Acute myeloid leukemia (AML) is a molecularly and clinically heterogeneous hematological malignancy. Although immunotherapy may be an attractive modality to exploit in patients with AML, the ability to predict the groups of patients and the types of cancer that will respond to immune targeting remains limited. This study dissected the complexity of the immune architecture of AML at high resolution and assessed its influence on therapeutic response. Using 442 primary bone marrow samples from three independent cohorts of children and adults with AML, we defined immune-infiltrated and immune-depleted disease classes and revealed critical differences in immune gene expression across age groups and molecular disease subtypes. Interferon (IFN)–γ–related mRNA profiles were predictive for both chemotherapy resistance and response of primary refractory/relapsed AML to flotetuzumab immunotherapy. Our compendium of microenvironmental gene and protein profiles provides insights into the immuno-biology of AML and could inform the delivery of personalized immunotherapies to IFN-γ–dominant AML subtypes.

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 (<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.

BackgroundThe PAM50-based (Prosigna) risk of recurrence (ROR) score and intrinsic subtypes are prognostic for women with high-risk breast cancer. We investigate the predictive ability of Prosigna regarding the effectiveness of cyclophosphamide-based adjuvant chemotherapy in premenopausal patients with high-risk breast cancer.MethodsProsigna assays were performed on the NanoString platform in tumors from participants in Danish Breast Cancer Group (DBCG) 77B, a four-arm trial that randomized premenopausal women with high-risk early breast cancer to no systemic treatment, levamisole, oral cyclophosphamide (C) or cyclophosphamide, methotrexate and fluorouracil (CMF).ResultsIn total, this retrospective analysis included 460 women (40% of the 1146 randomized patients). The continuous Prosigna ROR score was prognostic in the no systemic treatment group (unadjusted P < 0.001 for disease-free survival (DFS), P = 0.001 for overall survival (OS)). No statistically significant interaction of continuous ROR score and treatment on DFS and OS was found. A highly significant association was observed between intrinsic subtypes and C/CMF treatment for DFS (Pinteraction = 0.003 unadjusted, P = 0.001 adjusted) and OS (Pinteraction = 0.04). In the adjusted analysis treatment with C/CMF was associated with a reduced risk of DFS events in patients with basal-like (hazard ratio (HR) 0.14; 95% CI 0.06; 0.32) and luminal B (HR 0.48; 95% CI 0.27; 0.84) subtypes but not in patients with Human epidermal growth factor receptor-enriched (HR 1.05; 95% CI 0.56; 1.95) or luminal A (HR 0.61; 95% CI 0.32; 1.16) subtypes.ConclusionThe Prosigna ROR score and intrinsic subtypes were prognostic in high-risk premenopausal patients with breast cancer, and intrinsic subtypes identify high-risk patients with or without major benefit from adjuvant C/CMF treatment.Electronic supplementary materialThe online version of this article (10.1186/s13058-018-1012-0) contains supplementary material, which is available to authorized users.

Background. The therapeutic approach in patients (pts) with acute myeloid leukemia (AML) has not changed substantially in >30 years. The introduction of new treatment strategies, including immunotherapy, remains a priority. Flotetuzumab, a CD123 × CD3 bispecific DART immunotherapy, is being tested in a phase 1/2 study of relapsed/refractory (R/R) AML. We previously showed that AML pts with an immune-enriched and IFN-γ-dominant tumor microenvironment (TME) experience significantly shorter relapse-free survival, suggesting refractoriness to standard induction chemotherapy (Vadakekolathu J, et al. Blood 2017; 130: 3942A). Herein, we report that an IFN-γ-dominant TME, while predicting resistance to standard therapy, is favoring response of AML to flotetuzumab. Methods. Gene expression was analyzed in 78 bone marrow (BM) samples (36 at baseline, 27 post-cycle 1 and 15 post-cycle 2) from 40 pts with relapsed or refractory AML enrolled in a phase 1/2 clinical trial of flotetuzumab (NCT#02152956). Thirty-six baseline BM samples were included in the analysis, of which 34 from pts who were treated at the target dose of ≥500 ng/kg/day. The NanoString PanCancer IO360™ assay was used to assess the expression of 770 genes, including the levels of 14 immune cell types and of 32 immuno-oncology signatures, and their correlation with response to flotetuzumab. Data are presented as score means per group±SEM and analyzed by unpaired t-test. Results. Gene expression analysis of BM samples at baseline stratifies flotetuzumab-treated pts into 3 clusters within an immunological continuum: immune-depleted, immune-exhausted and immune-enriched (Fig. 1A). Pts with primary-refractory disease (refractory to ≥2 induction attempts, first CR of <6 months, or failure after ≥4 cycles of hypomethylating agents, HMA) showed prevalently an immune-infiltrated TME phenotype, which included higher inflammatory chemokine scores compared with relapse pts (3.27±0.22 vs 2.46±0.07, p=0.026). Within this group, chemotherapy-refractory and HMA-refractory pts further stratify into immune-enriched and immune-exhausted phenotypes, respectively. Specifically, HMA-refractory pts displayed features of immune exhaustion and adaptive immune resistance, including upregulation of TIGIT (5.55±0.34 vs 3.85±0.24, p=0.006), PD-L1 (3.55±0.18 vs 2.4±0.29, p=0.009) and Treg cells (4.87±0.23 vs 3.69±0.19, p=0.0009) together with a trend toward increasingly exhausted CD8+ T cells (CD244, EOMES, LAG3 and PTGER4) compared to chemotherapy-refractory pts (Fig. 1B). Responders to flotetuzumab showed a significantly higher IFN-γ signaling scores at baseline compared to non-responders (3.31±0.32 vs 2.27±0.11, p=0.0005), consistent with the greater frequency of responders in primary refractory pts compared to relapse pts. Accordingly, baseline IFN-γ signaling scores may be predictive of response to flotetuzumab therapy (AUC=0.841; Fig. 1C). Furthermore, comparison of post-cycle 1 BM samples to baseline samples showed treatment with flotetuzumab lead to increased immune cell infiltrate and immune activation scores, as reflected by a higher Tumor Inflammation Signature (Ayers M, et al. J Clin Invest 2017; 127: 2930-40) (6.49±0.20 vs 5.93±0.12, p=0.015) together with enhanced immunoproteasome (5.72±0.07 vs 5.23±0.10, p=0.0002) and IFN-γ signaling (3.38±0.23 vs 2.53±0.14, p=0.0015) scores. Flotetuzumab-induced TME gene activation was therefore reminiscent of an immune-enrichment rather than immune-exhaustion signature. Conclusions. We provide evidence for a range of immune gene expression profiles in AML, with primary refractory pts displaying an enhanced immune infiltration signature compared with relapse pts. Furthermore, an IFN-γ-related gene signature at baseline, a feature of primary refractory pts, was associated with clinical response to flotetuzumab. HMA-refractory pts showed an immune-rich but exhausted phenotype with PD-L1 expression, suggesting these pts may further benefit from flotetuzumab combination therapy with checkpoint inhibition (Rettig M, et al. Blood 2017; 130: 1365). Lastly, treatment with flotetuzumab further shifted the immune signature toward a more immune rich phenotype. The AML TME immune gene expression can influence susceptibility to therapy, with primary refractory AML showing an IFN-γ-dominant signature associated with response to flotetuzumab. Figure Figure. Disclosures Rutella: NanoString Technologies: Research Funding. Church:NanoString Technologies: Employment. Viboch:NanoString Technologies: Employment. Sullivan:NanoString Technologies: Employment. Hood:NanoString Technologies: Employment. Warren:NanoString Technologies: Employment. Cesano:NanoString Technologies: Employment. La Motte-Mohs:MacroGenics: Employment, Equity Ownership. Muth:MacroGenics: Employment. Lelièvre:Servier: Employment. Lowenberg:Royal Academy of Sciences and Arts, The Netherlands: Membership on an entity's Board of Directors or advisory committees; Agios Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; international Scientific Advisory Board, Institute Gustave Roussy, Paris: Membership on an entity's Board of Directors or advisory committees; Clear Creek Bio Ltd: Consultancy, Honoraria; Chairman Scientific Committee and Member Executive Committee, European School of Hematology (ESH, Paris, France): Membership on an entity's Board of Directors or advisory committees; Editorial Board "European Oncology & Haematology": Membership on an entity's Board of Directors or advisory committees; Editorial Board "The Netherlands Journal of Medicine": Membership on an entity's Board of Directors or advisory committees; Elected member, Royal Academy of Sciences and Arts, The Netherlands: Membership on an entity's Board of Directors or advisory committees; Supervisory Board, National Comprehensive Cancer Center (IKNL), Netherlands: Membership on an entity's Board of Directors or advisory committees; Chairman, Leukemia Cooperative Trial Group HOVON (Netherlands): Membership on an entity's Board of Directors or advisory committees; Editorial Board "International Journal of Hematology": Membership on an entity's Board of Directors or advisory committees; "Up-to-Date", section editor leukemia: Membership on an entity's Board of Directors or advisory committees; Astex: Consultancy; Celgene: Membership on an entity's Board of Directors or advisory committees. Davidson-Moncada:MacroGenics: Employment.