Impact of Immunophenotype and Cytogenetics in Early Assessment of Post Induction Response in Acute Myeloid Leukemia (AML)
Authors: Schneidewend Robert , Carlson Karen B. , Atallah Ehab , Michaelis Laura C. , Hosking Paul R. , Hari Parameswaran , Hamadani Mehdi , D'Souza Anita , Fenske Timothy S. , Pasquini Marcelo C. , Chitambar Christopher R. , Raj Renju V. , Johnston Kimberli , Essenmacher Amber , Zook Felicia 
Published: November 2019

Abstract
OBJECTIVE: The goal of this study was to determine whether flow cytometry and genetic estimation of blast populations in bone marrow samples from adult patients undergoing induction chemotherapy for AML were consistent with morphologic evaluation.
BACKGROUND: Early post induction evaluation is paramount to assess the need for additional treatment in patients with AML. Yet, low bone marrow cellularity lends uncertainty to evaluation on the level of residual disease. This study evaluates whether standard addition of multi-color flow cytometry and karyotype analysis by fluorescent in situ hybridization (FISH) examination can assist in determining residual disease and guide further therapy
METHODS: Fifty newly diagnosed adult patients with AML who received seven days of continuous infusion cytarabine and three days of anthracycline ('7+3') at a single institution between February, 2013 and March, 2015 were identified for this analysis. Patients were divided into three groups according to day-14 bone marrow biopsy evaluation: 1) those with ² 5% blasts; 2) those with ³ 5% blasts, and bone marrow cellularity > 20%; and 3) those with ³ 5% blasts, and bone marrow cellularity ² 20%. Results were considered concordant if the parameters being compared were both <5% or >5%.
Table 1. Patient Characteristics Number of Patients n = 50 Median Age 58.5 years (range 19-73 years) Male: Female 1.27:1 Risk stratification by ELN Low = 10 (20%) Intermediate-1 = 4 (8%) Intermediate-2 = 14 (28%) High Risk = 17 (34%) Indeterminate = 5 (10%) Antecedent MDS n = 14, 28% Therapy-related AML n = 2, 4% FLT-3 mutation (ITD or TKD) n = 8, 16% Overall response 56% CR, 18% CRi, 26% Induction Failure
RESULTS: Patient characteristics are described in Table 1. Informative results for FISH and flow cytometry were available in 56% and 86% of cases, respectively. Results from bone marrow evaluation of each subgroup are detailed in Table 2. Bone marrow interpretation based on flow cytometry versus morphologic estimation of myeloblasts was consistent in 60% (n = 26 of 43) of all mid-cycle samples. Immunophenotype demonstrated lower disease burden in the discordant cases (N=17). Among patients with chromosomal abnormalities (N=28), bone marrow myeloblast percentage based on FISH was consistent with morphologic evaluation in 60% ofsamples. FISH demonstrated higher disease burden in all discordant cases (N=11).
Table 2. Bone Marrow Evaluation ² 5% blasts ³ 5% blasts, and > 20% cellular > 5% blasts,and ² 20% cellular n, % total n = 26, 52% n = 6, 12% n = 18, 36% Mean age 53.46 63 51.5 European Leukemia Net (ELN) Risk-stratification Low = 8 Intermediate-1 = 2 Intermediate-2 = 5 High Risk = 8 Indeterminate = 3 Low = 0 Intermediate-1 = 0 Intermediate-2 = 2 High Risk = 2 Indeterminate = 2 Low = 2 Intermediate-1 = 2 Intermediate-2 = 6 High Risk = 7 Indeterminate = 1 Response to induction, all cycles CR = 16/26 = 62% CRi = 6/26 = 23% CR = 2/6 = 33% CRi = 1/6 = 17% CR = 10/18 = 56% CRi = 2/18 = 11% Consistent interpretation based on morphology and flow cytometry 22/23 = 96% 1/5 = 20% 3/15 = 20% Consistent interpretation based on morphology and cytogenetic/FISH analysis 10/14 = 71% 4/4 = 100% 3/10 = 30%
CONCLUSIONS:
Our data identifies a significant proportion of patients for whom the decision to administer a 2nd cycle of 7+3 chemotherapy is unclear based on day-14 bone marrow evaluation (i.e., > 5% myeloblasts, and ² 20% cellular). Standard immunophenotyping at day 14 did not assist interpretation of chemotherapy response, possibly because of sample hemodilution. However, when available, karyotyping with FISH demonstrated a better assessment of disease burden. Although the number of samples was too small to correlate with therapy response, our data suggests that future studies to quantify the leukemic clone by genetic analysis and correlate these results with clinical outcome may improve prognostication based on mid-cycle bone marrow responsiveness to chemotherapy.

Disclosures
Michaelis: Incyte: Membership on an entity's Board of Directors or advisory committees; CTI Biopharma: Membership on an entity's Board of Directors or advisory committees; Wyeth: Membership on an entity's Board of Directors or advisory committees; Pfizer: Equity Ownership. Hari:Celgene: Consultancy; Takeda: Consultancy; BMS: Consultancy; Janssen: Consultancy; Novartis: Consultancy; Spectrum: Consultancy; Sanofi: Consultancy. Hamadani:MedImmune: Consultancy; Cellerant: Consultancy; Takeda: Research Funding; Celgene: Consultancy. Fenske:Pharmacyclics: Honoraria; Millennium/Takeda: Research Funding; Celgene: Honoraria; Seattle Genetics: Honoraria.