Comparison of [18F]fluciclovine and [18F]FDG PET/CT in Newly Diagnosed Multiple Myeloma Patients

Purpose [18F]FDG PET/CT in multiple myeloma (MM) is currently the best technology to demonstrate patchy and extramedullary disease. However, [18F]FDG PET has some limitations, and imaging with alternative tracers should be explored. In this study, we aimed to evaluate the performance of [18F]fluciclovine PET compared to [18F]FDG PET in newly diagnosed MM patients. Procedures Thirteen newly diagnosed transplant eligible MM patients were imaged both with [18F]FDG PET/CT and [18F]fluciclovine PET/CT within 1 week in a prospective study. The subjects were visually assessed positive or negative for disease. The number of lesions and the SUVmax of selected lesions were measured for both tracers. Furthermore, tracer uptake ratios were obtained by dividing lesion SUVmax by blood or bone marrow SUVmax. Between-group differences and correlations were assessed with paired t-tests and Pearson tests. Bone marrow SUVs were compared to bone marrow plasma cell percentage in biopsy samples. Results Nine subjects were assessed positively by [18F]FDG PET (69%) and 12 positives by [18F]fluciclovine PET (92%). All positive subjects had [18F]fluciclovine scans that were qualitatively scored as easier to interpret visually than the [18F]FDG scans. The number of lesions was also higher; seven of nine subjects with distinct hot spots on [18F]fluciclovine PET had fewer or no visible lesions on [18F]FDG PET. The mean lesion SUVmax values were 8.2 and 3.8 for [18F]fluciclovine and [18F]FDG, respectively. The mean tumour to blood values were 6.4 and 2.0 for [18F]fluciclovine and [18F]FDG, and the mean ratios between tumour and bone marrow were 2.1 and 1.5 for [18F]fluciclovine and [18F]FDG. The lesion SUVmax and ratios were significantly higher for [18F]fluciclovine (all p < 0.01). Local [18F]fluciclovine SUVmax or SUVmean values in os ilium and the percentage of plasma cells in bone marrow biopsies were linearly correlated (p = 0.048). There were no significant correlations between [18F]FDG SUVs and plasma cells (p = 0.82). Conclusions Based on this pilot study, [18F]fluciclovine is a promising tracer for MM. The visual and semi-quantitative evaluations indicate that [18F]fluciclovine PET/CT can out-perform [18F]FDG PET/CT at diagnosis. Supplementary Information The online version contains supplementary material available at 10.1007/s11307-022-01734-0.


Introduction
The detection of minimal residual disease (MRD) has become increasingly important with the introduction of more effective treatment options for multiple myeloma (MM). Flow cytometry and next-generation sequencing, two different cell-sampling-based methods for detecting MRD in MM, are currently in use (1). However, given that about 80% of MM patients suffer from patchy manifestations of the disease (2), it is advantageous to include methods for whole-body assessments. PET imaging using 2-Deoxy-2-[ 18 F]-fluoroglucose ([ 18 F]FDG) is by The International Myeloma Working Group considered the best technology to demonstrate patchy and extramedullary disease in MM (3). [ 18 F]FDG PET has been shown to predict outcomes in MM (4)(5)(6), and it may be of special importance in response evaluation (7). [ 18 F]FDG PET negativity after treatment has been demonstrated as an independent predictor of prolonged survival for patients with conventionally defined complete response (5,(8)(9)(10).
However, [ 18 F]FDG PET has some limitations, such as that 8-10% of MM patients are [ 18 F]FDG negative at diagnosis (3,10,11). [ 18 F]FDG can also accumulate in areas of reparative inflammation and therefore contribute to false-positive assessments. Other PET tracers may therefore be of significant value. Previously studies have included for example the CXCR4-targeting [ 68 Ga]Gapentixafor (12,13), [ 11 C]acetate (14), Na[ 18 F]F (9,15), the CD38-targeted [ 64 Cu]-or [ 89 Zr]-DFO-daratumumab (16,17), and carbon-11-or fluorine-18 labeled choline (18)(19)(20). Also, the amino acid-based tracer [ 11 C]methionine is a promising non-[ 18 F]FDG tracer for MM, and it has shown higher sensitivity in comparison to [ 18 F]FDG to detect intra-and extramedullary MM (21,22). Unfortunately, the short half-life of carbon-11 (20 min) renders this tracer inconvenient for most centers. Anti-1-amino-3-[ 18 F]-fluorocyclobutane-1 -carboxylic acid ([ 18 F]fluciclovine) is an amino acid-based PET tracer analogous to leucine with a half-life of 110 min (23). It has demonstrated similar uptake patterns to [ 11 C]methionine (24) and is approved by the Food and Drug Administration for detection of prostate cancer recurrence in patients with elevated PSA levels. In a recent study for patients with prostate cancer, [ 18 F]fluciclovine first identified an incidental second primary neoplasm in 2.7% of the patients (25). The aim of the current study was to investigate the performance of [ 18

Study Design and Patient Characteristics
MM patients suitable for autologous stem cell transplantation (ASCT) treatment, of age 18 years and older, were eligible for inclusion in this prospective study. Fourteen newly diagnosed MM patients were screened and included in the trial; there were no screen failures. One of the fourteen patients originally included (number 06) withdrew before imaging, therefore thirteen patients were assessed both with [ 18 F]FDG PET/CT and [ 18 F]fluciclovine PET/CT. Age, sex, and disease risk factors of all subjects were recorded and are described in Table 1. In addition to imaging at diagnosis, both PET examinations were repeated 3 months after ASCT. The study was approved by the National Ethics Committee, and all subjects signed an informed consent form. The ClinicalTrials. gov Identifier is NCT03966443.

Image Acquisition and Reconstruction
All scans were performed on the same Discovery MI (GE Healthcare) PET/CT scanner. To establish optimum scanning parameters, [ 18 F]fluciclovine PET imaging was initially repeated at several time points, approximately 30, 60, and/or 120 min post injection (p.i.) in the first three subjects with 1-to 2.5 min acquisition per bed position. For subsequent subjects, whole body (WB) acquisitions of 1 min per bed position after 15 min p.i. were performed. A low-dose CT scan without contrast enhancement was performed for attenuation correction and anatomic information before the PET acquisition (120 kV, noise index 34.5). PET data were reconstructed using VP FX Q.Clear 400 with a matrix of 384, plus VP FX SharpIR, std filter 5, 16/3 subsets/iterations, and a matrix of 256. The latter reconstruction was used for SUV measurements. The reconstructed slice thickness was 2.79 mm.
For all but one of the patients (subject 08), the [

Image Analysis
Visual assessments were performed by two nuclear medicine physicians independently, and the final evaluation of positivity or negativity was reached in consensus. The [ 18 F] fluciclovine and [ 18 F]FDG examinations were assessed in random order. For [ 18 F]FDG PET/CT scans, the positive/ negative assessment was performed based on presence of active lesions and/or high uptake in bone marrow. The IMPe-TUs criteria were used as guidance (28). Also for the [ 18 F] fluciclovine PET/CT examinations was the overall assessment based on presence of active lesions and/or the appearance of bone marrow uptake (defined as diffuse uptake above physiological uptake). In addition, for positive scans, the images were later qualitatively scored as more, equally or less, easy to interpret compared to the [ 18 F]FDG images.
The approximate numbers of active lesions (areas of visually increased tracer uptake compared to background) were categorised as 0, 1, 2, 3, 4, 5, 6-10, 11-20, 21-50, 51-100 or > 100 lesions by an experienced nuclear medicine specialist. SUVs were measured using 1 cm 3 volumes of interest in liver (SUV mean ), mediastinal blood (SUV max ), and os ilium (SUV max and SUV mean ). The bone marrow regions were placed in areas avoiding focal hot spots, displaying as homogeneous uptake as possible. Also, for all subjects with active lesions, SUV max values were measured for three of the lesions demonstrating the highest uptake on the [ 18 F]fluciclovine PET examination and the same locations on the [ 18 F] FDG PET examination. Similarly, three of the lesions demonstrating the highest uptake on the [ 18 F]FDG PET examinations were also selected, and SUVs were measured from both image sets. Measurements for a total of up to six lesions were hence obtained for each subject. The measured locations were matched between the image sets in a rigorously manner by an experienced nuclear medicine physician. Tumour-tonormal tissue ratios were obtained by dividing lesion SUV max by blood or bone marrow SUV max . For the first three subjects with imaging at later time points, the first time point was used to measure SUVs. Syngo.via VB30 software (Siemens Healthineers) was used for the measurements.

Statistics
Paired t-tests and Pearson correlation tests were performed to investigate relationships between tumour SUVs and between tumour to blood or bone marrow ratios for [ 18 F] fluciclovine PET and [ 18 F]FDG PET. Pearson correlation tests were also used to investigate the correlation between the percentage of plasma cells and SUVs in the bone marrow. The first three subjects were excluded from all analyses as their [ 18 F]fluciclovine PETs were obtained at later time points than the rest. IBM SPSS version 25 was used for the statistical analyses.

Visual Assessments
Of the 13 subjects included, nine were assessed positive by  (Table 2). Typically, this was based on both bone marrow appearance and the number and interpretability of active lesions, but for three subjects, it was primarily based on bone marrow contribution (subjects 04, 10, and 13), and one subject with a single lesion with soft-tissue involvement in os frontalis had more clearly distinguishable uptake of [ 18

Acquisition Protocol
The first three subjects (01-03) were imaged at one or multiple time points ranging from approximately 30-120 min p.i. ( Table 2). Rapid washout from lesions was observed, and pronounced uptake in muscle was also found, increasing with time (Fig. 2). Therefore, WB imaging of later subjects was initiated already at 15 min p.i.. This avoided background accumulation in muscles.

Bone Marrow Biopsy and PET Results
Based on the eight patients with available biopsy results from crista iliaca and [ 18 F]fluciclovine imaging performed at 15 min p.i., there was a linear significant correlation between local [ 18 F]fluciclovine SUV max in os ilium and the percentage of plasma cells in bone marrow (p = 0.048; Fig. 4). There were no significant correlation between [ 18 F]FDG SUV and the percentage of plasma cells (p = 0.82 for SUV max ).

Discussion
In Accordingly, higher tumour SUVs and tumour to normal tissue ratios were also observed. Through investigations of imaging at different time points, we found acquisitions performed rapidly after injection of [ 18 F]fluciclovine to be optimal (Fig. 2). This is probably due to the bidirectional properties of the cellular transport system of leucine (29). While [ 18 F]fluciclovine imaging for prostate cancers was performed almost immediately after injection (30), the optimal time point for brain tumours is more uncertain and probably delayed (31). [ 11 C]methionine PET for MM appears to usually be performed at 20 min p.i. (20,22,32), but we were unable to find reports investigating different timing. Here, the [ 18 F]fluciclovine whole body images were obtained at 15 min p.i., but investigations of more immediate acquisitions could be interesting to pursue in a later study. The possibility of acquiring images earlier after administration of the radiopharmaceutical is nevertheless beneficial for several reasons. These include logistics for both patients and the nuclear medicine department as the time spent per patient is decreased. Furthermore, it will improve the image quality obtained for a certain absorbed dose, as early imaging allows decreased injected activity and/or acquisition time. Since the effective dose from the [ 18   week of each other, in random order, and will most likely be highly comparable. One of the main limitations of the current work, besides the study size, is the lack of standardised assessment criteria for [ 18 F]fluciclovine in MM. This is of course to be expected, as this pilot study is the first to explore the potential for this tracer. Still, especially pathological bone marrow uptake was challenging to characterise in a couple of cases, as normal physiological uptake of [ 18 F]fluciclovine is expected in the marrow (30). This may also have a heterogeneous nature. Our institution has an extensive experience with [ 18 F]fluciclovine for prostate cancer, and drew on this expertise for the current evaluations. While the majority of the [ 18 F]fluciclovine images were easy to characterise, it should be noted that a few cases were borderline (especially subject 04). A set of evaluation criteria for MM, possibly also relaying on measurements of bone marrow heterogeneity and uptake structure, should be established if this tracer is to be used more routinely.
The approximate numbers of lesions were clearly different between the tracers (Table 2 and Fig. 1). While this number will not alter the positive/negative status, a study has reported a higher number of [ 18 F]FDG foci to be associated with a less favourable prognosis (4), and an improved resolution of this number may therefore be relevant for stratification purposes. The semi-quantitative analyses for [ 18 (33)). Although the biopsy data is from untargeted iliac crest marrow, introducing a potential uncertainty (especially for patients with patchy bone marrow pattern), this is no different from the previous study mentioned and should also not impact the correlation for each tracer differently.
A potential disadvantage of [ 18 F]fluciclovine and other amino acid-based tracers is the unavailability for theragnostic applications; replacing the positron emitter with a therapeutic nuclide. Tracers such as the CXCR4-targeting [ 68 Ga]Ga-pentixafor (13), PSMA tracers (currently under investigation), and the CD38-targeted daratumumab (16) have this potential. However, many theragnostic tracers, especially antibodybased, can be subject to downregulation. A broader selection of tracers, including [ 18 F]fluciclovine or other amino acidbased tracers, should therefore be available to allow the best choice based on the clinical setting and stage.

Conclusion
Based on this pilot study, [ 18 F]fluciclovine is a promising PET tracer for MM patients. The visual assessments indicate that [ 18 F]fluciclovine PET performs better then [ 18 F]FDG PET at diagnosis, with both a higher number of active lesions present and improved interpretability of the bone marrow appearance. The semi-quantitative evaluations support this result, with higher SUVs, higher tumour to normal tissue values, and significant correlation with biopsy results observed only for [ 18 F]fluciclovine SUVs. A similar study with a larger number of subjects should be performed to assert the results from this pilot.
Funding Open access funding provided by University of Oslo (incl Oslo University Hospital)

Declarations
Ethics Approval and Informed Consent All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards. Informed consent was obtained from all individual participants included in the study.

Conflict of interest
The authors declare that they have no conflict of interests.
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