Maximum Lesion-to-contralateral Normal Brain Tissue Ration of 11C-methionine PET as a Prognostic Imaging Biomarker for Newly Diagnosed and Untreated Astrocytic Glioma

Purpose: This study aimed whether the uptake of amino tracer positron emission tomography (PET) can be used as an additional imaging biomarker to estimate the prognosis of glioma. Methods: Participants comprised 56 adult patients with newly diagnosed and untreated World Health Organization (WHO) grade(cid:0)-(cid:0) astrocytic glioma who underwent surgical excision and were evaluated by 11C-methionine PET prior to the surgical excision at Osaka City University Hospital from July 2011 to March 2018. Clinical and imaging studies were retrospectively reviewed based on medical records at our institution. Results: Preoperative Karnofsky Performance Status (KPS) only inuenced progression-free survival (PFS) (hazard ratio [HR] 0.20; 95% condence interval [CI] 0.10-0.41, p<0.0001), whereas histology (anaplastic astrocytoma: HR 5.30, 95%CI 1.23-22.8, p=0.025; glioblastoma: HR 11.52, 95%CI 2.27-58.47, p=0.0032), preoperative KPS ≥ 80 (HR 0.23, 95%CI 0.09-0.62, p=0.004), maximum lesion-to-contralateral normal brain tissue (LN max) ≥ 4.03 (HR 0.24, 95%CI 0.08-0.71, p=0.01), and isocitrate dehydrogenase (IDH) status (HR 14.06, 95%CI 1.81-109.2, p=0.011) were factors inuencing overall survival (OS) in multivariate Cox regression. OS was shorter in patients with LN max ≥ 4.03 (29.3 months) than in patients with LN max<4.03 (not reached; p=0.03). OS differed signicantly between patients with IDH mutant/LN max<4.03 and patients with IDH mutant/LN max ≥ 4.03. Conclusions: LN max using 11C-methionine PET may be used in prognostic markers for newly identied and untreated WHO grade(cid:0)-(cid:0) astrocytic glioma. IDH mutant astrocytoma showed lower-grade astrocytoma especially diffuse astrocytoma, whereas the majority of patients with IDH wild-type astrocytoma had high-grade glioma, especially glioblastoma (p = 0.009). Median LN mean and LN max were 2.46, and 4.03, respectively. Approximately 70% of patients with LN max ≥ 4.03 showed glioblastoma, whereas nearly 60% of patients with LN max < 4.03 showed diffuse astrocytoma (p < 0.0001). According to IDH status/LN max classication, 10 patients with diffuse astrocytoma, one patient with anaplastic astrocytoma, and one patient with glioblastoma were classied into the IDH mutant with LN max < 4.03 group. One patient with anaplastic astrocytoma, and two patients with glioblastoma were classied into the IDH mutant with LN max ≥ 4.03 group. Seven patients with diffuse astrocytoma, ve patients with anaplastic astrocytoma, and four patients with glioblastoma were classied into the IDH wild-type with LN max < 4.03 group. Two patients with diffuse astrocytoma, ve patients with anaplastic astrocytoma, and 18 patients with glioblastoma were classied into the IDH wild-type with LN max ≥ 4.03 group. methyltransferase; LN max using 11C-methionine PET offers a markers for estimating OS in patients with grade(cid:0)-(cid:0) astrocytoma. LN max can also be used as a prognostic imaging biomarker to estimate OS in addition to IDH status in IDH-mutated astrocytoma. 18F-FDOPA:


Introduction
Gliomas are the second most common primary brain tumors according to the 2012-2016 Central Brain Tumor Registry of the United States. [1] Approximately 48.3% of primary malignant brain tumors are glioblastomas, 16.7% are other astrocytomas, and 4.5% are oligodendrogliomas. [1] Overall incidence rates for diffuse astrocytoma, anaplastic astrocytoma, and oligodendroglioma have been decreasing recently, but overall incidence rates for glioblastoma have remained stable. [2] Although magnetic resonance imaging (MRI) has been one of the basic and less-invasive imaging modalities used in the management of glioma, brain PET imaging has recently been recommended. [3,4] We have previously reported a positive correlation between WHO grade and accumulation of 11C-methionine among astrocytomas, but that study did no analyze the relationship with prognosis. [5] Additional analysis was thus performed in the current study. Moreover, the clinical studies investigating the relationship between molecular analysis and uptake of amino acid PET in glioma patients are sparse, and detailed prognostic analyses of associations with molecular pro les and 11C-methionine PET uptake in glioma patients have not been fully completed. This study aimed to evaluate the association between 11Cmethionine uptakes, gene status, and prognosis in cases of newly diagnosed and untreated adult astrocytic glioma.

Patients
From July 2011 to March 2018, there were 66 adult patients and two patients under 18 years old with newly diagnosed and untreated WHO grade -glioma who underwent surgical tumor resection and preoperative 11C-methionine PET examination, as previously reported. [5] From this previous cohort, we included adult astrocytic glioma patients with IDH mutated-TERT promoter wild-type, or those with IDH wild-type in the present study. Finally, a total of 56 patients with astrocytic tumor were included in the present cohort. The 56 patients were comprised of 36 male and 20 female patients, with a mean age of 54.0 years (range, 21-82 years). All 11C-methionine PET was performed within one month prior to tumor resection in glioblastoma patients and within six months in patients with lower-grade glioma. Pathological diagnosis was determined according to the 2016 WHO classi cation for central nervous system tumors.
[6] This study was approved by the institutional review boards at the Graduate School of Medicine, Osaka City University (approval numbers: 2047 and 2020 − 115), and Osaka National Hospital (approval number: 0713). Genetic analyses were performed after obtaining written consent.

11C-methionine PET
An Eminence B PET scanner (Shimadzu, Kyoto, Japan) or Biograph-16 PET scanner (Siemens, Bon, Germany) was used for 11Cmethionine PET, according to previously reported procedures. [5,7] Mean and maximum lesion-to-contralateral normal brain tissue (L/N) ratios were determined by dividing the tumor standardized uptake value by the mean standardized uptake value of the normal contralateral region of the brain, as previously reported. [5] Genetic Analysis Genetic analysis was performed as previously described. [5] Genomic DNA was extracted from surgically resected tumor specimens using the DNeasy Blood & Tissue Kit (Qiagen, Valencia, CA, USA) or NucleoSpin Tissue (Machery-Nagel, Duren, Germany). Hotspot mutations of IDH1/2 (codon 132 of IDH1 and codon 172 of IDH2) and TERT promoter (termed C228 and C250) were examined using Sanger sequencing with a 3130xLGenetic Analyzer (Thermo Fisher Scienti c, Waltham, MA, USA) and Big-Dye® Terminator v1.1 Cycle Sequencing Kit (Thermo Fisher Scienti c, Waltham, MA, USA).

Survival Times
Progression-free survival (PFS) was de ned as the time in months between evaluation with 11C-methionine PET and tumor progression according to the Response Assessment in Neuro-oncology working group.
[8] Overall survival was de ned as the time in months between evaluation with 11C-methionine PET and death.
To compare the patients background characteristics of each group classi ed according to IDH status or LN max or both, we performed statistical analysis using Pearson's chi-square test. PFS and OS were analyzed using the Kaplan-Meier method. Survival date were evaluated using uni-and multivariate Cox regression analyses. The stepwise method was used to evaluate PFS and OS multivariate Cox regression analyses. Statistical signi cance was de ned at the level of p < 0.05. All statistical analyses were conducted using EZR software (Saitama Medical Center, Jichi Medical University, Saitama, Japan). [9] Results Patient Characteristics Patient characteristics are summarized in Table 1. Majority of patients with IDH mutant astrocytoma showed lower-grade astrocytoma especially diffuse astrocytoma, whereas the majority of patients with IDH wild-type astrocytoma had high-grade glioma, especially glioblastoma (p = 0.009). Median LN mean and LN max were 2.46, and 4.03, respectively. Approximately 70% of patients with LN max ≥ 4.03 showed glioblastoma, whereas nearly 60% of patients with LN max < 4.03 showed diffuse astrocytoma (p < 0.0001). According to IDH status/LN max classi cation, 10 patients with diffuse astrocytoma, one patient with anaplastic astrocytoma, and one patient with glioblastoma were classi ed into the IDH mutant with LN max < 4.03 group. One patient with anaplastic astrocytoma, and two patients with glioblastoma were classi ed into the IDH mutant with LN max ≥ 4.03 group. Seven patients with diffuse astrocytoma, ve patients with anaplastic astrocytoma, and four patients with glioblastoma were classi ed into the IDH wild-type with LN max < 4.03 group. Two patients with diffuse astrocytoma, ve patients with anaplastic astrocytoma, and 18 patients with glioblastoma were classi ed into the IDH wild-type with LN max ≥ 4.03 group.   Fig. 2A). Median OS was more favorable in patients with diffuse astrocytoma, anaplastic astrocytoma, and glioblastoma were not applicable, 27.1 months, and 20.5 months, respectively (p < 0.0001,

Discussion
The revised WHO 2016 classi cation of the central nervous system tumor requires the pathological diagnosis with molecular analysis to reach a diagnosis of glioma.
[6] This molecular information has been said to correlate with prognosis, whereas there is still a matter of debate whether imaging biomarkers help estimation of prognosis. Although MRI remains the gold standard for diagnosing glioma, its role in estimating prognosis is limited. [10] On the other hand, 11C-methionine PET using amino tracer might be useful to detect the tumor, predict the grade or genetic status or both, [5,[11][12][13][14] and distinguish tumor recurrence from radiation necrosis [15][16][17] in glioma patients. However, relatively few reports have investigated the relationship between the uptake of amino tracer using PET and prognosis in glioma.
Moreover, reports investigating prognosis of glioma patients in association with molecular analysis and PET in glioma have been limited.
[18-22] Thus, our goal in the present study was to determine whether 11C-methionine PET can be used as an additional imaging biomarker of prognosis.
In the present study, we excluded patients with oligodendroglioma, or those with IDH mutated-TERT promoter mutated, or both because oligodendroglioma is considered to show better prognosis than astrocytoma and is often accompanied by both IDH and TERT promoter mutations. Although TERT promoter mutation is often seen in oligodendroglioma and primary glioblastoma, prognoses differ markedly between oligodendroglioma and glioblastoma. [23,24] An argument has also been made regarding the association between uptake of 11C-methionine and oligodendroglioma. [14,[25][26][27][28] We have previously reported a positive correlation between WHO grade and the accumulation of 11C-methionine among astrocytomas, and a statistically higher uptake of 11C-methionine in oligodendroglioma than in diffuse astrocytoma. [5] Median survival rates in patients with diffuse astrocytoma, anaplastic astrocytoma, and glioblastoma in this study were 37. Recently, some reports have investigated the relationship between prognosis from molecular analysis and uptake of PET using 18F-uoroethyl-tyrosine (18F-FET) PET[18-20, 33] and 3,4-dihydroxy-6-18F-uoro-ethyl-L-phenylalanine (18F-FDOPA) PET [34]. Galldiks et al. in a study of photopenic IDH mutant gliomas reported that glioma with 18F-FET accumulation below the level of background healthy brain showed unfavorable outcomes, and thus should be treated more actively. [19] The utility of dynamic 18F-FET PET has also been reported.
[20] Suchorska et al. reported that longer minimal time-to-peak analysis using 18F-FET PET was associated with a favorable prognosis in IDH mutant astrocytomas. [20] A time-to-peak analysis ≥ 25 min was associated with longer PFS and OS in patients with IDH wild-type high-grade astrocytoma according to Bauer  Further study with a larger cohort is thus needed to assess the correlation between prognosis and molecular/imaging biomarkers with amino-tracer PET in patients with astrocytoma. Second, we did not take volumetric analyses into consideration in the current study, although some reports have suggested that metabolic tumor volume did not correlate with survival outcomes. [18,20,33,34,36] Conclusion LN max using 11C-methionine PET offers a markers for estimating OS in patients with grade -astrocytoma. LN max can also be used as a prognostic imaging biomarker to estimate OS in addition to IDH status in IDH-mutated astrocytoma.   Kaplan