The Impact of Molecular and Cytogenetic Clonal Evolution on Survival in Relapsed/Refractory AML

Background: Clonal diversity and evolution in acute myeloid leukemia (AML) are consequences of disease progression and play a pivotal role in the development of drug resistance and refractoriness to therapy. Much work has been done with characterizing clonal architecture in bulk tumors samples or single-cell DNA sequencing, but real-world data observing the impact of clonal evolution and responses to therapy remains scarce in the relapsed and refractory settings. Therefore, the purpose of this study was to analyze the impact of observed clonal evolution on survival. Patients & Methods: We retrospectively analyzed 81 patients with AML with relapsed or refractory disease from June 2018 to December 2020. Cytogenetic and molecular data were obtained at the time of diagnosis and the time of primary induction failure or relapse, when available. We identified a total of 24 patients that demonstrated evidence of clonal evolution following induction or salvage therapy. Baseline patient demographics were obtained, including cytogenetic risk and next-generation sequencing molecular profiling at diagnosis and throughout treatment alongside dates and types of induction regimens. We compared the survival of those with evidence of clonal evolution to matched groups without. The groups with and without clonal evolution were compared for baseline statistical differences using an unpaired t-test with Welch's correction and Kaplan-Meier survival analyses were computed and compared with log-rank tests using GraphPad Prism. The event for calculating the overall survival was the date of death with patients otherwise censored at the date of last contact. Results: Of the 24 patients with evidence of clonal evolution, we detected that evolution occurred in four (16.7%) patients at the time of primary induction failure (refractory to first induction), 19 (70.8%) at the time of first relapse, and 3 (12.5%) at the time of second relapse. The median age of patients with clonal evolution was 63 years (range: 23 - 74) with 17 (70.8%) males and 7 (29.2%) females. The most commonly occurring mutations were NPM1 (25.0%), FLT3-ITD (20.8%), and TP53 (20.8%). At the time of initial diagnosis, cytogenetic risk was favorable in one (4.2%), intermediate in 7 (29.2%), and adverse in 16 (66.7%). The majority (79.2%) of patients underwent first induction with 7+3 and 8 (33.3%) patients underwent allogenic SCT. At the time of clonal evolution and ignoring the adverse prognosis associated with relapse, one patient with favorable cytogenetics remained favorable, only one (14.3%) patient with intermediate cytogenetics shifted to adverse, and one (6.3%) patient in the adverse category shifted to intermediate by ELN criteria. Three (12.5%) patients out of 24 acquired FLT3-ITD and three (12.5%) acquired an additional cooperating mutation (NRAS, KRAS, or KIT) for a total of 25.0% acquiring a signaling pathway mutation. Out of 10 patients with either mutated TP53 or MLL, three (27.3%) acquired a cooperating mutation in FLT3 or RAS. We then compared the clonal evolution cohort with the 57 remaining relapsed/refractory patients without evidence of clonal evolution. There was no difference between the groups with respect to baseline characteristics, including age (p = 0.989), ECOG status at diagnosis (p = 0.4689), Charlson Comorbidity Index (CCI) score (p = 0.6454), or prior SCT (p = >0.999). The median overall survival (OS) of the clonal evolution group was 227 days and compared to 382 days in the non-evolution cohort (p = 0.843). Conclusion: We did not detect a significant OS difference between those with clonal evolution and those without. Independent of the adverse prognosis associated with relapsed or refractory disease, cytogenetic risk category appeared to remain the same with clonal evolution. Common trends during clonal evolution included acquisition of one or several cooperating mutations in FLT3 or RAS, both with and without the presence of mutated TP53 and MLL. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

NUP-98 Rearrangements Led to the Identification of Candidate Biomarkers for Primary Induction Failure in Pediatric Acute Myeloid Leukemia

Conventional chemotherapy for acute myeloid leukemia regimens generally encompass an intensive induction phase, in order to achieve a morphological remission in terms of bone marrow blasts (<5%). The majority of cases are classified as Primary Induction Response (PIR); unfortunately, 15% of children do not achieve remission and are defined Primary Induction Failure (PIF). This study aims to characterize the gene expression profile of PIF in children with Acute Myeloid Leukemia (AML), in order to detect molecular pathways dysfunctions and identify potential biomarkers. Given that NUP98-rearrangements are enriched in PIF-AML patients, we investigated the association of NUP98-driven genes in primary chemoresistance. Therefore, 85 expression arrays, deposited on GEO database, and 358 RNAseq AML samples, from TARGET program, were analyzed for “Differentially Expressed Genes” (DEGs) between NUP98+ and NUP98-, identifying 110 highly confident NUP98/PIF-associated DEGs. We confirmed, by qRT-PCR, the overexpression of nine DEGs, selected on the bases of the diagnostic accuracy, in a local cohort of PIF patients: SPINK2, TMA7, SPCS2, CDCP1, CAPZA1, FGFR1OP2, MAN1A2, NT5C3A and SRP54. In conclusion, the integrated analysis of NUP98 mutational analysis and transcriptome profiles allowed the identification of novel putative biomarkers for the prediction of PIF in AML.

Calibration of LgrassFlo, a new model of perennial grass phenology in response to temperature and photoperiod.

&lt;p&gt;In perennial grasses, the reproductive development encompasses several phenological events, such as apex induction, floral transition, heading and flowering, that deeply affect biomass production, forage quality and plant perenniality. Despite the importance of perennial grasses in agricultural systems and natural ecosystems, we still lack accurate models predicting the reproductive development and its consequences on plant growth and grassland management. Most of available models implements a fixed scheduling of the reproductive development expressed either in thermal time or in calendar time. The progressive completion of floral induction and the effects of environmental factors are generally poorly described. In addition, the vegetative and reproductive developments are represented as independent and successive phases. In the present work, we introduce the new model LgrassFlo, which simulates the reproductive development of perennial grasses in interaction with plant vegetative development and considering the effects of environmental conditions on floral induction.&lt;/p&gt;&lt;p&gt;LgrassFlo simulates the canopy as the dynamics of a collection of individual plants, each being composed of one or more tillers. The 3D description of leaf growth and tillering is based on a functional-structural plant model of perennial ryegrass (Lgrass). We developed a new model of floral induction describing the progression of the primary and secondary induction of each apex of the plant according to (i) the daily temperature, (ii) photoperiod and (iii) plant architecture. This model was coupled to Lgrass, the model ensemble being called LgrassFlo. During apex induction, LgrassFlo accounts for an increase in the rates of leaf primordia initiation and leaf elongation. After floral transition, we assume that the apex only initiates spikelet primordia and that internodes start to elongate. LgrassFlo simulates the date of floral transition, the final number of leaves and the heading date based on a 3D representation of plant architecture.&lt;/p&gt;&lt;p&gt;A specific experiment was carried out in order to calibrate LgrassFlo on data describing the vegetative and reproductive development of three &lt;em&gt;Lolium perenne&lt;/em&gt; cultivars contrasted for their precocity and exposed to four inductive conditions in growth chambers. The first three conditions consisted in a period allowing for primary induction (low temperature &amp;#8211; short day) followed by a period allowing for secondary induction (high temperature &amp;#8211; long day), the two periods being spaced by a non-inductive period (high temperature - short day) of 0, 3 or 6 weeks. In the fourth condition, plants were not exposed to conditions allowing for the primary induction. A set of vegetative and reproductive parameters were estimated for each individual plant of the experiment. The parameter values were independent of the experimental treatment but showed a large genetic diversity both between and within varieties. Using this calibration, LgrassFlo satisfactorily predicted the observed diversity in final leaf number and heading date.&lt;/p&gt;&lt;p&gt;The present model is a step forward towards a better prediction of perennial grass phenology in actual and future climatic conditions. In this respect, the model is being currently used to simulate the observed genetic diversity in the heading date of several Lolium perenne cultivars grown in contrasted temperate climates over the last 15 years.&lt;/p&gt;

292 Emergency airway management outside the operating room; a three year prospective service evaluation and quality improvement project

Aims/Objectives/BackgroundEmergency airway management outside a controlled theatre environment has been previously associated with a high rate of adverse events. Several initiatives to improve safety (such as video laryngoscopy, checklists, simulation training etc..) have been studied in isolation.It remains unclear as to whether these interventions have been embedded in the Emergency Department (ED) and whether they offer cumulative marginal gains in safety.Methods/DesignA prospective 3-year service evaluation delivered at a major trauma and neurosciences centre between 2016 and 2019. We designed a rolling quality improvement program to mitigate procedural airway risk through collaborative multidisciplinary team (MDT) working, education and transparent metrics.PDSA cycles included documentary guidance (including flowcharts and checklists), high fidelity simulation training, equipment redesign, prefilled medications and mandatory reporting items (figure 1).Abstract 292 Figure 1Abstract 292 Figure 2Primary induction agents selected throughout the study periodAbstract 292 Figure 3Results/ConclusionsWe analysed prospectively collected data on 1181 intubation episodes outside a theatre environment over a 39 month period, of which 575 (48.7%) were performed out of hours and 635 (53.8%) were performed in the ED.Bedside consultant presence and periprocedural checklist use both showed a sustained increase during the study period. Use of ketamine and thiopentone as primary induction agents increased and decreased, respectively (figure 2). Cricoid pressure and video laryngoscopy (VL) utilisation rates remained relatively static throughout, as did a first pass success (FPS) rate of between 83.0 to 93.5%.Composite major complications (including sustained hypotension and/or critical hypoxia) were significantly reduced during the study period, as demonstrated via statistical process chart (SPC) mapping (figure 3).In conclusion, we found a quality improvement program to be associated with a sustained reduction in the risk of major complications following emergency airway management. This improvement was not explained by simple direct changes in procedural care, such as the use of VL or technique changes resulting in improved FPS, but may have been influenced by unknown confounding variables.

TP53 abnormalities correlate with immune infiltration and associate with response to flotetuzumab immunotherapy in AML

Somatic TP53 mutations and 17p deletions with genomic loss of TP53 occur in 37% to 46% of acute myeloid leukemia (AML) with adverse-risk cytogenetics and correlate with primary induction failure, high risk of relapse, and dismal prognosis. Herein, we aimed to characterize the immune landscape of TP53-mutated AML and determine whether TP53 abnormalities identify a patient subgroup that may benefit from immunotherapy with flotetuzumab, an investigational CD123 × CD3 bispecific dual-affinity retargeting antibody (DART) molecule. The NanoString PanCancer IO360 assay was used to profile 64 diagnostic bone marrow (BM) samples from patients with TP53-mutated (n = 42) and TP53-wild-type (TP53-WT) AML (n = 22) and 45 BM samples from patients who received flotetuzumab for relapsed/refractory (R/R) AML (15 cases with TP53 mutations and/or 17p deletion). The comparison between TP53-mutated and TP53-WT primary BM samples showed higher expression of IFNG, FOXP3, immune checkpoints, markers of immune senescence, and phosphatidylinositol 3-kinase-Akt and NF-κB signaling intermediates in the former cohort and allowed the discovery of a 34-gene immune classifier prognostic for survival in independent validation series. Finally, 7 out of 15 patients (47%) with R/R AML and TP53 abnormalities showed complete responses to flotetuzumab (&lt;5% BM blasts) on the CP-MGD006-01 clinical trial (NCT #02152956) and had significantly higher tumor inflammation signature, FOXP3, CD8, inflammatory chemokine, and PD1 gene expression scores at baseline compared with nonresponders. Patients with TP53 abnormalities who achieved a complete response experienced prolonged survival (median, 10.3 months; range, 3.3-21.3 months). These results encourage further study of flotetuzumab immunotherapy in patients with TP53-mutated AML.