Impact Of Variant Allele Frequency Research Articles

Prognostic Impact of Variant Allele Frequency of Myelodysplasia-Related Gene Mutations Among Patients Newly Diagnosed As Acute Myeloid Leukemia, Myelodysplasia-Related According to the 5th World Health Organization Classification

Introduction: Myelodysplasia-related gene mutations have been newly included as diagnostic criteria for acute myeloid leukemia, myelodysplasia-related (AML-MR) in the new 5th World Health Organization (WHO) classification. Myelodysplasia-related gene mutations are defined as somatic mutations of eight genes - SRSF2, SF3B1, U2AF1, ZRSR2, ASXL1, EZH2, BCOR, and STAG2. According to the new classification, some cases diagnosed as AML, not otherwise specified (NOS), which has no prognostic significance, can be revised as AML-MR, which has poor prognosis regardless of variant allele frequency (VAF) of myelodysplasia-related gene mutations. The prognostic impact of VAF of myelodysplasia-related gene mutations in the patients newly diagnosed as AML-MR is not well established. In this study, we evaluated the proportion of the patients who were formerly diagnosed as AML, NOS and can be newly diagnosed as AML-MR according to the new classification, and the prognostic impact of VAF of myelodysplasia-related gene mutations. Methods: From March 2019 to February 2023, we retrospectively reviewed the medical records of the patients newly diagnosed as AML, NOS according to the revised 4th edition of WHO classification. We also reviewed the results of mutation profiles detected by next generation sequencing for identifying myelodysplasia-related gene mutations. The limit of detection for each variant was VAF 2%. We categorized patients into two groups; AML, NOS with myelodysplasia-related gene mutations (AML, NOS, MR+) group who can be newly diagnosed as AML-MR according to the new classification and AML, NOS without myelodysplasia-related gene mutations (AML, NOS, MR-) group. For comparing the survival rates between two groups, we performed Kaplan-Meier curve analysis. We also classified AML, NOS, MR+ group according to the VAF; 2-10% and ≥10% of somatic mutations and compared the survival rate. Results: We reviewed 98 patients newly diagnosed as AML, NOS. Among them, 55.1%(54/98) of patients had myelodysplasia-related gene mutations. The distribution of myelodysplasia-related gene mutations in these patients were as follows; ASXL1 (50.0%, 27/54), U2AF1 (29.6%, 16/54), BCOR (25.9%, 14/54), SRSF2 (25.9%, 14/54), STAG2 (13.0%, 7/54), SF3B1 (9.3% 5/54), EZH2 (7.4%, 4/54), and ZRSR2 (3.7%, 2/54). The diagnosis of these patients can be revised as AML-MR according to the new classification. Among these patients, variants of myelodysplasia-related gene mutations with VAF 2-10% and ≥10% were detected in 7 and 47 patients, respectively. The survival rate of AML, NOS, MR+ group (2 year survival rate, 32.0%) was inferior than that of AML, NOS, MR- group (3 year survival rate 44.6%), although the difference was not statistically significant ( p=0.069). Considering VAF of somatic mutations, the survival rate of patients with AML, NOS with VAF ≥10% myelodysplasia-related gene mutations (N=47) was significantly inferior than that of patients with AML, NOS without or with VAF 2-10% myelodysplasia-related gene mutations (N=51) ( p=0.020) (Figure 1). Conclusions: More than half of the patients formerly diagnosed as AML, NOS can be re-diagnosed as AML-MR according to the 5th WHO classification. More than 10% of VAF of myelodysplasia-related gene mutations can be associated with worse survival rate among patients newly diagnosed as AML-MR. These results suggest that VAF of myelodysplasia-related gene mutations has prognostic impact and should be considered when predicting survival rate.

BRAFV600 variant allele frequency predicts outcome in metastatic melanoma patients treated with BRAF and MEK inhibitors.

The prognostic impact of variant allele frequency (VAF) on clinical outcome in BRAFV600 mutated metastatic melanoma patients (MMPs) receiving BRAF (BRAFi) and MEK inhibitors (MEKi) is unclear. A cohort of MMPs receiving first line BRAFi and MEKi was identified by inspecting dedicated databases of 3 Italian Melanoma Intergroup centers. VAF was determined by next generation sequencing in pre-treatment baseline tissue samples. Correlation between VAF and BRAF copy number variation (CNV) was analysed in an ancillary study by using a training and a validation cohort of melanoma tissue samples and cell lines. Overall, 107 MMPs were included in the study. The VAF cutoff determined by ROC curve was 41.3%. At multivariate analysis, progression free survival (PFS) was significantly shorter in patients with M1c/M1d [HR 2.25 (95% CI 1.41-3.6, p<0.01)], in those with VAF >41.3% [(HR 1.62 (95% CI 1.04-2.54, p<0.05)], and in those with ECOG PS ≥1 [(HR 1.82 (95% CI 1.15-2.88, p<0.05)]. Overall survival (OS) was significantly shorter in patients with M1c/M1d [(HR 2.01 (95% CI 1.25-3.25, p<0.01)]. Furthermore, OS was shorter in patients with VAF >41.3% (HR 1.46 (95% CI 0.93-2.29, p=0.06), and in patients with ECOG PS ≥1 [(HR 1.52 (95% CI 0.94-2.87, p=0.14)]. BRAF gene amplification was found in 11% and 7% of samples in the training and validation cohort, respectively. High VAF is an independent poor prognostic factor in MMP receiving BRAFi and MEKi. High VAF and BRAF amplification coexist in 7-11% of patients.

Predictive values of mutational variant allele frequency in overall survival and leukemic progression of myelodysplastic syndromes.

The implication of mutational variant allelic frequency (VAF) has been increasingly considered in the prognostic interpretation of molecular data in myeloid malignancies. However, the impact of VAF on outcomes of myelodysplastic syndromes (MDS) has not been extensively explored. Targeted next-generation sequencing was performed in 350 newly diagnosed MDS cases. The associations of mutational VAF of each gene with overall survival (OS) and leukemia-free survival (LFS) were examined by multivariate Cox regression after univariate analysis. Shorter OS was independently associated with DNMT3A VAF (HR 1.020 per 1% VAF increase; 95% CI 1.005-1.035; p = 0.011) and TP53 VAF (HR 1.014 per 1% VAF increase; 95% CI 1.006-1.022; p = 0.001). LFS analyses revealed that TET2 VAF (HR 1.013 per 1% VAF increase; 95% CI 1.005-1.022; p = 0.003) and TP53 VAF (HR 1.012 per 1% VAF increase; 95% CI 1.004-1.021; p = 0.005) were independently associated with faster leukemic transformation. Furthermore, we established nomograms to predict OS and LFS, respectively,by integrating independent mutational predictors into the revised International Prognostic Scoring System. Our study highlights that VAF of certain genes should be incorporated into routine clinical prognostication of survival and leukemic transformationof MDS.

The Impact of Variant Allele Frequency in EGFR Mutated NSCLC Patients on Targeted Therapy.

EGFR mutations represent the most common currently targetable oncogenic driver in non-small cell lung cancer. There has been tremendous progress in targeting this alteration over the course of the last decade, and third generation tyrosine kinase inhibitors offer previously unseen survival rates among these patients. Nonetheless, a better understanding is still needed, as roughly a third of patients do not respond to targeted therapy and there is an important heterogeneity among responders. Allelic frequency, or the variant EGFR allele frequency, corresponds to the fraction of sequencing reads harboring the mutation. The allelic fraction is influenced by the proportion of tumor cells in the sample, the presence of copy number alterations but also, most importantly, by the proportion of cells within the tumor that carry the mutation. Mutations that occur early in tumor evolution, often called clonal or truncal, have a higher allelic frequency than late, subclonal mutations, and are more often drivers of cancer evolution and attractive therapeutic targets. Most, but not all, EGFR mutations are clonal. Although an exact estimate of clonal proportion is hard to derive computationally, the allelic frequency is readily available to clinicians and could be a useful surrogate. We hypothesized that tumors with low allelic frequency of the EGFR mutation will respond less favorably to targeted treatment.

138P The impact of variant allele frequency in EGFR mutated NSCLC patients on targeted therapy

138P The impact of variant allele frequency in EGFR mutated NSCLC patients on targeted therapy

The prognostic impact of variant allele frequency (VAF) in TP53 mutant patients with MDS: A systematic review and meta-analysis.

Tumor protein p53 (TP53) is frequently expressed in patients with myelodysplastic syndromes (MDS). Studies have already reported the poor prognostic impact of TP53 gene mutations in MDS patients. However, parts of this subgroup of patients with low-risk MDS still have relatively better survival and longer remission times. Therefore, we performed a meta-analysis to evaluate the prognostic difference intra-gene of variant allele frequency (VAF). The primary endpoint was overall survival (OS), and event-free survival (EFS) was selected as the secondary endpoint. We extracted the hazard ratio (HR) and 95% confidence interval (CI) for OS and EFS from univariate and multivariate Cox proportional hazard models. A total of 4003 MDS patients and 1278 TP53-mutated patients from 13 cohorts of 11 studies up to February 24, 2020, were included in our meta-analysis. Pooled HRs suggested that a high mutant VAF had an adverse impact on OS (HR=2.11, 95% CI: 1.48-3.01, P<.0001) but no impact on EFS (HR=15.57, 95% CI: 0.75-324.44, P=.003) in MDS patients. Twenty percent is a proper threshold to set (HR=2.02, 95% CI: 1.31-3.13, P=.001) and is a rough line between high clone burden and low clone burden, while 40% is an exact cutoff point (HR=2.11, 95% CI: 1.26-3.55, P<.0001) to guide diagnosis and treatment. Beyond the traditional binary classification of gene mutation, we aimed to find a way to divide mutant molecular markers more specifically by VAF to provide clinical therapeutic values. Our meta-analysis indicates that a high VAF is an independent, adverse prognostic factor for OS in TP53 mutant MDS patients. Patients with mediate/low-frequency parts who could be treated like wide-type patients have relatively better survival and may choose allogeneic hematopoietic stem cell transplantation as conditions permitting. Further prospective studies are needed in the future, and a large subgroup analysis of the same cutoff point subgroups is needed to obtain a more reliable basis for the impact of other mutant gene VAFs on the prognosis of MDS.

JAK2V617F Variant Allele Frequency Identifies Patients with Polycythemia Vera (PV) at High Risk for Venous Thrombosis

Background. Cardiovascular (CV) events are leading cause of morbidity and mortality in PV. Current risk stratification is based on variables predicting thrombotic risk, ie age >60y and history of thrombosis. Recent studies focused on additional thrombotic risk factors in PV, including generic CV factors and leukocytosis. PV patients (pts) are JAK2V617F mutated, and present wide heterogeneity in variant allele frequency (VAF); it was shown that a VAF >75% was associated with higher number of thrombotic events after diagnosis (Vannucchi AM, Leukemia 2007), but the prognostic role of JAK2 VAF is still debated.Aim. The aim of the study was to evaluate the impact of JAK2V617F VAF on rate of thrombosis in WHO-2016 defined PV pts.Patients and Method. In the CRIMM (Florence) database, a total of 577 pts with a JAK2VF VAF determined within 3 years from diagnosis, who met the 2016 WHO criteria for PV, were identified. All pts had information regarding thromboembolic events, including history of thrombosis, occurrence, type and date of thrombosis in the follow-up (FU) and presence of CV risk factors (smoking, hypertension, and diabetes mellitus). Thrombosis‐free survival (TFS) was determined from the time of diagnosis to the time the first thrombotic event occurred. Pts in whom thrombosis did not occur were censored at the time of last FU. Pre-receiver operating characteristic (ROC) plots were used to determine cutoff levels for continuous variables of interest. Differences in the distribution of continuous variables between categories were analyzed by Mann-Whitney or Kruskal-Wallis test. Pts' groups with nominal variables were compared by χ2 test. TFS was estimated by Kaplan Meier analysis; log rank test was used to compare TFS difference between groups. Cox proportional hazards regression was used for multivariable analysis. A two tailed P ≤ 0.05 was considered statistically significant.Results. The median age of pts at diagnosis was 61y, 308 (53.4%) were above 60y; 57.2% were males. All pts were mutated for JAK2V617F with a median VAF 43% (range 1-100%), 62% had at least one CV risk factor; 83 (14.4%) pts suffered from an episode of thrombosis within 3 yr from, or coincident with, diagnosis. The median FU was 7.3y (0.6-35.9y) during which 87 pts (15.1%) developed thrombosis. (50 arterial and 45 venous thrombosis). During the FU, 110 pts (19.1%) died.A JAK2VAF of ≥60% cutoff level, as determined by ROC analysis, correlated with measurements of stimulated erythropoiesis (higher hematocrit, lower mean cell volume and serum ferritin; all P<.01), leukocytosis (P<.0001), lower platelets count (P=.02) and elevated serum lactate dehydrogenase (LDH) (P<.03). Pts with ≥60% JAK2V617F VAF were at higher relative risk (RR) of having splenomegaly (RR 3.1; P<0.001), suffering from pruritus (RR 2.5; P<0.001) or constitutional symptoms (RR 1.9; P=0.01), harboring a BM fibrosis grade-1 (RR 3.1; P<0.001). Additionally, pts with a VAF>60% had greater risk to progress to PPV-MF (RR 8.5, P<.0001) and acute leukemia (RR 4.4, P=0.04) or to die (RR 3.8, P<0.0001). The JAK2VF VAF (continuous or ≥60%) did not correlate with occurrence of thrombosis at diagnosis, while the rate of thrombosis during FU was significantly increased in pts with VAF ≥60% (23.4% vs 11.0%, RR 2.4, 95%CI = 1.4-4.0; P<0.0001), more marked for venous (RR 3.7, 95%CI = 2.0-6.8; P<0.0001) than arterial (RR 1.8, 95%CI = 0.9-3.3; P=0.05) thrombosis. The impact of VAF on thrombosis during FU was then estimated according to the conventional risk category. In low risk pts (LR) (n=236), factors significantly associated with occurrence of FU thrombosis were CV risk factors (dyslipidemia (RR 3.3, P = 0.02) and hypertension (RR 1.8, P=0.048)), a G1 BM fibrosis (RR 5.3, P=0.006), presence of splenomegaly (RR, 3.2, P=0.001) or constitutional symptoms (RR 3.3, P=0.003) and a VAF ≥60% (RR 2.2, P = 0.024). In high risk pts (HR) (n=341), factors significant for FU thrombosis were splenomegaly (RR 2.0, P=0.03), elevated LDH (RR 4.0, P=0.009) and a VAF ≥60% (RR 2.3, P = 0.012). A VAF ≥60% was correlated with shortened venous TFS after diagnosis in HR (P = 0.01, HR = 3.2, 95%CI = 1.2−8.3; fig.) but not in LR pts (P = 0.20, HR = 1.1, 95%CI = 0.5−2.9).Conclusions. This study indicates that conventionally-defined high-risk PV pts with a JAK2V617F VAF ≥60% suffer from increased rate of venous events and might be worthwhile of more intensive antithrombotic prophylaxis. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Impact of variant allele frequency of mutant PTPN11 in AML: Single institution experience of 122 patients.

7046Background: Mutations in PTPN11, encoding tyrosine phosphatase SHP2, are present in 4-6% of AML. Largest report includes 27 patients (pts). Methods: Analysis included AML patients (pts) treated...

70 - Prognostic Impact of Variant Allelic Frequency of Molecular Mutations in Acute Myeloid Leukemia (AML) and Myelodysplastic Syndromes (MDS) on Allogeneic Hematopoietic Cell Transplant (HCT) Outcomes

70 - Prognostic Impact of Variant Allelic Frequency of Molecular Mutations in Acute Myeloid Leukemia (AML) and Myelodysplastic Syndromes (MDS) on Allogeneic Hematopoietic Cell Transplant (HCT) Outcomes