression were concordant between patient matched samples of LS and SCC. In addition, mdm-2 LI significantly correlated with dispersed pattern p53 LI suggesting a response to wild-type p53 protein accumulation. These findings support the hypothesis that neoplastic transformation occurs in sequential steps and compromises proteins involved in the cell cycle control. Concordance of p53 and mdm-2 protein expression in LS and adjacent SCC provides evidence that LS can act as a precursor lesion in the absence of morphologic atypia. Overexpression of mdm-2 with stabilization and inactivation of p53 protein may provide an alternate pathway for vulvar carcinogenesis.