Prognostic Value of O-(2-¹⁸F-Fluoroethyl)-l-Tyrosine PET and MRI in Low-Grade Glioma

Patient Population

Within the 7-y period of 1999–2005, a group of 33 consecutive patients with low-grade glioma were included in the study. All had newly diagnosed, previously untreated supratentorial low-grade glioma without signs of blood–brain barrier disruption on contrast-enhanced MRI. Patients with infratentorial, thalamic, and hypothalamic tumors; children; and patients with low-grade glioma with contrast enhancement on MRI were excluded because of their different biologic and prognostic behavior. All patients were eligible for elective treatment planning—that is, were in stable clinical and neurologic condition without signs or symptoms of increased intracranial pressure. Detailed information on the individual patients is given in Tables 1 and 2. The study was approved by the university ethics committee and federal authorities. All subjects gave written informed consent for their participation in the study.

Thirteen of the patients were female and 20 were male; age at diagnosis ranged from 27 to 65 y, with a mean of 40.9 y. Karnofsky performance status ranged from 90% to 100%, with a mean of 93%. Initial symptoms were seizures in 28 patients (85%), unspecific symptoms in 3 patients (9%), and focal neurologic deficits in 2 patients (6%).

The histologic diagnosis of low-grade glioma was established in all patients on tissue samples obtained by neuronavigated biopsy as described previously (17,18). In short, PET images were coregistered with MR images using dedicated software (MPI tool 3.28; ATV) and transferred to the neuronavigation system (Vector Vision; BrainLab). For coregistration, anatomic landmarks and external cutaneous fiducial markers were used in each examination. Neuronavigated biopsy samples were taken of FET-positive lesions from the area with maximal FET uptake. For FET-negative lesions, biopsy samples were taken only from areas showing an abnormal MRI signal.

Diffuse astrocytoma was diagnosed in 27 patients (82%), oligodendroglioma in 4 (12%), and oligoastrocytoma in 2 (6%). Subsequent first-line cytoreductive surgery was recommended in all patients with space-occupying tumors causing a clinically significant mass shift or tumors in noneloquent areas. According to these criteria, open surgery was offered to 20 patients, but 8 patients refused. Thus, surgery was performed on the remaining 12 patients. In 3 patients, resection was classified as “total” (>90% of the tumor volume was resected); in 6 patients, “subtotal” (50%–90% of the tumor mass was resected); and in 3 patients, “partial” (<50% of the tumor mass was resected). None of the patients received radiotherapy or chemotherapy as first-line therapy after biopsy or resection. Clinical and MRI follow-up was performed at regular intervals of 3–4 mo. The endpoints of course and outcome were tumor progression, malignant transformation to a glioma of WHO grade III or IV (high-grade glioma), and death of the patient.

Tumor progression was defined as clinical or radiologic deterioration. Symptoms and signs of clinical deterioration included new onset or aggravation of preexisting focal deficits or new onset of symptoms of increased intracranial pressure. Increased seizure activity was not regarded as a symptom of tumor progression. Radiologic deterioration was defined as a new onset of contrast enhancement within the tumor, significant enlargement of more than 20% of the initial tumor volume, or an increasing mass shift.

MRI

MRI was performed on a 1.5-T system (Sonata; Siemens). The imaging protocol consisted of a T1-weighted 3-dimensional magnetization-prepared rapid-acquisition gradient-echo sequence (field of view, 25 cm; matrix, 205 × 256; repetition time, 2,200 ms; echo time, 3.9 ms; inversion time, 1,200 ms; flip angle, 15; number of slices, 128; slice thickness, 1.5 mm; slice gap, 0 mm; number of averages, 1; time of acquisition, 6:38 min) before and 2 min after injection of 20 mL of gadolinium-diethylenetriaminepentaacetic acid (Magnevist; Schering) and a transversal fluid-attenuation inversion-recovery (FLAIR) sequence (field of view, 25 cm; matrix, 205 × 256; repetition time, 9,000 ms; echo time, 119 ms; inversion time, 2,500 ms; flip angle, 90; number of slices, 25; slice thickness, 5 mm; slice gap, 0 mm; number of averages, 2; time of acquisition, 4:32 min).

MRI scans were assessed by a senior neuroradiologist and 2 experienced senior neurosurgeons and scored by consensus. None of the tumors showed significant contrast enhancement after injection of gadolinium-diethylenetriaminepentaacetic acid. The tumor size was difficult to measure reliably because volumetric assessment was possible only for circumscribed tumors with well-defined borders—not for diffuse lesions. For semiquantitative evaluation, tumor size was estimated by the maximal cross-sectional diameter of the hypointense area on the T1-weighted images with respect to the hyperintensity on the FLAIR sequence. Tumors with a diameter greater than 30 mm were classified as large. According to their appearance on MRI, the tumors were classified as circumscribed in 24 patients (73%) and diffuse in 9 patients (27%). Circumscribed tumors were characterized by a homogeneous structure and sharp borders that were identical on T1- and T2-weighted images. Diffuse tumors showed an inhomogeneous signal pattern and poorly defined borders on MRI. In these tumors, the extent of hyperintensity on T2-weighted sequences was generally larger than the area of hypointensity on T1-weighted, indicating perifocal edema. Displacement of the brain midline by more than 2 mm was classified as a mass shift.

¹⁸F-FET PET

The patients were examined by ¹⁸F-FET PET within 6 wk after the initial diagnosis and no more than 3 wk before the neuronavigated biopsy. The amino acid ¹⁸F-FET was produced via phase transfer–mediated nucleophilic ¹⁸F-fluorination of N-trityl-O-(2-tosyloxyethyl)-l-tyrosine tert-butyl ester and subsequent deprotection (19). The uncorrected radiochemical yield was about 35% at a specific radioactivity of more than 200 GBq/μmol and a radiochemical purity of more than 98%. The tracer was administered as an isotonic neutral solution. PET studies were acquired 15–40 min after the intravenous injection of 200 MBq of ¹⁸F-FET. The measurements were performed on an ECAT scanner with 32 rings and an axial field of view of 15.5 cm (EXACT HR+; Siemens Medical Systems), in 3-dimensional mode. For attenuation correction, transmission scans with three ⁶⁸Ge/⁶⁸Ga rotating line sources were measured. After correction for random and scattered coincidences and dead time, image data were obtained by filtered backprojection in Fourier space using the ECAT 7.2 software (direct inverse Fourier transformation; Shepp filtering [2.48 mm in full width at half maximum]; pixel size, 2 × 2 × 2.4 mm). The reconstructed images were decay-corrected; the reconstructed image resolution was about 5.5 mm.

Preoperative MR images and ¹⁸F-FET PET images were coregistered and evaluated using regions of interest (ROIs) and dedicated software (MPI tool, version 3.28; ATV). For tumors with increased ¹⁸F-FET uptake, the transaxial slice showing the highest tumor intensity was chosen and an isocontour region around the tumor maximum was drawn automatically using a cutoff of 3 SDs above the average activity in the reference region. A larger reference ROI was placed on the contralateral hemisphere in an area of normal-appearing brain that included white and gray matter. Because one third of the low-grade glioma exhibited indifferent ¹⁸F-FET uptake, an objective positioning of the ROI on the PET scan based on threshold values was impossible. Therefore, a single irregular ROI was placed manually in an area of signal abnormality on the T1- and T2-weighted transversal MRI scan and transferred to the coregistered ¹⁸F-FET PET scan in each case. Mean and maximal lesion-to-brain ratios were calculated by dividing the mean and maximal ROI value (Bq/mL) of the lesion by the mean ROI value of normal brain in the ¹⁸F-FET PET scan. In a previous biopsy-controlled study, we found for tissue samples corresponding to normal and peritumoral tissue a mean lesion-to-brain ratio of 1.2 ± 0.4 for ¹⁸F-FET uptake (17,18). For prognostic purposes, however, a statistical preevaluation with different mean lesion-to-brain ratios identified a threshold value of 1.1 as optimal for predicting the course and outcome of the patients. Therefore, in this study, lesions with a mean lesion-to-brain ratio of 1.1 or less for tracer uptake were judged as ¹⁸F-FET–negative and lesions with a ratio of more than 1.1 as ¹⁸F-FET–positive. For the maximum lesion-to-brain ratios, a threshold value of 2.0 was chosen indicating hot-spot areas in the tumors.

Histopathology

Initial cytologic assessment was performed on touch preparations during biopsy to ensure representative tissue sampling for subsequent routine histologic preparations. The final diagnoses were established on formalin-fixed and paraffin-embedded tissue samples according to the WHO classification of tumors of the nervous system (20). Additional immunohistochemical analyses were performed with antibodies against the tumor suppressor protein p53 (Clone D07, primary antibody dilution: 1:100; Dako) and the proliferation-associated antigen Ki-67 (Clone Mib1, primary antibody dilution: 1:200; Dako) according to standard protocols. Immunohistochemical results were quantified using the following semiquantitative staining scores: for Ki-67, 0 (no positive tumor cells), 1 (<1% positive tumor cells), 2 (1%–5% positive tumor cells), or 3 (>5% positive tumor cells) and for p53, 0 (no positive tumor cells), 1 (individual positive tumor cells), 2 (moderate fraction of positive tumor cells [<50%]), or 3 (high fraction of positive tumor cells [≥50%]).

Statistical Analysis

The prognostic significance of various factors regarding the primary endpoints was determined using the Kaplan–Meier method and tested using censored linear rank tests, with each factor considered a separate, independent variable. The computations were done with the LIFETEST procedure of SAS system release 9.1 (SAS Institute Inc.). Probability values of less than 0.01 were considered significant. No correction for multiple statistical comparisons was done.

Two prognostically significant variables were identified that allowed for the definition of 3 prognostic groups. The homogeneity of the distributions of each of 3 variables—“time until progression,” “time until malignant transformation,” and “time until death”—over these groups was tested using the log-rank test. The rank tests for the association of these 3 variables with each of the other factors (Table 3) adjusted for group levels were not significant. Thereby, we excluded the possibility that the differences in clinical course and outcome between the 3 prognostic groups were influenced by a combination of other factors.

Prognostic Factors

An overview of the distribution of the clinical MRI, morphologic, PET metabolic, histologic, and therapeutic variables and the data on the patients' clinical courses and outcomes is given in Tables 1 and 2. The results of the statistical analysis are given in Table 3. A significant influence on the course of the disease was found for 2 prognostic variables only. The first of these—circumscribed versus diffuse tumor growth on baseline MRI—reached the highest level of significance, with P values of less than 0.0001 for time until progression, less than 0.0001 for time until malignant transformation, and 0.008 for time until death. The second of the significant variables was mean ¹⁸F-FET uptake on baseline PET, with P values of 0.001 for time until progression and 0.006 for time until malignant transformation. Statistical significance was not reached for time until death (P = 0.06), but it is important to note that all 6 deaths occurred in patients with ¹⁸F-FET–positive tumors.

Prognostic Groups

Through the combination of the 2 significant prognostic variables, 3 major prognostic groups were identified: patients having circumscribed tumors without ¹⁸F-FET uptake (Fig. 1), patients having circumscribed tumors with ¹⁸F-FET uptake, and patients having diffuse tumors with ¹⁸F-FET uptake (Fig. 1). None of the patients had a diffuse tumor without ¹⁸F-FET uptake, thus excluding the creation of a fourth subgroup. Among the first, second, and third prognostic group, the number of patients (11, 13, and 9, respectively) and the average time of follow-up per patient (32, 26, and 29 mo, respectively) were fairly similar.

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FIGURE 1. 

Examples of low-grade gliomas in prognostic groups A, B, and C. On the left are contrast-enhanced T1-weighted MR images, in the middle are T2-weighted MR images (FLAIR sequence), and on the right are ¹⁸F-FET PET images. Top row (prognostic group A) shows circumscribed tumors and no FET uptake, middle row (prognostic group B) shows circumscribed tumors with increased FET uptake, and bottom row (prognostic group C) shows diffuse tumors with increased ¹⁸F-FET uptake. Histopathology after biopsy showed diffuse astrocytoma of WHO grade II in each patient. None of the tumors showed contrast enhancement on T1-weighted MRI.

Statistical comparison of the 3 groups revealed significant differences in time until progression (P < 0.0001), time until malignant transformation (P < 0.0001), and time until death (P = 0.01) (Fig. 2). The rank tests for the association with each other factor proved that the differences between the 3 groups were not influenced by a combination of other factors. Figure 3 shows the Kaplan–Meier curves for time until progression in the 3 prognostic groups.

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FIGURE 2. 

Graph of follow-up results for each patient. Green portion of each bar represents progression-free survival after initial diagnosis of low-grade glioma. Yellow portion indicates time of first tumor progression. If histology of low-grade glioma remained unchanged, color of bar stays green. If malignant transformation to high-grade glioma occurred, bar becomes red. Time of patient death is indicated by black portion of bar. (A) Prognostic group A, with favorable clinical course, rare progression of tumor (18%), and no malignant transformation or death during follow-up. (B) Prognostic group B, with less favorable clinical course, frequent tumor progression (46%), and sometimes early tumor progression. Small subsets of patients showed malignant transformation of tumors (15%) and/or died (8%) during follow-up. (C) Prognostic group C, with rapid clinical course and poor outcome, 100% tumor progression, and usually early tumor progression. Most patients showed malignant transformation of tumors (78%) and died of the disease (56%).

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FIGURE 3. 

Example Kaplan–Meier plot of follow-up endpoint. Tumor progressed frequently (100%, n = 9) and early in prognostic group C, often (46%, n = 6) and sometimes early in prognostic group B, and rarely (18%, n = 2) and late in prognostic group A. Differences between the 3 groups were highly significant (P < 0.0001).