DOI: https://doi.org/10.21203/rs.3.rs-480178/v1
Background: Patients with low-grade gliomas (LGGs) harboring O6-methylguanine-DNA methyltransferase promoter nonmethyaltion (MGMT-non-pM) have a particularly short survival and are greatly resistance to chemotherapy. The objective of this study was to assess the efficacy of high-dose radiotherapy (RT) for LGGs with MGMT-non-pM.
Methods: 269 patients with newly diagnosed adult supratentorial LGGs from the multicenter Chinese Glioma Cooperative Group (CGCG) received postoperative RT during 2005-2018. MGMT promoter methylation analysis was conducted by pyrosequencing in all patients. Univariate and multivariable analyses were performed using the cox regression to determine the prognostic factors for overall survival (OS) and progression-free survival (PFS). RT dose-response on MGMT status defined subtypes was analyzed.
Results: On univariate analysis, the following were statistically significant favorable factors for both PFS and OS: oligodendrogliomas（p = 0.002 and p = 0.005）, high-dose RT (> 54 Gy) （p = 0.017 and p = 0.023） and 1p/19q codeletion （p <0.001 and p = 0.001）. On multivariable analyses, RT dose (> 54 Gy vs. ≤ 54 Gy) and IDH mutation were independently prognostic markers for OS (HR, 0.44; 95%CI, 0.21-0.92; p = 0.029; and HR, 0.43; 95%CI, 0.20-0.90; p = 0.025, respectively) and PFS (HR, 0.48; 95%CI, 0.27-0.90; p = 0.021; and HR, 0.52; 95%CI, 0.27-0.98; p = 0.044, respectively). High-dose RT was associated with longer OS (HR, 0.55; 95%CI, 0.32-0.93; p = 0.026) and PFS (HR, 0.57; 95%CI, 0.35-0.93; p = 0.026) than low-dose RT in MGMT-non-pM subtype. In contrast, no significant difference in either OS (p = 0.240) or PFS (p = 0.395) was observed with high-dose RT in the MGMT-pM subtype.
Conclusions: High-dose RT (> 54 Gy) is an independently protective factor for LGGs and associated with improved survival in patients with MGMT-non-pM.
Low-grade gliomas (LGGs) mainly refers grade II by the WHO grading system and are relatively uncommon, constituting approximately 10% of all primary brain tumors in adults [1–2]. Although often considered as “benign”, over half of these patients will develop tumor progression within 5 years and the rate of progression-free survival (PFS) at 10 years was 21–51% [3–4]. Therefore, postoperative radiotherapy (RT) is frequently utilized after surgical resection. Recently, molecular alterations, especially isocitrate dehydrogenase 1/2 mutation (IDH mutation) and chromosome arm 1p/19q codeletion (1p/19q codeletion), provide important diagnostic and prognostic information that can greatly improve diagnostic accuracy and management decision-making in patients with LGGs [5]. IDH mutation and 1p/19q codeletion are required for LGGs classification within the revised 2016 WHO guidelines. However, O6-methylguanine-DNA methyltransferase promoter methylation (MGMT-pM) was rarely reported in patients with LGGs, even though it accounts for about 79–92% in these patients [6–7]. Only one study RTOG (Radiation Therapy Oncology Group) 0424 has so far reported the association of MGMT status with survival of patients with LGGs [8]. In this study, MGMT status was an independent prognostic biomarker of high-risk, LGGs treated with radiotherapy combined with concomitant and adjuvant temozolomide (TMZ) chemothrapy. A survival benefit was observed in LGGs contained a methylated MGMT; Similar with glioblastoma [9], MGMT-non-pM confers a shorter OS (3 years vs. not reached) and PFS (2 years vs. not reached) compared with MGMT-pM in high-risk LGGs. Unfortunately, most of clinical trials tended to test new drugs as alternatives to TMZ for patients with MGMT-non-pM have failed. Bevacizumab plus irinotecan, paclitaxel poliglumex with RT, Cilengitide combined with TMZ, temsirolimus and procarbazine et al. have been proved to be not satisfactory in nonmethylated GBM [10–13]. Thus, new therapies for these patients are urgently needed.
Because of the requirements for long-term follow-up for patients with LGGs, most of studies on RT dose were conducted early, before the year 1990, and have many limitations in both diagnostic (computed tomography, CT) and treatment modalities (2D planning). However, modern technology (intensity-modulated radiation therapy, IMRT and magnetic resonance imaging, MRI) can greatly improve dose distribution of targeted field and reduce dose of adjacent structure. Therefore, we hypothesize that RT dose escalation might be effective in LGGs with MGMT-non-pM based on modern technology. In this study, we analyzed retrospectively the potential survival benefits of high-dose RT (> 54Gy) in 269 patients with LGGs containing the information of MGMT promoter methylation. These data provide evidence for making treatment decisions and designing clinical trials for LGGs based on MGMT status.
269 patients with newly diagnosed adult supratentorial LGGs (WHO II) were obtained from the multicenter Chinese Glioma Cooperative Group (CGCG) and the Chinese Glioma Genome Atlas (CGGA) in China during 2005–2018 (www.cgga.org.cn). Tumor histology was confirmed independently by two neuropathologists based on the 2007 WHO classification and the 2016 updated edition. The study protocol was approved by the Ethics Review Board of Tiantan Hospital in Beijing, China. Written informed consent was obtained from all participants. The patients had to be in good general condition as indicated by performance score after surgery: Karnofsky Performance Scores ≥ 60. Patient characteristics (stratified by the MGMT status) are summarized in Table 1.
|
characteristics |
n (%) |
MGMT-pM n (%) |
MGMT-non-pM n (%) |
|---|---|---|---|
|
Total |
269 |
115 (42.8) |
154 (57.2) |
|
Sex |
269 (100) |
||
|
Male |
153 (56.9) |
62 (53.9) |
91 (59.1) |
|
Female |
116 (43.1) |
53 (46.1) |
63 (40.9) |
|
Age (years) |
269 (100) |
||
|
≤ 40 |
154 (57.2) |
55 (47.8) |
99 (64.3) |
|
> 40 |
115 (42.8) |
60 (52.2) |
55 (35.7) |
|
Histopathology |
269 (100) |
||
|
A* |
221 (82.2) |
83 (72.2) |
138 (89.6) |
|
O |
48 (17.8) |
32 (27.8) |
16 (10.4) |
|
Seizure |
205 (72.6) |
||
|
Yes |
122 (59.5) |
33 (56.9) |
89 (60.5) |
|
No |
83 (40.5) |
25 (43.1) |
58 (39.5) |
|
Resection |
249 (92.6) |
||
|
Total |
116 (46.6) |
57 (54.3) |
59 (41.0) |
|
Subtotal |
133 (53.4) |
48 (45.7) |
85 (59.0) |
|
RT dose |
269 (100) |
||
|
High |
155 (57.6) |
64 (55.7) |
91 (59.1) |
|
Low |
114 (42.4) |
51 (44.3) |
63 (40.9) |
|
Chemotherapy |
261 (97.0) |
||
|
Yes |
88 (33.7) |
42 (38.9) |
46 (30.1) |
|
No |
173 (66.3) |
66 (61.1) |
107 (69.9) |
|
IDH muation |
251 (93.3) |
||
|
Yes |
206 (82.1) |
93 (92.1) |
113 (75.3) |
|
No |
45 (17.9) |
8 (7.9) |
37 (24.7) |
|
1p/19q codeletion |
162 (60.2) |
||
|
Yes |
63 (38.9) |
43 (50.0) |
20 (26.3) |
|
No |
99 (71.1) |
43 (50.0) |
56 (73.7) |
| *A: including astrocytoma and oligoastrocytoma which was eliminated from the 2016 WHO classification. | |||
All patients underwent surgical excision and postoperative three-dimensional conformal radiotherapy (3DCRT) or IMRT. The radiation fields were based on the postoperative T2 or FLAIR MRI-defined residual tumor and/or surgical cavity plus a 2 cm margin. The median dose was 55.8 Gy (range, 40–66 Gy) (1.8-2.0 Gy daily, 5 days per week). The distribution of doses in LGG patients was shown in Supplementary Fig. 1. All patients received RT at 4–8 weeks after surgery. The extent of resection was evaluated using preoperative and postoperative MRI. A total of 33.7% (88/261) of patients received RT followed by chemotherapy using carmustine, nimustine, or TMZ. In the first 2 years, follow-up and MRI were performed after RT every 6 months, and every 9–12 months thereafter until tumor progression.
DNA was extracted in formalin-fixed paraffin-embedded samples with a QIAamp DNA FFPE Tissue Kit (Qiagen, Hilden, Germany). Then 100 ng DNA was bisulfite converted with an Epitect Bisulfite kit (Qiagen, Hilden, Germany) according to the manufacturer’s protocol. The bisulfite-treated DNA was amplified and then sequenced by pyrosequencing. The amplification forward primer 5’-GTTTYGGATATGTTGGG ATAGTT-3’ and the biotinylated reverse primer 5’-biotin-ACRACCCAAACACTCA CCAA-3’. The methylation levels of CpG sites 75–78 were obtained with the sequencing primers 5’-GATATGTTGGGATAGT − 3’ or 5’-GTTTTTAGAAYGTTTT G-3’. The methylation levels of CpG sites 76–79 were detected with a commercial MGMT pyrosequencing kit (Qiagen, Hilden, Germany) with a PyroMark Q24 System (Qiagen, Hilden, Germany). Standardized positive and negative controls were included in all routine pyrosequencing testing, and every test was performed by 2 experienced molecular neuropathologists together (Supplementary Fig. 2).
The clinical features of the different groups were compared using the χ2 test with SPSS v22.0 (IBM, Armonk, NY, USA). OS and PFS curves were estimated by the Kaplan–Meier method and compared with use of the two-sided log-rank test. OS was calculated from the day of surgery to the date of the first event. The date of progression was defined as the date of the CT or MRI examination that confirmed progression or related neurologic symptoms. Cox proportional hazards regression was used to identify independent risk factors for OS and PFS. P < 0.05 (two-sided) was considered to indicate statistical significance.
Among all patients enrolled in this study, the median age was 38 years (range, 11–69 years), and the male-to-female ratio was 1.32:1 (153:116). Median follow-up time was 9.12 (7.93–10.30) years. There have been 79 deaths (29.4%) and 101 recurrences (37.5%) to date. Of the 269 samples, 221 (82.2%) were astrocytoma or oligoastrocytoma (oligoastrocytoma was essentially eliminated based on the molecular pathology on the updated WHO classification in 2016) and 48 (17.8%) were oligodendrogliomas. The 5-year OS and PFS rates were 80.6% and 73.3% in all patients. Median PFS was 11.4 years, and median OS was not yet reached. The baseline characteristics of patients, stratified by MGMT status, are reported in Table 1.
A dose of 54 Gy was extensively used in clinical decisions and trials of LGGs [1, 3, 8, 14]. Depending on the dose of 54 Gy, we divided patients into 2 groups: high dose (> 54 Gy,) and low dose (≤ 54 Gy). On univariate analysis, the following were statistically significantly favorable factors for both PFS and OS: oligodendrogliomas (p = 0.002 and p = 0.005), high-dose RT ༈p = 0.017 and p = 0.023༉ and 1p/19q codeletion ༈p < 0.001 and p = 0.001༉. IDH mutation༈p = 0.076༉and seizure symptom༈p = 0.074༉indicate a favorable prognosis, although the difference was not significant in analysis of OS and PFS, respectively. Multivariate analysis showed that high-dose RT (HR, 0.44; 95% CI, 0.21–0.92; p = 0.029; HR, 0.48; 95% CI, 0.27–0.90; p = 0.021, respectively) and IDH mutation (HR, 0.43; 95% CI, 0.20–0.90; p = 0.025; HR, 0.52; 95% CI, 0.27–0.98; p = 0.044, respectively) were significantly prognostic factors of both OS and PFS. 1p/19q codeletion ༈p = 0.072༉indicates a favorable prognosis, although the difference was not significant in analysis of OS (Table 2).
|
Variables |
n |
Univariate analyses |
Multivariate analyses |
||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
|
PFS |
OS |
PFS |
OS |
||||||||||
|
HR |
95% CI |
p |
HR |
95% CI |
p |
HR |
95% CI |
p |
HR |
95% CI |
p |
||
|
Age ≤ 40 vs. ༞40 |
269 115/154 |
1.08 |
0.72–1.61 |
0.73 |
1.03 |
0.65–1.62 |
0.907 |
1.09 |
0.60–1.97 |
0.78 |
1.32 |
0.68–2.57 |
0.417 |
|
Sex Male vs. female |
269 153/116 |
1.19 |
0.80–1.78 |
0.395 |
1.03 |
0.66–1.60 |
0.907 |
1.83 |
1.00-3.35 |
0.050 |
1.26 |
0.64–2.48 |
0.498 |
|
Histopathology A* vs. O |
269 221/48 |
3.20 |
1.55–6.59 |
0.002 |
3.32 |
1.44–7.64 |
0.005 |
2.20 |
0.78–6.18 |
0.135 |
1.88 |
0.58–6.05 |
0.290 |
|
Seizure Yes vs. no |
205 122/83 |
0.69 |
0.46–1.04 |
0.074 |
0.63 |
0.40–0.99 |
0.045 |
0.86 |
0.47–1.58 |
0.625 |
0.86 |
0.43–1.70 |
0.657 |
|
Resection Total vs. subtotal |
249 116/133 |
0.61 |
0.40–0.94 |
0.024 |
0.77 |
0.48–1.24 |
0.280 |
0.83 |
0.45–1.53 |
0.826 |
0.94 |
0.47–1.87 |
0.849 |
|
Chemotherapy Yes vs. no |
261 88/173 |
1.56 |
1.04–2.34 |
0.030 |
1.00 |
0.62–1.61 |
0.991 |
1.46 |
0.81–2.63 |
0.204 |
1.00 |
0.49–1.99 |
0.972 |
|
Dose > 54 Gy vs. ≤54 Gy |
269 155/114 |
0.62 |
0.42–0.92 |
0.017 |
0.60 |
0.38–0.93 |
0.023 |
0.48 |
0.27–0.90 |
0.021 |
0.44 |
0.21–0.92 |
0.029 |
|
IDH mutation Yes vs. no |
251 206/45 |
0.59 |
0.36–0.96 |
0.033 |
0.61 |
0.36–1.05 |
.076 |
0.52 |
0.27–0.98 |
0.044 |
0.43 |
0.20–0.90 |
0.025 |
|
1p/19q co-deletion Yes vs. no |
162 63/99 |
0.31 |
0.16–0.58 |
0.000 |
0.27 |
0.12–0.57 |
.001 |
0.43 |
0.19–0.97 |
0.042 |
0.41 |
0.15–1.08 |
0.072 |
|
MGMT pM Yes vs. no |
269 115/154 |
0.82 |
0.53–1.28 |
0.389 |
0.65 |
0.38–1.12 |
0.119 |
0.79 |
0.43–1.44 |
0.436 |
0.63 |
0.30–1.31 |
0.217 |
| * A including astrocytoma and oligoastrocytoma which was eliminated in the 2016 WHO classification | |||||||||||||
| PFS: progression-free survival; OS: overall survival; HR: hazard ratio; CI: confidence interval | |||||||||||||
MGMT promoter methylation was profiled in all patients. A significant protective effect on PFS and OS with a RT dose ≥ 54 Gy was observed in patients with MGMT-non-pM (HR, 0.57; 95% CI, 0.35–0.93; p = 0.026; and HR, 0.55; 95% CI, 0.32–0.93; p = 0.026, respectively) (Fig. 1A and B), but this was not the case in patients with MGMT-pM (p = 0.40 in PFS and p = 0.240 in OS) (Fig. 1C and D). Most clinical characteristics were comparable between groups (Supplementary Table 1). Among 261 patients, 88 received RT followed by chemotherapy (carmustine, nimustine, or TMZ). But patients with MGMT-pM did not receive benefit from the addition of chemotherapy (p = 0.058 in PFS and p = 0.195 in OS) (Supplementary Fig. 3A and B). Chemotherapy did also not improved the OS (p = 0.826) and PFS (p = 0.109) in patients with MGMT-non-pM (Supplementary Fig. 3C and D).
Gliomas with MGMT-non-pM are striking resistance to chemotherapy or targeted therapy. In our study, high-dose RT (> 54 Gy) was an independently protective factor of patients with LGGs. More importantly, patients with MGMT-non-pM can benefit from high-dose RT, but no benefit was observed with high-dose RT in patients with MGMT-pM. The results showed that replacement of TMZ chemotherapy by high-dose RT might be feasible for these patients with MGMT-non-pM. To the best of our knowledge, this is the first report on the relationship between RT dose and MGMT status. MGMT status could serve as the primary predictor of response to RT in LGGs.
MGMT is a DNA repair protein and a marker of resistance to the first line chemotherapeutic drug (TMZ). Methylated MGMT resulted in reduced protein and is a strong prognostic and predictive biomarker for benefit from TMZ chemotherapy in patients with GBM, especially in elderly patients [15–16]. Even in patients with treatment by only radiotherapy, MGMT-pM also confers a survival advantage [9, 17]. However, patients with MGMT-non-pM derive less benefit from TMZ or other alkylating agents and have a shorter median survival compared to those whose tumors are methylated (15.3 months vs. 21.7 months). Though many trials have tried to test new drugs as alternatives to TMZ, none of these was effective against unmethylated GBM. LGGs have relatively higher rates (75-92.5%) of MGMT-pM than GBM, but the association of MGMT status with survival of LGGs rarely reported. In RTOG 0424, MGMT-pM was found in 76% (57/75) of high-risk LGGs and was an independent prognostic biomarker based on RT and adjuvant TMZ chemotherapy. MGMT-non-pM was significantly associated with worse OS and PFS than MGMT-pM in high-risk LGGs [8]. However, the implication of MGMT status with respect to radio-chemotherapy sensitivity in patients with LGGs is not further studied.
Learning from the failed clinical trials in GBM with MGMT-non-pM, we hypothesize that RT dose escalation might be effective in these refractory tumors. Earlier retrospective studies have observed a dose-response relationship in LGGs. Although these studies were retrospective and had limited sample sizes (< 150 patients), they found that high-dose RT (> 52 Gy, > 53 Gy, or even > 55 Gy) confers a survival advantage compared with those who received low-dose RT (< 52 Gy, < 53 Gy, or even < 55 Gy) [18–20]. However, two randomized trials (the European Organisation for Research and Treatment of Cancer 22844 and the North Central Cancer Treatment Group 86-72-51) did not show an OS or PFS benefit to high-dose RT (59.4 Gy and 64.8 Gy) over low-dose RT (45 Gy and 50.4 Gy) [21–22]. The point to emphasize here is that these studies were activated in 1985 and 1986, respectively, and have many limitations in diagnostic and treatment modalities. Patients were treated in an era with older surgical, diagnostic instrument (CT scan) and radiation techniques (2D planning). Currently, IMRT and MRI are routinely used in clinical practice that has been a significant improvement in dose distribution of targeted field and dose limitation of adjacent structure [23]. More importantly, molecular pathology provides additional diagnostic and prognostic information that can greatly improve diagnostic accuracy and management decision-making. Therefore, it is needed to be reconsidered based on modern technology whether high-dose RT can obtained improved survival in some molecular subtypes. In our study, 269 patients with newly diagnosed LGGs received postoperative 3DCRT or IMRT. The results that RT dose is an independent prognostic factor for both OS and PFS indicated that the survival of patients with LGGs might be further improved by increasing RT dose in modern technology. High-dose RT was associated with longer PFS and OS in MGMT-non-pM subtype. In contrast, no significant difference in survival was observed with high-dose RT in the MGMT-pM subtype. The data showed that high-dose RT as alternatives to TMZ or other drugs might be effective in LGGs with MGMT-non-pM. The associations of MGMT status with RT dose were first reported in present study, our data is helpful in choice of therapeutic strategy for these refractory molecular subtypes. Although confirmation by prospective trials is needed, this study is also helpful in designing clinical trials for LGG based on MGMT status.
High-dose RT (> 54 Gy) was an independently protective factor for patients with LGGs. Patients with MGMT-non-pM may have improved survival upon administration of high-dose RT. Our findings will help to define the standard of care and assist decision-making as well as the design of prospective clinical trials for LGGs. However, the limitations of our retrospective study should be acknowledged. No information on quality of life was available. The adverse effects of high-dose RT must be assessed in further studies.
This study was reviewed and approved by the Institutional Review Board of Beijing Tiantan Hospital (Grant number: KY2013-017-01). Informed written consent from patients was waived by the Institutional Review Board of Beijing Tiantan Hospital due to the retrospective study design.
Not applicable
Not applicable.
All data were presented in the manuscript and supplementary materials.
The authors declare that they have no competing interests.
This work was supported by Beijing Natural Science Foundation (7192057), the Capital Health Development Research Project (2020-2-1072), and Beijing Hospitals Authority Youth Programme (QML20190506). The funding sources had no influence on the design, performance, or reporting of this study.
Study concept and design: Xiaoguang Qiu and Yanwei Liu; Data acquisition and analysis: Yanong Li, Peng Wang, Li Chen and Jin Feng; Statistics analysis: Yanwei Liu; Writing the first draft: Yanwei Liu, Yanong Li; Supervision study: Xiaoguang Qiu; Read and approved final version: All authors.
Thanks to the following members of Chinese Glioma Cooperative Group (CGCG) for providing gliomas patients data: Shouwei Li and Chenxing Wu (Sanbo Brain Hospital), Xuejun Yang (Tianjin Medical University General Hospital), Wei Yan (The First Affiliated Hospital of Nanjing Medical University), Hongjun Wang and Jinquan Cai (the Second Affiliated Hospital of Harbin Medical University), Wen Cheng (the First Affiliated Hospital of China Medical University)
1Department of Radiation Oncology, Beijing Tiantan Hospital, Capital Medical University, Beijing 100070, China; 2Departmentof Molecular Neuropathology, Beijing Neurosurgical Institute, Capital Medical University, Beijing 100070, China; 3National Clinical Research Center for Neurological Diseases, Beijing 100070, China.