P-03.04 Targeting CXCR4: implications for clonal dynamics and MRD assessment in acute myeloid leukemia

<h3>Background</h3> Acute Myeloid Leukemia (AML) is a difficult-to-treat disease with poor treatment outcomes. AML is characterized by its heterogeneous nature, which may enable the development of targeted therapies. The CXCR4/CXCL12 axis plays a crucial role in leukemic stem cell survival within the bone marrow microenvironment, making CXCR4 inhibitors a potential treatment approach. Measurable residual disease (MRD) assessment is important in AML, as post-remission MRD indicates higher relapse risk. Novel single-cell MRD (scMRD) workflows, such as the Tapestri® platform, offer potential breakthroughs by capturing genotypic and immunophenotypic changes at a single-cell level. For our analysis, we used bone marrow samples from a phase II clinical trial (BLAST, NCT02502968) investigating the effect of a novel CXCR4 inhibitor (BL-8040) in combination with conventional consolidation chemotherapy in AML patients in first clinical remission (CR). In this study, single-cell DNA and protein sequencing is used to assess MRD and clonal evolution in the context of CXCR4 inhibition. <h3>Materials and Methods</h3> We analyzed a cohort of 69 AML patients in first CR from the BLAST trial. For 11 patients, we analyzed paired samples from both first CR and relapse. We performed single-cell DNA and protein sequencing on a total of 250,000 cells across 23 multiplexed pools. Our methodology included 40 selected mutation hotspot genes and a 19-plex antibody oligonucleotide cocktail for AML-specific surface markers. <h3>Results</h3> Our analyses revealed a diverse genetic landscape with 25 different mutated genes in 69 patients. DNMT3A and NPM1 mutations were the most common alterations. Lower single-cell assessed MRD levels were consistently associated with improved overall survival (OS) and relapse-free survival (RFS). Patients with &lt;5% MRD showed significantly improved RFS (p = 0.0013), with median OS of 18 months and median RFS of 11.7 months. Interestingly, when analyzing RFS based on CXCR4 expression levels, we found that patients with high CXCR4 expression who received the CXCR4 inhibitor experienced lower relapse rates (p=0.036) compared to the placebo group. <h3>Conclusions</h3> In this AML cohort, our study provides insights into the genetic landscape and clonal dynamics of AML in remission, emphasizing the importance and potential of single-cell DNA and protein sequencing in refining MRD assessment compared to current techniques. In the completed clinical trial, the CXCR4 inhibitor did not show an overall benefit when added to chemotherapy; however, we demonstrated its potential to reduce relapse rates specifically in patients with high CXCR4 expression. Our findings suggest that the role of CXCR4 inhibition in AML treatment may involve multifaceted mechanisms, including effects on leukemic stem cells and possibly other CXCR4-expressing immune cells. Further research is needed to clarify these mechanisms and identify patient subgroups that may benefit from CXCR4 inhibition in AML treatment. <b>E. Ceran:</b> Other; Modest; Ipsen (Congress attendance DGHO). <b>A.K. Merbach:</b> None. <b>C. Rohde:</b> None. <b>R. Durruthy-Durruthy:</b> None. <b>A. Sciambi:</b> A. Employment (full or part-time); Modest; Mission Bio Inc. <b>M. Lotze:</b> None. <b>M. Wass:</b> None. <b>C. Röllig:</b> None. <b>R.F. Schlenk:</b> None. <b>M. Scheller:</b> None. <b>J. Steighardt:</b> None. <b>L.P. Müller:</b> None. <b>I. Gliko-Kabir:</b> A. Employment (full or part-time); Modest; BiolineRx. <b>C. Gromann:</b> None. <b>A. Wienke:</b> None. <b>C.D. Baldus:</b> None. <b>U. Platzbecker:</b> None. <b>H. Serve:</b> None. <b>M. Bornhäuser:</b> None. <b>C. Müller-Tidow:</b> None.