To determine the optimal concentration time factors for the fluoropyrimidines 5-fluorouracil (FU), 5-fluorouridine (FUR), and 5-fluoro-2'-deoxyuridine (FUdR) in regional chemotherapy, we tested these ...drugs against the colorectal carcinoma cell line HT 29 at various dosages and exposure times. The measure of cytotoxicity used was the degree of inhibition of colony formation in soft agar after drug treatment compared with untreated control cells. Colonies were visible after 6 days of growth in soft agar, so the initial evaluation of toxicity was done at this time. Additional colonies were found 10 and 16 days after the first evaluation, so the dishes containing the treated cells were also evaluated for this delayed growth phenomenon ("regrowth"), which we considered to be due to a cell growth inhibition effect of the drugs rather than a cytocidal effect. Exposure times of the cells to the drugs ranged from 5 min to 24 h and the doses, between 0.01 and 1000 micrograms/ml. The toxicity of FUdR was concentration-dependent, but its time dependence ceased after a relatively short exposure time. There was a cell population that was not susceptible to FUdR regardless of dose and exposure time; consequently, FUdR treatment was always accompanied by substantial regrowth of colonies. With FU and FUR, conditions could be achieved that resulted in complete cell death (no regrowth), but high concentrations and long exposure times were required with FU. With FUR, on the other hand, both cytostasis and cytotoxicity could be achieved with substantially lower doses and shorter exposure times than with FU. These results indicate that FUR has the potential to be an effective drug in chemotherapy protocols not involving systemic administration.
Diagnosis-to-Treatment Interval (DTI) remains associated with adverse clinical characteristics and outcome in newly diagnosed patients with diffuse large B-cell lymphoma treated on clinical trials.
...PURPOSE
A number of therapy combinations showing promise in nonrandomized studies in newly diagnosed diffuse large B-cell lymphoma (DLBCL) failed to demonstrate improvement over the control arm in randomized studies. Moreover, better than expected outcomes on the control (standard therapy arm) have been observed. Selection bias in clinical trials has consequences for scientific validity and applicability of study results to the general population. In an aggressive malignancy such as DLBCL, real or perceived urgency of initiation of therapy is weighed against the time required for trial consenting, screening, pathology review and biomarker assessment. The resulting exclusion of patients with rapidly progressive or symptomatic disease may lead to selection of patients with less aggressive disease enrolled on clinical trials. We recently examined the time from diagnosis to treatment initiation (DTI) and its association with clinical factors and outcome in a clinic-based observational cohort of patients with DLBCL from the US. Here we replicate those results in an independent clinical trial based cohort from Europe.
PATIENTS AND METHODS
Newly diagnosed patients with DLBCL were prospectively enrolled in the University of Iowa/Mayo Clinic SPORE Molecular Epidemiology Resource (MER) or the LYSA LNH-2003 clinical trials program. All enrolled cases from LYSA were used in analysis as intent to treat approach. Pathological diagnosis was centrally reviewed and all patients received anthracycline-based immunochemotherapy (IC) at initial diagnosis. DTI was defined as the time in days from date of first lymphoma-containing biopsy to the initiation of IC therapy. Associations of DTI with clinical factors and outcome were examined using the median DTI split (0-14 days) from the MER dataset and as using a continuous variable for DTI based on examination of functional form. Outcome was assessed using event-free survival at 24 months from diagnosis (EFS24) where events were defined as progression/relapse, initiation of new anti-lymphoma therapy, or death due to any cause.
RESULTS
990 patients were enrolled in the MER from 2002-2012 and 1446 patients were enrolled in the LYSA cohort from 2003-2009. Median (range) DTI was 15 days (0-155) days in the MER and 23 days (0-215) in LYSA. Short DTI (0-14 days) was strongly associated with adverse clinical factors including elevated LDH, poor performance status, presence of B symptoms, and higher aaIPI and IPI in both cohorts (all p<0.001) (table 1). Associations between DTI and sex (p>0.10) and DTI and age (p>0.068) were not statistically significant in both cohorts. There was no association between DTI and distance from enrollment center in the MER (p=0.99). Functional form analysis demonstrated a linear association between DTI and EFS24 in both cohorts. Longer DTI was associated with improved EFS24 in both the MER (per-week OR=0.80, 95%CI:0.74-.0.87, p<0.0001) and LYSA (OR=0.90, 95%CI:0.86-0.94, p<0.0001), and these associations remained significant after adjustment for IPI in MER (per-week OR=0.86, 95%CI:0.79-0.93, p=0.0001) and LYSA (OR=0.94, 95%CI:0.90-0.98, p=0.0053). Kaplan-Meier curves for DTI and EFS using a weekly grouping are shown in figure 1. In a combined cohort analysis, the association between longer DTI and improved EFS24 remained in a high risk (IPI 3-5) subset (OR=0.90, 95% CI: 0.85-0.96, p=0.0005). Association between DTI and EFS24 was similar in GCB (OR=0.83, 95% CI: 0.75-0.92, p=0.0004, N=505) and non-GCB (OR=0.89, 95% CI: 0.81-0.97, p=0.012, N=366) cell of origin subsets as determined via the Hans algorithm.
CONCLUSION
A short diagnosis-to-treatment interval is strongly associated with adverse clinical factors and poor outcome in newly diagnosed DLBCL. Conversely, patients with longer DTI have less aggressive clinical characteristics and better outcomes. This association has been replicated in a large cohort of patients enrolled on clinical trials. Clinical trials must take steps to avoid the bias of enrolling patients with expected good outcome due to their ability to delay treatment.
Display omitted
Ghesquières:Celgene and Mundipharma: Consultancy, Honoraria; Roche: Research Funding. Haioun:PFIZER: Consultancy, Honoraria; GILEAD: Consultancy, Honoraria; JANSSEN: Consultancy, Honoraria; Sandoz: Consultancy, Honoraria; Takeda: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Roche: Consultancy, Honoraria. Delarue:Gilead: Consultancy, Honoraria; BMS: Consultancy, Honoraria; Roche: Consultancy, Honoraria. Molina:Takeda: Other: travel support to the ASH meeting; Novartis: Honoraria; Merck Serono: Honoraria. Ansell:Bristol-Myers Squibb: Research Funding; Celldex: Research Funding; Seattle Genetics: Research Funding; Merck: Research Funding; Affimed: Research Funding. Cerhan:Janssen: Other: Scientific Advisory Board (REMICADELYM4001); Janssen: Other: Multiple Myeloma Registry Steering . Salles:morphosys: Consultancy, Honoraria; BMS: Consultancy; Servier: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; MSD: Consultancy, Honoraria; Kite: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Gilead: Consultancy, Honoraria; Celgene: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Roche: Consultancy, Honoraria, Research Funding. Witzig:Novartis: Research Funding; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Kura: Research Funding; Acerta: Research Funding. Tilly:Takeda: Consultancy, Honoraria; Immunogen: Honoraria; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Karyopharm: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Gilead: Honoraria. Nowakowski:celgene: Consultancy, Research Funding; Morphosys: Consultancy, Research Funding; Bayer: Consultancy, Research Funding; Genetech: Consultancy, Research Funding; Abbvie: Consultancy; pharmacyclics: Consultancy; Nanostring: Research Funding.
INTRODUCTION: For follicular lymphoma (FL) patients (Pts) in need of therapy, frontline treatment with bendamustine-rituximab (BR) has become a standard treatment option. The prospective randomized ...PRIMA trial demonstrated that rituximab maintenance (RM) after response to treatment with R-CHOP is safe and improves progression-free survival (PFS) vs observation (Obs). However, there remains lack of consensus regarding the application of RM after front-line BR; there are no randomized clinical trials demonstrating its safety/efficacy. Recently-presented data from the GALLIUM study comparing outcomes of FL Pts treated with either BR or B in combination with obinutuzumab (G) followed by RM or G maintenance showed a fatal adverse event (AE) rate of 4.7% and 5.9%, respectively with a median follow-up of 41 months (Hiddemann, EHA 2017). The aim of this study was to assess the efficacy and safety of RM vs. Obs after frontline BR for FL.
METHODS: We conducted a comprehensive multi-institutional retrospective analysis of outcomes of FL Pts (grade I/II & IIIA) initiated on therapy with BR between 2011-2016 followed by either RM or Obs. The decision to apply RM was based on preferences of individual treating physician and/or patient. Individual patient records were queried for baseline demographic, disease characteristics, and treatment history. Outcomes included adverse events during or after BR induction, response rates, PFS, overall survival (OS) and cause of death (COD).
RESULTS: We collected data on 640 Pts across 13 US academic medical centers. Median age was 60 years (range 21-88). 52% were male and 87% were Caucasian (N = 397 with available data). FLIPI was low (20%), intermediate (38%) and high risk (41%) (N=545), 21% had B symptoms (N = 422) and 23%, had bulky disease (N=408). 91% had grade I/II FL and 9% had grade IIIA (N=632). The median time from diagnosis to treatment was 1 month (range 0 - 135 months). With a median follow-up of 36 months, 63 Pts died (9.8%): COD was lymphoma (n=26), infection or multi-organ system failure (MOSF) (n=12), unknown (n=12) solid tumor (n=10), MDS (N=1) cardiovascular (n=1) and progressive multifocal leukoencephalopathy (n=1), representing a fatal AE rate of 2.3% or 4.1% if cases of unknown COD are excluded or included in determining fatal AE rate, respectively. Median time to death was 22 months (range 2-74 months). Among the 584 Pts who received ≥4 cycles of BR, responses to induction therapy were available in 394 cases. RM was more commonly applied in responding Pts than Obs: in the 260 Pts selected for RM, the complete and partial remission (CR, PR) rates were 71% and 28%, respectively and in the 134 Pts selected for Obs, the CR and PR rates were 58% and 30%, respectively, (P <0.001). Median number of cycles was 6 and median B dose was 90 mg/m2 and did not differ between RM and Obs Pts (P=0.14 for cycle, P=0.40 for dose). RM was administered q2 months (66%), q3 months (30%) or 4 doses q6 months (4%) for a mean of 18 months (range 3-24). The 3-year PFS for PR Pts was 80% and 44% for RM vs Obs, respectively (P=0.002). The 3-year PFS for CR Pts was 86% and 80% for RM vs. Obs, respectively (P=0.54). For Pts in PR/CR, multivariable analysis suggested that high FLIPI vs. low/intermediate was associated with worse PFS hazard ratio (HR) 2.47, 95% confidence interval (95% C.I.) 1.43-4.28), P=0.001 and OS HR 5.47 (95% CI 1.81-16.53), P=0.003. RM was associated with improvement in PFS for Pts in PR HR 0.36 (95% CI 0.18-0.71), P=0.003 but no benefit for Pts in CR HR 0.80 (95% CI 0.38-1.66), P=0.55. There was no association between use of RM and OS. Kaplan Meier estimates of PFS and OS (Figure) are depicted based on response (CR or PR) to BR induction.
CONCLUSIONS: In this large outcomes analysis of FL Pts treated with BR followed by RM, the fatal AE rate was comparable to that reported in GALLIUM when accounting for the real world nature of this unselected Pt cohort. RM was associated with a significant improvement in PFS vs. Obs in Pts who achieved PR after induction but no observed benefit for Pts in CR. In addition, we identified that the FLIPI was strongly prognostic for Pt survival for responding patients. Within the limitations inherent to retrospective analysis, these data suggest that FL Pts with partial response to BR induction therapy experience prolongation of PFS with RM, but consistent with the findings in the PRIMA study, there was no obvious impact on OS.
Display omitted
Nastoupil:Genentech: Honoraria, Research Funding; Abbvie: Honoraria, Research Funding; Celgene: Honoraria, Research Funding; Gilead: Honoraria; Janssen: Honoraria, Research Funding; TG Therapeutics: Honoraria, Research Funding; Karus Therapeutics: Research Funding. Cerhan:Janssen: Other: Scientific Advisory Board (REMICADELYM4001); Janssen: Other: Multiple Myeloma Registry Steering . Smith:Dohme Corp: Research Funding; Acerta: Research Funding; Portola Pharmaceuticals: Research Funding; Seattle Genetics: Research Funding; Sharp: Research Funding; Janssen: Research Funding; Genentech: Research Funding; Pharmacyclics LLC, an AbbVie Company: Research Funding; Merck: Research Funding. Lossos:Affimed: Research Funding. Portell:Roche/Genentech: Research Funding; Acerta: Research Funding; TG-Therapeutics: Research Funding; Infinity: Research Funding; AbbVie: Research Funding. Calzada:Seattle Genetics: Research Funding. Cohen:Bristol Myers Squibb: Research Funding; LAM Therapeutics, Inc: Research Funding; Bioinvent: Consultancy, Membership on an entity's Board of Directors or advisory committees; Abbvie: Consultancy, Membership on an entity's Board of Directors or advisory committees; Infinity: Consultancy, Membership on an entity's Board of Directors or advisory committees; Takada: Research Funding; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees. Ghosh:Gilead: Consultancy, Honoraria, Speakers Bureau; AbbVie: Consultancy, Honoraria, Speakers Bureau; TG Therapeutics: Consultancy, Honoraria, Research Funding; Kite Pharma: Consultancy, Honoraria; Celgene: Consultancy, Honoraria, Research Funding, Speakers Bureau; Seattle Genetics: Consultancy, Honoraria, Research Funding; Bristol Myers Squibb: Consultancy, Honoraria, Research Funding; Pharmacyclics: Consultancy, Honoraria, Research Funding, Speakers Bureau; Jassen: Consultancy, Honoraria, Research Funding. Caimi:Incyte: Equity Ownership; Seattle Genetics: Equity Ownership; Abbvie: Equity Ownership; Celgene: Speakers Bureau. Martin:Gilead: Consultancy, Other: travel expenses; Novartis: Consultancy; Janssen: Consultancy, Honoraria, Other: travel expenses; Celgene: Consultancy; Teva: Research Funding; Genentech: Consultancy. Evens:Kite Pharma: Consultancy; Celgene: Consultancy; Merck: Consultancy; Pharmacyclics: Consultancy; Amgen: Consultancy; Millennium: Consultancy; Novartis: Consultancy; Affimed: Consultancy; AbbVie: Consultancy; Seattle Genetics: Consultancy; • Spectrum Pharmaceuticals: Consultancy. Kahl:ADC Therapeutics: Research Funding; Seattle Genetics: Consultancy; Celgene: Consultancy; Gilead: Consultancy; Genentech: Consultancy.
Waldenström macroglobulinemia (WM) is a unique subset of lymphoplasmacytic lymphoma (LPL) that is defined by the presence of an LPL infiltrate in the bone marrow together with a monoclonal IgM ...protein in the serum. Family history is strongly associated with risk of WM, and the estimated heritability is high; however, the genetic basis for WM predisposition remains unknown. To identify susceptibility loci for WM, we conducted a two-stage genome-wide association study (GWAS) in over 500 WM/LPL cases and 4300 controls of European ancestry. In the discovery stage (stage 1), 217 unrelated WM cases (40% familial) and 3,798 controls were genotyped on the Illumina Omni Express or Illumina Omni2.5 platforms. After quality control filtering, the data were imputed using the Haplotype Reference Consortium panel and analyzed using logistic regression. Eleven promising loci (P <5.0 x 10-7) were selected for replication in 313 WM/LPL cases and 564 controls. Two novel loci were convincingly replicated in stage 2 (P<1.2 x 10-5) and reached genome-wide significance in the combined analysis of both stages: 6p25.3 (EXOC2, OR=21.14, P=1.36 x 10-54) and 14q32.13 (intergenic near TCL1, OR=4.90, P=8.74 x 10-19). A possible second independent signal was observed at 14q32.13 but did not reach genome-wide significance (P=4.66 x 10-7). When the analysis was limited to WM, results were slightly stronger but generally similar to those for WM/LPL combined. Dysregulated TCL1 expression in B cells enhances cell proliferation and survival, leading to cell transformation and mature B-cell tumors, and TCL1 has been shown to be aberrantly expressed in WM. EXOC2 interacts with Ral proteins to mediate oncogenic RAS signaling that is critical for cancer cell survival and proliferation. We found that cells transduced with the EXOC2 variant allele reporter pCS-EGFP-3'G showed significantly increased EGFP fluorescence compared to the wildtype, suggesting the variant is associated with increased EXOC2 protein levels. Cells harboring the EXOC2 variant allele were also observed to have significantly increased cell proliferation compared to wildtype cells. We further discovered that the E XOC2 variant abrogates a miRNA binding site, possibly contributing to gene expression changes for NF-κB pathway constituents. Together these newly discovered loci explain ~4% of the familial risk and suggest possible underlying biological mechanisms, significantly increasing our understanding of the genetic susceptibility of WM/LPL.
Cerhan:Janssen: Other: Scientific Advisory Board (REMICADELYM4001); Janssen: Other: Multiple Myeloma Registry Steering .
Introduction: Follicular lymphoma (FL) is the most common indolent non-Hodgkin lymphoma in the world and has a median age at diagnosis in the seventh decade. FL in young adults (YA; 40 years old or ...younger) is extremely rare. Currently, there are no standard approaches guiding treatment of YA patients with FL, and very little is known about disease characteristics and outcomes of YA patients with FL given limited research conducted in this vulnerable population.
To gain further insight into FL in YA, we analyzed the National LymphoCare Study (NLCS) to describe disease and patient characteristics, as well as features of treatment in YA patients with FL. We previously reported that 2-year progression-free survival (PFS) is an important survival endpoint in patients with FL undergoing chemo-immunotherapy. Hence, we also sought to characterize 2-year PFS in this age group and compare it to older cohorts.
Methods: Evaluablepatients were identified in the NLCS, and those between 18–40 years of age with newly diagnosed FL at any stage were classified as YA patients. Patients with mixed histology or transformed disease were excluded, as were patients with progression of disease prior to beginning first-line treatment. Survival probability was estimated by the Kaplan-Meier method. We estimated the association of age group with PFS using hazard ratios (HR) and 95% confidence intervals (CI) from multivariable Cox models.
Results: A total of164 YA patients with FL were analyzed, representing 6.2% of the NLCS population, similar to the observed frequency in the Surveillance, Epidemiology, and End Results (SEER) Program data (4.8% of all FL). Sixty nine percent of YA patients had advanced stage disease. The majority of patients (80%) had low-grade histology, and 50% had good risk disease according to the Follicular Lymphoma International Prognostic Index (FLIPI). Nineteen percent of patients (31/164) underwent watchful waiting, 12% received rituximab monotherapy, and 47% received chemo-immunotherapy (61% of whom received R-CHOP rituximab, doxorubicin, vincristine, prednisone). There was no significant difference in FLIPI score or other baseline disease characteristics compared to adult patients aged 41–60 years.
Eleven deaths occurred among YA with FL; only 5 of these were lymphoma related. Overall survival (OS) at 2 years was 97.4% (95% CI 93.3%, 99.0%), and at 5 years, 93.7% (88.3%, 96.7%), which was similar to patients aged 41–60 (97.2% 96.0%, 98.0% at 2 years, and 92.0% 90.1%, 93.5% at 5 years). After a median follow-up of 7.1 years, OS in YA FL was 92%. Through follow-up, there were 64 PFS events. The estimated 2-year PFS (95% CI) for YA and adults 41–60 was 75.9% (67.1%, 82.6%) and 80.9% (78.1%, 83.4%), respectively. After adjusting for FLIPI score, there was no difference in PFS for YA with FL requiring first-line treatment (excluding watchful waiting) compared to adults aged 41–60 years (HR=0.93; 95% CI 0.69, 1.25), and no difference in OS compared to adults aged 41–60 years (HR=1.19; 95% CI 0.64, 2.23).
Conclusions: In the largest cohort of YA patients with FL to date, we found few differences in outcomes compared to patients aged 41–60. FLIPI and other disease characteristics were similar to adults aged 41–60 years. There were no differences between YA FL and adults aged 41–60 in PFS for all treated patients. OS in the YA group of patients with FL was outstanding. YA patients with FL have reassuringly similar outcomes to patients aged 41–60. Fertility preservation and survivorship issues should be taken into consideration when defining management strategies, but otherwise these data support that YA patients with FL should not be approached differently from older adults with the same disease.
Byrtek:Genentech, Inc.: Employment, Equity Ownership. Dawson:Genentech, Inc.: Employment, Equity Ownership. Zhou:RTI-HS: Employee of RTI-HS, which has research contracts with Genentech Other. Flowers:Seattle Genetics: Consultancy; Spectrum: Consultancy, Research Funding; Sanofi: Research Funding; Abbott: Research Funding; Novartis: Research Funding; OptumRx: Consultancy; Millennium/Takeda: Research Funding; Janssen: Research Funding; Celgene: Research Funding; Allos: Consultancy. Farber:Gilead: Speakers Bureau; Janssen/Pharmacyclics: Speakers Bureau; Seattle Genetics: Speakers Bureau; Leukemia Lymphoma Society NJ Chapter: Membership on an entity’s Board of Directors or advisory committees; Genentech: Membership on an entity’s Board of Directors or advisory committees, Speakers Bureau; Celgene: Membership on an entity’s Board of Directors or advisory committees, Speakers Bureau; Alexion: Speakers Bureau, Stock ownership Other. Cerhan:Genentech, Inc.: LymphoCare Scientific Advisory Board Other. Link:Genentech, Inc.: Consultancy, Scientific Advisory Board for Genentech Other.
Performance status (PS) is a measurement of a patient's functional capabilities and has been validated as a prognostic indicator in patients with cancer. This study was conducted to compare patient ...and physician-rated Eastern Cooperative Oncology Group (ECOG) PS to determine the incidence of inter-observer variability and identify determinants of PS disagreement among a cohort of patients with hematologic malignancy.
Newly diagnosed patients with leukemia and lymphoma were prospectively enrolled in the University of Iowa/Mayo Clinic SPORE Molecular Epidemiology Resource (MER) from 2002-2008. At diagnosis, patients and their physicians were asked to independently rate ECOG PS. Those who reported PS within 1 month of diagnosis were included in this analysis. Chi-squared and Wilcoxon rank-sum tests were used to assess the association between PS disagreement and prognostic or demographic factors; Cox proportional hazards models and c-statistics were used to evaluate the association between PS and overall survival.
1269 patients were included with a median age of 61 years (range 18-91). Of these, 58% were male; 275 had chronic lymphocytic leukemia (CLL), 127 had Hodgkin lymphoma (HL), and 867 had non-Hodgkin lymphoma (NHL). Overall, 829 (65%) patients and physicians rated PS the same. Age greater than 60 was a significant predictor of disagreement overall (p<0.001), as well as in CLL (p=0.014) and NHL (p=0.004), but not in HL (p=0.65). We found that across disease subtypes, the level of disagreement increased with more aggressive disease. This finding was significant in those with HL (p=0.027) and NHL (p<0.0001) as International Prognostic Score (IPS) and International Prognostic Index (IPI) increased respectively. In CLL, the trend for Rai stage was suggestive but not statistically significant (p=0.27). The incidence of disagreement was highest among patients with NHL (37%), with a higher percentage of disagreement among those with aggressive (45%) vs. non-aggressive (25%) subtypes (p<0.001). Disagreement was lower in patients with CLL (27%) and HL (33%). There were no significant associations between patient gender or education level and PS rating among subtypes or overall.
Patient and physician-rated PS were both significant predictors of overall survival in univariate models and also adjusted for subtype and subtype specific risk score. The prognostic ability for PS was similar for both patient rated and physician-rated assessment overall (c-statistic=0.76 for both patient and physician rated PS) and in those with HL (c-statistic=0.85 for both patient and physician rated PS) and NHL (c-statistic=0.76 for both patient and physician rated PS). However, patient-rated PS was better for prognostication in CLL (c-statistic=0.75; p<0.0001), compared to physician-rated PS (c-statistic=0.67; p=0.002).
Patients with hematological malignancies and their physicians do not always rate PS the same, particularly in patients who are older or have more advanced or aggressive disease. These findings suggest the need for physicians to communicate with patients when determining PS, as PS is a strong predictor of survival.
No relevant conflicts of interest to declare.
KASCADE-Grande is an air-shower observatory devoted for the detection of cosmic rays with energies in the interval of 10 super(14) - 10 super(18) eV, where the Grande array is responsible for the ...higher energy range. The experiment comprises different detection systems which allow precise measurements of the charged, electron and muon numbers of extensive air-showers (EAS). These data is employed not only to reconstruct the properties of the primary cosmic-ray particle but also to test hadronic interaction models at high energies. In this contribution, predictions of the muon content of EAS from QGSJET II-2, SIBYLL 2.1 and EPOS 1.99 are confronted with the experimental measurements performed with the KASCADE-Grande experiment in order to test the validity of these hadronic models commonly used in EAS simulations.
Tumors arise and evolve by iterative steps of mutation, subclonal selection, and clonal expansion due to growth advantage of the fittest subclones and external mutation induction and selection ...pressure from radiation and drug therapies. Various studies have shown that greater clonal complexities in primary tumors are correlated with poor clinical outcome of tumor progression and/or drug resistance. Recently, we have reported that very few of the driver mutations, all of which were clonal, detected in primary diffuse large B-cell lymphoma (DLBCL) were associated with DLBCL outcome as measured by 24-month event-free survival (EFS24). In the current project, we identified and studied subclonal mutations in primary DLBCL tumors and assessed their associations with EFS24, as well as the role of activation-induced cytidine deaminase (AID) in genomic mutations occurred in DLBCL tumors.
The detection of subclonal mutations is still a significant bioinformatics challenge. In addition, identification of clonal mutations in samples with lower tumor purities faces similar challenges due to the low concentration of reads supporting the mutant alleles. The current methods of somatic mutation calling are not sufficiently sensitive to identify the low concentration mutations in tumor DNA sequencing data. We implemented a bioinformatics workflow for low-concentration and subclonal mutation detection which is based on the positional read pile-up data and a back-fill approach (PUB). The pile-up files were first generated using the re-aligned and re-calibrated BAM files after read alignment using Burrows-Wheeler Aligner (BWA). The variant positions with alternative bases were annotated by attributes defined in the Variant Quality Score Recalibration (VQSR). PUB then used a boosting method and a generalized linear model (GLM) to train a model of 'good quality' variants using common variants from HapMap, and prioritized and called clonal and subclonal variants based on the trained model. The VQSR attributes related to alternative allele depth were less-weighed in order to call subclonal mutations. The somatic mutations were then identified by Fisher's Exact Test using sequencing depths of the reference and alternative alleles from paired tumor and germline sequencing data.
The exome sequencing data of paired tumor and peripheral blood from 48 newly diagnosed DLBCL patients were analyzed using PUB. Thirty-six of the 48 patients achieved EFS24 with the other 12 patients experienced primary treatment failure. Most of the tumors studied had tumor purities between 40-70%. PUB identified substantially higher number of somatic mutations, both clonal and subclonal, compared to those detected using existing somatic callers. We observed that the prevalence of mutations in previously reported driver genes were higher using thresholds of mutation concentration ≥ 5% and Fisher's Exact Test p ≤ 0.05, including EZH2 (mutated in 26% of DLBCL tumors analyzed by PUB, compared to 12.7% as previously reported), MLL2 (72% vs. 31%), CD79B (22% vs. 14%), TNFRSF14 (36% vs. 20%), MEF2B (22% vs. 16.4%), CARD11 (46% vs. 21.8%), and MYD88 (18% vs. 11%). In addition, other genes involved tumorigenesis that have not been previously linked to DLBCL also harbored both clonal and subclonal mutations with substantial prevalence, including FGFR3 (12%), KIT (24%), and ATM (28%). Furthermore, association of genes displaying clonal and subclonal mutations with EFS24 identified potential biomarkers for DLBCL outcome. Among these, two genes EPGN and ASTE are involved in epithelial growth factor receptor signaling and had association p values of 0.0001 and 0.0016, respectively. The oncogene MAFB was also associated with EFS24 (P = 0.0016).
We searched for AID induced mutations among all identified variant positions and concluded that there was no evidence of AID site enrichments compared to a simulated data set.
In summary, we developed and applied a sensitive bioinformatics pipeline for the identification of both clonal and low concentration somatic mutations in primary DLBCL exome sequencing data which further revealed the clonal complexity of the primary DLBCL tumors. Several of the genes were identified as potential biomarkers for DLBCL outcome.
Maurer:Kite Pharma: Research Funding. Ansell:Bristol-Myers Squibb: Research Funding; Celldex: Research Funding. Link:Genentech: Consultancy, Research Funding; Kite Pharma: Research Funding. Cerhan:Kite Pharma: Research Funding.
Background: Peripheral T-cell lymphomas (PTCLs) comprise a heterogeneous group of relatively uncommon and generally aggressive non-Hodgkin lymphomas. Recent studies have begun to reveal their ...mutational landscape, such as the recurrent mutations affecting RHOA, IDH2, and TET2 in angioimmunoblastic T-cell lymphomas (AITLs) and related PTCLs. Here, we studied the exomes of 63 PTCLs to address several unanswered questions: (1) How frequently do PTCLs have mutations with a strong rationale for being actionable using clinically available targeted therapies? (2) What pathways are enriched in PTCLs that could point to additional targeted therapies? (3) What additional recurrent mutations exist that may provide insight into PTCL pathogenesis and classification?
Methods: We performed exome capture and sequencing in 63 frozen PTCL samples (with matched germline controls from 22), including 13 PTCLs, not otherwise specified; 6 AITLs; 37 anaplastic large cell lymphomas (ALCLs: 12 ALK+, 19 ALK-, and 6 cutaneous); 4 extranodal NK/T-cell lymphomas; 2 cutaneous T-cell lymphomas, and 1 enteropathy associated T-cell lymphoma. Following primary variant calling, data were filtered by removing variants that were: outside the capture region, polymorphic (0.1% in ExAC), seen in paired germline or 50 healthy controls, within repetitive elements, or synonymous. Potentially actionable variants required published evidence for altering protein function and demonstrated in vivo or in vitro sensitivity to an available targeted agent (including open clinical trials). Mutated cancer genes (COSMIC) were examined for enrichment of KEGG canonical signaling pathways. A recurrent mutation encoding MSC E116K was validated in tumor DNA and MSC protein was examined by immunohistochemistry.
Results: Of 25 unique variants with published evidence suggesting actionability (including ErbB, JAK/STAT, and RAS genes), at least one variant was present in 16/63 patients (25%). Variants in any COSMIC gene were seen in 55/63 samples (87%). Top genes were STAT3, RHOA, ARID1A, MLL2 (KMT2D), PTPRB, TP53, and TET2. The KEGG ErbB pathway was most significantly enriched for mutated genes in our dataset (EGFR, ERBB2, ERBB3, ERBB4, and others; q=3.88x10-3), followed by neurotrophin and mitogen-activated protein kinase signaling pathways. Among 25 recurrent non-synonymous variants, we identified MSC E116K with predicted gain-of-function exclusively in 3 ALK-ALCLs and 1 cutaneous ALCL. All mutated cases strongly expressed MSC protein. Among 96 cases, 13/18 ALK-ALCLs (72%) were positive for MSC, compared to 9/78 (12%) from other PTCL subtypes.
Conclusions: Mutations that were potentially actionable based on published variant-specific data were identified in 25% of PTCLs, indicating promise for the role of sequencing in individualizing therapy. Additionally, remaining variants were enriched for several targetable pathways, most significantly ErbB. A novel recurrent mutation, MSC E116K, was seen exclusively in ALK-ALCLs (systemic and cutaneous). MSC protein, a repressor of E2A-mediated transcription of targets such as MYC, was expressed in most ALK- ALCLs and generally was absent in other PTCL subtypes. This finding extends evidence for a genetic relationship between systemic and cutaneous ALCLs and suggests MSC may help distinguish ALK- ALCLs from other PTCLs.
Link:Genentech: Consultancy, Research Funding; Kite Pharma: Research Funding.
PDF
