89Zr-immunoPET-guided selection of a CD33xIL15 fusion protein optimized for antitumor immune cell activation and in vivo tumour retention in acute myeloid leukaemia.

PURPOSE: Immune cells are capable of eliminating leukemic cells, as evidenced by outcomes in hematopoietic cell transplantation (HCT). However, patients who fail induction therapy will not benefit from HCT due to their minimal residual disease (MRD) status. Thus, we aimed to develop an immunomodulatory agent to reduce MRD by activating immune effector cells in the presence of leukaemia cells via a novel fusion protein that chimerises two clinically tolerated biologics: a CD33 antibody and the IL15Ra/IL15 complex (CD33xIL15). METHODS: We generated a set of CD33xIL15 fusion protein constructs with varying configurations and identified those with the best in vitro AML-binding, T cell activation, and NK cell potentiation. Using 89Zr-immunoPET imaging we then evaluated the biodistribution and in vivo tumour retention of the most favourable CD33xIL15 constructs in an AML xenograft model. Ex vivo biodistribution studies were used to confirm the pharmacokinetics of the constructs. RESULTS: Two of the generated fusion proteins, CD33xIL15 (N72D) and CD33xIL15wt, demonstrated optimal in vitro behaviour and were further evaluated in vivo. These studies revealed that the CD33xIL15wt candidate was capable of being retained in the tumour for as long as its parental CD33 antibody, Lintuzumab (13.9 ± 3.1%ID/g vs 18.6 ± 1.1%ID/g at 120 h). CONCLUSION: This work demonstrates that CD33xIL15 fusion proteins are capable of targeting leukemic cells and stimulating local T cells in vitro and of concentrating in the tumour in AML xenografts. It also highlights the importance of 89Zr-immunoPET to guide the development and selection of tumour-targeted antibody-cytokine fusion proteins.

Improved image reconstruction of 89Zr-immunoPET studies using a Bayesian penalized likelihood reconstruction algorithm.

PURPOSE: The aim of this study was to evaluate the use of a Bayesian penalized likelihood reconstruction algorithm (Q.Clear) for 89Zr-immunoPET image reconstruction and its potential to improve image quality and reduce the administered activity of 89Zr-immunoPET tracers. METHODS: Eight 89Zr-immunoPET whole-body PET/CT scans from three 89Zr-immunoPET clinical trials were selected for analysis. On average, patients were imaged 6.3 days (range 5.0-8.0 days) after administration of 69 MBq (range 65-76 MBq) of [89Zr]Zr-DFO-daratumumab, [89Zr]Zr-DFO-pertuzumab, or [89Zr]Zr-DFO-trastuzumab. List-mode PET data was retrospectively reconstructed using Q.Clear with incremental ß-values from 150 to 7200, as well as standard ordered-subset expectation maximization (OSEM) reconstruction (2-iterations, 16-subsets, a 6.4-mm Gaussian transaxial filter, "heavy" z-axis filtering and all manufacturers' corrections active). Reduced activities were simulated by discarding 50% and 75% of original counts in each list mode stream. All reconstructed PET images were scored for image quality and lesion detectability using a 5-point scale. SUVmax for normal liver and sites of disease and liver signal-to-noise ratio were measured. RESULTS: Q.Clear reconstructions with ß = 3600 provided the highest scores for image quality. Images reconstructed with ß-values of 3600 or 5200 using only 50% or 25% of the original counts provided comparable or better image quality scores than standard OSEM reconstruction images using 100% of counts. CONCLUSION: The Bayesian penalized likelihood reconstruction algorithm Q.Clear improved the quality of 89Zr-immunoPET images. This could be used in future studies to improve image quality and/or decrease the administered activity of 89Zr-immunoPET tracers.

Dose escalation PET imaging for safety and effective therapy dose optimization of a bispecific antibody.

Selecting the dose for efficacy and first-in-human studies of bispecific antibodies (BsAbs) is a challenging process. Herein, positron emission tomography (PET) imaging with 89Zr-labeled IBI322, an anti-CD47/PD-L1 BsAb, was used to optimize the safety and effective therapy dose. By labeling with 89Zr, we aimed to assess the pharmacokinetics (PK), safety, and target engagement of IBI322 with dose escalation dynamic PET imaging in humanized transgenic animal models bearing MC38 tumors (knock-in of hCD47 and hPDL1). 89Zr-labeled IBI322 specifically accumulated in tumors with a tumor-to-muscle ratio of 12.37 ± 1.42 at 168 h (0.22 mg/kg) and the biodistribution of normal tissues from PET imaging could be used for preliminary safety prediction. According to the Pearson correlation analysis between the ELISA-quantified serum concentration and heart uptake (%ID/g) (r = 0.980), a modified Patlak model was proposed. The exploratory target-mediated 50% (0.38 mg/kg) and 90% (0.63 mg/kg) inhibitory mass doses were calculated with the current modified Patlak model. The preliminary pharmacodynamics (PD) study with 0.34 mg/kg revealed that the dose prediction was rational. In conclusion, dose escalation PET imaging with 89Zr-labeled antibodies is promising for PK/PD modeling and safety prediction, and helpful for determining rational dosing for preclinical and clinical trials of BsAbs.