Identification of CXCR4 Expression By Targeted PET/MR Imaging in Patients with Acute Myeloid Leukemia (AML)
Authors: Herhaus Peter , Habringer Stefan , Philipp-Abbrederis Kathrin , Vag Tibor , Gerngroß Carlos , Höllein Alexander , Altmann Torben , Müller-Thomas Catharina , Slotta-Huspenina Julia , Peschel Christian , Schwaiger Markus , Wester Hans-Jürgen , Verbeek Mareike , Spiekermann Karsten , Götze Katharina , Keller Ulrich 
Published: October 2019

Abstract
Introduction
AML is an aggressive hematological neoplasm originating from hematopoietic precursor or stem cells and is rapidly fatal if untreated. Although rates of complete remission (CR) approach 80% in patients with favorable risk profile according to European Leukemia Net classification, CR rates drop dramatically within the intermediate or adverse prognostic groups, and most patients relapse in spite of CR. The origin of refractory or relapsed disease is - according to several studies - the bone marrow (BM) niche that protects the leukemia initiating cells (LIC) from cytotoxic drugs.The chemokine receptor CXCR4 is a key factor mediating the crosstalk between the BM niche and LICs. CXCR4 is a G-protein coupled chemokine receptor with Stromal cell-derived factor-1α (SDF-1α) as its single known chemokine-ligand, and high expression is associated with poor prognosis in AML. Targeting the CXCR4/SDF-1αaxis with small peptide inhibitors or monoclonal antibodies has shown promising results in preclinical and clinical studies, thus rendering it an attractive target for future therapeutic strategies (Peled & Tavor, Theranostics, 2014). In this observational study we analyzed whether the new CXCR4-targeted positron emission tomography (PET)-probe [68Ga]-Pentixafor (Philipp-Abbrederis et al, EMBO Mol Med, 2015; Wester et al, Theranostics, 2015) is applicable for molecular imaging of patients with AML with the goal to develop a CXCR4- targeted peptide-receptor-radiotherapy (PRRT).
Methods
Using myeloid malignancies (MDS, active AML) as an exemplary CXCR4-expressing cancer entity we imaged 10 patients with the CXCR4 specific PET-probe [68Ga]-Pentixafor by whole body PET/magnetic resonance imaging (PET/MR) after they signed informed consent for this observational assessment. Maximum standardized uptake values (SUVmax) of the involved BM areas were compared to BM from patients without BM malignancy assessed in a different study. CXCR4 surface expression of leukemic BM cells of an unselected cohort of 67 patients with myeloid malignancies was analyzed by flow cytometry. CXCR4 expression in 14 established AML cell lines was measured by flow cytometry and quantitative PCR.
Results
Out of 10 patients who were imaged with [68Ga]-Pentixafor-PET/MR, 4 - all of them with the diagnosis of an AML - showed visually positive PET signal of the BM. Those areas correlated well with disease infiltration as determined by MR imaging and the meanSUVmax was significantly higher as compared to visually CXCR4 negative patients (meanSUVmax 8.23 ± 5.23 vs. 2.26 ± 0.47; p=0.036). One of the 10 patients presenting with proven extramedullary relapse showed a CXCR4 positive PET signal within the lesion (SUVmax 5.12), whereas the cytological non-involved BM was PET negative (meanSUVmax 1.82). CXCR4 surface expression in established AML cell lines was variable and ranged from low to high, and correlated with mRNA levels. These data reflect the different expression values we found in the cohort of patients with myeloid malignancies. There was a trend to higher CXCR4 surface expression on blasts when AML patients were compared to MDS patients (median fluorescent intensity CXCR4/isotype 1.73 ± 0.25 vs. 0.96 ± 0.06; p=0.063) that became significant when blast frequency exceeds 30% (≤30% vs. >30% blasts: 1.07 ± 0.11 vs. 2.08 ± 0.38; p=0.006). In this patient cohort we did not observe any correlation between CXCR4 expression and factors such as diagnosis (secondary vs. de novo AML), disease status (refractory or relapse vs. no pretreatment), molecular subtypes or cytogenetic aberrations.
Discussion
Imaging of CXCR4 by PET/MR with [68Ga]-Pentixafor is feasible in patients with AML. Due to the higher uptake in leukemic BM compared to control BM and its low uptake in other organs (e.g. liver, brain and gut), CXCR4 could serve as an attractive new PRRT-target in selected patients with AML. Such PRRT approaches are of special interest in the setting of allogeneic BM transplantation as conventional myeloablative conditioning regimens - although they are associated with a better relapse free survival - show substantial rates of transplant-related mortality and are therefore not applicable in many AML patients due to their age. In particular, therapeutic targeting of CXCR4 by PRRT provides a unique opportunity of integrating a LIC/niche-directed treatment into the conditioning regimen. Such clinical studies are in development.

Disclosures
Wester: Scintomics GmbH: Other: General Manager. Keller:Roche: Consultancy, Honoraria; Pfizer: Consultancy.