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. 2024 Jun 15:15:20406207241257904.
doi: 10.1177/20406207241257904. eCollection 2024.

Integrated genetic, epigenetic, and immune landscape of TP53 mutant AML and higher risk MDS treated with azacitidine

Amer M Zeidan  1 , Jan Philipp Bewersdorf  2   3 , Vanessa Hasle  4 , Rory M Shallis  2 , Ethan Thompson  4 , Daniel Lopes de Menezes  4 , Shelonidta Rose  4 , Isaac Boss  4 , Stephanie Halene  2 , Torsten Haferlach  5 , Brian A Fox  4
Affiliations

Integrated genetic, epigenetic, and immune landscape of TP53 mutant AML and higher risk MDS treated with azacitidine

Amer M Zeidan et al. Ther Adv Hematol. .

Abstract

Background: TP53 mutations are associated with an adverse prognosis in acute myeloid leukemia (AML) and higher-risk myelodysplastic syndromes (HR-MDS). However, the integrated genetic, epigenetic, and immunologic landscape of TP53-mutated AML/HR-MDS is not well defined.

Objectives: To define the genetic, epigenetic, and immunologic landscape of TP53-mutant and TP53 wild-type AML and HR-MDS patients.

Design: Post hoc analysis of TP53-mutant and TP53 wild-type patients treated on the randomized FUSION trial with azacitidine ± the anti-PD-L1 antibody durvalumab.

Methods: We performed extensive molecular, epigenetic, and immunologic assays on a well-annotated clinical trial dataset of 61 patients with TP53-mutated disease (37 AML, 24 MDS) and 144 TP53 wild-type (89 AML, 55 MDS) patients, all of whom received azacitidine-based therapy. A 38 gene-targeted myeloid mutation analysis from screening bone marrow (BM) was performed. DNA methylation arrays, immunophenotyping and immune checkpoint expression by flow cytometry, and gene expression profiles by bulk RNA sequencing were assessed at baseline and serially during the trial.

Results: Global DNA methylation from peripheral blood was independent of TP53 mutation and allelic status. AZA therapy led to a statistically significant decrease in global DNA methylation scores independent of TP53 mutation status. In BM from TP53-mutant patients, we found both a higher T-cell population and upregulation of inhibitory immune checkpoint proteins such as PD-L1 compared to TP53 wild-type. RNA sequencing analyses revealed higher expression of the myeloid immune checkpoint gene LILRB3 in TP53-mutant samples suggesting a novel therapeutic target.

Conclusion: This integrated analysis of the genetic, epigenetic, and immunophenotypic landscape of TP53 mutant AML/HR-MDS suggests that differences in the immune landscape resulting in an immunosuppressive microenvironment rather than epigenetic differences contribute to the poor prognosis of TP53-mutant AML/HR-MDS with mono- or multihit TP53 mutation status.

Trial registration: FUSION trial (NCT02775903).

Keywords: AML; MDS; TP53 mutation; gene expression; immune phenotype.

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Conflict of interest statement

AMZ received research funding (institutional) from Celgene/BMS, Abbvie, Astex, Pfizer, Medimmune/AstraZeneca, Boehringer-Ingelheim, Cardiff oncology, Incyte, Takeda, Novartis, Aprea, and ADC Therapeutics. AMZ participated in advisory boards, and/or had a consultancy with and received honoraria from AbbVie, Otsuka, Pfizer, Celgene/BMS, Jazz, Incyte, Agios, Boehringer-Ingelheim, Novartis, Acceleron, Astellas, Daiichi Sankyo, Cardinal Health, Taiho, Seattle Genetics, BeyondSpring, Cardiff Oncology, Takeda, Ionis, Amgen, Janssen, Epizyme, Syndax, Gilead, Kura, Chiesi, ALX Oncology, BioCryst, Notable, Orum, and Tyme. AMZ served on clinical trial committees for Novartis, Abbvie, Gilead, BioCryst, Abbvie, ALX Oncology, Geron, and Celgene/BMS. RMS participated in advisory boards, and/or had a consultancy with and received honoraria from Bristol Myers Squibb and Gilead Sciences, Inc.; divested equity interest in Curis Oncology. IB, ET, DLM, BF, VH, and SR are employees of Celgene, a Bristol Myers Squibb company, and may be shareholders. SH has received research funding received honoraria from FORMA Therapeutics. TH is the owner of Munich Leukemia Laboratory. All other authors have no relevant conflicts of interest to declare.

Figures

Figure 1.
Figure 1.
TP53 and co-occurring mutations. Panel (a) Illustrates the distribution and type of TP53 mutations as a lollipop plot. The majority of TP53 mutations were missense mutations localized in the p53 DNA-binding domain. Patients with TP53 mutations had a lower number of co-occurring mutations compared to TP53 wild-type patients (b; the central line shows the median, the ends of the boxes show the interquartile range (IQR) and the whiskers show the most distant point ⩽1.5 times the IQR). TP53 mutations occurred in isolation in 41.7% of patients, while TP53 wild-type patients frequently had several co-occurring mutations (c). The mutational and cytogenetic patterns by TP53 mutational status are shown in panel (d).
Figure 2.
Figure 2.
Baseline global DNA methylation status and immune phenotype by TP53 mutation status. Panel (a) shows the global DNA methylation score at baseline without statistically significant differences between TP53 wild-type, monohit or multihit TP53-mutant patients. Global DNA methylation score is shown as box-and-whisker plots with median and interquartile range. Distribution by TP53 mutation status was compared using Wilcoxon rank sum test. Figure panels (b–d) show flow cytometry from baseline bone marrow samples for AML patients (b), HR-MDS patients (c), and the combined patient cohort (d) by TP53 mutation status, respectively. Box-and-whisker plots show the median and interquartile range. Distribution by TP53 mutation status was compared using Wilcoxon rank sum test. AML, acute myeloid leukemia; HR-MDS, higher-risk myelodysplastic syndromes.
Figure 3.
Figure 3.
Gene expression pattern by TP53 mutation status in screening samples. Figure shows genes with statistically significant RNA expression levels with higher expression in TP53-mutant patients. TP53-mutant patients had higher expression of T-cell genes (e.g. IL7R), markers of proliferation (MKI-67), PD-L1 (CD274) expression compared to wild-type. Differences in gene expression between TP53-mutant and TP53 wild-type samples applied to both AML and HR-MDS patient cohorts. Box-and-whisker plots show the median and interquartile range. Distribution by TP53 mutation status was compared using Wilcoxon rank sum test. AML, acute myeloid leukemia; HR-MDS, higher-risk myelodysplastic syndromes.
Figure 4.
Figure 4.
Co-expression modules colored by differential expression in TP53 mutant versus wild-type and projected onto scRNA-seq data from van Galen et al. Figure shows the largest 53 co-expression modules from the bulk RNA-seq baseline samples and their logFC in TP53-mutant versus TP53 wild-type and expression in clusters of single cells. In panel (a), each node represents 25–57 genes which are co-expressed and labeled with the most central member of the module. Edges in the network are drawn if the central member has a Pearson correlation greater than or equal to 0.7 to any other node, and the thickness of the line is scaled to the correlation value. The color of each node is the log2 fold change of TP53 mutant versus TP53 wild-type with red showing modules which are higher in TP53-mutated samples and blue showing the opposite. Communities are shown by circles and defined by the graph structure. The circle labeled ‘singles’ is a collection of singleton modules, which are not connected to any other modules above a correlation of 0.7. The labels of the remaining communities are from (b), which is a heatmap showing the expression of each co-expression module from (a) in the cell types from the AML and healthy BM scRNA-seq data from van Galen et al. (GSE116256). The color of each element in the heatmap is based on the scaled mean of the expression of the genes within each module from (a), and the community names are based on the top cell type labels. AML, acute myeloid leukemia; BM, bone marrow; scRNA-seq, single-cell RNA sequencing.
Figure 5.
Figure 5.
Serial assessment of global DNA methylation status by TP53 mutation status. Figure shows the global DNA methylation score at baseline and after one cycle of treatment with azacitidine ± durvalumab for all patients (a). Patients experienced a decline in global DNA methylation scores independent of TP53 mutation status (b) without statistically significant differences between TP53 wild-type, monohit or multihit TP53-mutant patients neither at baseline nor after one cycle of treatment (c). This supports that HMAs exert an objective pharmacodynamic effect in patients independent of TP53 mutation status. Global DNA methylation score is shown as box-and-whisker plots with median and interquartile range. Distribution by TP53 mutation status was compared using Wilcoxon rank sum test. HMA, hypomethylating agent.
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