¹⁸F-FDG PET/CT of off-target lymphoid organs in CD19-targeting chimeric antigen receptor T-cell therapy for relapsed or refractory diffuse large B-cell lymphoma

From 12 patients with r/r DLBCL which were consecutively referred for PET/CT between March 2019 and February 2020, we retrospectively analyzed ten patients (Table 1). Patients were excluded if showing non-FDG avid indolent lymphoma lesions (n = 2). All patients underwent a baseline PET (PET1) before CAR-T-cell therapy, and received PET-based early and late response assessment at + 30 d (PET2, n = 7) and + 90 d (PET3, n = 5), if still alive and not showing clear evidence of progression on clinical examination or other imaging. Laboratory values including leukocyte count, lymphocyte count and C-reactive protein (CRP) were documented. ¹⁸F-FDG was administered in compliance with the German Medicinal Products Act, AMG §13.2b, and all subjects provided written informed consent. The institutional review board approved this retrospective analysis (No. 8610-BO-K-2019).

In all patients, lymphapheresis lead to successful production of Tisagenlecleucel as autologous CD19 CAR-T-cell product, albeit in two patients’ first attempt failed. All patients underwent lymphodepletion with fludarabine (25 mg/m²) and cyclophosphamide (250 mg/m²) for three consecutive days prior to infusion of Tisagenlecleucel. Fludarabine and cyclophosphamide were dose adjusted due to reduced kidney function in two patients. Details of CRS and ICANS [4], and CAR-T-cell doses are shown in Table 1. All patients received an extensive neurological work-up prior to CAR-T-cell infusion. During the first 10 days post-CAR-T infusion, patients were daily tested by a neurologist for signs and symptoms of neurotoxicity via clinical neurological examination and cognitive testing using the Montreal-Cognitive-Assessment (MoCA). Patients no. 5, 8 and 9 received PET-guided consolidation radiation therapy after PET2.

PET/CT acquisition and image reconstruction All studies were acquired using a standard PET/CT protocol (Supplemental Data). None of the patients suffered from diabetes. The mean blood glucose level at PET was 92 mg/dL (range 77–112 mg/dL).

Image analysis and calculation of imaging parameters. PET/CT images were analyzed using a dedicated workstation equipped with a commercial software package (syngo.via; V10B, Siemens Healthcare). All lesions suggestive for lymphoma were noted, and their localization (e.g., lymph node, bone marrow) was recorded. The largest CT diameter of each lesion was recorded, and bulky disease was defined as maximum lesion diameter ≥ 7.5 cm. For assessment of lymphoma metabolic parameters, each lesion was analyzed applying an isocontour volume-of-interest (VOI) including all voxels above 45% of the maximum using a three-dimensional segmentation and computerized volumetric technique. Mean and maximum SUVs (SUV_mean and SUV_max) were measured within all volumes of interest. This measurement also yielded the metabolic tumor volume (MTV). Next, MTV was multiplied by SUV_mean of the lesion, yielding the total lesion glycolysis (TLG). Both MTV and TLG of each lesion were summed up to calculate the whole-body MTV and whole-body TLG, respectively. For assessment of bone marrow and lymphoid organ activity (Fig. 1a), the bone marrow signal (averaged SUV_mean of 4 VOIs of 2 cm diameter placed within lumbar vertebrae L1 to L4), the spleen signal (averaged SUV_mean of 3 VOIs of 2 cm diameter), Waldeyer`s lymphatic ring signal (averaged SUV_max of left and right measurements), and the signal from lymph nodes unaffected by lymphoma (averaged SUV_max of 4 left and right inguinal and left and right cervical lymph nodes) were determined for each patient at PET1 and PET2, and used for further analyses.

Following CAR-T-cell therapy, treatment response was assessed using ¹⁸F-FDG PET/CT at + 30 d (PET2) and at + 90 d (PET3) according to the Lugano classification [8] criteria. A favourable outcome was defined as achieving partial or complete remission at PET3, whereas an unfavourable outcome was defined as progression, or death.

Categorical variables are presented with absolute and relative frequencies. Continuous variables are expressed as mean ± standard deviation (SD) and range. Normal distribution of data was verified using the Kolmogorov–Smirnov test. For group comparison, an unpaired t test (continuous variables) and Fisher’s exact test (categorical variables) were used. Differences in PET metabolic parameters, and their significance for treatment response and toxicity, were analysed. Statistical significance was established for P values < 0.05. Analysis was performed using GraphPad Prism^® (version 8.3 for Windows; Graphpad Software).

Two (20%) patients achieved complete remission, two (20%) patients partial remission, four (40%) patients had progressive disease, and two (20%) patients deceased within 3 weeks of therapy. Remission at PET3 required early metabolic response at PET2: all patients achieving a response (PR/CR) at PET3 had already shown an early metabolic response at PET2, whereas only one patient who had an unfavourable outcome had an early response to treatment (P = 0.0476) (Fig. 1c).

Baseline (PET1) parameters of lymphoma metabolism (P ≥ 0.1078), lymphoid organ metabolism (P ≥ 0.3653), and inflammatory markers (P ≥ 0.3801) were not significantly different between patients with unfavourable and favourable outcome (Supplemental Table 1). Bulky disease (P = 0.0476) was associated with unfavourable outcome.

Patients with unfavourable outcome demonstrated a significantly higher decrease in both spleen signal (− 42% ± 22% vs. -7% ± 10%; P = 0.0368) and lymph node signal (−44% ± 13% vs. + 4% ± 28%; P = 0.0470) between PET1 and PET2 (Fig. 2b). By contrast, the change in leukocyte count (P = 0.4434) or lymphocyte count (P = 0.3054) between baseline and + 30 d was not associated with outcome, and not associated with change in spleen or change in unaffected lymph node signal at PET (P ≥ 0.1310 in all cases). Moreover, there was a non-significant trend towards reduced metabolism (SUVs at PET2) in lymphoid organs and bone marrow in patients with unfavourable outcome (spleen, P = 0.0635; unaffected lymph nodes, P = 0.0784; Waldeyer’s lymphatic ring, P = 0.1423; bone marrow, P = 0.0654) (Fig. 2c).

Four (40%) patients developed neurotoxicity. Maximum SUV_max at baseline, but not MTV (P = 0.1481) or TLG (P = 0.1019), was significantly higher in patients who developed neurotoxicity (33.2 ± 8.8 (range 26.9–46.3) vs. 22.3 ± 6.2 (range 12.2–28.4); P = 0.0489) (Fig. 2d). No other baseline (PET1) parameter was associated with development of neurotoxicity (P ≥ 0.4039 in all cases). 4 (40%) patients developed a CRS. No baseline (PET1) parameter was associated with development of CRS (P ≥ 0.0822 in all cases).

In patients having received steroids (n = 2) and/or tocilizumab (n = 3) for treatment of toxicity, bone marrow signal (P = 0.6838), spleen signal (P = 0.5688), and lymph node signal (P = 0.4802) were not lower at PET2, and the % change between PET1 and PET2 was not different (e.g., change in lymph node signal, P = 0.4151).