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• Single Cell Sequencing of Sperm (scSperm)

  The aim of this project is to genotype and sequence single spermatozoa from two men, one in his twenties and the other in his seventies. The resulting data is used to quantify the mutations that have arisen in the gametes of both individuals in order to better understand the effect of aging on mutation rates and modes.Project Outline. In order to quantify mutations, semen from two individuals are sequenced. 48 single sperm cells are isolated from each individual, and their DNA is extracted. The resulting genomes are amplified using PicoPlex, GenomiPhi MDA, Repli-G MDA, and MALBAC. QC step is applied to check the quality of WGA DNA using standard Sequenom plex (26 SNPs). A subset of 32 amplification products which pass the intiall QC, are genotyped using Affymetrix SNP6 chips. 12 of the genotyped amplification products are also sequenced. In addition, one multi-cell sample per individual is sequenced as a reference and for validation purposes.Altogether, 12 single cell sperm genomes and two multi-cell genomes are sequenced, coming to a total of 14 genomes. Of the single cell sperm genomes, 2 are sequenced to 50x coverage, and the other 10 to 25x coverage. Both multi-cell genomes are sequenced to 25x coverage.

  Dataset EGAD00001001216

• Dataset for liposarcoma-RNA

  Rare cancer sequencing data of 40 runs in tumor/control pairs, which were uploaded to umbrella studies. The sequencing was always paired

  Dataset EGAD00001008854

• Cell-free DNA Methylome Analysis Enables Early Preeclampsia Prediction

  This dataset contains in solution target-enrichment bisulfite sequencing of placental tissue, buffy coat and plasma DNA from pregnant women. Blood samples were taken for cell-free DNA (cfDNA) DNA extraction from 64 women at the time of early-onset preeclampsia (PE) diagnosis, or from 38 controls (uncomplicated pregnancies) at a similar gestational age that did not develop preeclampsia subsequently. Among these subjects, plasma samples from 7 PE patients and 6 controls were also subjected to oxidative bisulfite sequencing. Placental tissues from 11 PE and 26 control subjects after delivery, and buffy coat from 16 PE and 16 control subjects at the same time of cfDNA sampling were profiled. A discovery cohort for early PE assessment in the first trimester was collected. In this cohort, cfDNA from 75 pregnancies that went on to develop early-onset PE and from 124 matched controls were collected and methylome sequencing were carried out. An independent validation cohort to validate early PE assessment with methylome profiling was collected as well. This validation cohort includes cfDNA samples from 61 PE and 136 control pregnancies.

  Dataset EGAD00001010159

• Target sequencing of 27 cancer-predisposing genes and 13 renal cell carcinoma-related genes in Japanese renal cell carcinoma patients

  Identifying causative genes via genetic testing is useful for the screening, prevention, and treatment of cancer. Several hereditary syndromes are known to occur in renal cell carcinoma (RCC). However, these evidences came from the European population; it remains unclear how the RCC-related genes and other cancer-predisposing genes contribute to the development RCC in the Japanese population. A case-control study of 14 RCC-related genes and 26 cancer-predisposing genes was performed using 1,563 Japanese patients with RCC and 6,016 controls. Patients were divided into two major histological subtypes of clear cell RCC (ccRCC) or non-clear cell RCC (nccRCC). Gene-based analysis of germline pathogenic variants in patients with each subtype and cancer-free subjects was performed. Following quality control, 1,532 patients with RCC and 5,996 controls were further analyzed. For ccRCC, 52 of 1,283 (4.05%) patients carried pathogenic variants mainly in the cancer-predisposing genes such as TP53 (P = 1.73 �� 10-4; OR, 5.8; 95% CI, 2.2���15.7). Approximately 80% of patients with pathogenic variants in TP53 had p.Ala189Val that was specific in East Asian population. For nccRCC, 14 of 249 (5.62%) patients carried pathogenic variants mainly in the RCC-related genes such as BAP1 (P = 6.27 �� 10-5; OR, Inf; 95% CI, 10.0���Inf), and FH (P = 6.27 �� 10-5; OR, Inf; 95% CI, 10.0���Inf). Our study showed different and population-specific contributions of risk genes between ccRCC and nccRCC in Japanese for better-personalized medicine.

  Study JGAS000414

• Exome sequencing of uterine leiomyosarcomas

  Uterine leiomyosarcomas (ULMSs) are aggressive smooth muscle tumors associated with poor clinical outcome. Despite previous cytogenetic and molecular studies, their molecular background has remained elusive. To examine somatic variation in ULMS, we performed exome sequencing on 19 tumors. Altogether 43 genes were mutated in at least two ULMSs. Most frequently mutated genes included tumor protein P53 (TP53; 6/19; 33%), alpha thalassemia/mental retardation syndrome X-linked (ATRX; 5/19; 26%), and mediator complex subunit 12 (MED12; 4/19; 21%). Unlike ATRX mutations, both TP53 and MED12 alterations have repeatedly been associated with ULMSs. All the observed ATRX alterations were either nonsense or frameshift mutations. ATRX protein levels were reliably analyzed by immunohistochemistry in altogether 44 ULMSs, and the majority of tumors (23/44; 52%) showed clearly reduced expression. Loss of ATRX expression has been associated with alternative lengthening of telomeres (ALT), and thus the telomere length was analyzed with telomere-specific fluorescence in situ hybridization. The ALT phenotype was confirmed in all ULMSs showing diminished ATRX expression. Exome data also revealed one nonsense mutation in death-domain associated protein (DAXX), another gene previously associated with ALT, and the tumor showed ALT positivity. Aberrant expression of both TP53 and ATRX were associated with poor overall survival. In conclusion, exome sequencing revealed that TP53, ATRX, and MED12 are frequently mutated in ULMSs. ALT phenotype was commonly seen in tumors, indicating that ATR inhibitors, which were recently suggested as possible new drugs for ATRX-deficient tumors, could provide a potential novel therapeutic option for ULMS.

  Study EGAS00001001612

• Circumventing intratumoral heterogeneity to identify potential therapeutic targets in hepatocellular carcinoma. Details please contact lifuqiang@genomics and zhaoxin@genomics.cn.

  Background and aims: Intratumoral heterogeneity (ITH) challenges identifying mutations with target therapy potential whereas circulating cell-free DNAs (cfDNAs) could reflect nearly the entire mutation spectrum in given tumors. We investigated how to minimize the limit of ITH for profiling hepatocellular carcinoma (HCC).Methods: Thirty-two multi-regional HCC samples from five patients were subjected to whole exome sequencing (WES) and targeted deep sequencing (TDS). ITH extent was measured by the average percentage of non-ubiquitous mutations (present in parts of tumor regions). Matched cfDNAs were also analyzed by WES and TDS. Profiling efficiencies of single tumor specimen and cfDNA were compared and the one better depicted mutational landscape was selected to screen therapeutic targets.Results: We found variable extents of ITH in HCCs and observed branched and parallel evolution patterns. ITH level decreased at higher sequencing depth of TDS than that measured by WES (28.1% vs 34.9%, P < 0.01) but it remained unchanged upon additional samples analyzed. TDS of single tumor specimen detected an average of 70% the total mutations in HCC. Although more mutations were detected in cfDNA under TDS than WES, an average of 47.2% total HCC mutations uncovered by cfDNA suggested tissue outperform cfDNA and the latter may serve as alternative in profiling HCC genome. Consequently, TDS of single tumor tissue in 66 patients and cfDNAs in four unresectable HCCs identified 38.6% (26/66 and 1/4) patients bearing therapeutic targets.Conclusions: TDS of single tumor specimen could largely circumvent ITH to uncover mutations indicative of target therapy in HCC.

  Study EGAS00001002207

• Genomic consequences of aberrant DNA repair mechanisms stratify ovarian cancer histotypes

  Aberrant DNA repair processes are a hallmark of human tumours. How these deficiencies variously impact the genomes of ovarian cancer between and within histotypes remains unknown. We studied the whole genome pointmutation and structural variation patterns of 133 tumours (59 high grade serous (HGSC), 35 clear cell (CCOC), 29 endometrioid (ENOC), and 10 adult granulosa cell tumours (GCT)) as a substrate for class discovery in ovarian cancer. Ab-initio clustering of integrated point mutation and structural variation signatures revealed seven novel subgroups, comprising between and within histotype stratification. Prevalence of fold-back inversions (FBI) co-localised with high level amplification events divided HGSC into two prognostically significant subgroups. This finding was recapitulated in two additional independent cohorts (total n=576 cases), and transcended gene based mutation status and gene expression as prognostically relevant features of HGSC. CCOC were divided into one group harbouring a signature reflective of active genome editing via APOBEC enzymes (26%) and a complementary group with an age-relatedmutational signature. ENOC were divided intomicrosatellite instable (MSI) cases (28%) with a distinctmismatch repairmutation signature and an outlying mutation rate, with the remainder of cases distributed amongst the remaining six groups. We suggest the FBI, MSI, and APOBEC groups represent biologic strata that could direct treatment strategies. Taken together, our work establishes the efficacy of the somatic genome as a biomarker to stratify ovarian cancers, simultaneously identifying patients that may benefit from emerging therapeutics and distinct subgroups within classic ovarian cancer histotypes.

  Study EGAS00001002390

• Tremelimumab +/- durvalumab in combination with paclitaxel as immune induction in metastatic urothelial cancer: clinical and translational results of the ICRA trial

  DESCRIPTION: New therapeutic strategies for therapy-refractory metastatic urothelial carcinoma (mUC) patients are still needed. In the phase I-II ICRA trial, we evaluated whether paclitaxel plus tremelimumab (T; anti-CTLA-4), with or without durvalumab (D; anti-PD-L1), could induce a tumor response in mUC patients who had already received platinum chemotherapy and immune checkpoint inhibition (ICI). After a dose-escalation safety phase, patients were randomized between three arms: A (n=20 after expansion) paclitaxel plus T750mg (cycles 2-8); B (n=12) paclitaxel plus T300mg (cycle 2) plus D1500mg (cycles 2-13); C (n=12) T750mg (cycles 1-7). The objective response rate in arm A was 26% (88% confidence interval (CI) = [14,36%]) versus 17% (88% CI = [3,42%]) in arm B and 8% (88% CI = [0.3,33%]) in arm C. Grade 3-4 treatment-related adverse events occurred in (A) 45%, (B) 75%, and (C) 25% of patients. Murine experiments showed attenuated tumor outgrowth for paclitaxel plus anti-CTLA-4 compared to monotherapy with either agent. Transcriptomic analyses showed upregulation of inflammation signatures in on-treatment tumor tissue biopsies. This study demonstrated encouraging antitumor activity in therapy-refractory mUC treated with paclitaxel plus high-dose anti-CTLA-4 and suggests immune induction may still be possible after failure to anti-PD-(L)1 therapy. PROJECT GOAL: We aimed to explore both potential biomarkers for response to combination ICB, as well as treatment dynamics to gain a better understanding of the effects of ICB on the urothelial cancer tumor microenvironment (TME).

  Study EGAS50000001574

• Cerebrospinal fluid cfDNA sequencing for classification of central nervous system glioma

  Primary central nervous system (CNS) gliomas can be classified by characteristic genetic alterations. In addition to solid tissue obtained by surgery or biopsy, cell-free DNA (cfDNA) derived from cerebrospinal fluid (CSF) is an alternative source of material for genomic analyses. Experimental design: We performed targeted next-generation sequencing (NGS) of CSF cfDNA in a representative cohort of 85 patients presenting with with suspicion of primary or recurrent glioma at two neurooncological centers. Copy-number variation (CNV) profiles, single nucleotide variants (SNVs), and small insertions/ deletions (indels) were combined into a molecular-guided tumor classification. Comparison with the solid tumor was performed for 38 cases with matching solid tissue available. Results: Cases were stratified into four groups: glioblastoma (n = 32), other glioma (n = 19), non-malignant (n = 17) and no confirmed diagnosis available (n = 17). We introduce a molecular-guided tumor classification, enabling identification of tumor entities and/ or cancer specific alterations in 75.0 % (n = 24) of glioblastoma and 52.6 % (n = 10) of other glioma cases. The overlap between CSF and matching solid tissue was highest for CNVs (26-48 %) and SNVs at pre-defined gene loci (44 %), followed by SNVs/indels identified via uninformed variant calling (8-14 %). A molecular-guided tumor classification was made 23.5 % (n = 4) of cases with no confirmed diagnosis available. Conclusions: We developed a workflow for targeted sequencing of CSF cfDNA as well as a strategy for interpretation and reporting of sequencing results based on a molecular-guided tumor classification in glioma.

  Study EGAS50000000060

• Exome-wide mutation analysis of cell-free DNA to simultaneously monitor the full spectrum of cancer treatment outcomes

  Purpose: Cell-free DNA (cfDNA) offers a non-invasive approach to monitor cancer. Here we develop a method using whole-exome sequencing (WES) of cfDNA for simultaneously monitoring the full spectrum of cancer treatment outcomes, including MRD, recurrence, evolution, and second primary cancers. Experimental Design: Three simulation datasets were generated from 26 cancer patients to benchmark the detection performance of MRD/recurrence and second primary cancers. For further validation, cfDNA samples (n=76) from cancer patients (n=35) with six different cancer types were used for performance validation during various treatments.Results: We present a cfDNA-based cancer monitoring method, named cfTrack. Taking advantage of the broad genome coverage of WES data, cfTrack can sensitively detect MRD and cancer recurrence by integrating signals across known clonal tumor mutations of a patient. In addition, cfTrack detects tumor evolution and second primary cancers by de novo identifying emerging tumor mutations. A series of machine learning and statistical denoising techniques are applied to enhance the detection power. On the simulation data, cfTrack achieved an average AUC of 99% on the validation dataset and 100% on the independent dataset in detecting recurrence in samples with tumor fraction ≥0.05%. In addition, cfTrack yielded an average AUC of 88% in detecting second primary cancers in samples with tumor fraction ≥0.2%. On real data, cfTrack accurately monitors tumor evolution during treatment, which cannot be accomplished by previous methods.Conclusion: Our results demonstrated that cfTrack can sensitively and specifically monitor the full spectrum of cancer treatment outcomes using exome-wide mutation analysis of cfDNA.

  Study EGAS00001005906

• PD-L1 blockade in combination with carboplatin as immune induction in metastatic lobular breast cancer: the GELATO-trial

  Invasive lobular breast cancer (ILC) is the second most common histological breast cancer subtype but published data on trials specific for ILC are so far lacking. Translational research revealed that a subset of ILCs may be immune-related and more sensitive to DNA-damaging agents such as platinum. In murine ILC models, synergy between immune checkpoint blockade and platinum has been observed. Here, we tested this concept in the phase II, GELATO-trial (NCT03147040), in which patients with metastatic ILC were treated with weekly carboplatin (AUC 1.5) as immune induction treatment for 12 weeks and atezolizumab (PD-L1 blockade; every three weeks) from the third week onwards until disease progression. Four out of 23 evaluable patients had a partial response (17%, 95%CI 5-39%) and two patients had stable disease for at least 24 weeks, resulting in a clinical benefit rate of 26% (95%CI 10-48%). Out of these six patients, four patients had triple-negative ILC (TN-ILC). In serial biopsies of metastatic lesions, we observed higher CD8 T-cell infiltration, expression of immune checkpoints, and exhausted T cells upon carboplatin/PD-L1 blockade. This is the first report of a clinical trial specifically for ILC and we demonstrate promising anti-tumor activity of atezolizumab with carboplatin as immune induction, in particular for TN-ILC. While activity of carboplatin/PD-L1 blockade in classical ER+ ILC was limited, our translational data yield important insights for the design of highly needed clinical trials in ILC.

  Study EGAS00001006902

• Drug Signatures for Prediction and Mitigation of Toxicity

  The overall goal of the drug signatures for prediction and mitigation of toxicity study is to use genomic and proteomic high-throughput measurements as the basis for computational analysis that integrates network analyses with structural constraints and dynamical models in multiple cell types to identify signatures that predict toxicity induced by 55 FDA-approved chemotherapeutic drugs and potential mitigation of this toxicity. Specifically, we have recruited and pre-screened healthy individuals to be included in our study. Ninety six male and female individuals that satisfied the initial pre-screening for the study and their sex, age, race/ethnicity recorded through an enrollment questionnaire were consented. Eighty five underwent a formal and thorough medical health screening. The evaluation involved assessment of all inclusion and exclusion criteria, a full medical history, measurement of weight, height, waist and hip circumference, heart rate, blood pressure, respiratory rate, and oxygen saturation, a physical exam and an electrocardiograph (ECG). Blood was drawn for analysis of clinical relevant parameters. A pregnancy test was included for female participants. All blood draws and pregnancy tests were sent to a certified clinical laboratory for clinical analysis. Of the 85 subjects that were screened 42 (48.3%) were deemed eligible for final inclusion. Of these 42, one declined to undergo skin biopsy and/or venesection and one subject didn’t show up for the scheduled biopsy, leaving 40 clinically healthy subjects on which a skin biopsy and venesection was performed. Fibroblast lines were established from the skin biopsies. Induced pluripotent cells (iPSC) were generated from the established fibroblast lines of these 40 clinically healthy racially diverse male and female study participants who ranged in age from 22 and 61 years through reprogramming using either the mRNA or Sendai-virus methods. One clone from each of the 40 iPSC line was fully characterized for normal karyotype, short-tandem repeat matching to the original fibroblast line (authentication), pluripotency by select pluripotent marker expression via immunocytochemistry and by mRNAseq-based PluriTest analysis, and whole-genome sequencing.

  Study phs002088

• Characterization of mutational signatures in human cancer cell lines reveals episodic APOBEC mutagenesis

  Multiple signatures of somatic mutations have been identified in human cancer genomes. To investigate whether mutational signatures continue to be generated, and if so their temporal patterns of activity, subsets of cell lines were cultured in vitro for extended periods and subjected to single cell cloning and whole genome or exome sequencing or directly to single cell whole genome sequencing. As expected, signatures of past exogenous exposures, such as tobacco smoke and ultraviolet light, were not generated in vitro. In contrast, signatures of normal and defective DNA repair and replication continued to be generated at essentially constant mutation rates. Signatures of APOBEC cytidine deaminase DNA-editing activity exhibited a distinctive pattern with substantial fluctuations in mutation rate over time and episodic bursts of mutations. The initiating factors for these bursts are unclear although retrotransposon mobilisation may play a role. This cell line set now constitutes a comprehensive resource of live experimental models of mutational processes of both known and unknown aetiologies potentially retaining the patterns of activity and regulatory influences operative in human cells in vivo.

  Dataset EGAD00001004202

• Mult-omics Palbociclib Resistance Study in HR+/HER2– Metastatic Breast Cancer

  This study is multi-omics study of a Asian longitudinal metastatic breast cancer (MBC) cohort treated with palbociclib plus endocrine therapy. It contains NGS of baseline (BL) and progressive disease (PD) from 70 patients, consisting of 79 tumor/normal matched whole exome sequencing (WES) from 62 patients and 90 tumor whole transcriptome sequecing samples (WTS) from 70 patients. There were 56 BL biopsies profiled by WES and 64 by WTS; 23 PD biopsies were profiled by WES and 26 by WTS. Twenty and 23 patients had paired BL and PD biopsies profiled by WES and WTS, respectively.

  Dataset EGAD00001008314

• A Study to Assess the Cardiovascular, Cognitive, and Subjective Effects of Atomoxetine in Combination with Intravenous Methamphetamine

  The Study to Assess the Cardiovascular, Cognitive, and Subjective Effects of Atomoxetine in Combination with Intravenous Methamphetamine started in 2007 and ended in 2011. It was a small pilot study that included methamphetamine abusing individuals and healthy control participants aged 18-50 years. Participants who qualified over the telephone were scheduled to visit the London Laboratory at the UCLA Semel Institute for Neuroscience and Human Behavior for the In-Person Screening Phase. The screening procedures, usually scheduled over two visits, determined if participants were eligible to complete the subsequent study visits. Both groups completed identical screening procedures, including a SCID diagnostic interview to rule out any major psychiatric disorders. Methamphetamine users also answered questionnaires specific to their drug use. Urine toxicology for marijuana, opiates, cocaine, methamphetamine, and benzodiazepines were monitored at the beginning and throughout the study. Additionally, participants had blood samples to test for Rapid Plasma Reagin (exposure to syphilis), purified protein derivative (PPD), HIV and Hepatitis-C (HCV). Positive test results were relayed by a study physician and resulted in termination from continuing in the study. The inpatient portion of the study lasted a total of 24 days. To confirm compliance with abstinence from illicit and prescription drug use for the duration of the study, urine and breath alcohol testing was performed for both inpatient (methamphetamine users) and outpatient (healthy control) groups. Each subject was randomized to receive either atomoxetine (80 mg daily) or placebo. During inpatient days of the study, a study physician oversaw research subjects daily and experimental procedures were conducted in the General Clinical Research Center (GCRC). During drug administration sessions, heart rate and blood pressure were assessed at frequent intervals. Experimental sessions lasted around three hours and were conducted at approximately the same time of day for a given participant. Lunch was provided to participants at 11:30am with the prohibition of caffeine on study days involving methamphetamine administration. Participants were provided with smoke breaks as needed to assure that acute nicotine withdrawal did not negatively impact performance on cognitive testing. The following outlines the day to day procedures for a methamphetamine inpatient: Phase I Inpatient Study (15 days total) Days 1-3: On the day of admission, participants received blood tests and an EKG in order to determine that they had no occult medical conditions which would make their participation in the study medically contraindicated. A three day washout period was provided at the beginning of the study to establish an appropriate methamphetamine-free baseline for testing. Days 4-6: Participants completed cognitive test batteries to assess inhibitory control including self-report measures and cognitive testing, as well as up to three MRI scanning sessions. Day 7: On day 7, at 9AM and 1PM, participants received methamphetamine in a single-blinded condition. It was administered as two IV infusions of 15mg separated by a 60min break. The total dose of IV methamphetamine was 30mg. Participants were clinically evaluated by continuous cardiac telemetry, serial EKG and measurement of vital signs, during and after infusions, in order to determine the medical safety of methamphetamine infusion in the absence of study medication. Day 8: On day 8 we evaluated the safety and residual effects of the methamphetamine administered on the previous day. Days 9-10: After demonstrating medical and psychiatric stability of the initial methamphetamine infusion, participants were then randomized to receive placebo or atomoxetine (0 or 40mg) under double-blind conditions, which would be given at 8AM on both study days. Days 11-12: On days 11 and 12, participants remained in the GCRC and took two doses of test compound atomoxetine (0 or 40mg) at 8AM and 8PM. On the morning of day 12, participants completed a series of cognitive testing. Day 13: On day 13, participants remained in the GCRC, and took two doses of the test compound atomoxetine (0 or 40mg) at 8AM and 8PM. At 9AM and noon, they received methamphetamine under double-blind conditions administered non-contingently as two infusions of 0 or 15mg by IV (intravenously), with each infusion separated by 60min. The total dose of IV methamphetamine that each participant received was 30mg. As before, participants were clinically evaluated by continuous cardiac telemetry, serial EKG and measurement of vital signs, during and after infusions, in order to determine the medical safety of methamphetamine infusion in those participants receiving study medication. Day 14: On day 14 at 8AM, participants received their final dosing of the first test compound (atomoxetine 0 or 40mg). At 9AM and 1PM, participants selected between methamphetamine infusion or money using double-blind conditions in a multiple-choice self-administration paradigm. Participants were able to take up to two infusions of 0 or 15mg of methamphetamine, separated by 60min. The total dose of IV methamphetamine that each participant received on day 14 was 30mg. Participants were clinically evaluated by continuous cardiac telemetry, serial EKG and measurement of vital signs, during and after infusions, in order to determine the medical safety of methamphetamine infusion in those participants receiving study medication. At 5pm on day 14 discharge laboratory studies were drawn. Day 15: On day 15, a final EKG was performed as well as a review of discharge laboratory studies, including review of the EKG and continued medical and psychiatric stability. Upon clearance from a physician, participants were discharged on day 15. Participants spent at least 2 weeks (14 days) out of the hospital prior to returning to the UCLA GCRC for inpatient Phase II. Phase II Inpatient Study (9 days total) Day 1-2: On the day of re-admission, participants received blood tests and an EKG. A two day washout period was provided at the beginning of the study to establish an appropriate methamphetamine free condition for safe administration of the second test compound condition. Days 3-4: On days 3 and 4, given medical stability, participants were then switched to the opposite test compound (atomoxetine 0 or 40mg) under double-blind conditions, in the same manner as in Days 9-10 in Phase I. Days 5-6: Same procedure as in Days 11-12 in Phase I but with opposite medication condition. Day 7: Same procedure as in Day13 in Phase I but with opposite medication condition. Day 8: Same procedure as in Day14 in Phase I but with opposite medication condition. Day 9: Same procedure as in Day14 in Phase I but with opposite medication condition. For healthy control participants, the study consisted of 30 days or fewer of out-patient testing. Healthy control participants had an extensive screening, including a history and physical exam performed by a study physician, an EKG, and laboratory studies performed to assess for medical or psychiatric conditions. Control participants meeting criteria were invited back to UCLA to undergo cognitive testing, and some participants received fMRI scans. Participants were required to refrain from illicit and prescription drug use for the duration of the study, which was confirmed with urine toxicology and breath alcohol level testing on examination days. Caffeine was restricted on study days for least 2 hours prior to cognitive testing and MRI scans. Participants were permitted to drink their normal daily intake of caffeine, given the caveat above. On study days, cigarette smoking was permitted to match conditions used with methamphetamine users. The incorporation of smoke breaks during the test day assured that acute nicotine withdrawal did not negatively impact performance on cognitive tests or neuroimaging. The following outlines procedures for healthy control participants: For normal controls, participation involved one day of outpatient screening, and one day of baseline cognitive testing and possible MRI scans. After these procedures were completed, participants were randomized to the first test compound (atomoxetine 40mg or placebo), to be taken once daily in the morning for 2 days. It was then increased to twice daily for the next 3 days, followed by a final dose (40mg or placebo) on the sixth day of study medication. Participants were also asked to bring their pill packages to assess compliance with the study medication. Additionally, control participants completed a series of repeated-measures cognitive testing (on placebo or atomoxetine, respectively) and possibly two fMRI sessions (separated by a short break). After at least a four-day washout period, control participants were instructed to start the opposite test compound (atomoxetine 40mg or placebo), again to be taken once daily in the morning for two days, then twice daily for 3 days, reaching the final dose (40mg or placebo) of the second test compound in the morning. Participants were again asked to bring their pill packages to assess compliance and had blood drawn to assess atomoxetine levels. Control participants then completed another series of cognitive testing and fMRI testing sessions. They were subsequently discharged from the study. Normal control participants returned two weeks after completion of both study phases in order to complete assessments to their physical and psychological status following treatment.

  Study phs001195

• Low mutation burden and frequent loss of CDKN2A/B and SMARCA2, but not PRC2, define pre-malignant neurofibromatosis type 1-associated atypical neurofibromas

  Neurofibromatosis type 1 (NF1) is a tumor-predisposition disorder caused by germline mutations in NF1. NF1 patients have an 8-16% lifetime risk of developing a malignant peripheral nerve sheath tumor (MPNST), a highly aggressive soft-tissue sarcoma, often arising from pre-existing benign plexiform neurofibromas (PN) and atypical neurofibromas (ANF). ANF are distinct from both PN and MPNST, representing an intermediate step in malignant transformation. In the first comprehensive genomic analysis of ANF, we performed tumor/normal exome sequencing (ES) of 16 ANFs. In addition, we conducted ES of three MPNSTs, copy-number meta-analysis of 26 ANF and 28 MPNST, and whole transcriptome RNA-seq analysis of five ANF and five MPNST. We identified low mutation burden (median 1, range 0-5) in the exomes of ANF (only NF1 somatic mutations were recurrent), and frequent deletions of CDKN2A/CDKN2B (69%) and SMARCA2(42%) loci. We determined that polycomb repressor complex 2 (PRC2) genes EED or SUZ12 were frequently mutated, deleted or downregulated in MPNST but not in ANF. Our gene expression study revealed upregulated NRAS, MDM2, CCND1/CCND2/CCND3 and CDK4/CDK6 in both ANF and MPNST, and overexpression of EZH2 in MPNST only. We conclude that the PN-ANF transition is primarily driven by the deletion of CDKN2A/CDKN2B in addition to already present inactivated NF1. Further progression from ANF to MPNST likely involves broad chromosomal rearrangements and frequent inactivation of the PRC2 genes, loss of the DNA repair genes, and copy-number increase of signal transduction, cell cycle and pluripotency self-renewal genes.

  Study phs001993

• Study of Controlled Human Malaria Infections to Evaluate Protection After Intravenous or Intramuscular Administration of PfSPZ Vaccine in Malaria-Naive Adults

  The study was performed to evaluate whole-blood transcriptomic profiles via RNA-seq before, during, and after immunization with the PfSPZ Vaccine in malaria-naïve, health adults enrolled in VRC 314 (NCT02015091). VRC 314 was designed as an open-label evaluation of the safety, tolerability, immunogenicity, and protective efficacy of the PfSPZ Vaccine. This study was designed to substantiate the initial results with the IV vaccination route for protection against CHMI that was observed in VRC 312 (NCT01441167). Based on the potential importance of dose and schedule in optimizing sustained immunity with this vaccine, an increase in PfSPZ IV dosage on schedules of 3 to 5 vaccinations was evaluated for protection against CHMI conducted early (about 3 weeks) and late (about 24 weeks) after completion of vaccinations. To assess if a higher dose given by another route confers protection, one group received PfSPZ IM, with half of the amount administered in each arm on a schedule with 4 vaccination. The primary objectives of the study were related to the safety and tolerability of vaccinations by the IV and IM routes of administration and protection against Plasmodium falciparum (Pf) challenge performed via a well-established CHMI procedure early (2-4 weeks) after completing schedules of 3 to 5 vaccinations. The secondary objective was related to the durability of protection at 20-26 weeks after the last vaccination, and exploratory objectives were related to the immunogenicity of the PfSPZ Vaccine and identifying potential immune correlates of protection. To further determine potential molecular correlates of immunogenicity and/or protection, RNA-seq was performed on whole blood collected from subjects in all dose groups at time points before, during, and after immunization.

  Study phs002423

• Discovery and Characterization of Genetic Risk Loci in Sjogren's Syndrome

  The Sjögren's Genetics Network (SGENE) is an international collaboration focused on identifying and understanding the genetic variations that influence Sjögren’s pathology. Collectively, SGENE collaborators from 26 different sites (5 in the United States; 21 in foreign countries) have recruited a large cohort of geographically diverse participants of European ancestry. Recruitment, sample collection, and genotyping were performed at unique SGENE sites in compliance with local institutional review board approval. High-density SNP genotyping data were collated by the Oklahoma Medical Research Foundation (OMRF) and used to perform a GWAS of Sjögren's of European ancestry. Participants underwent extensive medical assessment for Sjögren's Syndrome using internationally accepted American-European Consensus Group (AECG) classification criteria or the American College of Rheumatology/European League Against Rheumatism classification criteria. Data quality control, GWAS imputation using the Haplotype Reference Consortium, panel version 1.1, and dbGaP preparation/posting were performed at the OMRF. To facilitate research focused on understanding the genetics of Sjögren's syndrome, the Genomic Summary Results from this GWAS are being made available herein using a total of n=3232 Sjögren's cases and n=17,481 controls. A total of n=8962 controls were obtained from our SGENE collaborators or from our previous study (Lessard, et al. Nature Genetics 2013. PMID: 24097067). To achieve adequate case-to-control ratios for the GWAS analyses, population-based control GWAS data were also obtained from dbGaP (phs000428.v2.p2; phs000672.v1.p1; phs000196.v3.p1; phs000187.v1.p1). In accordance with IRB approval, subject consents, and data sharing restrictions, we are also making available the individual-level genotyping data for a subset of Sjögren's cases (n=1,199) that were genotyped at the OMRF.

  Study phs002723

• Genetic Aberrations and Subclonal Structure Impact Chronic Lymphocytic Leukemia

  Large-scale whole-exome sequencing (WES) of primary tumor samples enables the unbiased discovery of recurrent putative driver events and patterns of clonal evolution. We report the identification of 44 recurrently mutated genes and 11 recurrent CNVs through the WES of 538 chronic lymphocytic leukemia (CLL) and matched germline DNAs. These include previously unrecognized cancer drivers (e.g., RPS15, IKZF3), and collectively identify nuclear export, MYC activity and MAPK signaling as central pathways affected by somatic mutation in CLL. A clonality analysis of this large dataset further enabled the reconstruction of temporal relationships between these driver events. Several drivers were associated with shorter progression-free survival (PFS) in 280 samples that were collected prior to uniform treatment with front line chemo-immunotherapy, with mature follow up of greater than 10 years. Direct comparison between matched pretreatment and relapse CLL from 59 samples demonstrated marked clonal evolution occurring in more than 95% of these patients. Distinct patterns of clonal evolution in relationship to specific gene alteration were observed, suggesting a hierarchy of fitness amongst mutations. Thus, large WES datasets of clinically informative samples enable the discovery of novel driver genes as well as the network of relationships between the drivers and their impact on disease relapse and clinical outcome.Additionally, we performed RNA-seq for 268 CLL samples (including 26 follow-up samples) and used them to identify expression subtypes of CLL. RRBS for 30 of these samples was also generated. In an integrative analysis of genetic, transcriptomic, and epigenetic data, incorporating known and newly identified subtypes of CLL, we built new models to improve patient prognostication.

  Study phs000922

• Genomic platform specific polygenic risk scores impact breast cancer risk stratification

  Background: At present, there is no consensus on which genotyping platform should serve as the standard for clinical polygenic risk score (PRS) implementation. Previous studies have compared the overall performance and concordance of different genotyping and sequencing technologies; however, these analyses have generally averaged the results over the whole genome. We evaluated differences in a 313-variant breast cancer PRS (PRS313) across genomic platforms and their impact on risk stratification. Methods: We compare PRS313 derived from genotyping arrays (Global Screening Array [GSA], OncoArray-500K [OncoArray], Global Diversity Array [GDA], custom Axiom_PrecipV1 array [ThermoFisher]) and low-coverage genome sequencing (lc-WGS) in 2 cell lines and 92 individuals. Probes are designed for all variants on ThermoFisher (success rate: 259/313). Sanger sequencing profiles indels. Concordance of high-risk classification (PRS313scoresum > 0.6) across platforms is assessed using Kappa statistics. Results: In saliva samples, indel concordance with Sanger sequencing varies widely (Kappa: 0.007-1.000). PRS313-ThermoFisher is predictable from other platforms using linear models, despite systematic differences. Greater agreement is observed between arrays with high imputation overlap (e.g., GDA ~ GSA slope=0.986). Agreement in high-risk classification before mean correction is moderate (Fleiss's Kappa=0.552) and improves after mean correction (Kappa=0.650). Arrays with similar designs show higher agreement before mean correction (Kappa=0.745). Mean correction narrows high-risk proportions from 4-45% to 15-21%. Overall, 26 of 92 samples are classified as high risk on at least one platform, but only 7 are high risk across all. When restricting to identical variants across all platforms for PRS313 calculation, the corresponding number of high-risk individuals are 24 and 11. Conclusion: Our findings demonstrate that platform-specific variability can influence PRS313 estimates to potentially reclassify individuals around clinically relevant thresholds.

  Study EGAS00001008439

• Genetic characterization of B-cell prolymphocytic leukemia: a hierarchical prognostic model involving MYC and TP53 abnormalities - WXS

  B-PLL is a rare disease, whose molecular pathology is largely unknown. We report here the cytogenetic and molecular findings in a large series of 34 B-PLL. Karyotype (K) was complex (≥3 abnormalities) in 73%, and highly complex (HCK≥5) in 45%. The most frequent chromosomal aberrations were: translocation targeting the MYC gene [t(MYC)] (62%), 17p deletion including TP53 gene (38%), trisomy 18/18q (30%), 13q14 deletion (29%), trisomy 3 (24%), trisomy 12 (24%) and 8p deletion (23%). Whole-Exome Sequencing performed in 16 patients revealed recurrent mutations in TP53 (6/16, 38%), MYD88 (n=4), BCOR (n=4), MYC (n=3), SF3B1 (n=3), FAT1 (n=3), SETD2 (n=2), CHD2 (n=2), CXCR4 (n=2) and BCLAF1 (n=2). The main group of patients (21/34, 62%) had a t(MYC) associated with a higher percentage of prolymphocytes (p=0.03), CD38 expression (p<0.001), lower K complexity (p=0.0004), mutations in MYC and in genes involved in RNA metabolism and chromatin remodeling. Principal component analysis of gene expression data showed that patients with t(MYC) clustered together. A second group with MYC gain (5/34, 15%), was associated with HCK≥5 (p=0,01) and trisomy 3 (p=0,008). Altogether, 26/34 patients (76%) had a MYC activation, translocation or gain, that were mutually exclusive. We identified 3 distinct cytogenetic prognostic groups (p=0.0006): lower risk: absence of MYC activation (median not reached); intermediate risk: MYC activation without del17p (125 months); high risk: MYC activation + del17p (11 months). Our results show that cytogenetic analysis is a useful diagnostic tool in B-PLL that improves prognostic stratification.

  Study EGAS00001003275

• PCGP Ph-like ALL

  Background: BCR-ABL1-like, or Ph-like acute lymphoblastic leukemia is characterized by a gene expression profile similar to BCR-ABL1 positive ALL, genetic alterations of lymphoid transcription factor genes, and poor outcome. Sequencing of small numbers of Ph-like ALL cases has identified genetic alterations activating kinase signaling suggesting Ph-like ALL may be amenable to treatment with tyrosine kinase inhibitors. However, the spectrum of genetic alterations in childhood and adult Ph-like ALL is incompletely understood. Methods: We performed genomic profiling of 1736 B-ALL cases and next-generation sequencing for 160 Ph-like cases. We examined the functional effects of chimeric fusion proteins in mouse cell lines. Results: The frequency of Ph-like ALL rose from 11% in children to 26% in young adults, and was associated with very poor outcome. Kinase-activating alterations were identified in 90% of Ph-like cases, most commonly fusions involving 10 kinase or cytokine receptor genes (ABL1, ABL2, CRLF2, CSF1R, EPOR, JAK2, NTRK3, PDGFRB, PTK2B and TYK2), and mutations involving FLT3, IL7R and SH2B3. Expression of ABL1, ABL2, CSF1R and JAK2 fusions resulted in cytokine-independent proliferation of cell lines and activation of pSTAT5. Cells expressing ABL1, ABL2, CSF1R and PDGFRB fusions were sensitive to dasatinib, and JAK2 fusions to ruxolitinib. Conclusions: Ph-like ALL is characterized by a diverse range of genomic alterations that converge on a limited number of kinase signaling pathways amenable to inhibition with currently available tyrosine kinase inhibitors. Trials identifying Ph-like ALL and testing the efficacy of tyrosine kinase inhibitor therapy are warranted

  Study EGAS00001000654

• Recurrent somatic DICER1 mutations in non-epithelial ovarian tumors

  Background: Germline truncating mutations in DICER1, an RNase III-type endoribonuclease essential for processing of microRNAs, are seen in families with the pleuropulmonary blastoma-family tumor and dysplasia syndrome; these individuals have a propensity to develop non-epithelial ovarian tumors (i.e. sex-cord stromal and germ cell tumors). Methods: We sequenced the whole transcriptomes or exomes of 14 non-epithelial ovarian tumors and noted closely clustered mutations in the RNase IIIb domain of DICER1 in four cases. Using Sanger sequencing, we then analyzed additional ovarian tumors for mutations in this region. The impact of the mutations on the enzymatic activity of Dicer1 was determined using in vitro RNA cleavage assays. Results: RNase IIIb domain DICER1 mutations were found in 30/102 non-epithelial ovarian tumors (29%), predominantly Sertoli-Leydig cell tumors (26/43, 60%), including 4/4 from individuals with germline DICER1 mutations. The mutations were restricted to metal binding sites within the RNase IIIb catalytic centres critical for miRNA interaction and cleavage, and were somatic in all 14 cases where germline DNA was available for testing. Of 266 epithelial ovarian and endometrial cancers tested, one case, an ovarian carcinosarcoma, had a hotspot mutation. In vitro, the mutant DICER proteins showed reduced RNase IIIb activity but retained RNase IIIa activity. Conclusions: Somatic missense mutations of the RNase IIIb domain of DICER1 are common in non-epithelial ovarian tumors. Genetic, pathologic, and functional evidence suggest that these mutations do not obliterate DICER1 function, rather, they alter it in specific cell types, a novel mechanism through which perturbed microRNA expression is oncogenic.

  Study EGAS00001000135

• Genetic characterization of B-cell prolymphocytic leukemia: a hierarchical prognostic model involving MYC and TP53 abnormalities - RNA-seq

  B-cell prolymphocytic leukemia (B-PLL) is a rare disease, whose molecular pathology is largely unknown. We report here the cytogenetic and molecular findings in a large series of 34 B-PLL. Karyotype (K) was complex (≥3 abnormalities) in 73%, and highly complex (HCK≥5) in 45%. The most frequent chromosomal aberrations were: translocation targeting the MYC gene [t(MYC)] (62%), 17p deletion including TP53 gene (38%), trisomy 18/18q (30%), 13q14 deletion (29%), trisomy 3 (24%), trisomy 12 (24%) and 8p deletion (23%). Whole-Exome Sequencing performed in 16 patients revealed recurrent mutations in TP53 (6/16, 38%), MYD88 (n=4), BCOR (n=4), MYC (n=3), SF3B1 (n=3), FAT1 (n=3), SETD2 (n=2), CHD2 (n=2), CXCR4 (n=2) and BCLAF1 (n=2). The main group of patients (21/34, 62%) had a t(MYC) associated with a higher percentage of prolymphocytes (p=0.03), CD38 expression (p<0.001), lower K complexity (p=0.0004), mutations in MYC and in genes involved in RNA metabolism and chromatin remodeling. Principal component analysis of gene expression data showed that patients with t(MYC) clustered together. A second group with MYC gain (5/34, 15%), was associated with HCK≥5 (p=0,01) and trisomy 3 (p=0,008). Altogether, 26/34 patients (76%) had a MYC activation, translocation or gain, that were mutually exclusive. We identified 3 distinct cytogenetic prognostic groups (p=0.0006): lower risk: absence of MYC activation (median not reached); intermediate risk: MYC activation without del17p (125 months); high risk: MYC activation + del17p (11 months). Our results show that cytogenetic analysis is a useful diagnostic tool in B-PLL that improves prognostic stratification.

  Study EGAS00001003274

• IMCISION DNAseq

  Standard-of-care (SOC) surgery for locoregionally advanced head and neck squamous cell carcinoma (HNSCC) is intensive and results in 30‒50% five-year overall survival. Anti-PD1 immune checkpoint blockade (ICB) durably improves survival rates in recurrent/metastatic HNSCC. We report on the non-randomized phase Ib/IIa IMCISION trial (NCT03003637) of 32 HNSCC patients treated with two doses neoadjuvant nivolumab (NIVO MONO, n=6, phase Ib arm A) or two doses of nivolumab plus a single dose of ipilimumab (COMBO, n=26, 6 in phase Ib arm B and 20 in phase IIa) prior to surgery and, if indicated, adjuvant radiotherapy. Neoadjuvant ICB was feasible in all phase Ib patients, meeting the phase Ib primary feasibility endpoint. One phase IIa patient had progressive disease precluding surgery. Primary tumor pathological response (phase IIa primary endpoint), defined as the % change in viable tumor cells from baseline biopsy to on-treatment resection, was evaluable in 29 patients. . We observed a major pathological response (MPR, 90‒100% response) in 8/23 (35%) evaluable COMBO and 1/6 (17%) NIVO MONO patients. None of the patients with MPR after NIVO MONO or COMBO developed a tumor relapse after 24.0 months median follow-up. FDG-PET-based total lesion glycolysis identified MPR patients prior to surgery. A baseline AID/APOBEC-associated mutational profile and an on-treatment decrease in hypoxia signature expression were observed in MPR patients. Our data indicate that neoadjuvant NIVO MONO and COMBO are safe in HNSCC, and that particularly COMBO ICB shows encouraging effectivity. IMCISION provides rationales for future HNSCC trials aiming to pre-operatively select patients with a likely MPR after neoadjuvant ICB for de-escalation of SOC.

  Study EGAS00001005466

• National Eye Institute (NEI) Age-Related Eye Disease Study (AREDS)

  The Age-Related Eye Disease Study (AREDS) was initially designed as a long-term multi-center, prospective study of the clinical course of age-related macular degeneration (AMD) and age-related cataract. In addition to collecting natural history data, AREDS included a clinical trial of high-dose vitamin and mineral supplements for AMD and a clinical trial of high-dose vitamin supplements for cataract. AREDS participants were 55 to 80 years of age at enrollment and had to be free of any illness or condition that would make long-term follow-up or compliance with study medications unlikely or difficult. On the basis of fundus photographs graded by a central reading center, best-corrected visual acuity and ophthalmologic evaluations, 4,757 participants were enrolled in one of several AMD categories, including persons with no AMD. The clinical trials for AMD and cataract were conducted concurrently. AREDS participants were followed on the clinical trials for a median time of 6.5 years. Subsequent to the conclusion of the clinical trials, participants were followed for an additional 5 years and natural history data were collected. The AREDS research design is detailed in AREDS Report 1. AREDS Report 8 contains the mainline results from the AMD trial; AREDS Report 9 contains the results of the cataract trial. Blood samples were also collected from 3,700+ AREDS participants for genetic research. Genetic samples from 600 AREDS participants (200 controls, 200 Neovascular AMD cases, and 200 Geographic Atrophy cases) were selected using data available in March 2005 and then were evaluated with a genome-wide scan. These data, as well as selected phenotypic data, were made available in the dbGaP. DNA from AREDS participants, which is currently being stored in the AREDS Genetic Repository, is available for research purposes. However, not all of the 3,700+ AREDS participants who submitted a blood sample currently have DNA available. In addition to including the data from the genome-wide scan on the 600 original samples, this second version of the AREDS dbGaP database provides a comprehensive set of data tables with extensive clinical information collected for the 4,757 participants who participated in AREDS. The tables include information collected at enrollment/baseline, during study follow-up, fundus and lens pathology, nutritional estimates, quality of life measures and measures of morbidity and mortality. In November 2010, over 72,000 high quality fundus and lens photographs of 595 AREDS participants (of the original 600 selected for the genome-wide scan) were made available in the AREDS dbGaP. In addition to the genome-wide scan data, the fundus and lens grading data for these participants are also available through the AREDS dbGaP. Details about the ocular photographs that are available may be found in the document "Age-Related Eye Disease Study (AREDS) Ocular Photographs". In January 2012, a measure of daily sunlight exposure was added in a separate "sunlight" table. Furthermore, the "followup" table has been revised. The visual acuity for the right eye was inadvertently missing at odd-numbered visits (01, 03, 05, etc.). This data is now part of the table. In February 2014 over 134,500 high-quality fundus photographs (macular field F2) of 4613 AREDS participants were added to the existing AREDS dbGaP resource. The AREDS dbGaP image archive already contains over 72,000 high quality fundus and lens photographs of 595 AREDS participants for whom dbGaP-accessible genotype data exist. Information about the available ocular photographs found in the document "Age-Related Eye Disease Study (AREDS) Ocular Photographs" has been updated with an addendum. It is hoped that this resource will better help researchers understand two important diseases that affect an aging population. These data may be applied to examination and inference on genetic and genetic-environmental bases for age-related diseases of public health significance and may also help elucidate the clinical course of both conditions, generate hypotheses, and aid in the design of clinical trials of preventive interventions. Definitions of Final AMD Phenotype Categories Please see phd001138.1 for a detailed description of how AREDS participants' final AMD phenotype was categorized. User's Guide for AREDS Phenotype Data A detailed User's Guide for the AREDS phenotype data is available. This User's Guide is meant to be a comprehensive document which explains the complexities of the AREDS data. It is recommended that all researchers using AREDS phenotype data make use of this User's Guide.

  Study phs000001

• A Genome-Wide Association Study in Patients Experiencing Musculoskeletal Adverse Events on NCIC CTG Trial MA.27 Evaluating Aromatase Inhibitors as Adjuvant Therapy in Early Breast Cancer. A Collaboration Between the NIH Pharmacogenetics Research Network and the RIKEN Yokohama Institute Center for Genomic Medicine.

  Source of Patients The source of the patients for this genome-wide case control study was MA.27, which was conducted as a multi-cooperative group effort under the auspices of the NCI Breast Cancer Intergroup of North America. The NCIC Clinical Trials Group (CTG) serves as the coordinating group, with participation by the NCI-sponsored North Central Cancer Treatment Group, Eastern Cooperative Oncology Group (ECOG), Southwest Oncology Group, and Cancer and Leukemia Group B (CALGB). MA.27 involved postmenopausal women with histologically confirmed and completely resected invasive breast cancer with surgical margins clear of invasive carcinoma and DCIS in the following TMN categories (AJCC Version 6): pT1, pT2, pT3; pNx, pN0, pN1, pN2, pN3 (only when the sole basis is presence of 10 or more involved axillary nodes); MO. The primary tumor must have been ER and/or progesterone receptor positive. Patients were stratified by lymph node status at diagnosis, prior adjuvant chemotherapy, and trastuzumab use and were randomized to 5 years of adjuvant therapy with anastrozole or exemestane. The trial was activated on May 26, 2003, and reached its accrual objectives on July 31, 2008, after the randomization of 6827 North American patients, with the majority (79%) providing DNA and consent for genetic testing. Non-North American patients were also entered by the International Breast Cancer Study Group but they did not contribute DNA. From 2003 to December 21, 2004, patients also underwent a second randomization to celecoxib 400 mg twice daily or placebo but, after the entry of 1,622 patients, this treatment was discontinued because of reports of increased cardiovascular risk associated with celecoxib. Case Definition for Musculoskeletal Adverse Events Cases were defined to have had at least one of the following six MSAE): 1. pain - joint, 2. pain - muscle, 3. pain - bone, 4. arthritis, 5. joint - function, 6. musculoskeletal - other. Cases were required to have at least grade 3 toxicity, according to the NCI Common Terminology Criteria for Adverse Events v3.0, or go off treatment for any grade of the six MSAEs; and were required to have had an MSAE within the first two years (i.e., a MSAE occurring after 2 years was not be considered a case). Subjects were excluded from the case group who met the case definition while on celecoxib or in the three months after stopping celecoxib but subjects were allowed to be cases if they developed their first MSAE more than three months after stopping celecoxib. Study Design The design was a nested matched case-control study with two controls selected for each case. Random matching from the potential control group (i.e., risk-set sampling such that controls had at least the same length of followup as the timing of the case event) was done by matching the case on the following factors: 1. treatment arm (exemestane vs. anastrozole, blinded), 2. prior adjuvant chemotherapy (yes vs. no), 3. age at start of treatment (+/- 5 years), 4. Celecoxib allocation (yes vs. no). Attempts were made to exactly match each case with two controls according to the above criteria. If, however, controls could not be exactly matched, "close" matching was used to define a distance between each case and all potential controls in an optimal matching algorithm. Race is a potential confounding factor, and 94% of the MA.27 population is Caucasian, 3% are Black, 1% Asian, 1% American Indian, and 1% are of unknown race. Because there are so few non-Caucasian participants, was restricted to only Caucasians. Following identification of cases and controls, the list was sent to the CALGB Pathology Coordinating Office (PCO) at Ohio State University Medical Center for plating of the samples. Other patient factors that might influence the association of SNP genotypes with a MSAE, either as confounding factors or as effect modifiers, were included as investigational covariates. These include 1. body mass index (BMI), 2. bisphosphonate use, 3. fractures in past 10 years, 4. baseline ECOG performance status, 5. prior hormone replacement therapy, 6. prior adjuvant radiotherapy, and 7. prior taxane treatment. An Honest Broker Process was utilized to ensure patient confidentiality. Genotype Assays and Quality Control A GWAS was performed with the Illumina Human610-Quad at the RIKEN Center for Genomic Medicine. The DNA for all of the samples were adjusted to a concentration of 50 to 100 ng/µL in a final volume of 50 µL. The 96-well plates were designed to have a unique distribution pattern with x duplicates per plate to evaluate genotype concordance. Samples with <98% call rate were removed from analyses. SNPs with ambiguous call will be excluded using the following criteria: 1. low signal intensity of each allele, 2. low cluster separation score, 3. SNP call rate <0.98, 4. SNPs with Hardy Weinberg Equilibrium p-value <0.001 were excluded. X-chromosome SNPs were used to validate female sex and quality control measures were examined across the different submitting centers. For all pairs of subjects, the percentage of matching genotypes were computed to screen for unexpected duplicate samples. All SNPs found to be associated with MSAE were subjected to visual inspection of the cluster plots to verify that the genotype clustering was adequate.

  Study phs000210

• CIDR: Collaborative Study on the Genetics of Alcoholism Case Control Study

  This is a case-control study of alcoholism, in which the subjects have been drawn from the Collaborative Study on the Genetics of Alcoholism (COGA), a large, ongoing family-based study that includes subjects from seven sites around the US. COGA has gathered detailed, standardized data on study participants, including diagnostic and neurophysiological assessments. This sample has already proved successful in identifying several genes that influence the risk for alcoholism and neurophysiological endophenotypes, which have been independently replicated. COGA data were included as part of two Genetic Analysis Workshops, and the phenotypes are familiar to the genetics community. Alcoholic probands were recruited from treatment facilities, assessed by personal interview, and after securing permission, other family members were also assessed. A set of comparison families was drawn from the same communities as the families recruited through an alcoholic proband. Assessment involved a detailed personal interview developed for this project, the Semi-Structured Assessment for the Genetics of Alcoholism (SSAGA), which gathers detailed information on alcoholism related symptoms along with other drugs and psychiatric symptoms. Many participants also came to the laboratories for electroencephalographic studies. Neurophysiological features that have been shown to be useful endophenotypes for which we have linkage and in some cases association results are included on a subset of the case-control sample: the beta power of the resting electroencephalogram (EEG), the P3(00) amplitude of the visual event-related potential (ERP), and the theta and delta event-related oscillations (EROs) underlying the P3 (See Porjesz et al., 2005; Porjesz and Rangaswamy, 2007 for reviews). A brief description of COGA is in Edenberg, H. J. (2002) The Collaborative Study on the Genetics of Alcoholism: an update. Alcohol Res Health 26, 214-218., Bierut, LJ, NL Saccone, JP Rice, A Goate, T Foroud, HJ Edenberg, L Almasy, PM Conneally, R Crowe, V Hesselbrock, T-K Li, JI Nurnberger, Jr, B Porjesz, MA Schuckit, J Tischfield, H Begleiter, and T Reich (2002) Defining alcohol-related phenotypes in humans: The Collaborative Study on the Genetics of Alcoholism. Alcohol Res Health 26, 208-213. Edenberg HJ and Foroud T (2006) The genetics of alcoholism: identifying specific genes through family studies. Addiction Biology 11, 386-396. This case-control sample of biologically unrelated individuals was drawn from COGA subjects. All cases meet DSM-IV criteria for alcohol dependence. Controls are individuals who have consumed alcohol, but did not meet any definition of alcohol dependence or alcohol abuse, nor did they meet any DSM-IIIR or DSM-IV definition of abuse or dependence for other drugs (except nicotine). All cases and controls have undergone identical clinical assessments. Many individuals in this case-control sample have not previously been genotyped. The Collaborative Study on the Genetics of Alcoholism (COGA) has four Co-Principal Investigators: B. Porjesz, V. Hesselbrock, H. Edenberg, L. Bierut. COGA includes nine different centers where data collection, analysis, and storage take place. The nine sites and Principal Investigators and Co-Investigators are: University of Connecticut (V. Hesselbrock); Indiana University (H.J. Edenberg, J. Nurnberger Jr., T. Foroud); University of Iowa (S. Kuperman); SUNY Downstate (B. Porjesz); Washington University in St. Louis (L. Bierut, A. Goate, J. Rice); University of California at San Diego (M. Schuckit); Howard University (R. Taylor); Rutgers University (J. Tischfield); Southwest Foundation (L. Almasy). Q. Max Guo serves as the NIAAA Staff Collaborator. This national collaborative study is supported by the NIH Grant U10AA008401 from the National Institute on Alcohol Abuse and Alcoholism (NIAAA) and the National Institute on Drug Abuse (NIDA). Funding support for genotyping, which was performed at the Johns Hopkins University Center for Inherited Disease Research, was provided by the National Institute on Alcohol Abuse and Alcoholism, the NIH GEI (U01HG004438),and the NIH contract "High throughput genotyping for studying the genetic contributions to human disease" (HHSN268200782096C). COGA has over 250 publications listed at www.niaaagenetics.org

  Study phs000125

• High-depth whole genome sequencing of 26 premalignant breast lesions

  Ductal carcinoma in situ (DCIS) is a non-obligate precursor lesion of breast cancer, representing 20-25% of newly diagnosed breast cancer lesions. It is currently not possible to predict which DCIS patients will progress to invasive breast cancer. The samples deposited here are part of a larger study using high-depth whole-genome sequencing (WGS) to investigate the somatic mutation genomic landscape of DCIS. Comprehensive evaluation of DCIS genomes aims to uncover biological insights into breast cancer progression and potential opportunities to stratify DCIS patients for personalised treatment options or active surveillance. Due to specific restrictions imposed by the ethical approval at sample collection, the use of the germline data is restricted to filtering of somatic mutation calls only and cannot be used outside this purpose.

  Study EGAS50000001559

• scRNA-seq of total bone marrow and T cells from multiple myeloma long-term survivors

  scRNA This dataset contains 50 scRNA-seq samples from bone marrow aspirates of 11 multiple myeloma patients experiencing long-term survival and 3 healthy donors. For each donor, total bone marrow and CD3+ T cells were sequenced. For multiple myeloma patients, paired samples were collected at initial diagnosis and between 7-17 years after first-line therapy. Bone marrow mononuclear cells were isolated by Ficoll density gradient centrifugation. For sorting of total bone marrow cells singlet, live cells were gated and sorted, for sorting of T cells CD45+, CD3+ cells were gated and sorted on either FACSAria Fusion or FACSAria II. Single-cell RNA sequencing were generated using 10x Genomics single-cell RNAseq technology (Chromium Single Cell 3’ Solution v2) according to the manufacturer’s protocol and sequenced on an Illumina HiSeq4000 (paired end, 26 and 74 bp). Bulk RNA Singlet, live CD3+CD4- CXCR3+CD8+ and CD3+CD4- CXCR3-CD8+ cells were sorted from 7 bone marrow and 3 peripheral blood samples of 7 multiple myeloma patients using a FACSAria Fusion machine. Bulk-RNA sequencing libraries were generated using the SMART Seq Stranded Total RNA-Seq kit (Takara) and sequenced using the Illumina NovaSeq 6000 platform (2 x 100 bp).

  Dataset EGAD00001010025

• Influence of the Microbiome on Epigenetic Mechanisms in IBD

  Live CD4 T cells were sorted from inflamed and non-inflamed tissue samples of IBD patients or from healthy and IBD blood samples. ATAC-Seq libraries were generated from live CD4 T cells sorted from i) inflamed and non-inflamed tissue samples, ii) healthy and IBD blood samples, or from iii) CD4 T cell subsets polarised from healthy blood samples. After isolating crude nuclei, live CD4+ T cells were treated with Tagment DNA buffer and Tagment DNA Enzyme (Nextera DNA Library Prep Kit, Illumina), and then the DNA was purified by MinElute PCR Purification Kit (Qiagen). Transposed DNA fragments were amplified using specific adapters followed by purification with MinElute PCR Purification Kit (Qiagen). Fragments from 240-360pb were selected in the PippinHT system (Sage Science). The quality of the library and its DNA concentration were assessed by Bioanalyzer instruments (Agilent Technologies) and ultimately submitted for sequencing using Illumina HiSeq 2500 sequencer, V4 chemistry. On the other hand, single cell RNA-Seq libraries were generated exclusively from inflamed and non-inflamed tissue samples of Crohn’s disease patients. Briefly, live CD4 T cells were captured and encapsulated before cDNA amplification using the 10X Genomics Chromium Platform. Samples were prepared as outlined by 10x genomics Single Cell 3’ Reagent Kits v2 user guide. Samples were sequenced on a HiSeq 2500 with the following run parameters: Read 1 – 26 cycles, read 2 – 98 cycles, index 1 – 8 cycles.

  Dataset EGAD00001011066

• Smoking-dependent expression alterations in nasal epithelium reveal immune impairment linked to germline variation and lung cancer risk.

  Lung cancer is the leading cause of cancer-related death in the world. In contrast to many other cancers, a direct connection to lifestyle risk in the form of cigarette smoke has long been established. More than 50% of all smoking-related lung cancers occur in former smokers, often many years after smoking cessation. Despite extensive research, the molecular processes for persistent lung cancer risk are unclear. CT screening of current and former smokers has been shown to reduce lung cancer mortality by up to 26%. To examine whether clinical risk stratification can be improved upon by the addition of genetic data, and to explore the mechanisms of the persisting risk in former smokers, we have analyzed transcriptomic data from accessible airway tissues of 487 subjects. We developed a model to assess smoking associated gene expression changes and their reversibility after smoking is stopped, in both healthy subjects and clinic patients. We find persistent smoking associated immune alterations to be a hallmark of the clinic patients. Integrating previous GWAS data using a transcriptional network approach, we demonstrate that the same immune and interferon related pathways are strongly enriched for genes linked to known genetic risk factors, demonstrating a causal relationship between immune alteration and lung cancer risk. Finally, we used accessible airway transcriptomic data to derive a non-invasive lung cancer risk classifier. Our results provide initial evidence for germline-mediated personalised smoke injury response and risk in the general population, with potential implications for managing long-term lung cancer incidence and mortality.

  Dac EGAC50000000169

• Whole Genome Sequencing of a Triple Negative Breast Cancer Patient: Matched Primary Tumor, Normal, Metastasis and Xenograft samples

  Breast Cancer Subject Participant ID 700064 (Source Sample names: 6888 and 206). We used massively parallel DNA sequencing technologies to screen entire genomes, in an unbiased manner, for genetic changes associated with tumor growth and metastasis. We describe the complete genome sequence analysis of four DNA samples from a 44-year old African-American patient with basal-like breast cancer: peripheral blood, the primary tumor, a brain metastasis that developed within a year of initial therapy, and a first-passage xenograft derived from the primary tumor. A total of 50 validated mutations were discovered within coding, RNA, or splice site sequences. Of these, 20 mutations were abundantly present in all three tumors, including mutations in CSMD1 and JAK2. These two genes subsequently were found to be mutated in other breast tumors. The metastasis contained two de novo mutations not present in the primary tumor, and was significantly enriched for 20 shared mutations, suggesting that they may be involved in the metastatic process. The xenograft contained no unique coding, RNA, or splice site mutations and retained all primary tumor mutations, albeit at different frequencies. However, a significant increase in copy number alterations was observed in the xenograft as compared to the primary tumor. We validated 28 large deletions and six inversions, as well as seven translocations in at least one of the three tumor samples. Among them, a 26 kb deletion in MECR was solely identified, assembled, and validated in the brain metastasis and two overlapping large deletions on chromosome 5 encompassing CTNNA1, a potential tumor suppressor gene, were identified in all three tumors. The differential mutation frequencies and structural variation patterns between primary and metastatic tumors suggest that metastatic tumors may arise from minor subpopulations of cells within the primary. Namely, the metastatic and xenografting processes apparently select for cells harboring a distinct subset of the primary tumor mutation repertoire.

  Study phs000245

• Genetic analysis of HLA and immune escape genes in Diffuse Large B-cell Lymphoma

  Fifty percent of diffuse large B-cell lymphoma (DLBCL) cases lack cell-surface expression of the class-I major histocompatibility complex (MHC-I), thereby escaping immune recognition by cytotoxic T cells. In order to comprehensively identify the mechanisms involved in MHC-I loss, we first performed immunophenotypic analysis of both MHC class-I and -II in 657 cases across the spectrum of B-NHL, revealing that loss of MHC-I, but not MHC-II, is preferentially restricted to DLBCL. We then used whole exome and targeted deep-sequencing to examine genes involved in MHC-I expression in 74 DLBCL samples representative of MHC-I positive and negative cases. We show here that somatic biallelic or monoallelic inactivation of B2M and/or HLA-I is present in 80.9% (34/42) of MHC-I negative tumors. Furthermore, 68.8% (22/32) of MHC-I positive DLBCLs also harbored monoallelic HLA-I genetic alterations (MHC-I positivemono) that lead to allelic imbalance, suggesting allele-specific inactivation. Both MHC-I negative and MHC-I positivemono cases showed a significantly higher mutational burden as well as a higher number of inferred neo-antigens, suggesting co-selection of HLA-I loss and sustained neo-antigen production. Interestingly, the analysis of > 500.000 individuals in two databases revealed homozygosity of germline HLA-I genes in 26-38% of DLBCL patients, a frequency significantly higher than that observed in any other cancer type. In mice, germinal-center B cells lacking HLA-I expression did not progress to lymphoma and were counterselected in the context of oncogene-driven lymphomagenesis, suggesting that additional events are needed to license immune evasion. These results suggest a multi-step process of HLA-I loss including both germ-line and somatic events in DLBCL development, and have direct implications for the pathogenesis and immunotherapeutic targeting of this disease.

  Study EGAS00001005054

• RNA-seq data for study 'Smoking-dependent expression alterations in nasal epithelium reveal immune impairment linked to germline variation and lung cancer risk.'

  RNA-seq data from nasal and bronchial tissues in 649 subjects, many with lung cancer. Lung cancer is the leading cause of cancer-related death in the world. In contrast to many other cancers, a direct connection to lifestyle risk in the form of cigarette smoke has long been established. More than 50% of all smoking-related lung cancers occur in former smokers, often many years after smoking cessation. Despite extensive research, the molecular processes for persistent lung cancer risk are unclear. CT screening of current and former smokers has been shown to reduce lung cancer mortality by up to 26%. To examine whether clinical risk stratification can be improved upon by the addition of genetic data, and to explore the mechanisms of the persisting risk in former smokers, we have analyzed transcriptomic data from accessible airway tissues of 487 subjects. We developed a model to assess smoking associated gene expression changes and their reversibility after smoking is stopped, in both healthy subjects and clinic patients. We find persistent smoking associated immune alterations to be a hallmark of the clinic patients. Integrating previous GWAS data using a transcriptional network approach, we demonstrate that the same immune and interferon related pathways are strongly enriched for genes linked to known genetic risk factors, demonstrating a causal relationship between immune alteration and lung cancer risk. Finally, we used accessible airway transcriptomic data to derive a non-invasive lung cancer risk classifier. Our results provide initial evidence for germline-mediated personalised smoke injury response and risk in the general population, with potential implications for managing long-term lung cancer incidence and mortality.

  Dataset EGAD50000000333

• CNS Embryonal tumors

  The dataset represents a total of 58 DNA samples from 16 male and 12 female pediatric patients affected with embryonal central nervous system tumors. The samples were subject to whole genome sequencing, WGS, [48 samples, (representing 12 male and 11 female individuals)] and whole exome sequencing, WES, [10 samples, (representing 4 male and 1 female individuals)]. One tumor tissue sample and one peripheral blood sample were analyzed from each of 26 patients, whereas two tumor tissue samples and one peripheral blood sample were analyzed from two patients. The WGS samples were sequenced 2x150 bp paired-end on an Illumina HiSeqX v2.5 instrument, and the WES samples were sequenced 2x100 bp paired-end on an Illumina HiSeq 2500 instrument. The FASTQ files generated were aligned to the human reference genome sequence GRCh38/hg38 using bwa-mem, with the ALT-aware option turned on. Sorting of reads and marking of PCR duplicates was performed with GATK. Base quality score recalibration and joint realignment of reads around insertions and deletions (indels) were conducted using GATK tools. The dataset consists of 58 files in the CRAM format (lossless compression) with a total file size of ~8,8 TB. All CRAM files but one, are derived from one sequence run and one sample. P4551_227N_P4552_112N is a CRAM file where 2 sequence runs (P4551_227N and P4552_112N) from peripheral blood samples from the same individual, P019, were aligned into one single CRAM file. Additional genomic and molecular data (FASTQ, BAM, IDAT, and VCF files) and limited clinical data can be requested by ethically approved projects conducting research in the field of pediatric cancer.

  Dataset EGAD50000000298

• Meningioma Exome

  Meningioma Exome

  Dataset EGAD00001000099

• CRLF2 sequencing project Exomes

  CRLF2 sequencing project Exomes

  Dataset EGAD00001000077

• Whole Exome and Target Sequencing Data in 75 Samples from 5 Hepatocellular Carcinoma Patients.

  Whole Exome and Target Sequencing Data in 75 Samples from 5 Hepatocellular Carcinoma Patients. The sequencing was performed by Illumina HiSeq 4000. Background and aims: Intratumoral heterogeneity (ITH) challenges identifying mutations with target therapy potential whereas circulating cell-free DNAs (cfDNAs) could reflect nearly the entire mutation spectrum in given tumors. We investigated how to minimize the limit of ITH for profiling hepatocellular carcinoma (HCC).Methods: Thirty-two multi-regional HCC samples from five patients were subjected to whole exome sequencing (WES) and targeted deep sequencing (TDS). ITH extent was measured by the average percentage of non-ubiquitous mutations (present in parts of tumor regions). Matched cfDNAs were also analyzed by WES and TDS. Profiling efficiencies of single tumor specimen and cfDNA were compared and the one better depicted mutational landscape was selected to screen therapeutic targets.Results: We found variable extents of ITH in HCCs and observed branched and parallel evolution patterns. ITH level decreased at higher sequencing depth of TDS than that measured by WES (28.1% vs 34.9%, P < 0.01) but it remained unchanged upon additional samples analyzed. TDS of single tumor specimen detected an average of 70% the total mutations in HCC. Although more mutations were detected in cfDNA under TDS than WES, an average of 47.2% total HCC mutations uncovered by cfDNA suggested tissue outperform cfDNA and the latter may serve as alternative in profiling HCC genome. Consequently, TDS of single tumor tissue in 66 patients and cfDNAs in four unresectable HCCs identified 38.6% (26/66 and 1/4) patients bearing therapeutic targets.Conclusions: TDS of single tumor specimen could largely circumvent ITH to uncover mutations indicative of target therapy in HCC.

  Dataset EGAD00001003278

• Clonal selection after gene therapy in sickle cell disease

  While gene therapy (GT) provides a potentially curative treatment option for patients with sickle cell disease (SCD), the occurrence of myeloid malignancies in clinical trials has prompted concern. To interrogate potential mechanisms underlying increased cancer risk, we used hematopoietic stem cell (HSC) clonal tracking by whole genome sequencing (WGS) to map the somatic mutation and clonal landscape of 2,592 gene modified as well as unmodified single stem and progenitor cells from six SCD patients undergoing gene therapy (7-26 years old, average 12.7× depth). Pre-GT phylogenetic trees in SCD were highly polyclonal and mutation burdens per cell were elevated in some, but not all, patients. Post-GT, no clonal expansions were identified. However, an increased frequency of driver mutations associated with myeloid neoplasms or clonal hematopoiesis (DNMT3A- and EZH2-mutated clones in particular) were seen in both genetically modified and unmodified cells suggested positive selection of mutant clones during gene therapy. This work sheds light on the mutation landscape and HSC clonal dynamics in gene therapy for SCD and highlights enhanced fitness of some HSCs harboring pre-existing driver mutations following gene therapy. Future studies should define the long-term fate of mutant clones including any contribution to expansions associated with myeloid neoplasms.

  Dataset EGAD00001010914

• Therapeutic Trial of Potassium and Acetazolamide in Andersen-Tawil Syndrome

  Andersen-Tawil syndrome (ATS) is an ion channel disorder that causes episodes of muscle weakness and potentially life-threatening heart arrhythmias. The majority of ATS cases are caused by a mutation in the KCNJ2 gene, which is linked to potassium channels in the heart, brain, and skeletal muscle; other cases are presumed to be caused by an undetermined gene lesion. To date, the treatment for ATS has been largely anecdotal, and no treatments have been formally assessed in a controlled clinical trial. This study will determine whether potassium supplements and/or acetazolamide, which is a diuretic medication, affect the duration of muscle weakness and heart rhythm abnormalities in people with ATS. Participation in this study will last about 11 months. Participants will first attend a 3-day inpatient visit that will include a medical history, physical examination, blood work, heart rhythm testing by an electrocardiogram (ECG) and Holter monitor, strength testing, a health questionnaire, and daily potassium supplementation. Participants will also track the number and length of weakness episodes that they experience while in the hospital. On the last day of the inpatient visit, participants will be provided with multiple bottles containing either potassium or placebo. Participants will then return home for an 18-week treatment period that will consist of six 3-week-long treatments of either potassium or placebo, with the treatment schedule being randomly determined. Upon completing the first 18-week treatment period, participants will attend a second 3-day inpatient visit that will include the same tests and procedures as the first. The only difference will be that participants will receive acetazolamide along with potassium. This will be followed by a second 18-week treatment period that will consist of six 3-week-long treatments of either acetazolamide or placebo. At the end of the second treatment period, participants will fill out another health questionnaire. Throughout both 18-week treatment periods, participants will phone in daily to track any muscle or heart problems. They will also provide blood samples on a weekly basis. At Weeks 2, 5, 8, 11, 14, and 17 of both treatment periods, participants will wear a Holter monitor for 24 hours and then mail it in. A final outpatient visit will occur 8 weeks after the end of the second treatment period and will include heart rhythm testing, muscle strength testing, and blood work.

  Study phs001316

• Final results and ctDNA analyses from the randomized phase 3 IMpassion031 trial evaluating peri-operative atezolizumab for early-stage triple-negative breast cancer

  In IMpassion031, adding atezolizumab to neoadjuvant chemotherapy significantly improved pathologic complete response (pCR) rate (primary endpoint) for stage II/III triple-negative breast cancer (TNBC). Patients with stage II/III TNBC were randomized 1:1 to neoadjuvant chemotherapy plus peri-operative atezolizumab or pre-operative placebo. Descriptive secondary endpoints included event-free, disease-free, and overall survival. Long-term outcomes favored the atezolizumab group (event-free survival HR 0.76, 95% CI: 0.47–1.21). Among patients without pCR, 14/70 (20%) atezolizumab-treated and 33/99 (33%) placebo-treated patients received additional adjuvant therapy, frequently capecitabine. In exploratory analyses, pCR was prognostic for long-term outcome. Patients with baseline ctDNA-negative status (6%) had excellent long-term outcomes. Most patients (87%) had cleared ctDNA at surgery. ctDNA-positive status at surgery identified a subset of non-pCR patients with poorest prognosis. Long-term safety was consistent with primary results. Adding atezolizumab to chemotherapy for stage II/III TNBC is associated with more favorable long-term outcomes. ctDNA dynamics provide prognostic value beyond pCR. ClinicalTrials.gov identifier: NCT03197935

  Study EGAS50000000974

• OpACIN-neo – A Multicenter Phase 2 Study to identify the Optimal neo-Adjuvant Combination scheme of Ipilimumab and Nivolumab – Whole exome sequencing

  The outcome of patients with macroscopic stage III melanoma is poor. Neoadjuvant treatment with ipilimumab plus nivolumab at the standard dosing schedule induced pathological responses in a high proportion of patients in two small independent early-phase trials. However, toxicity of the standard ipilimumab plus nivolumab dosing schedule was high, preventing its broader clinical use. The aim of the OpACIN-neo study (NCT02977052) to identify a dosing schedule of ipilimumab plus nivolumab that is less toxic but equally effective. 86 patients with macroscopic stage III melanoma were randomised to one of three dosing schedules: arm A, two cycles of ipilimumab 3 mg/kg plus nivolumab 1 mg/kg once every 3 weeks; arm B, two cycles of ipilimumab 1 mg/kg plus nivolumab 3 mg/kg once every 3 weeks; or arm C, two cycles of ipilimumab 3 mg/kg once every 3 weeks directly followed by two cycles of nivolumab 3mg/kg once every 2 weeks. Within the first 12 weeks, grade 3–4 immune-related adverse events were observed in 12 (40%) of 30 patients in group A, six (20%) of 30 in group B, and 13 (50%) of 26 in group C. Pathological responses occurred in 24 (80%) patients in group A, 23 (77%) in group B, and 17 (65%) in group C. The 2-year estimated relapse-free survival was 84% for all patients, 97% for patients achieving a pathologic response and 36% for nonresponders (p<0.001). High tumor mutational burden (TMB) and high interferon-gamma-related gene expression signature score (IFN-γ score) were associated with pathologic response and low risk of relapse; pathologic response rate (pRR) was 100% in patients with high IFN-γ score/high TMB, patients with high IFN-γ score/low TMB or low IFN-γ score/high TMB had pRRs of 91% and 88%; while patients with low IFN-γ score/low TMB had a pRR of only 39%. These data demonstrate long-term benefit in patients with a pathologic response and show the predictive potential of TMB and IFN-γ score.

  Study EGAS00001004832

• OpACIN-neo – A Multicenter Phase 2 Study to identify the Optimal neo-Adjuvant Combination scheme of Ipilimumab and Nivolumab – RNA sequencing

  The outcome of patients with macroscopic stage III melanoma is poor. Neoadjuvant treatment with ipilimumab plus nivolumab at the standard dosing schedule induced pathological responses in a high proportion of patients in two small independent early-phase trials. However, toxicity of the standard ipilimumab plus nivolumab dosing schedule was high, preventing its broader clinical use. The aim of the OpACIN-neo study (NCT02977052) to identify a dosing schedule of ipilimumab plus nivolumab that is less toxic but equally effective. 86 patients with macroscopic stage III melanoma were randomised to one of three dosing schedules: arm A, two cycles of ipilimumab 3 mg/kg plus nivolumab 1 mg/kg once every 3 weeks; arm B, two cycles of ipilimumab 1 mg/kg plus nivolumab 3 mg/kg once every 3 weeks; or arm C, two cycles of ipilimumab 3 mg/kg once every 3 weeks directly followed by two cycles of nivolumab 3mg/kg once every 2 weeks. Within the first 12 weeks, grade 3–4 immune-related adverse events were observed in 12 (40%) of 30 patients in group A, six (20%) of 30 in group B, and 13 (50%) of 26 in group C. Pathological responses occurred in 24 (80%) patients in group A, 23 (77%) in group B, and 17 (65%) in group C. The 2-year estimated relapse-free survival was 84% for all patients, 97% for patients achieving a pathologic response and 36% for nonresponders (p<0.001). High tumor mutational burden (TMB) and high interferon-gamma-related gene expression signature score (IFN-γ score) were associated with pathologic response and low risk of relapse; pathologic response rate (pRR) was 100% in patients with high IFN-γ score/high TMB, patients with high IFN-γ score/low TMB or low IFN-γ score/high TMB had pRRs of 91% and 88%; while patients with low IFN-γ score/low TMB had a pRR of only 39%. These data demonstrate long-term benefit in patients with a pathologic response and show the predictive potential of TMB and IFN-γ score.

  Study EGAS00001004833

• Brain Arteriovenous Malformation Genetics Study

  The Predictors of Spontaneous Cerebral AVM Hemorrhage study was designed to identify genetic markers that predict adverse outcomes in brain arteriovenous malformation (BAVM) patients, examine race-ethnicity influences on BAVM hemorrhage risk, and use genome-wide association analysis to identify novel genetic predictors of AVM and hemorrhage using the Affymetrix GeneChip. We performed the first genome-wide association study (GWAS) (PMID: 26818729) of sporadic BAVM to investigate association of common single nucleotide polymorphisms (SNPs) with risk of sporadic BAVM in the international, multicenter Genetics of Arteriovenous Malformation (GEN-AVM) consortium. The study included a Caucasian discovery cohort of 515 BAVM cases and 1,191 controls genotyped using Affymetrix genome-wide SNP arrays. The Phase 1 discovery cohort comprised of 338 BAVM cases (154 male) and 504 controls genotyped using the Affymetrix Genome-Wide Human SNP Array 6.0. The molecular data (individual genetic data) for the Phase 1 discovery cohort 338 BAVM cases (self-reported Caucasian) recruited at University of California, San Francisco, or Kaiser Permanente of Northern California (KPNC) are included in the dbGaP Study Report: Brain Arteriovenous Malformation Genetics Study, with the additional BAVM case phenotypic and sample attribute characteristics: sex, race, age at BAVM diagnosis, hemorrhagic presentation, BAVM size. Written informed consent was obtained from all participants and the study was approved by the respective Institutional Review Boards. The molecular dataset for this study in dbGaP is based on the genotypes obtained for the initial set of 906,600 SNPs for cases and controls called together using Birdseed v2 algorithm in Affymetrix Genotyping Console Software. The final QC'd dataset includes 717,335 markers as SNPs with MAF<0.01 or deviating from Hardy-Weinberg equilibrium (P<0.00001) in controls were excluded. After QC, the average genotyping call rate was 99%. The discovery phase GWAS identified 57 top BAVM-associated SNPs which were tested in a replication cohort including 608 BAVM cases and 744 controls. The molecular dataset for these 338 BAVM cases was also included in a report investigating common and rare genome-wide copy number variation for association with BAVM (PMID: 24098321), which included a total sample size of 371 sporadic Caucasian BAVM cases and 563 Caucasian controls. This study identified a CNV region on 1p36.13 (NBPF1 locus) that was significantly enriched with duplications in BAVM cases compared to controls, although this association did not replicate in an independent cohort of 184 sporadic BAVM cases and 182 controls. This resource will help researchers to better understand the genetic risk factors for BAVM and/or hemorrhage. These data may be applied to examine the genetic contributions to arteriovenous malformation, stroke, and related vascular disorders and may help generate hypotheses regarding the genetics of cerebrovascular disease.

  Study phs002069

• Multi-Modal Single-Cell, Spatial, and Genomic Analyses of Human Non-Small Cell Lung Cancer Brain Metastases

  We performed single nuclei RNA-sequencing (snRNA-seq) with matched T cell receptor sequencing (TCR-seq), and pool matched low pass whole genome sequencing (WGS) of 12 treatment-naive non-small cell lung cancer (NSCLC) primary tumors (PTs) and 31 treatment-naive NSCLC brain metastases (BMs). In total, we recovered 277,206 cell transcriptomes in 43 samples. Single-nuclei suspension from frozen NSCLC metastases were generated using methods published in Slyper et al. (Nature Medicine, 2020, PMID:32405060) with modifications. The tissue was first cut in 20 µm sections using a cryostat and stored on dry ice until the next step. Then the tissue was put on wet ice and washed once with 5 ml PBS to remove OCT. After this step nuclei were extracted by vigorous pipetting in salt-tris buffer with Tween-20 (TST) followed by a high-volume wash in salt-tris buffer (ST). Nuclei were counted and 10,000-14,000 and were loaded using a Chromium controller (10X genomics) and Chromium Single Cell 5' Reagents (V1.1 and V2). Complementary DNA libraries were generated according to manufacturer's instructions, with the modification of one additional cycle of amplification. Matched single nuclei T cell receptor (snTCR) sequencing libraries were prepared from amplified cDNA libraries using either Chromium Single Cell V(D)J Enrichment Kit for human T cells Reagent Kit, v1.1, or single cell human TCR amplification kit. Final snTCR sequencing libraries were prepared using Single Index Kit T Set A for V1.1 libraries or Library construction kit and Dual Index Kit TT set A for V2 libraries. Sequencing libraries were quantified using Tapestation D5000 reagents and a 2200 TapeStation and pooled for sequencing. Pool matched low pass whole-genome sequencing (lp-WGS) was performed in addition to snRNA-seq to improve the identification of the malignant and non-malignant cell compartments. Nuclei from snRNA-seq sample preparations were collected by centrifugation (500g, 5 min), snap frozen after removing excess ST buffer, and stored at -200C until further processing. If insufficient nuclei (0C for 9 minutes. Adapters were not diluted, and the optional PCR (5 cycles) and cleanup were performed. After library cleanup with AMPure beads (Beckman Coulter, Brea, CA), libraries were quantified with Tapestation D5000 HS tapes (Agilent) and stored at -200C until sequencing.Using multiplexed immunofluorescence in an independent cohort of treatment-naïve pairs of primary tumors and brain metastases from the same patients with NSCLC, we validated genomic and tumor-microenvironmental findings and identified a cancer cell population characterized by neural features that strongly enriched in brain metastases. This comprehensive analysis provides insights into human NSCLC brain metastasis biology and serves as an important resource for additional discovery.

  Study phs003865

• Atezolizumab Plus Personalized Neoantigen Vaccination in Patients with Urothelial Cancer: a Phase 1 Trial

  This is a phase 1 single center clinical trial evaluating a multi-peptide personalized neoantigen vaccine, PGV001, in combination with atezolizumab for patients with urothelial cancer. Patients were enrolled in either the adjuvant or metastatic setting. Those in the adjuvant setting included patients considered high risk of recurrence (pathological stage ypT2-T4 or ypN+ for those treated with prior neoadjuvant chemotherapy or pT3-T4 or pN+ for those not treated with prior neoadjuvant chemotherapy). Those in the metastatic setting included patients progressing after prior chemotherapy or those with at least stable disease after chemotherapy treated as “switch maintenance” therapy. The primary endpoint was feasibility (including the number of neoantigens identified per patient, vaccine production time, the proportion of patients consenting to the vaccine development phase for whom a vaccine product was prepared, and the proportion of patients eligible for the treatment phase who completed the priming cycle 1 of PGV001 plus atezolizumab) and safety. Secondary endpoints included the objective response rate, progression-free survival or disease-free survival, overall survival, and immunomodulatory effects of treatment. Urothelial tumor tissue and normal tissue or peripheral blood specimens were utilized for DNA sequencing. HLA typing was performed utilizing genomic DNA isolated from whole blood, and RNA sequencing was performed utilizing tumor tissue specimens. Neoantigen prediction and final selection for incorporation into the vaccine was determined by the OpenVax computational pipeline. 12 patients were initially enrolled into the vaccine development phase and 10 patients initiated the treatment phase. PGV001 was administered with poly-ICLC and tetanus peptide on subsequent days for up to 10 doses, including an initial priming cycle 1. Atezolizumab was administered every 3 weeks for up to 1 year in the adjuvant setting or 2 years in the metastatic setting. The vaccine was feasible and safe, the median vaccine production time was 20.3 weeks, and 100% of patients that initiated the treatment phase completed the priming cycle. The most common treatment-related adverse events were local grade 1 injection site reactions, fever, and fatigue. Three of the 4 patients in the adjuvant setting remained disease-free at median follow-up of 39.5 months. There was a 40% objective response rate among patients with measurable disease in the metastatic setting. The single patient treated as “switch maintenance” therapy remained with no evidence of disease at 39.4 month follow-up. All patients demonstrated on-treatment neoantigen-specific T cell reactivity, as measured by ELISPOT and flow cytometry assays. Overall, PGV001 plus atezolizumab was feasible, safe, and warrants further investigation. Whole exome sequencing from tumor tissue and normal tissue or whole blood specimens and RNA sequencing from tumor tissue specimens will be available through dbGaP.

  Study phs003922

• Constitutional and somatic genomic rearrangements coherently restructure chromosome 21 in acute lymphoblastic leukaemia

  Changes in gene dosage are a major driver of cancer1, engineered from a finite, but increasingly well annotated, repertoire of mutational mechanisms2-6. These processes operate over levels ranging from individual exons to whole chromosomes, often generating correlated copy number alterations across hundreds of linked genes. An example of the latter is the 2% of childhood acute lymphoblastic leukemia (ALL) characterized by recurrent intrachromosomal amplification of megabase regions of chromosome 21 (iAMP21)7,8 To dissect the interplay between mutational processes and selection on this scale, we used genomic, cytogenetic and transcriptional analysis, coupled with novel bioinformatic approaches, to reconstruct the evolution of iAMP21 ALL. We find that individuals born with the rare constitutional Robertsonian translocation between chromosomes 15 and 21, rob(15;21)(q10;q10)c, have ~2700-fold increased risk of developing iAMP21 ALL compared to the general population. In such cases, amplification is initiated by chromothripsis involving both sister chromatids of the dicentric Robertsonian chromosome. In contrast, sporadic iAMP21 is typically initiated by breakage-fusion-bridge (BFB) events, often followed by chromothripsis or other rearrangements. In both sporadic and iAMP21 in rob(15;21)c individuals, the final stages of amplification frequently involve large-scale duplications of the abnormal chromosome. The end-product is a derivative chromosome 21 or a derivative originating from the rob(15;21)c chromosome, der(15;21), respectively, with gene dosage optimised for leukemic potential, showing constrained copy number levels over multiple linked genes. In summary, the constitutional translocation, rob(15;21)c, predisposes to leukemia through a novel mechanism, namely a propensity to undergo chromothripsis, likely related to its dicentric nature. More generally, our data illustrate that several cancer-specific mutational processes, applied sequentially, can co-ordinate to fashion copy number profiles over large genomic scales, incrementally refining the fitness benefits of aggregated gene dosage changes.

  Dataset EGAD00001000658

• University of Illinois at Chicago (UIC) Autism Centers of Excellence (ACE) Exome Sequencing Analysis

  Autism spectrum disorder (ASD) is highly heritable with recent sibling recurrence risk estimates of 19% overall and 26% in males. The heritable phenotype of hyperserotonemia, or elevated levels of platelet serotonin (5HT), in ~35% of people with ASD is a well-established biomarker. The efficacy of drugs like risperidone and potent serotonin transporter inhibitors at treating behaviors related to Insistence on Sameness, along with evidence of the contribution of hyperserotonemia to autism susceptibility collectively support the hypothesis that dysfunction in the 5HT system is a significant etiological target for investigation of the genetic component to autism. ASD genetic architecture is complex; common variants of large effect do not contribute substantially to overall ASD risk, but there are clearly common variants with small effects and rare genetic/genomic variation of larger effect among ASD genetic risk factors. The NIH ARRA Autism Sequencing Consortium, including Dr. Sutcliffe and Dr. Cook among others, has initiated exome sequencing studies to identify more of the rare variants contributing to ASD. Data from our group and others reveal numerous rare de novo and inherited sequence mutations, and the number of de novo and other functional mutations that are found to affect molecules encoding 5HT signaling and its regulation further reinforces our hypothesis that regulation of serotonin levels is important in autism genetic susceptibility. Integrin receptor signaling pathways were prominently featured among identified de novo mutations, thus defining a set of interrelated gene networks that control 5HT signaling. We propose to extend our findings to date by conducting exome sequencing on ACE subjects, for whom we also have 5HT biochemical data, to identify rare variants (including CNVs) in 5HT-related gene networks. In total we propose to have exome sequencing completed on 523 samples. The purpose of the proposed research is two-fold: 1) we will conduct more extensive investigations to determine the support for the hypothesis that genetic variation at genes regulating platelet serotonin levels affect susceptibility to ASD and 2) we will combine the exome sequence data from these studies with sequence data for autism being accumulated in a variety of other research projects to further the goal of identifying and characterizing the genetic component to ASD.

  Study phs000712

• Jackson Heart Study (JHS) Cohort

  The Jackson Heart Study (JHS) is a large, community-based, observational study whose participants were recruited from urban and rural areas of the three counties (Hinds, Madison and Rankin) that make up the Jackson, MS metropolitan statistical area (MSA). Participants were enrolled from each of 4 recruitment pools: (1) random, 17%, (2) volunteer, 30%, (3) currently enrolled in the Atherosclerosis Risk in Communities (ARIC) Study, 31%, and (4) secondary family members, 22%. The final cohort of 5,306 participants included 6.59% of all African American Jackson MSA residents, aged 35-84, during the baseline exam (N-76,426, US Census 2000). Among these, approximately 3,600 gave consent that allows genetic research and deposition of data into dbGaP. Major components of three clinic examinations (see study history) include medical history, physical examination, blood/urine analytes, and interview questions on areas such as physical activity, stress, coping and spirituality, racism and discrimination, socioeconomic position, and access to health care. Extensive clinical phenotyping includes anthropometrics, electrocardiography, carotid ultrasound, ankle-brachial blood pressure index, echocardiography, CT chest and abdomen for coronary and aortic calcification, liver fat, and subcutaneous and visceral fat measurements, and cardiac MRI. At 12-month intervals after the baseline clinic visit (Exam 1), participants have been contacted by telephone to update information, confirm vital statistics, document interim medical events, hospitalizations and functional status, and obtain additional sociocultural information. Questions about medical events, symptoms of cardiovascular disease and functional status are repeated annually. Ongoing cohort surveillance includes abstraction of medical records and death certificates for relevant International Classification of Diseases (ICD) codes and adjudication of nonfatal events and deaths. CMS data are currently being incorporated into the dataset. Note: Regarding date variables warehoused in the Jackson Heart Study (JHS) Cohort on dbGaP, the coordinating center has developed an algorithm that will systematically review and de-identify any of the (nearly 100) date-related variable types stored in the data package. To simultaneously minimize (i) de-identifiability of the data and (ii) impact on analyses utilizing sensitive data elements, a participant-level random number was generated to avoid the necessity of sharing any potentially sensitive data. The coordinating center maintains an archived linkage of these data in their raw form and regularly reviews ad hoc requests to utilize the raw data on a project-by-project basis. The JHS Cohort is utilized in the following dbGaP substudies. To view genotypes, other molecular data, and derived variables collected in these substudies, please click on the following substudies below or in the "Substudies" section of this top-level study page phs000286 JHS Cohort. phs000402 ESP Heart GO JHS phs000498 JHS Allelic Spectrum Seq phs000499 JHS CARephs001069 MI Gen JHS phs001098 T2D GENES Exome Seq phs001356 Exome Chip phs002256 Cardiometabolic Renal Proteomics phs002369 Metabolomics Insulin Resistance

  Study phs000286