Study of 18 F-fluciclovine PET for serial assessment of glioblastoma tumor volumes during surgery and radiotherapy

Abstract

Purpose

Recent evidence supports incorporating 18 F-Fluciclovine PET for glioblastoma treatment planning and monitoring, as it better captures tumor infiltration compared to conventional MRI. However, the relationship between PET- and MRI-defined tumor volumes remains unclear, particularly in the post-treatment setting. This study prospectively compares tumor volumes on MRI and PET at multiple timepoints throughout the treatment course and evaluates volumetric changes with therapy.

Methods

We prospectively enrolled 8 adults with IDH-wildtype glioblastoma treated with surgery and chemoradiation between September 2019 and 2021. Participants underwent paired 18 F-Fluciclovine PET/CT and conventional MRI at four timepoints: preoperatively, pre-radiation, and at one- and six-months post-radiation. Biological tumor volume (BTV) from PET, FLAIR, and post-contrast T1volumes (T1CV) were segmented. Volumetric changes were compared using the Friedman test.

Results

Participants (5 males, median age 63 years [IQR 54,66]) showed significantly larger BTVs compared to T1CV at diagnosis (median BTV = 27.2mL vs. T1CV = 13.3mL, adjusted P =.03), pre-radiation (BTV = 25.2mL vs. T1CV = 6.9mL, adjusted P =.03), and at one-month post-radiation (BTV = 27.3mL vs. T1CV = 12.1mL, P =.04). BTVs closely approximated yet were slightly smaller than their corresponding FLAIR volumes at all timepoints. After surgery, the median decrease in BTV (-4.4%) was significantly smaller than T1CV (-66.5%, P =.046), with a similar nonsignificant trend observed post-radiation (P =.50).

Conclusion

Glioblastoma BTVs consistently exceed post-contrast T1 volumes and closely approximate FLAIR abnormalities throughout treatment. BTVs decline more gradually post-treatment, indicating persistent hypermetabolic tumor burden. Thus, 18 F-Fluciclovine PET can serve as an adjunct to conventional MRI in glioblastoma treatment planning and monitoring.

Supplementary Information

The online version contains supplementary material available at 10.1007/s11060-025-05146-2.

Introduction

Glioblastoma remains the most aggressive primary malignant brain tumor in adults despite the use of multimodal therapies and advances in cancer precision therapeutics. Patients harboring this tumor have a 1-year relative survival of 42.9% and a 5-year relative survival of 6.9%, among the worst of all cancers [1]. Although conventional MRI sequences constitute the mainstay of treatment planning and monitoring [2], clinicians and researchers have long acknowledged the limitations of these sequences in assessing the true extent of disease in the CNS [3].

Functional molecular imaging with PET provides complementary non-invasive metabolic information about gliomas beyond current conventional MRI capabilities. The synthetic amino acid radiotracer 18 F-Fluciclovine (anti-1-amino-3-[18 F]fluorocyclobutane-1-carboxylic acid or 18 F-FACBC), currently FDA-approved for detecting prostate cancer recurrence with an orphan drug designation for the diagnosis of gliomas [4], offers several biological and practical advantages compared to other radiolabeled compounds. It has a substantially lower background uptake in normal grey matter compared to 18 F-FDG (2-deoxy-2-[18 F]fluoro-D-glucose), which enhances the accuracy of delineation of gliomas. Additionally, it is not required to be exclusively produced at centers with on-site cyclotrons given its longer half-life compared to 11 C-MET (L-[methyl-11 C]methionine) (110 versus 20 min), thus enabling a broader distribution and clinical use [5, 6]. Moreover, its transport is mediated by two amino acid transporters ASCT2 and LAT1, compared to 11 C-MET, 18 F-FET (O-(2-[18 F]fluoroethyl)-L-tyrosine) and 18 F-FDOPA (6-[18 F]fluoro-L-3,4-dihydroxyphenylalanine) which only rely on the LAT1 transport system. As tumors often upregulate multiple pathways, this dual transport would allow 18 F-Fluciclovine to target a wider range of metabolic profiles, thus improving the detection accuracy of gliomas [7–10].

Multiple studies of 18 F-Fluciclovine PET in glioblastoma patients demonstrated radiotracer uptake beyond the contrast-enhancing tumor on conventional MRI. For instance, Karlberg et al. reported that hypermetabolic volumes defined with 18 F-Fluciclovine PET were 1.5 to 2.8 times larger and more concordant with true disease extent than volumes on post-contrast T1-weighted (T1C) MRI (n = 11) [11]. Moreover, studies with image-guided sampling of these hypermetabolic, non-contrast-enhancing areas demonstrated a near-perfect positive predictive value for tumor in these regions [12, 13]. However, these studies primarily assessed hypermetabolic volumes in the preoperative setting.

The knowledge gap this study seeks to address is the use of 18 F-Fluciclovine in the post-treatment setting. To date, only one exploratory study by Fatania et al. compared 18 F-Fluciclovine PET and T1C volumes following surgery and during chemoradiation, but conclusions were limited by small sample size, early study termination, as well as relatively stringent definitions of 18 F-Fluciclovine PET volumes [14]. Given the infiltrative nature of glioblastoma and the survival benefit associated with resecting and irradiating non-contrast-enhancing tumor while preserving healthy brain tissue [15], it is essential to elucidate the relationship between hypermetabolic volumes and non-contrast-enhancing volumes on T2-weighted FLAIR.

Thus, the purpose of this study is to prospectively compare tumor volumes defined by 18 F-Fluciclovine PET, T1C, and FLAIR at multiple timepoints throughout the treatment course of a set of glioblastoma patients, and evaluate the volumetric changes observed as a function of therapy. We hypothesize that hypermetabolic volumes will consistently exceed T1C volumes but remain smaller than FLAIR volumes across all timepoints. Additionally, we expect that tumor volumes delineated by 18 F-Fluciclovine will decrease without completely resolving following surgery and radiation, consistent with the known behavior and biology of the disease.

Materials and methods

This is a Health Insurance Portability and Accountability Act-compliant, Institutional Review Board-approved prospective cohort study conducted at a comprehensive cancer center between September 1, 2019, and September 31, 2021 (Protocol Number 2018 − 0869). Eligible patients undergoing care at our institute were approached by the clinical or research staff to obtain written informed consent prior to enrollment.

Participants enrollment

Patients included in this study (1) were at least 18 years of age, (2) had a clinically suspected or biopsy-confirmed diagnosis of glioblastoma, isocitrate dehydrogenase-wildtype, and (3) were candidates for standard-of-care maximal safe tumor resection and adjuvant chemoradiation as determined by a multidisciplinary team of neuro-oncologists, neurosurgeons, radiation oncologists and neuroradiologists. Patients were excluded if they (1) were pregnant, (2) had a known allergy to gadolinium-based contrast agents or 18 F-Fluciclovine, (3) renal failure as evidenced by an eGFR less than 60 mL/min/1.73m², or (4) pacemakers, electronic stimulation, metallic foreign bodies, and devices and/or other conditions deemed not MRI safe. Patients were also excluded (5) if histopathologic examination of resected tumor did not reveal high-grade histologic features such as central necrosis or microvascular proliferation. A total of 8 participants were enrolled in this study, which was substantially lower than the recruitment target of 25 patients due to the unexpected cessation of 18 F-Fluciclovine radiopharmaceutical distribution and the COVID-19 pandemic. The median follow-up time was 591 days (interquartile range 282–998). The ensuing article adheres to the STROBE (Strengthening the Reporting of Observational Studies in Epidemiology) checklist for cohort studies (Table S1, Online Resource) [16].

Clinical and histo-molecular data

Basic demographic characteristics such as age, sex, Karnofsky Performance Status at diagnosis; histo-molecular data including histologic grade, O6-methylguanine-DNA methyltransferase promoter (MGMTp) methylation status and EGFR amplification status; extent of resection reported by board-certified neuroradiologists at our institute on standard-of-care early postoperative MRI, within 48–72 h of surgery; [15] and cumulative radiotherapy dose were collected throughout the follow-up period. Overall survival (OS) was assessed from the date of preoperative MRI to the date of death or right-censoring.

Clinical imaging

Timeline

Participants underwent serial MRIs and 18 F-Fluciclovine PET/CTs of the brain at four timepoints throughout disease course: (1) within 14 days prior to initial tumor resection i.e. at the preoperative timepoint, (2) within 7 days prior to initiation of radiation therapy i.e. at the pre-radiation timepoint, (3) within 28 ± 5 days following radiation therapy completion i.e. at the early post-radiation timepoint, (4) at 6 months ± 14 days following radiation therapy completion i.e. at the late post-radiation timepoint. All MRI and 18 F-Fluciclovine PET/CT scans were completed within 3 days of one another.

MRI acquisition

MRI scans were performed on a 3T clinical scanner (MR750, GE Healthcare, Milwaukee, WI, USA) using an 8-channel brain coil. Images from a 3D T2-weighted FLAIR sequence and a 3D T1-weighted GRE sequence acquired before and after contrast administration of 0.1 mmol/kg gadobutrol (Gadavist, Bayer HealthCare), respectively, were used for this study. The 3D FLAIR sequence was acquired as sagittal slices (TR = 7,000 ms, TR = 89 ms, TI = 2,069 ms, echo train length = 140, in-plane resolution = 1.1 mm x 1.1 mm, slice thickness = 0.5 mm with 50% resolution), and reformatted to 1.1-mm slices in the axial and coronal planes. The 3D GRE sequence was acquired as axial slices (TR = 5.8 ms, TE = 2.5 ms, flip angle = 20°, in-plane resolution = 1.1 mm x 1.0 mm, slice thickness = 0.7 mm with 50% resolution), and reformatted to 1.1-mm slices in the sagittal and coronal planes.

18 F-fluciclovine PET/CT acquisition and reconstruction

All patients were administered 5 ± 15 mCi of 18 F-Fluciclovine intravenously. Following 3 to 5 min in a quiet room, all patients were positioned supine on a GE Discovery 690 PET/CT system with arms next to their body. Position aids and warm blankets were used to ensure patient comfort during the scan time. Patient imaging then followed and consisted of a CT scout of the whole head, a CT scan for anatomical landmarking and attenuation correction, and a PET scan consisting of one bed position. The total scan time including patient positioning, radiotracer injection, CT and PET acquisition was about 45 min. The CT acquisition parameters were 120 kVp, with 0.5 s rotation, a pitch of 0.984, 3.75 mm slice thickness with 3.27 mm interval, and a 40 mm beam collimation (0.625 × 64). Tube current modulation was utilized to minimize patient radiation exposure. The CT data was then reconstructed with 512 matrix with a standard filter using a 25 cm FOV. The PET data, on the other hand, was acquired in 30 min with TOF and reconstructed using OSEM with 2 iterations and 32 subsets with a 6.4 mm gaussian filter and point spread function modeling using a 256 matrix and a 25 cm FOV. All PET data correction for attenuation, scatter, and randoms were applied during image reconstruction using manufacturer software on the scanner console.

Volumetric data

Tumor volumes on T1C, FLAIR and 18 F-Fluciclovine PET/CT were delineated using MIM software version 7.2 (MIM Software Inc., Cleveland, OH).

First, a research fellow manually segmented tumors on T1C and FLAIR. Enhancing blood vessels, venous sinuses, meninges, and areas of intrinsic T1 shortening, if any, were manually excluded from the T1C volume (T1CV) after careful examination of pre-contrast T1-weighted sequences. Areas of necrosis were included in the preoperative T1CV, consistent with previously published segmentation approaches [17], as necrotic burden has been shown to significantly correlate with survival in glioblastoma [18, 19]. Postoperative contrast enhancement about the surgical tract and resection cavity, along with the cavity itself were also excluded from the T1CV as well as the FLAIR volume (FLAIRV) to ensure accurate quantification of residual or recurrent tumor burden. Final contours were audited by a board-certified neuroradiologist to enhance accuracy and minimize interobserver variability, with only minor adjustments performed.

Independently, a second research fellow, blinded to the information provided by the anatomic MRI sequences, contoured the metabolically active tumors on 18 F-Fluciclovine PET/CT to derive the biological tumor volume (BTV) using the PETEdge + tool of MIM. This tool does not rely on a fixed uptake threshold or tumor-to-background ratio; it rather relies on a validated gradient approach that calculates spatial derivatives along tumor radii, then defines the tumor edge on the basis of derivative levels and continuity [20, 21]. This tumor mask was subsequently mirrored onto the contralateral normal appearing brain, and standardized uptake values of the tumor and background were derived. Final contours were reviewed by a board-certified neuroradiologist with experience in nuclear imaging with only minor adjustments performed.

Radiation target volume delineation

Radiation planning volumes were independently defined as gross tumor volume, clinical tumor volume and planning target volume. The institutional approach for glioblastoma contours is such that the gross tumor volume includes the resection cavity, the T1CV and portions of the FLAIRV if considered grossly abnormal. The clinical tumor volume is a 2 cm expansion from the gross tumor volume to encompass all FLAIR changes. Furthermore, the planning target volume margins are 3 mm with image-guided radiation therapy. Treatment planning is a two-volume simultaneously integrated boost approach with prescription dose of the gross tumor volume to 60 Gy and the clinical tumor volume to 50 Gy.

Statistical analysis

Data was analyzed with SPSS version 29.0 (IBM SPSS, inc., Chicago, IL). Qualitative and quantitative variables were summarized as frequency (percent) and median (interquartile range), respectively. The primary endpoint was to examine the relationship between tumor volumes on anatomic MRI and 18 F-Fluciclovine PET/CT at each timepoint, and across timepoints throughout disease course. Median BTV, T1CV, and FLAIRV were compared with the Friedman test followed by post-hoc Bonferroni-adjusted pairwise comparisons at the preoperative and pre-radiation timepoints. To avoid underpowered statistical testing secondary to attrition, only BTV and T1CV were compared with the paired Wilcoxon signed rank test at the early post-radiation timepoint; these volumes were qualitatively compared without formal statistical testing at the late post-radiation timepoint. BTV-to-T1CV and FLAIR-to-BTV ratios were also reported. Additionally, changes in volume between consecutive timepoints were computed as percentages then stratified by extent of resection of the contrast-enhancing tumor [15]. An exploratory Kaplan-Meier analysis stratified by median pre-radiation residual BTV was conducted, and median OS were compared with the Breslow test. Significance level was set at 5%.

Results

Participant characteristics

Of the 102 adult patients with clinically suspected or biopsy confirmed glioblastoma who visited our center during the study period and were candidates for surgical resection followed by chemoradiation, 9 consented to participate in the study. One participant was excluded after the preoperative timepoint due to lack of necrosis or microvascular proliferation on postoperative pathology; thus, 8 participants were included in the analysis. While all of them underwent 18 F-Fluciclovine PET/CT and MRI at the preoperative timepoint, 6/8 (75.0%) were imaged with both modalities at the pre-radiation timepoint, 5/8 (62.5%) at the early post-radiation timepoint, and 2/8 (25.0%) at the late post-radiation timepoint. While 2 out of the 6 patients passed away prior to completing all 4 study timepoints, the remaining 4 patients opted to complete treatment with standard-of-care imaging monitoring at an outside facility. Reasons for attrition are further detailed in the study flow diagram (Fig. 1). The median age at enrollment was 63.3 years (54.3, 66.4) and 5/8 (62.5%) of participants were male. The median preoperative Karnofsky Performance Status was 90; 3 participants underwent complete resection (CR), 1 underwent near-total resection (NTR) and 4 underwent subtotal resection (STR). Molecular analysis revealed MGMTp-methylated in 4/8 (50.0%), and MGMTp-unmethylated in 3/8 (37.5%) participants. The median cumulative radiotherapy dose received by participants was 60.0 Gy (interquartile range: 45.0, 60.0). The median OS for the cohort was 383 days (95%CI 0, 1075). These baseline characteristics are illustrated in Table 1, and the distribution of tumor and background standardized uptake values at the four study timepoints is outlined in Table S2 (Online Resource).

Fig. 1.

Study flow diagram showcasing participants enrollment and serial imaging with MRI and 18 F-Fluciclovine PET/CT. Reasons for attrition are also outlined in the diagram. (*) Out of the 9 patients who underwent dual imaging at the pre-operative timepoint, 8 participants were included in the analysis. Abbreviations: TMZ = temozolomide, OSH = outside hospital

Table 1.

Baseline characteristics of study participants (n = 8)

Variable	Median (interquartile range) or Frequency (percent)
Age at diagnosis (years)	63.3 (54.3, 66.4)
Sex	5 males (62.5%); 3 females (37.5%)
Karnofsky Performance Status at diagnosis	90 (90, 90)
Extent of resection	3 CR (37.5%); 1 NTR (12.5%); 4 STR (50.0%)
Histologic grade	8 Grade 4 (100.0%)
MGMTp methylation status	4 methylated (50.0%); 3 unmethylated (37.5%); 1 unknown (12.5%)
EGFR status	3 amplified (37.5%); 2 wildtype (25.0%); 3 unknown (37.5%)
Cumulative RT dose (Gy)	60.0 (45.0, 60.0)*
Overall survival (days)	383 (228, 1246) (95% CI: 0, 1075)

(*) Six out of eight patients received the conventional 60.0 Gy radiotherapy regimen. One patient passed away before completing radiotherapy and had only received 2.67 Gy, and another patient received the hypofractionated regimen with a cumulative dose of 40.05 Gy given their advanced age. Abbreviations: CR = complete resection, NTR = near-total resection, STR = subtotal resection, RT = radiotherapy, MGMTp = O6-methylguanine DNA methyltransferase promoter

Relationship between BTV, T1C and FLAIR volumes at key timepoints

Volumes on 18 F-Fluciclovine PET/CT and MRI differed at the preoperative timepoint (P =.002); post-hoc pairwise comparison revealed the median BTV = 27.2mL (20.1, 38.5) was twice as high as the median T1CV = 13.3mL (6.4, 23.4) but did not differ from median FLAIRV = 28.1mL (25.2, 44.1) (adjusted P =.03 and 0.95, respectively). Indeed, the median BTV-to-T1CV ratio was 2.0 (1.6, 2.3) while the median FLAIRV-to-BTV ratio was 1.5 (1.0, 1.7). Similarly, volumes on 18 F-Fluciclovine PET/CT and MRI differed at the pre-radiation timepoint (P =.01); post-hoc pairwise comparison revealed the median BTV = 25.2mL (9.2, 42.1) was about 3 times higher than median T1CV = 6.9mL (0, 14.0) but did not differ from median FLAIRV = 26.8mL (10.7, 39.9) (adjusted P =.03 and 1.0, respectively). In fact, the median BTV-to-T1CV ratio was 3.1 (2.3, ∞) whereas the median FLAIRV-to-BTV ratio was 1.0 (0.9, 1.2). Furthermore, the median BTV = 27.3mL (5.1, 47.2) at the early post-radiation timepoint was also higher than the median T1CV = 12.1mL (0, 15.0) (P =.04). In fact, the median BTV-to-T1CV ratio was 3.4 (2.3, ∞). The median FLAIRV at this timepoint was 27.5mL (2.9, 48.8) and the median FLAIRV-to-BTV ratio was 1.0 (1.0, 1.1). Finally, the median BTV at the late post-radiation timepoint was 1.0mL (0.3, 1.7) while there was no measurable contrast-enhancement attributable to residual tumor on T1C. The median FLAIRV at this timepoint was 5.0mL (4.3, 5.6) and the median FLAIRV-to-BTV ratio was 12.4 (2.5, 22.4). These results are outlined in Fig. 2, and a representative case is illustrated in Fig. 3.

Fig. 2.

Boxplots representing tumor volumes on conventional MRI and 18 F-Fluciclovine PET/CT at the four study timepoints. (*) indicates P <.05, ns = not significant, na = not assessed. Abbreviations: RT = radiotherapy, T1C = post-contrast T1-weighted MRI, BTV = biological tumor volume

Fig. 3.

74-year-old female participant with glioblastoma, MGMT promoter methylated, centered in the right frontal centrum semiovale and abutting the ipsilateral corticospinal tract. Karnofsky Performance Status at diagnosis was 90. The participant underwent near-total resection of the contrast-enhancing tumor followed by concurrent chemoradiation and subsequent cycles of temozolomide. Imaging with18F-Fluciclovine PET/CT was performed at 4 timepoints during her disease course. The BTV is represented in magenta while the contralateral normal brain is outlined in light blue. Axial PET scans (top row) show a decreased BTV from 19.0 mL preoperatively, to 9.16 mL pre-radiation, to 5.06 and 1.74 mL at one and six months after radiotherapy, respectively. A similar pattern can be appreciated on conventional MRI (middle and bottom rows). It is worth noting that contrast-enhancing volume outlined in green is consistently lower than BTV, while the FLAIR volume depicted in yellow approximates BTV (graph). The participant passed away 1,246 days after diagnosis. Abbreviations: MGMT = O6-methylguanine DNA methyltransferase, BTV = biological tumor volume

Changes in volumes with surgery

The median time between surgery and the pre-radiation timepoint was 21 days (20, 21). When comparing volumes between the preoperative and pre-radiation timepoints, the median decrease in BTV= -4.4% (-49.5, 50.2) was significantly lower than the median decrease in T1CV= -66.5% (-100.0, 26.6) (P =.046). Of note, a negative percentage indicates a decrease in volume. The median change in FLAIRV between these timepoints was − 31.3% (-70.2, 43.8). Subgroup analysis by extent of resection revealed a nonsignificant trend towards a smaller decrease in BTV compared to T1CV. In fact, patients who underwent CR or NTR had a median change in BTV= -49.5% (-50.7, -28.3) compared to a median change in T1CV= -100.0% (-100.0, -100.0) (P =.11). On the other hand, patients who underwent STR had a median increase in BTV = 50.2% (24.3, 72.5) compared to a median increase in T1CV = 26.6% (-3.2, 53.2) (P =.285). Changes in volumes with surgery are highlighted in Fig. 4a.

Fig. 4.

Boxplots representing changes in volumes with surgery (Fig. 4a) and radiotherapy (Fig. 4b) on conventional MRI and 18 F-Fluciclovine PET/CT stratified by extent of resection. A negative change indicates a decrease in tumor volume, while a positive change translates into an increase in tumor volume. (*) indicates P <.05, na = not assessed. Abbreviations: CR = complete resection, NTR = near-total resection, STR = subtotal resection, T1C = post-contrast T1-weighted MRI, BTV = biological tumor volume

Changes in volumes with radiation

There was no difference between the change in BTV=-25.0% (-35.0, -18.9) and in T1CV = 0% (-10.5, 0) between the pre-radiation and early post-radiation timepoints, although a non-significant trend towards a greater decrease in BTV was observed (P =.50). The median change in FLAIRV between these timepoints was − 31.1% (-51.0, -5.9). Subgroup analysis similarly revealed a non-significant trend towards a greater decrease in BTV=-34.9 (-44.8, -25.0) compared to T1CV = 0% (0, 0) for patients who underwent CR or NTR (P =.18). However, this was not evident for STR patients who had a change in BTV=-19.0% (-27.0, 19.1) and a change in T1CV= -10.5% (-11.3, -1.8) (P = 1.00). Only 2 patients who underwent CR received a late post-radiation scan; while there was no measurable contrast-enhancement attributable to tumor on T1C, the BTV further decreased by -77.7% (-90.2, -65.1) (P =.18). Changes in volumes with radiation are summarized in Fig. 4b.

Survival by absolute residual BTV

Kaplan-Meier analysis revealed a nonsignificant trend towards improved survival in patients who underwent CR or NTR compared to STR (median OS was 799 days compared to 228 days, P =.23). Similarly, there was a nonsignificant trend towards improved survival in patients with a pre-radiation BTV below 26mL i.e. the median BTV at the pre-radiation timepoint (median OS was 1,246 days compared to 383 days, P =.24). Of note, there was no association between MGMTp methylation status and either stratum in both survival analyses (P = 1.00 and P = 1.00, Fisher’s exact test). Survival curves are illustrated in Fig.S1 (Online Resource).

Discussion

In this prospective study, we evaluated how hypermetabolic tumor volumes on 18 F-Fluciclovine PET compared with T1C and FLAIR volumes at multiple timepoints during the management of glioblastoma. Our findings highlight three key observations: (1) BTVs consistently exceeded T1CVs in the preoperative and peri-radiation settings, (2) BTVs approximated but were slightly smaller than their corresponding FLAIRVs at most timepoints, and (3) residual BTV persisted after surgery and radiation which could have implications for OS.

Our data indicates that BTVs were approximately two and three times larger than T1CVs at the preoperative and pre-radiation timepoints, respectively; however, they did not significantly differ from FLAIRVs. These findings support a growing body of evidence suggesting 18 F-Fluciclovine PET may detect infiltrative tumor beyond contrast-enhancing regions. In fact, Karlberg et al. reported that in newly diagnosed or recurrent gliomas, BTVs were 1.5 to 2.8 times larger and enclosed more than 98% of T1CVs [11]. Similarly, researchers consistently identified viable tumor upon image-guided biopsy of hypermetabolic areas beyond contrast enhancement [12, 13]. Moreover, our observation that FLAIRVs tend to only marginally surpass BTVs further supports the notion that FLAIR signal abnormalities additionally encompass peritumoral edema or reactive changes. However, infiltrating tumor cells might still exist at low density in this area such that 18 F-Fluciclovine uptake would be insufficient to be detected on PET. Wakabayashi et al. reported that the sensitivity of high-grade glioma detection in non-contrast enhancing, PET-negative areas of FLAIR signal abnormalities was 65.5% [22]. Hence, 18 F-Fluciclovine PET enhances our ability to delineate glioblastoma and should be viewed as an adjunct imaging modality to MR sequences, providing complementary, albeit imperfect, information on the extent of tumor infiltration beyond the contrast-enhancing tumor core.

Our results underscore that CR or NTR led to a significantly steeper decrease in T1CV compared to BTV, indicating that resection was focused on the contrast-enhancing tumor. These results highlight the importance of incorporating 18 F-Fluciclovine PET as an adjunct to conventional MRI when planning surgical resection. In fact, Wakabayashi et al. found that 18 F-Fluciclovine PET imaging increased the extent of surgical resection in 11 out of 23 high-grade glioma patients by identifying tumor foci otherwise undetected by T1C alone, thus decreasing postoperative tumor burden and potentially improving survival [22]. Surprisingly, some study participants undergoing STR had an increase in T1CV. Given that post-operative contrast enhancement was excluded to the best of our ability during segmentation, this increase could be explained by the growth of contrast-enhancing tumor outside of the resection cavity between surgery and the pre-radiation timepoint, which was 21 days (20, 21). The significantly more pronounced increase in BTV in this subgroup of participants reflects the additional growth of non-contrast-enhancing yet hypermetabolic tumor, offering a more comprehensive assessment of post-operative, pre-radiation tumor burden to the treatment team. Although 18 F-Fluciclovine uptake is highly specific to tumor, it is not unreasonable to partially attribute radiotracer uptake in the pre-radiation BTV to postoperative inflammation, particularly if it occurs about the resection cavity. A study by Oka et al. found that 14 C-FACBC uptake was relatively high in activated lymphocytes of rodents, which contribute to subacute postoperative inflammation [23]. However, no studies with 18 F-Fluciclovine PET-guided biopsies of hypermetabolic areas in resected gliomas have been conducted to validate these pre-clinical findings.

Our study highlights a significantly greater reduction in 18 F-Fluciclovine-defined volumes relative to T1CVs between the pre- and early post-radiation timepoints, particularly in participants who underwent CR or NTR where minimal-to-no residual contrast-enhancing tumor remained postoperatively. This suggests that BTV declines at a significantly lower rate than T1CV throughout glioblastoma treatment course, with a more pronounced reduction after radiation compared to surgery (25.0% versus 4.4%). At our institution, T2-weighted FLAIR signal abnormalities are included in target volume delineation for radiotherapy [24], resulting in irradiation of the non-enhancing, infiltrative tumor that could not be resected. This corresponds to the non-contrast enhancing hypermetabolic tumor on 18 F-Fluciclovine PET which was 3 times larger than T1CV at the pre-radiation timepoint. Although formal statistical analysis was not performed at the late post-radiation timepoint due to small sample size, the BTV further decreased and there was no measurable contrast enhancement attributed to residual tumor at the late post-radiation timepoint.

Additionally, there was a nonsignificant trend for improved OS in patients who underwent CR or NTR compared to STR. Although our study was not powered to detect survival differences, this nonsignificant trend re-emphasizes the well-established independent prognostic role of extent of resection in high-grade gliomas [25–27]. Multiple recent studies highlighted that post-operative absolute residual T1CV more accurately risk-stratifies patients with glioblastoma given the wide range of T1CVs at diagnosis [15, 28]. Since 18 F-Fluciclovine uptake is more specific to tumor cells compared to gadolinium uptake, we attempted to correlate post-operative pre-radiation BTV with survival in our cohort. We were also able to observe a nonsignificant trend towards improved OS in patients with lower residual BTV prior to radiotherapy. Fatania et al. similarly attempted to risk-stratify 6 glioblastoma patients by residual BTV and found that a smaller postoperative BTV relative to T1CV (and dynamic contrast-enhanced MRI volume) tends to correlate with longer OS (P =.89) [14]. It is important to note that Fatania et al. used a relatively stringent standardized uptake value threshold to define their BTV i.e. 3 times the maximal standardized uptake value of the contralateral normal brain versus PETEdge + derived hypermetabolic volumes in our study. Hence, their BTVs were overall smaller as they only captured tumor regions with the highest 18 F-Fluciclovine uptake; this could explain why patients with long OS in their cohort exhibited smaller BTV compared to T1CV. Larger cohorts are needed to evaluate the prognostic impact of residual BTV.

Our study has several limitations. First, our cohort size was small which restricted the generalizability of our findings. It also limited formal statistical testing between BTV and FLAIRV at the early and late post-radiation timepoints. The lack of significant difference between these volumes at the preoperative and pre-radiation timepoints is probably due to low power, rather than an actual equivalence of volumes. It is well known that FLAIR signal abnormalities harbor edema along with microscopic tumor cells infiltrating the brain parenchyma with relatively low density and proliferation rate; these cells would not uptake a sufficient amount of 18 F-Fluciclovine leading to false-negative results on PET imaging [7, 8]. Thus, FLAIRVs are expected to be larger than BTVs. Second, we observed an attrition rate of 75% at the late post-radiation timepoint, with 4 out of the 6 patients lost to follow-up opting to complete chemoradiation and/or adjuvant chemotherapy at an outside institution closer to their homes, where 18 F-Fluciclovine PET was not available, in the setting of the COVID-19 pandemic. This high attrition rate similarly hampered any formal analysis at this stage. Such analysis could help further investigate the role of 18 F-Fluciclovine PET volumes in differentiating progression from pseudo-progression and support the existing literature on this ongoing debate [29]. Third, image-guided tissue sampling of different tumor regions was not performed. While our study did not aim to evaluate the diagnostic accuracy of 18 F-Fluciclovine PET versus conventional MRI, such data could support the existing literature and further reinforce our conclusions. This would be particularly helpful if combined with a spatial analysis of FLAIRV and BTV, which would not only quantify the extent of overlap and spatial mismatch between these two volumes but also assess for viable tumor in FLAIR-hyperintense areas that do not uptake 18 F-Fluciclovine, or vice versa. Fourth, advanced MRI sequences such as diffusion-weighted imaging and dynamic susceptibility contrast were not compared to 18 F-Fluciclovine PET in his study. Given their growing clinical use and superior accuracy in assessing treatment response relative to conventional MRI [30], comparing these modalities with 18 F-Fluciclovine PET would more adequately demonstrate its value in current clinical practice. Finally, medications affecting the extent of contrast enhancement and edema, such as steroids and bevacizumab, along with their dosage and duration of intake were not incorporated in the analysis. Future studies should account for these factors given their impact on the T1CV and FLAIRV.

Our findings encourage further prospective evaluation of 18 F-Fluciclovine PET for guiding surgical resection and radiotherapy target delineation in glioblastoma. Larger cohorts are also needed to validate our volumetric analysis, further understand the biological and clinical implications of the volumetric differences observed between conventional and advanced molecular modalities, and assess the prognostic role of residual BTV. These cohorts should have standardized conventional, advanced MRI and 18 F-Fluciclovine PET protocols, as well as standardized definitions of BTV.

Conclusions

In conclusion, glioblastoma tumor volumes delineated on 18 F-Fluciclovine PET consistently exceed post-contrast T1 volumes and closely approximate FLAIR abnormalities throughout the treatment course. Moreover, PET-defined volumes decrease more gradually after surgery and radiation, indicating persistent viable tumor in the post-treatment setting. Larger prospective studies are needed to firmly establish the role of 18 F-Fluciclovine PET as an adjunct modality in glioblastoma treatment planning and monitoring.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Abbreviations

BTV

Biological tumor volume

CR

Complete resection

FLAIRV

FLAIR volume

MGMTp

O6-methylguanine-DNA methyltransferase promoter

NTR

Near-total resection

OS

Overall survival

STR

Subtotal resection

T1C

Post-contrast T1-weighted MRI

T1CV

T1C volume

Author contributions

Study conceptualization and design: S.A.D., J.M.J., O.M., H.L., M.W., D.S., L.F., D.N.Y., J.S.W., S.D.F., M.K.G., Data collection: S.A.D., J.M.J, R.M.M.M., S.A., O.M., H.L., M.W., D.S., L.F., M.K.G., Data cleaning and analysis: S.A.D., J.M.J, R.M.M.M., S.A., M.K.G., Results interpretation: S.A.D., J.M.J., R.M.M.M., S.A., O.M., H.L., M.W., D.S., L.F., D.N.Y., J.S.W., S.D.F., M.K.G., Creation of initial draft: S.A.D., J.M.J., R.M.M.M., S.A., O.M., H.L., M.K.G., Revisions and creation of final draft: S.A.D., J.M.J., R.M.M.M., S.A., O.M., H.L., M.W., D.S., L.F., D.N.Y., J.S.W., S.D.F., M.K.G.

Funding

This study was partially funded by Blue Earth Diagnostics which provided the radiotracer doses used for 18 F-Fluciclovine PET imaging. Jason M. Johnson, MD received research funding from the Diagnostic Imaging Clinical Research Committee at The University of Texas MD Anderson Cancer Center, which supported the 18 F-Fluciclovine PET/CT scans. Conventional MRI scans were conducted as part of routine clinical care and did not require additional funding.

Data availability

The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.

Declarations

Ethics approval and consent to participate

This study was performed in line with the principles of the Declaration of Helsinki. Approval was granted by the Institutional Review Board of The University of Texas MD Anderson Cancer Center (Protocol Number 2018 − 0869). Written informed consent was obtained from all individual participants included in the study.

Consent for publication

The authors affirm that human research participants provided informed consent for publication of the images in Fig. 3.

Competing interests

The authors declare no competing interests.