Clinical Utility of Molecular Profiling in Higher Risk MDS Treated with Azacitidine: Can We Tailor Therapy Accordingly?

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1984-1984 ◽  
Author(s):  
David Sallman ◽  
Eric Padron ◽  
Jinming Song ◽  
Mohammad Omar Hussaini ◽  
Christine Vaupel ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Research Funding ◽  
Somatic Mutations ◽  
Multivariable Analysis ◽  
Molecular Profiling ◽  
Molecular Profile ◽  
Cell Transplant ◽  
Hematopoietic Stem ◽  
Myeloid Malignancies ◽  
The Impact

Abstract Background Hypomethylating agent (HMA) therapy represents the standard of care for patients with higher risk myelodysplastic syndromes although only 50% of patients respond to treatment. Recent evidence from molecular profiling through next-generation sequencing (NGS) in myeloid diseases has been conflicting as to the value of somatic mutations as a biomarker for response to HMA. TET2 mutations, predominantly in the absence of ASXL1 mutations (or variant allele frequency (VAF) < 10%), have been shown to predict HMA response in MDS and chronic myelomonocytic leukemia (CMML) (Itzykson et al., 2011; Bejar et al., 2014 (80% decitabine)) while DNMT3A mutations predict response to frontline HMA treatment in acute myeloid leukemia (AML) (Coombs et al., 2016). Therefore, our goal was to identify molecular predictors of response and outcomes to azacitidine in myeloid malignancies. Patients and Methods Genetically profiled higher-risk MDS, CMML and oligoblastic AML (20-30% blasts) cases were retrospectively identified from the Moffitt Cancer Center MDS database. We evaluated gene mutations associated with DNA methylation (TET2, DNMT3A, IDH1, IDH2, and WT1) and up to 19 additional genes. NGS was performed prior to the initiation of HMA in all patients. The lower limit of VAF detection was set at 5% and the minimum depth of coverage at each position was 500X. Clinical variables and outcomes of MDS patients were characterized at the time of sample procurement. Fisher's exact and t-tests were used for comparative analyses. Kaplan-Meier curves were used to estimate overall survival and analyzed from the date of mutation identification. Multivariate Cox regression models were created to adjust for clinical characteristics. Results From May 2013 to February 2016, a total of 77 patients with NGS for somatic mutations prior to HMA therapy were identified with a median age of 70 years and male predominance (66%). Of the cohort, 97% of patients (n=75) were treated with azacitidine with 17% of patients (n=13) proceeding to allogeneic hematopoietic stem cell transplant (AHSCT). A total of 86% of patients (n=66) had at least one pathogenic mutation. Mutations in DNA methylation occurred in 43% of patients (n=33) while TET2 mutation without clonal ASXL1 mutations occurred in 13% of patients (n=10). At a median follow up 17 months, the median OS of the entire cohort was 12.5 months. Patients with a DNA methylation mutation had a median OS that was not reached (NR) vs a median OS of 11.5 months in wildtype (WT) patients (HR 0.38, 95% CI 0.21 to 0.75; P = 0.005), which remained significant when censoring patients at time of AHSCT (P = 0.002). TP53 mutant (MT) patients (n=14, 18% of cohort) had a median OS of 7.9 months vs 15.4 months in WT patients (HR 3.69, 95% CI 3.04 to 28.8; P = 0.0001). The presence of DNA methylation or TP53 mutation in comparison to wildtype patients significantly stratified prognosis in azacitidine treated patients (Figure 1A, P = 0.0001). In multivariable analysis incorporating age and revised international prognostic scoring system (IPSS-R) category, DNA methylation (HR 0.45, 95% CI 0.21 to 0.96; P = 0.04) and TP53 (HR 2.34, 95% CI 1.04 to 5.26; P = 0.04) mutation status remained predictive for survival. Notably, response rates in DNA methylation mutant patients were similar to WT patients (40% versus 42%) with no difference in treatment duration. However, the overall response rate of TET2 MT/ASXL1 WT patients was 70% versus 36% in the rest of the cohort (P =0.08) and 0% in TET2 MT/ASXL1 MT patients (0/6, P =0.01) with a significantly longer duration of treatment in the TET2 MT/ASXL1 WT cohort (median number of cycles 7.5 versus 4; P = 0.002). Additionally, TET2 MT/ASXL1 WT patients had longer survival (median OS NR vs 12.2 months; P = 0.047). When censoring for transplant, the impact of TET2 MT/ASXL1 WT genoptype was significantly strengthened with 70% of patients alive past 15 months (median OS NR vs 9.9 months; HR 0.24, 95% CI 0.19 to 0.75; P = 0.007; Figure 1B). Conclusion In patients with myeloid malignancies, molecular profiling via NGS can predict outcomes to azacitidine therapy. Patients with mutations of DNA methylation have improved OS whereas OS is poor in TP53 MT patients. Most importantly, TET2 MT/ASXL1 WT identifies a genotypic subgroup with particularly good outcomes when treated with HMA without AHSCT and potentially challenges the early timing of AHSCT for higher risk patients and this molecular profile. Figure 1 Figure 1. Disclosures Padron: Novartis: Honoraria; Incyte: Research Funding; CTI: Honoraria, Research Funding; KALOBIOS: Research Funding. Vaupel:Genoptix, a Novartis Company: Employment. Hall:Genoptix, a Novartis Company: Employment. Komrokji:Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Consultancy, Speakers Bureau; Boehringer-Ingelheim: Research Funding; Incyte: Consultancy.

scholarly journals Clonal Suppression of TP53 Mutant MDS and Oligoblastic AML with Hypomethylating Agent Therapy Improves Overall Survival

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 1817-1817 ◽  
Author(s):  
David A Sallman ◽  
Najla Al Ali ◽  
Seongseok Yun ◽  
Eric Padron ◽  
Jinming Song ◽  
...  
Keyword(s):  
Overall Survival ◽  
Research Funding ◽  
Multivariable Analysis ◽  
Response Rates ◽  
International Prognostic Scoring System ◽  
Cancer Center ◽  
Cell Transplant ◽  
Hematopoietic Stem ◽  
Male Predominance ◽  
Hypomethylating Agent

Abstract Background Hypomethylating agent (HMA) therapy represents the standard of care for patients with higher risk myelodysplastic syndromes (MDS) although only 50% of patients respond to treatment. Recent evidence from molecular profiling through next-generation sequencing (NGS) in myeloid diseases has been conflicting as to the value of somatic mutations as a biomarker for response to HMA. In particular, there have been conflicting data on response rates and outcomes in TP53 mutant (MT) MDS and acute myeloid leukemia (AML) based on azacitidine versus decitabine (Welch et al., NEJM 2016; Garcia-Manero et al., NEJM 2017). However, the TP53 mutant cohorts in these studies were small (median 23 patients, range 13-39) and heterogeneous based on treatment status (treatment naïve versus relapse/refractory). Therefore, our goal was to characterize outcomes of TP53 mutant MDS patients who received frontline HMA therapy. Patients and Methods TP53 MT MDS and oligoblastic AML (20-30% blasts) cases were retrospectively identified from the Moffitt Cancer Center MDS database. All patients had NGS of TP53 and up to 53 additional genes performed prior to the initiation of HMA. The lower limit of VAF detection was set at 5% and the minimum depth of coverage at each position was 500X. Clinical variables and outcomes of MDS patients were characterized at the time of sample procurement. Fisher's exact tests were used for comparative analyses. Kaplan-Meier curves were used to estimate overall survival (OS) and analyzed from the date of HMA initiation. Response rates and outcomes of TP53 MT patients were compared to a cohort of wildtype (WT) patients (n=63). Results From May 2013 to May 2018, a total of 71 patients with TP53 mutant MDS were identified with a median age of 68 years (39-82) and male predominance (66%). Fourteen patients (20%) had multiple mutations in TP53. Of the cohort, 82% of patients (n=58) were treated with azacitidine (88% (n=51) with AZA monotherapy; 12% (n=7) with AZA in combination (2 pts with lenalidomide and 5 pts with investigational agents)) with 18% (n=13) receiving decitabine. The median # of HMA cycles was 4 (range 1-33). Thirteen pts (18%) proceeded to allogeneic hematopoietic stem cell transplant (HSCT). Of the cohort, 18% (n=13) obtained complete remission (CR) with 39% (n=28) overall response rate (ORR). There was no difference in CR or ORR in pts treated with AZA vs DAC (P=0.24 and P=0.2, respectively). At a median follow up 20 months, the median OS of the entire cohort was 9.7 months. There was no difference in median OS between AZA vs AZA combo vs DAC (7.6 vs 15.2 vs 12.5 months; P = 0.44; Figure 1A). TP53 variant allele frequency (VAF > 20% vs ≤ 20%) was not predictive of outcomes to HMA (7.8 vs 10.4 months, P = 0.36). However, TP53 MT patients who had clonal response to HMA (i.e. VAF < 5%; n=19 (27%)) had improved OS (14.5 vs 7.5 months; HR 0.33, 95% CI 0.18 to 0.59; P = 0.001; Figure 1B). In multivariable analysis incorporating age, revised international prognostic scoring system (IPSS-R) category, HSCT status, or type of HMA, TP53 clonal clearance remained an independent covariate for improved OS (HR 0.34, 95% CI 0.16 to 0.72; P = 0.005). Pts who underwent HSCT (n=13) had a trend for improved OS (14.5 months vs 7.9 months; P = 0.09). Notably in transplanted pts who had serial TP53 NGS (n=7), pts who achieved a VAF < 5% had significant improved OS (16.3 months vs 8.9 months; P=0.03). Compared to higher risk MDS/AML TP53 WT patients treated with HMA, there was no difference in CR (18% vs 14% (P = 0.64) or ORR rates (39% vs 40%). In contrast, TP53 MT patients had significantly inferior OS with HMA therapy (9.7 vs 15.4 months; HR 2.14, 95% CI 1.32. to 3.27; P = 0.001; Figure 1C). Conclusion In this large cohort of higher risk MDS and oligoblastic AML pts who received frontline HMA therapy, TP53 MT patients have significantly inferior OS with no significant differences in response rates or outcomes by HMA. TP53 MT patients who achieve maximum clonal suppression with HMA treatment (i.e. VAF < 5%) have improved OS as well as improved outcome with HSCT. Novel therapy targeting TP53 mutation is needed to improve outcomes. Figure 1. Figure 1. Disclosures Sallman: Celgene: Research Funding, Speakers Bureau. Sweet:Agios: Consultancy; Jazz: Speakers Bureau; Astellas: Consultancy; Phizer: Consultancy; Phizer: Consultancy; Astellas: Consultancy; Jazz: Speakers Bureau; Celgene: Honoraria, Speakers Bureau; Novartis: Consultancy, Honoraria, Speakers Bureau; Novartis: Consultancy, Honoraria, Speakers Bureau; Agios: Consultancy; BMS: Honoraria; Celgene: Honoraria, Speakers Bureau; BMS: Honoraria. List:Celgene: Research Funding. Komrokji:Novartis: Honoraria, Speakers Bureau; Celgene: Honoraria, Research Funding; Novartis: Honoraria, Speakers Bureau; Novartis: Honoraria, Speakers Bureau; Celgene: Honoraria, Research Funding; Novartis: Honoraria, Speakers Bureau.

Somatic Mutations Implicated In Myeloid Malignancies Are Frequent In Idiopathic Aplastic Anaemia and Its Relevance To Disease Classification and Treatment- a Comprehensive Analysis Of 150 Patients

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 803-803 ◽  
Author(s):  
Jie Jiang ◽  
Austin G Kulasekararaj ◽  
Alexander E Smith ◽  
Azim M Mohamedali ◽  
Shreyans A Gandhi ◽  
...  
Keyword(s):  
Stem Cell ◽  
Somatic Mutations ◽  
Conflicts Of Interest ◽  
Hot Spot ◽  
Clonal Evolution ◽  
Molecular Evidence ◽  
Cell Transplant ◽  
Hematopoietic Stem ◽  
Monosomy 7 ◽  
Myeloid Malignancies

Abstract Background Acquired aplastic anemia (AA) is an immune mediated disease, characterized by severe quantitative defects in stem cell number leading to a hypocellular marrow and peripheral blood cytopenias. A major complication of AA following immunosuppressive therapy (IST) is the emergence of clonal hemopoiesis, manifesting as paroxysmal nocturnal hemoglobinuria (PNH), myelodysplastic syndrome (MDS) and acute myeloid leukaemia. The nature of genetic defects in such cases evolving to clonal hemopoiesis has not been addressed hitherto. It is likely that the hematopoietic stress leads to telomere attrition, chromosomal instability and susceptibility to acquire mutations in genes several years after the initial diagnosis. We postulated the existence of mutations leading to clonal haemopoiesis in a subset of AA and thereby helping in early prediction. Methods We undertook a massively parallel targeted sequencing of 832 genes implicated in regulating hematopoiesis initially in a cohort of 57 patients with acquired AA. We also performed single nucleotide polymorphism-array karyotyping, telomere measurement, PNH flow cytometry and correlated it with clinical characteristics. A second cohort of 93 patients was sequenced at a greater depth for ASXL1 and DNMT3A. Results The median age of AA patients was 44 years (range 17-87 years). The diagnosis was non severe AA (NSAA) 64, severe AA (SAA) 52, very severe AA (VSAA) 27 and pure red cell aplasia (PRCA)/amegakaryocytic thrombocytopenia 7. PNH-type cells and cytogenetic aberrations were seen in 52% and 9% of patients respectively. IST was administered in 72% of patients with 55% receiving ATG containing regimens. Allogeneic hematopoietic stem cell transplant was performed in 25% (n=37). Clonal evolution to MDS/AML was observed in 12% of patients. Somatic mutations in genes usually implicated in myeloid malignancies were detected in 15% (22/150) of AA. No particular sub-group of AA patients had a predilection to harbour somatic mutations, although a slightly higher incidence was noted in SAA and NSAA, compared to VSAA (22% vs. 14%, p=0.1).Thirty-five (23%) AA patients had either morphological (n=18) and/or molecular (n=22) evidence of evolution to MDS. Of the 18 patients with morphological evolution to MDS, ASXL1 and TET2 mutations were observed in 5 patients. ASXL1 (n=11) mutations were the most common somatic variation and correlated with older age, evolution to MDS and the presence of monosomy 7. Although there was no predilection for a particular ‘hot spot’, all mutations occurred in exon 12 and were uniformly heterozygous. DNMT3A (n=7) and BCOR(n=2) were the other frequent mutations. The median allele burden of somatic mutations was 30%. AA with somatic mutations were older (51 vs. 38 years, p<0.07), had a longer disease duration (44 vs. 9 months, p< 0.06), shorter telomere lengths (median T/S length, 1.3 vs. 0.8, p<0.008) and were less likely to achieve complete remission to IST (10% vs. 34%) compared to those without mutations. Conclusion Up to 25% of AA patients have morphological or molecular evidence of clonal evolution with a fifth of AA patients harboring mutations in genes implicated in myeloid malignancies, with both diagnostic and therapeutic implications. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Impact of Race and Geographic Location on Outcomes in Allogeneic Transplant

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 17-18
Author(s):  
Audrey M. Sigmund ◽  
Qiuhong Zhao ◽  
Justin Jiang ◽  
Patrick Elder ◽  
Don M. Benson ◽  
...  
Keyword(s):  
Board Of Directors ◽  
Research Funding ◽  
Ohio State University ◽  
Cell Transplant ◽  
State University ◽  
Advisory Committees ◽  
Hematopoietic Stem ◽  
Significant Difference ◽  
The Ohio State University ◽  
The Impact

Introduction: Allogeneic hematopoietic stem cell transplant (allo-HCT) is a potential curative therapy for a variety of both malignant and nonmalignant hematologic disorders. However, allo-HCT is costly and requires highly specialized, technologically advanced care that is only available in select healthcare centers across the country. Due to its cost and limited availability, minority populations are at risk for healthcare disparities in access to and outcomes of allo-HCT. Prior studies have focused on the impact of health disparities, including race, and geographic residence at time of transplant, on allo-HCT outcomes with variable results. The aim of this study was to evaluate the impact of race and location of residence on outcomes of allo-HCT at one major referral institution. Methods: We performed a retrospective cohort study of patients that underwent allo-HCT at the Ohio State University from 1984 to 2018. The impact of demographic factors including race and place of primary residence were assessed. Patients were divided into race defined as Caucasian, African American (AA), and other. They were also grouped by zip code into rural, suburban, and urban groups. Rural was defined as less than 1000 people per square mile, suburban between 1000-3000 people per square mile, and urban greater than 3000 people per square mile. 2018 population estimates were used. Patients were then stratified into 7 groups based on year (yr) of transplant for analysis. Group (gp) 1 included 1984-1988, gp 2 1989-1993, gp 3 1994-1998, gp 4 1999-2003, gp 5 2004-2008, gp 6 2009-2013, and gp 7 2014-2018. Primary endpoints were progression free survival (PFS) and overall survival (OS). PFS and OS were calculated using Kaplan Meier Curves and compared using log-rank test between race and residence groups. Results: A total of 1,943 patients were included in the study. Of these patients, median age at time of transplant was 50 years old (range 18-76), and 59.6% were male. AML/MDS patients made up the majority of the cohort at 46.3%, with the other most common diagnoses being non-Hodgkin's lymphoma (14.2%), acute lymphocytic leukemia (11.8%), and chronic myeloid leukemia (10.1%). Most patients (94.3%) identified as Caucasian, while 4.6% identified as AA, and 1.1% other. The majority of patients lived in a rural area at the time of transplant with 63.4% rural, 22.9% suburban, and 13.8% urban. There was no significant difference in OS or PFS between Caucasian and AA patients (Figure 1A and B; p=0.15, 0.21). Median OS for AA was 1.9 yrs [95% confidence interval (CI): 0.8-3.6] as compared to 2.3 yrs (95% CI: 1.9-2.9) for Caucasians, with 5 -yr OS of 33 vs. 42% and 10-yr OS of 21 vs. 36% for AA and Caucasian, respectively. Median PFS was 0.9 (95% CI: 0.5-2.7) and 1.3 yrs (95% CI 1.1-1.6), with 5 -yr PFS of 30 vs. 37% and 10-yr PFS of 21 vs. 32% for AA and Caucasian, respectively. There also was no significant difference in OS or PFS between rural, urban, and suburban patients (Figure 2A and 2B; p=0.39, 0.17), with median OS in the three groups 2.2 (95%CI: 1.7-2.9), 2.9 (95%CI: 1.6-4.5), and 2.2 (95% CI: 1.6-3.6) yrs, and 5-yr OS of 40 vs. 43 vs. 43% and 10-yr OS of 33 vs. 39 vs. 39%, respectively. Median PFS were 2.2 (95%CI: 1.7-2.9), 2.9 (95%CI: 1.6-4.5), and 2.2 yrs [95% CI: 1.6-3.6], with 5-yr PFS of 36 vs. 40 vs. 38% and 10-yr PFS of 30 vs. 37 vs. 35%, respectively. Conclusion: Our study suggests that once patients undergo allo-HCT, there is no significant difference in outcomes between patients based on race or residence. This finding suggests that while these underserved populations may initially have less access to specialized care for HCT, if they ultimately undergo allo-HCT, outcomes are similar to their counterparts. Our study did show a significantly lower rates of allo-HCT performed in non-Caucasian races (94% Caucasians vs 4.6% AA and 1% other), which may reflect disparities in access to care in these groups as well as a lack of donors. Further research is needed to assess the barriers for these underserved patients to undergo transplant and to help ameliorate these barriers. Disclosures Chaudhry: Sanofi: Consultancy, Membership on an entity's Board of Directors or advisory committees. Bumma:Amgen: Speakers Bureau; Sanofi: Speakers Bureau. Khan:Amgen: Consultancy; Janssen: Consultancy. Devarakonda:Janssen: Consultancy. Vasu:Janssen: Membership on an entity's Board of Directors or advisory committees; Omeros: Membership on an entity's Board of Directors or advisory committees; Kiadis Inc: Other: Kiadis has obtained exclusive licensing requirements from The OHio State University. Jaglowski:Kite, a Gilead Company: Consultancy, Research Funding; Juno: Consultancy; Novartis: Consultancy, Research Funding; CRISPR: Consultancy. William:Merck: Research Funding; Celgene: Consultancy, Honoraria; Dova: Research Funding; Seattle Genetics: Research Funding; Incyte: Research Funding; Guidepoint Global: Consultancy; Kyowa Kirin: Consultancy, Honoraria. Mims:Syndax Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Kura Oncology: Membership on an entity's Board of Directors or advisory committees; Leukemia and Lymphoma Society: Other: Senior Medical Director for Beat AML Study; Agios: Consultancy; Novartis: Speakers Bureau; Jazz Pharmaceuticals: Other: Data Safety Monitoring Board. Brammer:Seattle Genetics, Inc.: Speakers Bureau; Celgene Corporation: Research Funding. Efebera:Celgene: Research Funding; Pharmacyclics: Research Funding; Takeda: Honoraria, Speakers Bureau; Ohio State University: Current Employment.

scholarly journals TP53 Mutations and Outcome in Patients with Myelodysplastic Syndromes (MDS)

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4336-4336 ◽  
Author(s):  
Karam Al-Issa ◽  
Mikkael A. Sekeres ◽  
Alek d Nielsen ◽  
Babal Jha ◽  
Bartlomiej P Przychodzen ◽  
...  
Keyword(s):  
Myelodysplastic Syndromes ◽  
Research Funding ◽  
International Prognostic Scoring System ◽  
Entire Group ◽  
Prognostic Impact ◽  
Cell Transplant ◽  
Transactivation Domain ◽  
Hematopoietic Stem ◽  
Tp53 Mutations ◽  
Myeloid Malignancies

Abstract Objectives: TP53, a tumor suppressor gene, is frequently mutated in myeloid malignancies. The negative impact of TP53 mutations in myelodysplastic syndromes (MDS) has been described previously but there is controversy regarding the prognostic impact of the mutation's characteristics (location, type, passenger vs. driver, and others). Methods: We sequenced DNA samples from 732 patients (pts) with MDS and related myeloid malignancies for the presence of TP53 mutations and 61 other genes that have been described as recurrently mutated in MDS. Overall survival (OS) was measured from the time of diagnosis to time of death or last follows up. Variant allele frequencies (VAFs) adjusted by zygosity were used to define clonal architecture of driver clones. Results: Of 80 mutations detected in 73 (10%) pts, 66 (88%) were missense, 7 (9%) were nonsense, and 7 (9%) were frame shift deletions/insertions. Pts with TP53 mutations had a higher WBC (4.6 vs. 3.9 X 109/L, p = .04), higher bone marrow blast % (median 9 vs. 3, p =0.1), and a higher risk category by revised International prognostic Scoring System (56% vs. 27%, p =.01) compared to pts with TP53 wild type. TP53 mutations were commonly occurred with TET2 (16%), PRPF8 (13%), ASXL1 (11%), DDX54 (8%), DNMT3A (8%), and IDH2 (8%). The mean VAF for TP53 was 41.9 (5-100). TP53 mutations were defined as drivers in 20% of samples, passengers in 40%, and mosaic in 40%. Mutation positions included: 19 (24%) in the DNA binding domain, 2 (3%) in the transactivation domain, 1 (1%) in the tetramerization domain and 58 (72%) other. With a median follow up of 16.4 months, the median OS for the entire group was 8.24 months. Patients with TP53 as driver mutations had a worse OS compared with patients with TP53 as passenger mutations (median, 2.2 vs. 13.0 months, respectively, p =.02). Similarly, OS by TP53 VAF categorized as low (<25%), intermediate (25-50%), and high (>50%) was 12.4, 8.5, and 3.4 months, respectively. Among patients with available treatment data, 29 patients were treated with the hypomethylating agents, azacitidine or decitabine, 17% responded (60% CR, and 40% PR), and OS was similar compared to patients treated with other therapies. Patients treated with Hematopoietic stem cell transplant (HCT) had superior OS compared to pts not receiving HCT (median, 14.9 vs. 8.9 months, respectively, p =.05). Conclusions: TP53 mutations are associated with poor outcome in MDS, but the outcome varies depending on the type of mutation and VAF. Treatment with HCT remains a valid treatment option in a subset of patients but novel treatment strategies are desperately needed. Disclosures Mukherjee: Novartis: Consultancy, Honoraria, Research Funding; Ariad: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding.

Landscape of Mutations in Relapsed Acute Myeloid Leukemia

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 2367-2367
Author(s):  
Samuli Eldfors ◽  
Mika Kontro ◽  
Kimmo Porkka ◽  
Olli Kallioniemi ◽  
Caroline Heckman
Keyword(s):  
Drug Resistance ◽  
Research Funding ◽  
Somatic Mutations ◽  
Sequence Data ◽  
Donor Cell ◽  
Driver Mutations ◽  
Cell Transplant ◽  
Cancer Genes ◽  
Hematopoietic Stem ◽  
Germline Variants

Abstract While the majority of acute myeloid leukemia (AML) patients respond to induction chemotherapy, disease recurrence and drug resistance is common. Recently, mutations underlying AML pathogenesis have been extensively characterized by sequencing large numbers of samples obtained at diagnosis. However, mutations driving disease progression and drug resistance in relapsed AML are not well characterized. In addition, understanding the clonal composition of relapsed AML is compounded by interference of donor cell variants present in those patients who have received an allogeneic hematopoietic stem cell transplant (alloHSCT). In this study we sought to identify mutations and copy number aberrations associated with development of drug resistant AML, and at the same time develop methods to identify and filter out donor variants. For the study we analyzed samples from patients who had relapsed after therapy (N=18) by exome sequencing. This included a set of patients where diagnosis and relapse samples were available (n=10), and one patient with diagnosis, remission and relapse samples. All patients had received prior chemotherapy and a subset had relapsed after receiving an allogeneic hematopoietic stem cell transplant (alloHSCT, n=6). Four patients had secondary AML that had developed after treatment for earlier hematologic malignancy. Tumor DNA was from bone marrow mononuclear cells and germline DNA from matched skin biopsies. Exome libraries were prepared then sequenced with the Illumina HiSeq instrument. Sequence data was processed and somatic variants identified as described previously (Koskela et al., NEJM, 2012). We identified relapse specific and relapse enriched somatic mutations by comparing mutation profiles of diagnosis and relapse samples. Donor derived germline variants in chimeric samples from patients relapsing after alloHSCT were identified with a bioinformatic methodology utilizing the dbSNP population variant database. Somatic mutations called from chimeric samples were filtered for common population variants present in the donor’s genome. Rare donor derived population variants that have not been previously described were identified as variants not present in the patient’s germline genome and which had similar tumor variant allele frequencies as the common donor derived variants. We estimated the level of chimerism based on the variant allele frequencies of all donor derived variants. In chimeric samples, the number of donor derived variants vastly exceeded the number of somatic mutations in AMLs (Fig 1). Donor cell content varied widely ranging from close to 100% in a post transplant remission sample to 10-40% in relapse samples. In post-transplant samples, we identified on average 6800 donor germline variants within the exome-capture regions, many of which occurred within cancer genes which could potentially be misinterpreted as driver mutations. Many recurrent driver mutations in cancer genes were identified in the relapse samples: FLT3 (n=6, 33%), DNMT3A (n=4, 22%), NPM1 (n=2, 11%), WT1 (n=2, 11%), TP53 (n=2, 11%), CBL (n=2, 11%), NRAS (n=1, 6%), KRAS (n=1, 6%), IDH1 (n=1, 6%), PHF6 (n=1, 6%) and PTPN11 (n=1, 6%). In several cases, we observed that relapse-specific driver mutations occurred in the same genes or pathways that already had initial mutations at diagnosis. For example, one patient’s AML had a FLT3-ITD at diagnosis; at relapse an activating mutation in CBL and a loss of function mutation in PTPN11 were acquired. Both CBL and PTPN11 act downstream of FLT3 (Fig 2). In two patients with a heterozygous WT1 mutation at diagnosis, we found additional WT1 mutations or deletion of the remaining wild type allele in the relapse sample, suggesting full loss of normal WT1 function contributes to disease progression. Our results suggest that AML progression and drug resistance may be caused by strengthening aberrant signaling through pathways already affected by a mutation present at diagnosis. Hence, the pattern of mutual exclusivity of mutations to genes affecting the same pathway, which has been observed in diagnostic samples, does not occur at relapse. On the contrary, in several cases the relapse specific mutations affected genes in pathways already affected at diagnosis. In addition, we show that donor derived germline variants can be identified and filtered from exome sequence data. Figure 1 Figure 1. Disclosures Porkka: BMS: Honoraria; BMS: Research Funding; Novartis: Honoraria; Novartis: Research Funding; Pfizer: Research Funding. Kallioniemi:Medisapiens: Consultancy, Membership on an entity's Board of Directors or advisory committees.

AMC-053: Pilot Study of an Oncolytic Viral Strategy Using Bortezomib with ICE +/- Rituximab for Relapsed/Refractory HIV+ Lymphomas

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 786-786 ◽  
Author(s):  
Erin Gourley Reid ◽  
David Looney ◽  
Frank Maldarelli ◽  
Ariela Noy ◽  
David H. Henry ◽  
...  
Keyword(s):  
Viral Load ◽  
Research Funding ◽  
B Cell Lymphoma ◽  
Proteasome Inhibition ◽  
Small Sample ◽  
Cell Transplant ◽  
Hematopoietic Stem ◽  
Viral Loads ◽  
Dose Levels ◽  
The Impact

Abstract Introduction: Both HIV+ Hodgkin and non-Hodgkin lymphomas have higher rates of latent infection by the gamma-herpesviruses (GHVs), Epstein-Barr virus (EBV) and Kaposi sarcoma-associated herpes virus (KSHV), than corresponding lymphomas in the HIV-seronegative population. Bortezomib, a proteasome inhibitor, induces lytic activation of both EBV and KSHV. Lytic activation of GHVs latently infecting lymphoma cells is hypothesized to beneficial both for direct tumor cell lysis as well as increased cytotoxic immune response due to viral lytic gene products. Furthermore, preclinical studies found proteasome inhibition impairs HIV infectivity via preservation of human anti-retroviral APOBEC3G, suggesting a novel therapeutic strategy to control HIV. Given that therapy of relapsed or refractory HIV-associated lymphoma (R/R-HAL) results in modest rates of remission, we sought to capitalize on the high viral association within HAL using an oncolytic strategy with bortezomib. Objectives: The primary objective of this study was to evaluate safety and overall response rate (ORR) of R/R-HAL to bortezomib combined with ifosfamide, carboplatin, etoposide +/- rituximab (ICE/R). The secondary objectives of this study were to estimate the impact of bortezomib on lytic activation of EBV and KSHV, using peripheral blood mononuclear cell (PBMC) viral loads, and on HIV using single copy plasma viral loads; to report overall survival at 1 year (1yr-OS); and to correlate EBV and KSHV viral load changes with lymphoma response. Methods: A 3+3 dose escalation design with a 7-day lead-in period of bortezomib alone prior to bortezomib + ICE/R allowed for assessment of early effects of bortezomib on viral loads. Bortezomib was given intravenously on day 1 and 8 of each cycle at one of 4 dose levels: 0.7, 1, 1.3 or 1.5 mg/m2. Standard dose ICE +/- R began day 8 of cycle 1 (28-day cycle); ICE +/- R began day 1 of all subsequent cycles (21-day cycle). Rituximab was included in the regimen only for CD20+ lymphoma. Binomial proportions were used to estimate ORR. The product-limit (Kaplan-Meier) method was used to estimate 1yr-OS. The Wilcoxon signed rank test was used to evaluate changes in viral loads. Results: Twenty-three subjects were enrolled from 7 sites within the AIDS Malignancy Consortium (AMC). More than 90% of enrolled subjects were men and half were minorities; at baseline, 20/23 were on antiretroviral therapy, median CD4 count was 315/µL and median HIV viral load was undetectable. Mean age was 50 years. Over half of subjects had stage IV HAL; the majority had diffuse large B-cell lymphoma (DLBCL) (n=15), 2 had primary effusion (PEL), 3 had plasmablastic and 2 had Hodgkin lymphoma. Figure 1 summarizes grade 3-4 toxicities of the 22 subjects evaluable for adverse events during the dose-limiting toxicity period (cycles 1+2). The maximum tolerated dose was not reached at the highest dose cohort studied (bortezomib 1.5mg/m2). Responses occurred in 14/22 subjects initiating protocol therapy: 5 complete and 9 partial responses (PR). Of the 20 subjects who completed 2 or more cycles, the ORR was 80%. Nine of the responders underwent auto-hematopoietic stem cell transplant after protocol therapy. 1yr-OS was 55%. After bortezomib alone, median values of EBV PBMC viral load measured on day 8 were 2x greater than baseline. However, paired analysis did not confirm significant change in this small sample (n=16 evaluable), and there was no correlation found between change in EBV viral load and response. The 2 subjects with known KSHV+ lymphoma (PEL) each had more than a 1-log increase in day 8 KSHV viral load compared with baseline. Both of these subjects attained a PR from protocol therapy. Conclusions: Addition of bortezomib to ICE/R in R/R HAL is feasible with ORR (80%) and 1yr-OS (55%) comparing very favorably with a prior AMC retrospective report of ICE/R in R/R HAL (n=31, ORR 32%, 1yr-OS 38%, Bayraktar 2012). Evaluation of data collected from individual subjects suggests GHV lytic activation may occur with bortezomib alone. However, the lower bortezomib dose levels used to treat the bulk of study subjects and the limited study sample size limit our power to confirm this conclusion. We plan to further explore the effects of proteasome inhibition on GHVs. Evaluation of the impact of bortezomib on HIV replication is pending and will be presented at the meeting. Disclosures Reid: Millennium: Research Funding. Sparano:Takeda: Research Funding.

scholarly journals Effect of Early Post-Transplantation Tacrolimus Concentration on the Risk of Acute Graft-Versus-Host Disease in Allogenic Stem Cell Transplantation

Cancers ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 613
Author(s):  
Nidhi Sharma ◽  
Qiuhong Zhao ◽  
Bin Ni ◽  
Patrick Elder ◽  
Marcin Puto ◽  
...  
Keyword(s):  
Stem Cell ◽  
Graft Versus Host Disease ◽  
Tacrolimus Concentration ◽  
Cell Transplant ◽  
Hematopoietic Stem ◽  
Host Disease ◽  
Graft Versus Host ◽  
The Impact ◽  
Risk Of Relapse ◽  
Acute Graft

Acute graft versus host disease (aGVHD) remains a leading cause of morbidity and mortality in allogeneic hematopoietic stem cell transplant (allo-HSCT). Tacrolimus (TAC), a calcineurin inhibitor that prevents T-cell activation, is commonly used as a GVHD prophylaxis. However, there is variability in the serum concentrations of TAC, and little is known on the impact of early TAC levels on aGVHD. We retrospectively analyzed 673 consecutive patients undergoing allo-HSCT at the Ohio State University between 2002 and 2016. Week 1 TAC was associated with a lower risk of aGVHD II–IV at TAC level ≥10.15 ng/mL (p = 0.03) compared to the lowest quartile. The cumulative incidence of relapse at 1, 3 and 5 years was 33%, 38% and 41%, respectively. TAC levels at week 2, ≥11.55 ng/mL, were associated with an increased risk of relapse (p = 0.01) compared to the lowest quartile. Subset analysis with acute myeloid leukemia and myelodysplastic syndrome patients showed significantly reduced aGVHD with TAC level ≥10.15 ng/mL at week 1 and a higher risk of relapse associated with week 2 TAC level ≥11.55 ng/mL (p = 0.02). Hence, achieving ≥10 ng/mL during the first week of HCT may mitigate the risk of aGVHD. However, levels (>11 ng/mL) beyond the first week may be associated with suppressed graft versus tumor effect and higher relapse.

scholarly journals Absence of Evidence Implicating Hematopoietic Stem Cells As Common Progenitors for DLBCL Mutations

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4107-4107
Author(s):  
Max Jan ◽  
Florian Scherer ◽  
David M. Kurtz ◽  
Aaron M Newman ◽  
Henning Stehr ◽  
...  
Keyword(s):  
Stem Cells ◽  
T Cells ◽  
B Cells ◽  
Hematopoietic Stem Cells ◽  
Research Funding ◽  
Phylogenetic Trees ◽  
Somatic Mutations ◽  
Hematopoietic Stem ◽  
Tumor Biopsies ◽  
Myd88 L265p

Abstract Background: Pre-leukemic hematopoietic stem cells (HSC) have been implicated in AML (Jan et al STM 2012) and also for several lymphoid leukemias including ALL, HCL, and CLL. Separately, relapse of ALL following CD19 CAR-T cell therapy has been associated with lymphomyeloid lineage switch. Finally, healthy persons with clonally expanded HSCs are at increased risk of hematologic malignancies including lymphomas, and in mouse DLBCL models we previously demonstrated the oncogenic sufficiency of BCL6 overexpression in HSC (Green et al 2014 Nat Comm). Nevertheless, the cellular origin of DLBCL in the majority of patients is not definitively known. We sought to investigate the presence of mutations found in DLBCL within matched HSCs. Methods: We deeply genotyped somatic mutations in diagnostic biopsy tissues of 16 patients with DLBCL using CAPP-Seq to a median sequencing depth of 1100x (Newman et al 2014 Nat Med; Scherer et al 2015 ASH). We then profiled each patient for evidence implicating HSCs using somatic mutation lineage tracing, in either direct or indirect fashion. For direct evaluation, we used highly purified, serially FACS-sorted HSCs from grossly uninvolved bone marrow (BM) (n=5; Fig 1a-b). For indirect assessment, we either profiled serial tumor biopsies (n=13), or interrogated sorted cells from terminally differentiated blood lineages (n=7), including peripheral CD3+ T cells, CD14+ Monocytes, and B cells expressing a light-chain discordant to that of tumor isotype. HSCs and differentiated lineages were then interrogated by direct genotyping, using 3 highly sensitive orthogonal quantitative methods, including Myd88 L265P droplet digital PCR (n=6), BCL6 translocation breakpoint qPCR (n=4), and DLBCL CAPP-Seq profiling of 268 genes (n=5). We used the theoretical limit of detection (LOD) genotyping performance for CAPP-Seq (0.001%, Newman et al 2016 Nat Biotech), and established analytical sensitivity of our custom MYD88 ddPCR via limiting dilution (~1%). These LODs met or exceeded the expected limit of sorting impurity by FACS (~1%). For 6 patients experiencing one or more DLBCL relapse, we deeply profiled 13 serial tumor biopsies by CAPP-Seq, and then assessed overlap in somatic mutations and VDJ sequences in biopsy pairs as additional indirect evidence implicating HSCs. Results: We obtained a median of ~2000 sorted HSCs and ~1700 sorted cells from differentiated lineages, and genotyped each population using one or more of the 3 direct genotyping methods described above. Three patients with sufficient cell numbers were profiled both by CAPP-Seq and either ddPCR (n=2) or qPCR (n=1). Surprisingly, we found no evidence implicating HSCs either directly or indirectly in any of the 16 patients, regardless of the assay employed or the cell types/lineages genotyped (e.g., Fig 1b). In 2 patients with MYD88 L265P mutations, we found evidence for MYD88+ B-cells with discordant light chains by ddPCR (~0.1%) potentially implicating common lymphoid precursors (CLPs), but found no evidence for similar involvement of T-cells or monocytes. In 6 DLBCL patients experiencing relapse, tumor pairs profiled by CAPP-Seq (median depth 957) shared 93% of somatic mutations (75-100%, Fig 1c). Such pairs invariably shared clonal IgH VDJ rearrangements (4/4, 100%), thus implicating a common progenitor arising in later stages of B-cell development, not HSCs. Conclusions: We find no evidence to implicate HSCs in the derivation of DLBCL. While formal demonstration of absence of pre-malignant HSCs in DLBCL would require overcoming practical and technical limitations (including number of available HSCs, sorting purity, and genotyping sensitivity), the pattern of shared somatic alterations at relapse makes this highly unlikely. We speculate that unlike lymphoid leukemias, the cell-of-origin for most DLBCLs reside later in B-lymphopoiesis, beyond CLPs. Figure. (a) HSC sorting from BM by FACS (b) Allele frequencies of mutations found by CAPP-Seq in an examplary DLBCL case (x-axis) compared to the same variants in HSCs (y-axis). (c) Phylogenetic trees of DLBCL patients experiencing relapse (n=6) with tumor pairs sequenced by CAPP-Seq. Shown are the evolutionary distances between (i) germline and common inferrable progenitor (CIP) illustrating the fraction of shared mutations between tumor pairs, and (ii) CIP and both diagnostic (tumor 1) and relapse tumors (tumor 2) indicating unique mutations to each tumor. Figure. (a) HSC sorting from BM by FACS (b) Allele frequencies of mutations found by CAPP-Seq in an examplary DLBCL case (x-axis) compared to the same variants in HSCs (y-axis). (c) Phylogenetic trees of DLBCL patients experiencing relapse (n=6) with tumor pairs sequenced by CAPP-Seq. Shown are the evolutionary distances between (i) germline and common inferrable progenitor (CIP) illustrating the fraction of shared mutations between tumor pairs, and (ii) CIP and both diagnostic (tumor 1) and relapse tumors (tumor 2) indicating unique mutations to each tumor. Disclosures Newman: Roche: Consultancy. Levy:Kite Pharma: Consultancy; Five Prime Therapeutics: Consultancy; Innate Pharma: Consultancy; Beigene: Consultancy; Corvus: Consultancy; Dynavax: Research Funding; Pharmacyclics: Research Funding. Diehn:Novartis: Consultancy; Quanticel Pharmaceuticals: Consultancy; Roche: Consultancy; Varian Medical Systems: Research Funding.

scholarly journals Molecular Analysis of Diamond Blackfan Anemia and Genotype-Phenotype Correlation: Experience from the Canadian Inherited Marrow Failure Registry

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3621-3621
Author(s):  
Omri Avraham Arbiv ◽  
Bozana Zlateska ◽  
Robert J. Klaassen ◽  
Conrad Fernandez ◽  
Rochelle Yanofsky ◽  
...  
Keyword(s):  
Ribosomal Protein ◽  
Research Funding ◽  
Gene Panel ◽  
Ribosomal Protein Genes ◽  
Severe Phenotype ◽  
Hematopoietic Stem ◽  
Diamond Blackfan Anemia ◽  
Genetic Groups ◽  
Number Of Patients ◽  
The Impact

Abstract Background/Objectives: Diamond Blackfan anemia (DBA) is an inherited disorder characterized by chronic hypoproductive anemia, physical malformations, and an increased risk of malignancies. At least 12 DBA genes have been identified, which include various ribosomal protein genes and the transcription factor GATA1. The aims of our study were (1) to identify the mutation spectrum of DBA patients, utilizing a cohort of patients enrolled on the Canadian Inherited Marrow Failure Registry (CIMFR) and (2) to determine whether specific hematological abnormalities, malformations, and outcomes are associated with specific mutations. Methods: Patients were enrolled on the CIMFR, which is a multicenter cohort study of inherited bone marrow failure syndromes (IBMFS). Genetic testing was performed using one or more of the following tests: Sanger sequencing, next generation sequencing (NGS) DBA gene panel, a comprehensive NGS IBMFS gene panel developed in our laboratory, or comparative genetic hybridization (CGH). Severity of the hematological disease was dichotomized according to a patient's requirement for chronic treatment: those who were maintained on corticosteroids, blood transfusions, or received a hematopoietic stem cell transplantation were considered to have a more severe phenotype than those who did not require hematological treatment. Chi-square tests with a Fisher's exact test correction were used to compare genetic groups with at least 5 patients on observed phenotypes. Results: 71 patients with DBA have been enrolled in our registry. A causal mutation has been identified in 36 of these patients, with the following rates: RPS19 (n=11), RPL11 (n=7), RPL5 (n=6), RPS26 (n=5), RPL35a (n=2), RPS24 (n=2), and one of each RPS7, RPS29, RPS17. Remarkably, a substantial number of patients in our population-based cohort (19.4%) had mild hematological phenotype requiring no therapy. Patients with RPL11 mutations tended to have a less severe DBA phenotype, while patients with RPS19 mutations tended to have a more severe phenotype (p=0.04). In terms of non-hematological malformations, we found no differences in cardiac, stature and craniofacial malformations across the groups compared (all p>0.1). However, patients with RPL5 mutations had significantly more hand malformations (p=0.02), and patients with RPS26 mutations had more genitourinary malformations (p=0.04). To control for the impact of mutation severity on the observed phenotype, we compared the prevalence of mutations that are predicted to result in truncated or lack of protein from the respective allele (large copy-number variation, nonsense, or indel frameshift) to mutations that are predicted to be hypomorphic or affect function (splicing, indel/inframe and, missense) between mutation categories. There were no differences among genetic groups in the severity of their mutations (p=0.58). Conclusions: Mutations in a wide spectrum of ribosomal protein genes underlie DBA cases in Canada, which approximate those observed by other registries in Western countries. Patients with DBA caused by RPL11 mutations tended to have a milder hematological phenotype, while patients with RPS19 mutation tended to have a more severe phenotype. Mutations in RPS26 and RPL5 are associated with genitourinary and hand malformations, respectively. Our findings may help improve counseling of DBA patients and their family. Future studies are needed to replicate our results and determine whether these findings can help personalize care. Disclosures Lipton: Ariad: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Teva: Consultancy, Research Funding; Bristol-Myers Squibb: Consultancy, Research Funding; Novartis Pharmaceuticals: Consultancy, Research Funding.

scholarly journals Immunosuppressive Compounds Affect the Fungal Growth and Viability of Defined Aspergillus Species

Pathogens ◽  
2019 ◽  
Vol 8 (4) ◽  
pp. 273 ◽  
Author(s):  
Stanislaw Schmidt ◽  
Michael Hogardt ◽  
Asuman Demir ◽  
Frauke Röger ◽  
Thomas Lehrnbecher
Keyword(s):  
Fungal Growth ◽  
Invasive Fungal Disease ◽  
Inhibitory Effect ◽  
Immunosuppressive Drugs ◽  
Aspergillus Species ◽  
Cell Transplant ◽  
Hematopoietic Stem ◽  
Number Of Patients ◽  
Species Specific ◽  
The Impact

Immunosuppressive drugs are administered to a number of patients; e.g., to allogeneic hematopoietic stem cell transplant recipients. Immunosuppressive drugs impair the immune system and thus increase the risk of invasive fungal disease, but may exhibit antifungal activity at the same time. We investigated the impact of various concentrations of three commonly used immunosuppressive compounds—cyclosporin A (CsA), methylprednisolone (mPRED), and mycophenolic acid (MPA)—on the growth and viability of five clinically important Aspergillus species. Methods included disc diffusion, optical density of mycelium, and viability assays such as XTT. MPA and CsA had a species-specific and dose-dependent inhibitory effect on the growth of all Aspergillus spp. tested, although growth inhibition by MPA was highest in A. niger, A. flavus and A. brasiliensis. Both agents exhibited species-specific hyphal damage, which was higher when the immunosuppressants were added to growing conidia than to mycelium. In contrast, mPRED increased the growth of A. niger, but had no major impact on the growth and viability of any of the other Aspergillus species tested. Our findings may help to better understand the interaction of drugs with Aspergillus species and ultimately may have an impact on individualizing immunosuppressive therapy.

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