Presence Of TP53 Mutations Research Articles

Genetic profiling-based prognostic prediction of patients with advanced small-cell lung cancer in large scale analysis

ObjectivesComprehensive genomic analysis of small-cell lung cancer (SCLC) revealed various genetic alterations. However, obtaining suitable samples for genetic analysis is difficult in advanced SCLC. Thus, the prognostic effect of genetic alterations on the outcome of SCLC patients has not been well investigated. Therefore, this study evaluated the effect of genetic alterations on the survival of SCLC patients. Materials and methodsWe collected samples obtained from 220 patients with advanced SCLC before cancer treatment. Genomic DNA extracted from the samples was subjected to a 1.499 Mb-sized custom panel that captured all exons of 244 cancer-related genes, and the captured DNA was analyzed through next-generation sequencing. The associations between genetic alterations and overall survival were evaluated. ResultsGenetic analysis was successful in 204 samples (93%). Genetic alterations in the PI3K/AKT/mTOR pathway and inactivating mutations inTP53 and RB1 were detected in 14 (7%), 150 (74%), and 85 (42%) of the tumors. In extensive disease (ED, N = 126) patients, multivariate analysis revealed that the presence of genetic alterations in the PI3K/AKT/mTOR pathway was significantly associated with unfavorable survival [hazard ratio (HR), 2.14; 95% CI 1.02–4.06; P = 0.04]. In limited disease (LD, N = 78) patients, the presence of TP53 mutation and the absence of RB1 mutation were significantly associated with unfavorable survival (HR, 2.41; 95% CI 1.21–5.34; P = 0.01, and HR, 0.45; 95% CI 0.25–0.79; P < 0.01, respectively). ConclusionsSequencing-based genetic profiling is feasible and useful to predict the prognosis in advanced SCLC. Genetic alterations in the PI3K/AKT/mTOR pathway, TP53 mutations and RB1 mutations were associated with prognosis in SCLC patients. The genetic alterations associated with the prognosis were different between ED-SCLC and LD-SCLC.

Influence of TP53 Mutation on Survival in Patients With Advanced EGFR-Mutant Non-Small-Cell Lung Cancer.

TP53 mutation (MT) in epidermal growth factor receptor (EGFR) -MT non-small cell lung cancer (NSCLC) is associated with poor response to targeted therapy; however, its impact on survival is not clearly established. We performed an analysis of patients with stage IV EGFR MT NSCLC with available gene sequencing data. Associations between baseline characteristics; molecular profile, including TP53 MT; and survival outcomes were assessed. We identified 131 consecutive patients with EGFR MT; 81 (62%) had a TP53 MT, and 55 (42%) had other coexisting oncogenic MTs. Emergent EGFR T790M MT was observed in 42 patients (32%). Overall survival (OS) was longer for younger patients (P = .003), never smokers (P = .002), those with Eastern Cooperative Oncology Group performance status 0 to 1 (P = .004), and emergent T790M MT (P = .018). TP53 MT (P = .021) and other coexisting oncogenic MTs (P = 0.011) were associated with inferior OS. In a multivariable regression analysis adjusted for age, smoking, Eastern Cooperative Oncology Group performance status, and the presence of TP53 MT (P = .063) and other coexisting MTs (P = .064) did not achieve statistical significance. Patients with EGFR T790M/TP53 double MT had worse OS compared with patients with T790M MT alone (46.4 months v 82.9 months). In our series, five patients transformed to small-cell lung cancer (5.6%). All had TP53 MT. In four patients, allelic fraction of TP53 MT increased at the time of transformation. The presence of TP53 and other coexisting MTs in EGFR MT NSCLC were associated with inferior OS, including patients with emergent T790M MT. An increase in TP53 mutation allelic fraction may potentially be a useful clinical predictor of small-cell transformation.

Abstract 937: Longitudinal circulating-tumor DNA profiling of EGFR-mutated non-small cell lung cancer patients treated with EGFR-tyrosine kinase inhibitors

Abstract Background: In EGFR-mutant non-small cell lung cancer (NSCLC), progression disease (PD) under 1st-generation EGFR tyrosine kinase inhibitors (EGFR-TKIs) is driven by the EGFR T790M mutation in about 50% of cases. This mutation is targeted with osimertinib, a specific and potent inhibitor that also showed superior efficacy to that of 1st-generation EGFR-TKIs in the 1st-line treatment EGFR+ NSCLC. Strategy: We performed a longitudinal circulating-tumor DNA (ctDNA) analysis to evaluate 1) the dynamics of sensitizing or resistant mutations over time and 2) the genomic alterations associated with EGFR-TKI resistance, in NSCLC patients treated with either 1st-generation EGFR-TKIs (n=24) or with osimertinib (n=20). ctDNA sequencing was performed using InVisionSeq™, an amplicon-based NGS (36-gene panel) assay. Results: Sensitivity was 80% and 100% for the detection of EGFR-activating mutations and the EGFR T790M mutation, respectively, at PD in ctDNA vs tissue biopsies. The type of progression appears to influence the detection of EGFR mutations in plasma. Patients (pts) with exclusively brain or thoracic PD had significantly lower allelic fractions (AFs) of EGFR-activating mutations than pts with systemic PD (P=0.006). AFs of both EGFR-activating and EGFR T790M mutations were highest in patients with progressive metastases in liver or bone. High cell-free DNA load was associated with the presence of TP53 mutations regardless of the time under treatment (n=248 samples). The total number of mutations detected in ctDNA was higher in pts treated with osimertinib vs. 1st-generation EGFR-TKIs (P=0.004), suggesting a potential increase of tumor heterogeneity over time. We observed that the AFs of mutant ctDNA were correlated with response to treatment. Complete clearance of EGFR T790M (AF detection limit: 0.01%) was observed in pts who presented a partial response to osimertinib (n=17), of these, EGFR-activating mutations were found in 4/17 pts (AF:0.05-1.03%). PD was evidenced in 36/44 pts. Emergence of mechanisms of resistance, before the confirmation of clinical PD, was evidenced in 5 and 3 pts under erlotinib and osimertinib, respectively. PD to 1st-generation EGFR-TKIs was mainly driven by EGFR T790M (86%), followed by mutations in PIK3CA or PTEN. We detected the emergence of heterogeneous potential mechanisms of resistance to osimertinib in 3/9 cases, including mutations in EGFR, KRAS, BRAF as well as HER2 and MET amplification. We did not identify genomic drivers of resistance in 16/36 pts, despite evidence of clinical PD. These pts had either exclusive brain or thoracic progression. ctDNA WES will be performed in these samples. Conclusion: Our results suggest that ctDNA sequencing might be a complementary, noninvasive tool to monitor response to treatment and heterogeneous mechanisms of resistance in NSCLC pts treated with EGFR inhibitors. Citation Format: Sandra Ortiz-Cuaran, Aurélie Swalduz, Camille Léonce, Solène Marteau, Séverine Martinez, Gilles Clapisson, Virginie Avrillon, Luc Odier, Lionel Falchero, Pierre Fournel, Emma Green, Clive Morris, Maurice Pérol, Pierre Saintingy. Longitudinal circulating-tumor DNA profiling of EGFR-mutated non-small cell lung cancer patients treated with EGFR-tyrosine kinase inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 937.

Methylation of O6-methylguanine DNA methyltransferase promoter is a predictive biomarker in Chinese melanoma patients treated with alkylating agents

Background: The O6-methylguanine-DNA methyltransferase gene ( MGMT ) promoter methylation status can be used to predict the prognosis of patients with various cancers following treatment with alkylating agents. Moreover, MGMT promoter methylation often coexists with TP53 gene mutation. However, MGMT has not been identified as a biomarker of melanoma. Therefore, this study systematically analyzed the prognostic role of MGMT and the correlation between MGMT methylation and TP53 mutation in non-Caucasian patients with melanoma. Methods: This study involved tumor samples and clinical data collected from 205 melanoma patients treated with alkylating agents at Peking University Cancer Hospital & Institute. The MGMT promoter methylation and TP53 mutation status were analyzed respectively using methylation-specific polymerase chain reaction and polymerase chain reaction followed by Sanger sequencing. Additionally, MGMT protein expression in tumor samples was assessed via immunohistochemistry. Results: MGMT promoter methylation was detected in 97 (47%) of the 205 tumor samples, and was significantly associated with a loss of MGMT protein expression (P=0.021). MGMT promoter methylation was also significantly associated with the presence of TP53 mutation (P=0.004). Regarding prognosis, patients without MGMT promoter methylation exhibited worse overall survival outcomes, compared to those with methylation (hazard ratio: 1.443; 95% confidence interval: 0.731–2.342; P=0.015). Conclusions: MGMT promoter methylation appears to coexist frequently with TP53 mutation. Patients harboring MGMT promoter methylation within tumors may benefit from therapy with alkylating agents.

Прогностическое значение молекулярно-генетических и клинических характеристик медуллобластом группы SHH

SHH group of medulloblastoma is a heterogeneous tumor cohort. The neoplasms differ by biological characteristics as well as clinical features and prognosis of the disease. Purpose of the study is the analysis of clinical, molecular and genetic features for prognosis defining in patients with SHH group medulloblastoma. 28 patients with SHH group medulloblastomas were included in the study. The MB molecular group verification was performed in parallel by Nanostring gene expression profiling and immunohistochemical assessment of tumor samples. The detection of TP53 gene mutations was carried out with Sanger sequencing. The prognostic impact of the clinical, molecular and genetic factors of the disease was analyzed by calculating 5-years event-free survival (EFS). The median of follow up time achieved 38.9 months. All patients harboring the TP53 mutation (n = 3) had dismal outcome (two patients died from the progression of the disease, one patient has secondary tumor). Children from older age group (&gt; 3 years) had more adverse events comparing to younger children (&lt; 3 years): EFS 49.2 ± 31.3% vs. 78.8 ± 13.9%. The relapse of the disease occurred in 7 patients (25%). Notably, that in younger children the second line treatment was effective. The presence of TP53 mutations as well as age above 3 years are associated with poor prognosis in SHH group medulloblastomas. The novel molecular and genetics markers are needed for precise prognosis defining.

The Driver Mutational Landscape of Ovarian Squamous Cell Carcinomas Arising in Mature Cystic Teratoma.

Purpose: We sought to identify the genomic abnormalities in squamous cell carcinomas (SCC) arising in ovarian mature cystic teratoma (MCT), a rare gynecological malignancy of poor prognosis.Experimental design: We performed copy number, mutational state, and zygosity analysis of 151 genes in SCC arising in MCT (n = 25) using next-generation sequencing. The presence of high-/intermediate-risk HPV genotypes was assessed by quantitative PCR. Genomic events were correlated with clinical features and outcome.Results: MCT had a low mutation burden with a mean of only one mutation per case. Zygosity analyses of MCT indicated four separate patterns, suggesting that MCT can arise from errors at various stages of oogenesis. A total of 244 abnormalities were identified in 79 genes in MCT-associated SCC, and the overall mutational burden was high (mean 10.2 mutations per megabase). No SCC was positive for HPV. The most frequently altered genes in SCC were TP53 (20/25 cases, 80%), PIK3CA (13/25 cases, 52%), and CDKN2A (11/25 cases, 44%). Mutation in TP53 was associated with improved overall survival. In 8 of 20 cases with TP53 mutations, two or more variants were identified, which were bi-allelic.Conclusions: Ovarian SCC arising in MCT has a high mutational burden, with TP53 mutation the most common abnormality. The presence of TP53 mutation is a good prognostic factor. SCC arising in MCT share similar mutation profiles to other SCC. Given their rarity, they should be included in basket studies that recruit patients with SCC of other organs. Clin Cancer Res; 23(24); 7633-40. ©2017 AACR.

Phase II Study of CPX-351 (Cytarabine: Daunorubicin) Liposome Injection in Patients (Pts) with Newly Diagnosed Acute Myeloid Leukemia (AML) at High Risk for Induction Mortality

Phase II Study of CPX-351 (Cytarabine: Daunorubicin) Liposome Injection in Patients (Pts) with Newly Diagnosed Acute Myeloid Leukemia (AML) at High Risk for Induction Mortality

The prognostic value of TP53 mutations in hypopharyngeal squamous cell carcinoma

BackgroundTP53 is the most frequently mutated gene in human cancers. Previous studies reported that TP53 mutations correlated with poor prognoses in patients with head and neck squamous cell carcinoma (HNSCC). However, the relationship between TP53 mutations and hypopharyngeal squamous cell carcinoma (HPSCC) is not known. The current study aimed to evaluate TP53 mutation status as a predictive biomarker in patients with HPSCC.MethodsWe retrospectively reviewed the clinical charts of 57 HPSCC patients treated with initial surgery between 2008 and 2014. TP53 mutation status was determined by Sanger sequencing, and patients were classified into wild-type, missense mutation, and truncating mutation groups. Additionally, p53 expression was determined using immunohistochemistry in surgical specimens.ResultsTP53 mutations were identified in 39 (68%) patients. The 3-year disease-specific survival (DSS) rate of wild-type, missense mutation, and truncating mutation group were 94%, 61%, and 43%, respectively. The TP53 mutation group displayed significantly worse DSS and overall survival rates than the wild-type group (P = 0.01 and P = 0.007, respectively). Multivariate analyses revealed that the presence of TP53 mutations and ≥4 metastatic lymph nodes were independent adverse prognostic factors for HPSCC. p53 immunopositivity was detected in 22 patients, including 5 (28%) and 17 (71%) patients in the wild-type and missense mutation groups, whereas none of the patients with truncating mutation exhibited p53 immunopositivity (P = 0.0001).ConclusionThe TP53 mutation status correlated with poor prognosis in surgically treated HPSCC patients. Specifically, truncating mutations which were not detected by p53 immunohistochemistry were predictive of worst survival.

Favorable impact of allogeneic stem cell transplantation in patients with therapy-related myelodysplasia regardless of TP53 mutational status.

Therapy-related myelodysplastic syndrome is a long-term complication of cancer treatment in patients receiving cytotoxic therapy, characterized by high-risk genetics and poor outcomes. Allogeneic hematopoietic cell transplantation is the only potential cure for this disease, but the prognostic impact of pre-transplant genetics and clinical features has not yet been fully characterized. We report here the genetic and clinical characteristics and outcomes of a relatively large cohort of patients with therapy-related myelodysplastic syndrome (n=67) who underwent allogeneic transplantation, comparing these patients to similarly treated patients with de novo disease (n=199). The 5-year overall survival was not different between patients with therapy-related and de novo disease (49.9% versus 53.9%; P=0.61) despite a higher proportion of individuals with an Intermediate-2/High International Prognostic Scoring System classification (59.7% versus 43.7%; P=0.003) and high-risk karyotypes (61.2% versus 30.7%; P<0.01) among the patients with therapy-related disease. In mutational analysis, TP53 alteration was the most common abnormality in patients with therapy-related disease (n=18: 30%). Interestingly, the presence of mutations in TP53 or in any other of the high-risk genes (EZH2, ETV6, RUNX1, ASXL1: n=29: 48%) did not significantly affect either overall survival or relapse-free survival. Allogeneic stem-cell transplantation is, therefore, a curative treatment for patients with therapy-related myelodysplastic syndrome, conferring a similar long-term survival to that of patients with de novo disease despite higher-risk features. While TP53 alteration was the most common mutation in therapy-related myelodysplastic syndrome, the finding was not detrimental in our case-series.

Complex analysis of the TP53 tumor suppressor in mantle cell and diffuse large B-cell lymphomas.

Mutations and deletions of the tumor suppressor TP53 gene are the most frequent genetic alterations detected in human tumors, though they are rather less frequent in lymphomas. However, acquisition of the TP53 mutation was demonstrated to be one of the characteristic markers in mantle cell lymphoma (MCL) and diffuse large B-cell lymphoma (DLBCL) and prognostic value of the TP53 status has been recognized for these diseases. We present the complex analysis of the TP53 aberrations in 57 cases of MCL and 131 cases of DLBCL. The TP53 status was determined by functional analyses in yeast (FASAY) followed by cDNA and gDNA sequencing. The level of the p53 protein was assessed by immunoblotting and loss of the TP53-specific locus 17p13.3 was detected by FISH. Altogether, we detected 13 TP53 mutations among MCL cases (22.8%) and 29 TP53 mutations in 26 from 131 DLBCL cases (19.8%). The ratio of missense TP53 mutations was 76.9% in MCL and 82.8% in DLBCL. The frequency of TP53 locus deletion was rather low in both diseases, reaching 9.3% in MCL and 15.3% in DLBCL. The presence of TP53 mutation was associated with shorter overall survival (OS) and progression-free survival (PFS) in MCL. Among DLBCL cases, the TP53 mutations shortened both OS and PFS of patients treated with R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine and prednisone) and decreased both OS and PFS of patients with secondary DLBCL disease.

Activity of Hypomethylating Agents in the Treatment of Therapy-Related Myelodysplastic Syndrome

▪INTRODUCTION: Therapy-related myelodysplastic syndromes (t-MDS) are secondary malignancies that develop in patients who have received cytotoxic chemotherapy and/or radiation therapy for a primary cancer. t-MDS tend to have poor outcomes, with few available therapies and poor responses to conventional chemotherapies.PATIENTS AND METHODS: 263 patients diagnosed with t-MDS and treated with hypomethylating agents (HMA) between July 2001 and December 2015 at a single institution were retrospectively reviewed. Clinical and demographic data were obtained from clinical records. Response was assessed by modified 2006 International Working Group (IWG) criteria. Statistical analyses were performed with the IBM SPSS Statistics 23.0 software. Overall survival (OS) and leukemia free survival (LFS) were defined as the time between treatment onset and death or leukemia (or last contact), respectively. Data were summarized using median, range, and percentage. Censored endpoints were estimated by the nonparametric Kaplan-Meier method and compared using the log-rank test. All tests were 2-sided with significance set at p<0.05.RESULTS: Baseline characteristics are shown in Table 1. 56% of the patients were male. Median age at diagnosis was 66 years (range 13-87). According to the MDS International Prognostic Score System (IPSS), 17 patients (7%) belonged to the low risk group, 77 (29%) to the intermediate-1 risk group, 132 (50%) to the intermediate-2 risk group, and 34 (13%) to the high risk group. Following the Revised International Prognostic Score System (IPSS-R), 6 (2 %), 43 (16%), 20 (7%), 80 (30%), and 100 (38%) belonged to the very low, low, intermediate, high, and very high risk groups, respectively. Following the previously published scoring system for t-MDS, 57 (22%), 120 (46%), 63 (24%) belonged to the good, intermediate, and poor risk groups, respectively. The most common IPSS-R cytogenetic group was complex karyotype with more than 3 abnormalities, which was detected in 112 patients (43%), followed by the good risk group, which was present in 58 patients (22%). Next generation sequencing (NGS) analyzing a panel of 28 or 53 genes was available in 48 patients. The most frequent mutation observed was TP53 (present in 23 patients [48%]), followed by TET2 (present in 8 patients [7%]) and ASXL1 and DNMT3A (present in 3 patients each [6%]). Previous therapy received for the first neoplasm was chemotherapy alone in 123 patients (47%), radiotherapy alone in 27 (10%), or both in 111 patients (42%).A total of 73 (28%) patients received azacitidine in monotherapy (AZA) for the treatment of their t-MDS, 97 (37%) decitabine in monotherapy (DAC), and 92 (35%) were treated with combinations. Overall response rate (ORR) was 52% (137/263), including 33% (87/263) complete response (CR). No difference in terms of OS were observed between those patients treated with AZA vs DAC in monotherapy (p=0.75). The median duration of response was 6 months (range 1-69 months). Median OS was 13 months (Fig. 1A), and transformation to AML occurred in 126 patients (47.9%). There were statistically significant OS differences between responders and non-responders: 17 vs. 9 months, respectively (p<0.01) (Fig.1B).Univariate analysis revealed that concentration of hemoglobin (< 8 g/dl vs. ≥ 8 g/dl; p=0.003), number of platelets (<50x103/mm3 vs. ≥50x103/mm3; p< 0.001), presence of complex karyotype (p< 0.001), and presence of TP53 mutation (p=0.003) were all predictive of survival. After including all the significant factors in the univariate analysis in the multivariate model, only the presence of TP53 mutation was identified as independently associated with shorter survival in patients with t-MDS treated with HMA (HR[IC95%]: 3,44[1.53-7.34]) (Fig. 1C).CONCLUSION: Despite the poor prognosis of patients with t-MDS (median OS 12 months), HMA are clinically effective in the treatment of those patients, providing an ORR of 52% with a CR rate of 33%. Better OS were observed in responders when compared to non-responders, as well as in those patients without TP53 mutations. These results need to be confirmed in a larger numbers of patients. [Display omitted] [Display omitted] DisclosuresJabbour:ARIAD: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Novartis: Research Funding; BMS: Consultancy. Cortes:ARIAD: Consultancy, Research Funding; Bristol-Myers Squib: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Teva: Research Funding. DiNardo:Abbvie: Research Funding; Agios: Research Funding; Celgene: Research Funding; Daiichi Sankyo: Research Funding; Novartis: Research Funding. Konopleva:Calithera: Research Funding; Cellectis: Research Funding.

Prognostic Significance of Genomic Alterations in Mantle Cell Lymphoma

BackgroundWhole genome, whole exome, and targeted sequencing have identified various genomic alterations in mantle cell lymphoma (MCL). There remains limited data correlating genomic alterations with clinical outcome in MCL. We describe genomic alterations in MCL using a targeted sequencing platform and correlate these alterations with clinical outcome.MethodsGenomic DNA was isolated from formalin-fixed paraffin-embedded tissue specimens from 55 cases of MCL and screened for clinically relevant mutations by deep-coverage targeted sequencing. Adaptor ligated sequencing libraries were captured by solution hybridization using two custom bait sets targeting 585 biologically significant cancer-related genes for DNA-Seq. Captured libraries were sequenced to high depth (Illumina HiSeq), averaging >300x. Twenty-eight tumor specimens had corresponding matched normal specimens that facilitated identification of somatic mutations with a high degree of confidence. For the remaining 27 unmatched cases, somatic mutations were inferred according to annotated-based rules. Briefly, non-synonymous variants were considered somatic if they met any of the following criteria: 1) previously reported in the COSMIC database at least 5 times or more, 2) previously reported in published MCL genomic data, or 3) truncating mutations including nonsense mutations, frameshift mutations and splicing mutations that are not at the 3’ end of the genes. The sequencing data were also used for detecting copy number changes including amplifications of established oncogenes or homozygous deletions of known tumor suppressor genes. Overall survival (OS) analyses were performed using Kaplan-Meier and log-rank tests.Results55 cases of newly diagnosed MCL treated at MSKCC from 1999 to 2006 with available specimens were analyzed. Median age of diagnosis was 59 years (range 24-79) with a male predominance (76%). Advanced disease (III/IV) was seen in 93% of cases and an elevated Ki-67 (>=30%) was noted in 41% of patients. Initial treatment included intensive therapy (chemotherapy + ASCT/Allo)(55%), radioimmunotherapy (131I-tositumomab followed by chemotherapy) (20%),bendamustine and an anti-CD20 monoclonal antibody (rituximab or ofatumumab) (11%), single agent anti-CD20 monoclonal antibody (5%), initial observation (7%), and other (2%).The most frequent alterations, above a level of 4%, are shown in Figure 1. The most commonly altered gene was ATM, a tumor suppressor involved in DNA damage repair, and observed in 42% of cases. In addition, known driver mutations in MCL were found, such as TP53 (16%) and CCND1 (11%). Also an amplification of CCND1 was observed in one patient. Alterations in chromatin modifying genes (e.g. KMT2D, MEF2B, WHSC1, SMARCA4) were common, occurring collectively in 38% of the cases. Recurrent alterations in NOTCH1 were identified in 9% of cases. Alterations in BIRC3 (7%) and TRAF2 (4%), key components of the alternative NF-ĸB pathway and mediators of apoptosis, were identified. Homozygous deletion of BIRC3 was also noted in one patient. Homozygous deletion of CDNK2A was noted in 9% of cases. Recurrent promoter mutations were observed in TERT (Telomerase Reverse Transcriptase) gene (13%), which encodes an enzyme involved in cell immortality.With a median follow-up of 7 years, the presence of TP53 mutations was significantly associated with inferior survival compared to TP53-unmutated patients (P<0.001), Figure 2. Notably, homozygous deletion of CDNK2A and mutations in NOTCH1 and KMT2D were not prognostically significant in our series.Conclusion:This analysis further describes the mutational landscape in MCL. Our results are congruent with prior data demonstrating the prognostic relevance of p53 mutations. Several of the genomic alterations identified may have therapeutic significance. For example, TRAF2 and BIRC3 mutations, in the alternative NK-κB pathway, are hypothesized to be associated with ibrutinib resistance. Loss of CDNK2A may confer sensitivity to PRMT5 inhibition. In future,we plan to analyze additional cases of MCL to elucidate the biologic and clinical significance of the observed genetic alterations with the aim of developing mechanism-based, biologically-targeted therapy in MCL. [Display omitted] [Display omitted] DisclosuresKumar:Celgene: Honoraria, Other: Scientific Advisory Board; Celgene: Research Funding; Adaptive Biotechnologies: Research Funding; Seattle Genetics: Research Funding; Pharmacyclics: Research Funding. Zelenetz:Gilead Sciences: Research Funding.

Genetic Alterations Predict Outcomes in Patients with Myelodysplastic Syndrome Receiving Allogeneic Hematopoietic Stem Cell Transplantation

▪Background: Allogeneic hematopoietic stem cell transplantation (HCT) is the only potentially curative therapy for myelodysplastic syndrome (MDS). However, mortality after HCT for MDS is high, with deaths attributable to both relapsed disease and transplant-related complications. The genetic lesions that drive MDS pathogenesis have the potential to predict the outcome of patients following HCT.Methods: We performed targeted sequencing of 127 genes on pre-HCT blood samples from 1514 patients who received allogeneic HCT for MDS between 2005 and 2014. All MDS patients enrolled in the Center for International Blood and Marrow Transplant Research (CIBMTR) Repository were considered for inclusion. Patients were excluded only if blasts were >/= 20% or if they had diagnosis of CMML or MDS/MPN overlap. The median age of patients was 59 years (range 0.4 - 77 years), including 241 (16%) < 40 years of age. Donors were 8/8 HLA-matched unrelated donors (n = 862, 57%), HLA-identical siblings (n = 181, 12%), < 8/8 HLA-matched unrelated donors (n = 296, 20%), or umbilical cord blood (n = 174, 11%).Results:TP53 mutations were independently associated with poor overall survival (HR 1.71, 95% CI 1.45-2.02, p < 0.0001) and a shorter time to relapse (HR 2.34, 1.85-3.00, p < 0.0001). In a multivariable model the adverse prognostic significance of TP53 mutations was independent of recipient age, performance status, hematologic status at time of HCT, bone marrow blast count, and karyotype. RAS pathway mutations were independently associated with a shorter time to relapse (HR 1.73, 1.34-2.23, p < 0.0001) and JAK2 V617F mutations were associated with higher transplant-related mortality (TRM) (HR 2.13, 1.38-3.27, p = 0.0006).Patients with therapy-related MDS (t-MDS) comprised 21% of the cohort (n = 311) and had significantly worse survival than patients with primary MDS (HR 1.4, 1.2-1.6, p = 0.0002). Only TP53 (OR 3.8, p <0.0001) and the p53 regulator PPM1D (OR 4.8, p < 0.0001) were selectively mutated in t-MDS. Overall survival was inferior in t-MDS patients with TP53 mutations (HR 1.7, 1.3-2.3, p = 0.0004), but similar in t-MDS patients without TP53 mutations compared to patients with primary MDS (HR 1.1, 0.9-1.4, p = 0.37).In univariate analysis, young recipient age was powerfully associated with good post-transplant survival. However, we identified a group of adult patients under 40 years old with compound heterozygous mutations in the Shwachman-Diamond Syndrome (SDS)-associated SBDS gene that had remarkably poor prognosis and no long-term survivors (HR 6.0, 1.6-23.0, p = 0.009). All patients with biallelic SBDS mutations had somatic TP53 mutations, suggesting a biological synergy between SBDS and TP53 mutations in clonal MDS transformation of genetically-defined SDS.Finally, we used recursive partitioning to identify 6 distinct MDS subgroups based on the association between clinical and genetic variables and post-HCT survival, then evaluated each subgroup to identify links to relapse or TRM (Figure 1). Consistent with our multivariable model, the presence of TP53 mutations was the most important prognostic variable, with mortality driven by relapse and not influenced by conditioning intensity. In patients age 40 and older without TP53 mutations, two subgroups had poor outcomes: those with RAS pathway mutations, with median OS of 11 months and high risk of relapse (p = 0.001), and those with JAK2 mutations, with median OS of 6 months and high risk of TRM (p < 0.001). Among patients under 40 years old without TP53 mutation, we identified two subgroups based on evaluation of three high-risk clinical features (t-MDS, platelets < 30 x 109/L at HCT, or bone marrow blasts at diagnosis >/= 15%). Patients < 40 years old with at least one high-risk feature had 3 year OS of 49.0%, associated with high TRM (p < 0.001), while those with no high-risk features had 3 year OS of 81.4%.Conclusions: We found that TP53 mutations are the most important predictor of prognosis in allo-HCT for MDS, independent of other clinical and genetic variables, and that the adverse outcome in TP53 mutated MDS is not influenced by conditioning intensity, donor type, or graft source. TP53 mutations account for the characteristically poor outcomes of clinically distinct subgroups, including t-MDS and genetically-defined SDS. We present a multivariable model that identifies 6 distinct MDS subgroups with differential post-HCT survival and differences in relapse and TRM. [Display omitted] DisclosuresLindsley:Takeda Pharmaceuticals: Consultancy; MedImmune: Research Funding. Mar:H3 Biomedicine: Other: Spouse's employment.

P53 Protein Expression Correlates with Adverse Outcome in Low-Risk Myelodysplastic Syndromes

BackgroundThe presence of diverse spectrum of mutations in myelodysplastic syndromes (MDS) has largely revolutionized the understanding of the disease. Although certain mutations have doubtful prognostic significance, the adverse impact of TP53 mutation is irrefutable in all publications. We and others have shown the strong correlation between presence of TP53 mutation and overexpression of p53 by immunohistochemistry (IHC). This evidence provides the rationale for the evaluation of p53 overexpression which is available in routine diagnostic workup, as a surrogate marker to predict TP53 mutations. Additionally, we predicted that the overexpression of p53 might be an independent prognosticator, even in the absence of TP53 mutations or RPS14 haploinsufficiency, in low risk MDS.In our study,we analyzed the p53 protein expression in a cohort of 277 'lower risk' MDS patients and compared the clinico-pathological features, overall survival and risk of progression and evolution to AML with respect to p53 IHC status.MethodsThis study enrolled 277 adult low risk MDS patients seen at King’s College Hospital between March 2009 and February 2016. All marrow samples were assessed for p53 expression (intensity and proportion of cells) and scored using a Modified Quick Score (MQS). MQS ≥2 was chosen to define p53-positive staining. The median follow-up was 18.7 months [95% confidence interval (CI), 16.5-21.0 months].ResultsAccording to WHO categories, the predominant group was RCMD (78%). Patients were either IPSS low (n=134, 48%) or int-1 (n=91, 33%). We included 52 (19%) pts for whom the IPSS category was not available, due to the lack or failed cytogenetic data, but these were low-risk WHO categories with no excess of blasts. IPSS-R categories were very low 103 (37%), low 77 (28%), intermediate 38 (14%) and high risk 7 (3%). Overall, 19 (7%) patients progressed to AML and 41 (15%) patients died of which 25 (61%) were disease-related deaths.Of the 277 patients, 148 (53%) showed p53 protein expression with MQS ≥ 2 and were considered "p53-positive" patients (p53+). The p53 stain intensity was negative in 125 (45%), weak in 84 (30%), moderate in 40 (14%) and strong in 28 (11%). Seventy six (28%) patients had >5% p53 staining cells.p53 expression correlated with a higher age at diagnosis (median age 64 vs. 60 years, p=0.01). lower haemoglobin levels (9.8 vs 11 g/dL, p=0.002), but a higher platelets count (139 vs 99 x109/L, p =.003) (Table 1). This significance persisted even on exclusion of patients with 5q- and MDS/MPN. P53+ did not correlate with any cytogenetic risk group or degree of fibrosis.Sequencing data was available in 121 patients with 63 harbouring somatic mutations. Among these patients, only 3 showed the TP53 gene mutation; all of them were classified as positive when assessed for p53 stain (MQS≥2), showing that the nuclear staining reflects underlying TP53 mutations in such cases. Additionally, presence of SF3B1 mutation correlated strongly with p53+ (p=0.02).Patients with p53+ had a significantly shorter OS compared with those with p53- (figure 1a) with 2yr and 4yr survival probability of 87% &68% vs 95% & 90% , respectively (p=0.03 ).Analysis of progression-free survival (progression to AML, cytogenetic evolution, increase of blasts or fibrosis) showed that patients with p53 expression had a significantly shorter PFS compared to patients without p53 expression (Figure 1b): 2yr & 4yr PFS 70% & 54% vs 85% &75% respectively, p=0.01). Multivariate analysis confirmed the independent prognostic value of p53 expression for PFS (p=0.013, HR 2.6, 95% CI 1.2-5.2).ConclusionIn conclusion, our study showed that p53 overexpression by IHC, represented as MQS cutoff ≥ 2, correlates with features of poor prognosis, such as more advanced age at diagnosis, lower haemoglobin levels and shorter overall survival, adversely impacting progression-free survival.p53 IHC in MDS patients may represent an important, easily available, cheap and applicable prognostic tool and it should be considered for auxiliary analysis when determining the therapeutic options for a patient. The p53+ predicts poor outcome, especially disease progression, independent of the TP53 mutation status, indicating an alternate yet undefined pathway for p53 overexpression. [Display omitted] [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

TP53 Alterations Correlate with Response to VEGF/VEGFR Inhibitors: Implications for Targeted Therapeutics.

TP53 tumor-suppressor gene mutations are among the most frequent abnormalities in cancer, affecting approximately 40% of patients. Yet, there is no accepted way to target these alterations in the clinic. At the same time, antagonists of VEGFR or its ligand are best-selling oncology drugs, with multiple, expensive compounds approved. Although only a subset of patients benefit from these antiangiogenesis agents, no relevant biomarker has been identified. Interestingly, TP53 mutations upregulate VEGF-A and VEGFR2. We prospectively enrolled 500 patients, to be interrogated by comprehensive genomic profiling (CGP) (next-generation sequencing, 236 genes), and to be matched, whenever possible, with targeted agents. Herein, we analyze outcomes based on VEGF/VEGFR inhibitor treatment and presence of TP53 mutations. Of the 500 patients, 188 (37.6%; with ≥1 alteration) were treated; 106 (56% of 188) had tumors that harbored TP53 mutations. VEGF/VEGFR inhibitor therapy was independently associated with improvement in all outcome parameters [rate of stable disease (SD) ≥6 months/partial and complete remission (PR/CR); (31% versus 7%; TP53-mutant patients (who received no other molecular-matched agents) treated with versus without VEGF/VEGFR inhibitors), time-to-treatment failure, and overall survival (multivariate analysis: all P ≤ 0.01)] for the patients harboring TP53-mutant cancers, but improvement was not seen in any of these parameters for patients with TP53 wild-type neoplasms. We conclude that TP53 mutations predict sensitivity to VEGF/VEGFR inhibitors in the clinic. TP53 alterations may therefore be a ready biomarker for treatment with antiangiogenesis agents, a finding of seminal importance across the cancer field. Mol Cancer Ther; 15(10); 2475-85. ©2016 AACR.

Synergy between histone deacetylase inhibitors and DNA-damaging agents is mediated by histone deacetylase 2 in colorectal cancer.

Previous studies have associated the overexpression of histone deacetylase 2 (HDAC2) and the presence of TP53 mutations with the progression to advanced stage drug resistant colorectal cancer (CRC). However, the mechanistic link between HDAC2 expression and the TP53 mutational status has remained unexplored. Here, we investigated the function of HDAC2 in drug resistance by assessing the synergistic effects of DNA-targeted chemotherapeutic agents and HDAC inhibitors (HDACis) on two TP53-mutated colorectal adenocarcinoma CRC cell lines (SW480 and HT-29) and on the TP53-wild type carcinoma cell line (HCT116 p53+/+) and its TP53 deficient sub-line (HCT116 p53−/−). We showed that in the untreated SW480 and HT-29 cells the steady-state level of HDAC2 was low compared to a TP53-wild type carcinoma cell line (HCT116 p53+/+). Increased expression of HDAC2 correlated with drug resistance, and depletion by shRNA sensitised the multi-drug resistance cell line HT-29 to CRC chemotherapeutic drugs such as 5-fluorouracil (5-FU) and oxaliplatin (Oxa). Combined treatment with the HDACi suberoylanilide hydroxamic acid plus 5-FU or Oxa reduced the level of HDAC2 expression, modified chromatin structure and induced mitotic cell death in HT-29 cells. Non-invasive bioluminescence imaging revealed significant reductions in xenograft tumour growth with HDAC2 expression level reduced to <50% in treated animals. Elevated levels of histone acetylation on residues H3K9, H4K12 and H4K16 were also found to be associated with resistance to VPA/Dox or SAHA/Dox treatment. Our results suggest that HDAC2 expression rather than the p53 mutation status influences the outcome of combined treatment with a HDACi and DNA-damaging agents in CRC.

Reduced MUTYH, MTH1, and OGG1 expression and TP53 mutation in diffuse-type adenocarcinoma of gastric cardia

The effects of oxidative stress in adenocarcinomas of gastric cardia (AGCs) have not been fully elucidated. With a strict definition of AGC, we examined the immunohistochemical expressions of inducible nitric oxide synthase; 8-hydroxy-deoxyguanosine; and the base excision repair enzymes such as MUTYH, MTH1, and OGG1, and TP53 mutational status. Sixty-three cases of AGC were characterized by younger patient age (P = .0227) and more frequent venous invasion (P = .0106) compared with the adenocarcinomas of pylorus (APs). 8-hydroxy-deoxyguanosine was accumulated (P = .0011), whereas MUTYH (P = .0325) and OGG1 (P = .0007) were decreased, in the AGCs compared with the adjacent mucosa, but these differences were not detected in the APs. Among the AGCs, lower expressions of MUTYH (P = .0013) and MTH1 (P = .0059) were each significantly associated with diffuse-type histology. A lower expression of OGG1 was correlated with higher T-stage (P = .0011), lymphatic invasion (P = .004), and lymph node metastasis (P = .0094). In addition, the presence of TP53 mutation was associated with diffuse-type histology (P = .0153) and a lower level of MUTYH (P = .0221). The AGCs also showed a relatively high rate of a transversion-type mutation of TP53 (50%), whereas all TP53 mutations in the APs were transition type. Age 62years or older (P = .0073), diffuse-type histology (P = .0020), and TP53 mutation (P = .0066) were each associated with worse survival in the AGC patients. Our results indicate that oxidative stress accumulation and a downregulation of base excision repair enzymes may play an important role in the pathogenesis of AGC, in particular diffuse-type AGCs. Diffuse-type AGC might involve molecular pathways different from those of other subsets of gastric cancer.

Cell-free DNA as a molecular tool for monitoring disease progression and response to therapy in breast cancer patients.

Due to the spatial and temporal genomic heterogeneity of breast cancer, genomic sequencing obtained from a single biopsy may not capture the complete genomic profile of tumors. Thus, we propose that cell-free DNA (cfDNA) in plasma may be an alternate source of genomic information to provide comprehensive data throughout a patient's clinical course. We performed a retrospective chart review of 100 patients with stage 4 or high-risk stage 3 breast cancer. The degree of agreement between genomic alterations found in tumor DNA (tDNA) and cfDNA was determined by Cohen's Kappa. Clinical disease progression was compared to mutant allele frequency using a two-sided Fisher's exact test. The presence of mutations and mutant allele frequency was correlated with progression-free survival (PFS) using a Cox proportional hazards model and a log-rank test. The most commonly found genomic alterations were mutations in TP53 and PIK3CA, and amplification of EGFR and ERBB2. PIK3CA mutation and ERBB2 amplification demonstrated robust agreement between tDNA and cfDNA (Cohen's kappa=0.64 and 0.77, respectively). TP53 mutation and EGFR amplification demonstrated poor agreement between tDNA and cfDNA (Cohen's kappa=0.18 and 0.33, respectively). The directional changes of TP53 and PIK3CA mutant allele frequency were closely associated with response to therapy (p=0.002). The presence of TP53 mutation (p=0.0004) and PIK3CA mutant allele frequency [p=0.01, HR 1.074 (95% CI 1.018-1.134)] was excellent predictors of PFS. Identification of selected cancer-specific genomic alterations from cfDNA may be a noninvasive way to monitor disease progression, predict PFS, and offer targeted therapy.

The Complete Mutatome and Clonal Architecture of Del(5q)

Cytogenetic abnormalities are found in around half of MDS patients (pts) and have both clinical impact and may be subtype-defining, e.g. in 5q-syndrome. Interstitial deletion of the long arm of chr.5 [del(5q)] is the most common aberration (almost 20% of cases with abnormal cytogenetics). Del(5q) is heterogeneous, occurring as a sole abnormality or in combination, with the deleted region often truncated within or extended and/or beyond the CDR boundaries. Isolated del(5q) is frequently shorter and confers a more favorable prognosis with regard to survival and lenalidomide (LEN) responsiveness, while del(5q) in the context of a complex karyotype (CK) imparts a poor prognosis. In addition to chromosomal lesions, somatic mutations can contribute to the pathogenesis of MDS, including del(5q). We theorized that recognition of molecular defects in MDS with del(5q) may clarify the pathogenic mechanisms behind this lesion and help explain the clinical heterogeneity.We analyzed 225 pts with myeloid neoplasia and del(5q) using WES (n= 107 samples) and targeted multiplexed PCR (top 60 most frequently mutated genes) (n =133 samples); serial analysis was performed in 15 pts studied at ≥2 time points, 11 during LEN therapy and 4 upon relapse/progression. A total of 116 samples had a CK with other lesions such as -7/del(7q) found in 31% cases, and 18% had -17/del(17p).WES (average depth >60x) was followed by a bioanalytic pipeline, detecting ≥1 mutated gene in 71% of cases. Candidate somatic alterations were found in 357 genes and selected for further analysis. When focused on hemizygous mutations within the retained 5q allele, CSNK1A1 mutations were the most common, found in 4 pts, while other genes were only sporadically affected. Among heterozygous mutations on the non-deleted portion of del(5q) and other chromosomes (Chr), we found several novel mutations, in addition to TP53 (n=26), DNMT3A (n=8), PRPF8 (n =8), RUNX1 (n=5), TET2 (n=5), and ASXL1 (n=4), among others. Furthermore, LOH/haploinsuffciency of genes on 7q (e.g., LUC7L2, CUX1, EZH2 and MLL3) appears to be a common defect seen in pts with non-isolated del(5q), suggesting synergistic functional defects. When functionally grouping gene mutations, DNA methylation family (8 cases) and transcription factor mutations (29 cases) were associated with advanced disease (AD) and a CK. Heterozygous mutations in TP53 (34%) or deletions involving the TP53 locus (23%) resulted in total of 42% of cases carrying either TP53 LOH or mutation. TP53 lesions were more common in pts with AD vs. low risk. (21 vs. 5 p =.0008). In contrast, TP53 mutations are found in 8-10% of cases of MDS.A total of 34 pts were treated with LEN and subgrouped into responders (n=17) vs. refractory (n=9) with an overall response rate of 65%. When mutational profiles were compared, the presence of TP53 mutations did not preclude responsiveness to LEN. CK was present in 12% of responders vs. 67% of refractory pts. The most frequent Chr abnormalities were -7/7q (0% vs. 67% in responders vs. refractory) and 17p-(6% vs. 67% in responders vs. refractory) suggestive of their role in LEN resistance.In addition to cross sectional analysis, our WES study using paired Germline/tumor samples followed by deep sequencing facilitated analyses of clonal architecture by examining clonal dynamics over time. Assessment of del(5q) clone size by allelic imbalance combined with clonal burden by VAF allowed us to reconstruct the clonal hierarchy: in 73% of cases, del(5q) appeared to be the initial defect followed by subsequent mutations (e.g., TP53, DNMT3A, IDH2). In contrast, in 24% of cases, TP53, RUNX1, JARID2, were the primary defect followed by a subclonal del(5q) events. Serial samples collected before and after therapy demonstrated that responses were associated with decreased clonal burden for del(5q) but persistence of certain mutations. In refractory cases, persistent subclonal lesions and the appearance of new lesions were associated with progression. For example, pts with TP53, LAMB4, EPHA6 progressed and acquired additional lesions such as CSMD2 or KCND2, and did not see the disappearance of TP53 alterations upon treatment.In conclusion, no unifying somatic defect was found in pts with del(5q) regardless if the deletion event was primary or subclonal. Most commonly associated lesions were not present on the retained 5q alleles but rather other chr yet modified clinical behavior, including responsiveness to LEN. DisclosuresBejar:Celgene: Consultancy, Honoraria; Alexion: Other: ad hoc advisory board; Genoptix Medical Laboratory: Consultancy, Honoraria, Patents & Royalties: MDS prognostic gene signature. Sekeres:Celgene Corporation: Membership on an entity's Board of Directors or advisory committees; TetraLogic: Membership on an entity's Board of Directors or advisory committees; Amgen: Membership on an entity's Board of Directors or advisory committees.

Abstract 3082: miR-155-5p expression is associated with response to neoadjuvant paclitaxel treatment of locally advanced breast cancer

Abstract Introduction Resistance to chemotherapy remains a prominent obstacle to cure in cancer. We have previously shown that genetic defects in key pathways involved in processes like apoptosis, cell-cycle arrest and senescence may be involved in resistance to chemotherapy in vivo. MicroRNAs (miRNAs) are known to be crucial regulators of gene expression, and to potentially affect the response to drugs, including chemotherapy. Here, we aimed to explore the potential role of miRNAs in mechanisms of breast cancer chemoresistance in-vivo, by analyzing miRNA expression in breast cancer patients with recorded responses to chemotherapy. Patient materials and Methods Breast tumour biopsies were collected before chemotherapy in a prospective clinical study, where patients were randomized to receive epirubicin or paclitaxel monotherapy (n = 223). Information about the response to therapy, as defined by the UICC criteria, in addition to long-term outcome, was available for all patients. The global miRNA expression levels were analyzed in 50 breast tumour biopsies by deep sequencing. The patients were selected based on their response to first-line chemotherapy. To verify the results obtained by miRNA sequencing, selected miRNAs were quantified in all patients by qPCR. Results No global miRNA patterns were observed when performing unsupervised clustering of miRNA sequencing data. Among samples from paclitaxel-treated patients, we identified three differentially expressed miRNAs between response groups; the expression of miR-155 was significantly lower (p = 0.005) in patients with poor response to paclitaxel, and the expression of both miR-181a and miR-181b were significantly higher (p = 0.003 and 0.012, respectively) in patients with poor response to paclitaxel. qPCR data validated the initial association identified between high miR-155-5p expression levels and response to paclitaxel treatment. Intriguingly though, high levels of miR-155 were also associated with a poor prognosis in the study population as a whole (p = 0.012), as well as with the presence of mutations in TP53 (p &amp;lt; 0.001). Conclusion Here, we show that breast cancer patients responding to paclitaxel treatment have higher expression levels of miR-155-5p. Also, patients with elevated levels of miR-155-5p expression have a poor prognosis and a higher frequency of TP53 mutations. Citation Format: Anne Hege Straume, Einar Birkeland, Stian Knappskog, Per Eystein Lønning. miR-155-5p expression is associated with response to neoadjuvant paclitaxel treatment of locally advanced breast cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3082. doi:10.1158/1538-7445.AM2015-3082