The average overall survival (OS) and progression-free survival (PFS) of the population studied were 23.70 (STD 18.64) and 9.93 (STD 11.86) months, respectively. The average preoperative Karnofsky performance status (KPS) was 83.48 (STD 15.83), while the average postoperative KPS was 80.11 (STD 16.51).
In total, 63.64% of the sample was male with an average age of 54.28 years (STD 13.86), average OS of 23.46 months (STD 17.71), PFS 8.70 months (STD 12.10), and postoperative KPS of 79.10 (STD 19.43), while 36.36% was female with an average age of 60.31 years (STD 12.18), average OS of 24.12 (STD 20.77), PFS 12.06 (STD 11.06), and postoperative KPS of 81.87 (STD 9.81). We found no statistically significant differences between males and females in terms of outcome.
Patients under 50 years of age had an OS of 22.76 months (STD 13.59), a PFS of 9.61 months (STD 9.83), a preoperative KPS of 83.07 (STD 23.93), and a postoperative KPS of 82.06 (STD 17.02); patients aged 51 to 75 years had an OS of 26.19 months (STD 21.74), a PFS of 11.11 (STD 13.71), a preoperative KPS of 84.4 (STD 11.48), and an average postoperative KPS of 78.26 (STD 17.6); patients over 75 years of age had an OS of 13.80 months (STD 6.41), a PFS of 4.40 months (STD 1.81), a preoperative KPS of 78 (STD 8.36), and a postoperative KPS of 84 (STD 5.47), Figure 1
Patients with preoperative KPS greater than or equal to 80 points had an OS of 26.44 months (STD 20.31) and a PFS of 11.52 months (STD 13.04); patients with preoperative KPS between 50 and 80 points had an OS of 13.25 months (STD 4.06) and a PFS of 4.62 months (STD 2.92); patients with preoperative KPS less than 50 points (only 1.32% of our population) had an average OS of 15.66 months (STD 7.63) and a PFS of 4 months (STD 1.81).
Half of our population started with epileptic seizures. The mean age of patients who started with seizures was 52.40 years (STD 12.58), OS 26.36 months (STD 22.05) vs. 21.04 (STD 14.52) with p
-value = 0.350 and a mean PFS of 8.86 months (STD 11.56) vs. 11 (STD 12.33) in the control group. Preoperative KPS in patients with seizures was 81.13 (STD 19.87) vs. 85.95 (STD 9.95) of the controls with a p
-value = 0.324. Postoperative KPS in patients with seizures was 80.22 (STD 18.15) vs. 80 (STD 15.11) of the controls and a p
-value = 0.964, Figure 2
Patients undergoing gross total resection (GTR) had an average OS of 27.61 months (STD 20.38) and a PFS of 11.87 (STD 13.52). The control group underwent a subtotal resection (STR) with an OS of 14.38 months (STD 8.57) and a PFS of 5.30 months (STD 3.77), Figure 3
. The Student t
-test showed a statistically significant difference between the OS of the two groups (27.61 months vs. 14.38 months, p
-value = 0.030), while the difference in PFS was remarkable but not significant (11.87 months vs. 5.31 months, p
-value = 0.091).
The difference between postoperative KPS in the GTR and STR groups was not statistically significant. However, the distribution of the sample showed that the STR group presented medium–high postoperative KPSs (between 60 and 100), while in the GTR group there were patients with low postoperative KPSs, as shown in Figure 4
Patients with wild type IDH1 gliomas had an OS of 22.63 months (STD 17.67), while for the mutated IDH1 group it was 38.33 (STD 29.73), p-value = 0.016. PFS in the wild-type group was 10.07 months (STD 12.25) vs. 8 months in the mutated group (STD 4.00), p-value = 0.774. Preoperative KPS for the wild type group was 83.50 (STD 16.06), while for the mutated IDH1 group it was 83.33 (STD 15.27), p = 0.980. Postoperative KPS for the wild-type group was 70 (STD 36.05), while for the mutated IDH1 group it was 82.31 (STD 12.45), p-value = 0.105.
Patients with MGMT methylation (Figure 5
) had an OS of 31.95 months (STD 5.19), while for patients without its methylation it was 16.83 months (STD 2.01), p
-value = 0.005. The mean preoperative KPS in patients with promoter methylation was 87.75 (STD 11.75), whereas in patients without methylation it was 79.78 (STD 18.12), p
-value = 0.100. The mean postoperative KPS in patients in the methylation group was 77.50 (STD 19.15), whereas in the methylation-free group it was 82.29 (DS 13.98) p
-value = 0.343.
Patients with ATRX loss (Figure 6
) had an OS of 30.75 months (STD 29.62), while patients without mutation had an OS of 22.13 months (STD 15.42) p
-value = 0.241. The PFS of the group with the ATRX loss was 14.12 months (STD 17.51), while in the control group it was 9 months (STD 10.33), p
-value = 0.274. The group of patients with ATRX loss had a preoperative KPS of 82.50 (STD 11.64) vs. a KPS of 83.71 (STD 16.77) in the normal type, p
= 0.847. The ATRX loss group had a postoperative KPS of 76.25 (STD 16.85) vs. a KPS of 80.97 (STD 16.55) of the normal type ATRX group, p
-value = 0.470.
Patients with EGFR amplification/EGFRvIII mutation (Figure 7
) had a mean OS of 19.83 months (STD 16.31) vs. 32 months (STD 21.15) of the control group with p
-value = 0.035. The mean PFS in the EGFR amplification/EGFRvIII mutation group was 7.4 months (STD 7.84), whereas in the control group it was 15.75 (STD 16.79) p
-value = 0.055. Preoperative KPS was 81.50 (STD 17.27) in the EGFR amplification/EGFRvIII mutation group vs. 88.07 (STD 11.08), p
-value = 0.214, while average postoperative KPS was 80.51 (STD 12.94) vs. 77.14 (STD 22.97), p
-value = 0.421.
In total, 82.67% of our population presented with TP53 mutation, with an OS of 21.13 months (STD 13.42), a PFS 8.96 months (STD 11.05), and a postoperative KPS 80.97 (STD 16.77). In the 17.33% without TP53 mutation, we found an OS of 27 (STD 19.33), a PFS of 11.21 (STD 12.45), and a KPS of 78.35 (STD 19.35). No statistically significant differences were found between the two groups (OS p-value = 0.156).
The sample with Ki-67 ≤ 10% had an average OS of 31.69 months (STD 19.17), an average PFS of 13.15 months (STD 13.52), preoperative KPS of 90 (STD 11.28), and postoperative KPS of 76.92 (STD 22.13); in patients with 10% < Ki-67 ≤ 20%, we found an average OS of 28.46 months (STD 24.68), an average PFS of 10.93 months (STD 13.02), preoperative KPS 79 (STD 21.23), and postoperative KPS 75.66 (STD 24.70); the population with a Ki-67 > 20% had an average OS of 16.33 months (STD 11.89), a PFS 7.26 months (STD 8.95), preoperative KPS 84.33 (STD 12.08), and postoperative KPS 81.33 (STD 9.90). Comparing the group with Ki-67 ≤ 10% and the group with Ki-67 > 20%, we observed an OS of 31.69 vs. 16.33 months respectively with p
-value = 0.021, Figure 8
Multivariate analysis showed that more than 50% of the OS of our population depended on the variables examined (R2 = 0.496, F(9,34) = 3.723, p = 0.002). The number of months between the first procedure and the recurrence of disease was significantly associated with OS (B = 0.313, t = 3.213, p = 0.003). The percentage of Ki-67 showed an association with OS tending to statistical significance (B = −0.025, t = −1.816, p = 0.078).
The independent variables examined as a whole also statistically significantly correlated for about 50% with PFS (R2 = 0.496, F(9,32) = 3.493, p = 0.004).
We examined, in a consecutive single-operator series of 122 GBM patients surgically treated from 2013 to 2017 at Sapienza University of Rome, the correlation between sex, age, preoperative KPS, presenting with seizures, and extent of resection (EOR) with OS, PFS, and postoperative KPS, along with the prognostic value of mutations of IDH1, MGMT, ATRX, EGFR, and TP53 genes and of Ki67.
Our study, therefore, carried out a systematic and complete analysis of the main prognostic factors clinical and molecular of GBM on a very homogeneous patient series treated by a single first operator in a single institution. Simultaneous and multivariate analysis allowed us to investigate the correlation with the prognosis and quality of life.
Our results showed that sex did not affect prognosis. Sex influences survival only when combined with the methylation state of the MGMT promoter: women with a methylated phenotype have a higher OS than men with the same phenotype [10
OS and PFS were instead significantly higher in the group of patients younger than 75 years of age (age ≥ 75 years was an independent negative prognostic factor). Also, the group of patients with preoperative high KPS (KPS ≥ 80) showed a significantly better prognosis.
Presenting with seizures was found to be remarkable, as well. OS and PFS were, in fact, higher in the group with seizures than in the group without seizures [11
]. This finding might be related to the higher chance of an early diagnosis being more frequent in cortical lesions and in mutated IDH1 gliomas. The mutation of IDH1 (see below) leads, as is well known, to the formation of 2-HG (2-hydroxyglutarate). 2-HG has a molecular structure similar to glutamate and is able to bind and activate N
-aspartate receptors (NMDA), thus resulting in a possible reduction in seizure threshold (this mutation is associated with about 70%–88% of low-grade gliomas with these being more epileptic than high-grade ones). Antiepileptic therapy could also have a sensitizing role in chemotherapy treatment, as pointed out by Vecth et al. [11
]. Recently, it has been found that by combining valproic acid (VPA) with TMZ, survival rates improve in adults with GBM as well as children with brain tumors other than GBM. This could possibly be explained by the chemotherapy-sensitizing properties of VPA, including the inhibition of histone deacetylase, leading to improved survival [11
]. The numerous side effects, drug interactions, and the consequent poor handling of this drug compared to others such as Levetiracetam (LEV), used for our patients, must be, however, taken into account. Moreover, LEV can provide a survival benefit in patients with GBM who receive TMZ [12
]. Other authors suggested that the clinical therapeutic efficacy of TMZ in GBM might be potentiated through the combination with LEV and the enhancement of apoptotic pathways [13
]. Nonetheless, a combined analysis of survival association with antiepileptic drug use at the beginning of chemo-radiotherapy and TMZ, performed in a pooled patient cohort (n
= 1869) of four contemporary randomized clinical trials in newly diagnosed GBM, showed no outcome improvement for either LEV or VPA use [14
]. It is therefore necessary to carry out a prospective study to clarify whether the use of LEV or VPA associated with TMZ is able to determine a real improvement in the outcome, and which of the two drugs is more effective. Our study confirmed that the increase in OS in patients treated with LEV, though present, had no statistical significance and it was rather due to the characteristics of the tumors presenting with seizures: early diagnosis and cortical location, which makes these lesions more accessible.
As far as surgery is concerned, the GTR group had significantly higher OS and PFS than the STR group, as widely reported in current literature [15
]. Patients with lesions in eloquent areas were treated with STR in order to avoid neurological deficits resulting in a postoperative KPS and quality of life worsening (Figure 8
). In our series, none of the patients in the STR group exceeded 36 months of OS, while in the group of patients treated with GTR there were survival rates of up to 85 months (none of the patients were treated with GTR in reintervention after being treated with STR, as reported by Block O et al. [16
]). When STR was performed, residual disease volume seemed to be the most important factor influencing OS and PFS [17
], being—in our series—an independent prognostic factor. We did not find any difference in terms of survival in patients with residual tumor when the extent of resection exceeded 90%. Our results showed that survival was markedly greater not only in patients treated with GTR as a first intervention, but also in patients who were treated with GTR in reintervention. GTR is hence an independent prognostic factor for survival, even in reintervention cases and even if associated with lower postoperative KPS values in comparison to STR.
Multivariate analysis of prognostic factors showed that the number of months between tumor removal and disease recurrence was the independent variable most specifically related to OS (B = 0.313, t = 3.213, p
= 0.003) and was also related to postoperative KPS. This parameter summarizes the validity of the treatment and can be influenced by the presence of tumor residue [18
] and by resistance to treatment with temozolomide [19
]. This parameter may, therefore, be fundamental in the selection of recurrences amenable to a new surgical treatment and in the choice of a new chemotherapy line.
We also evaluated prognostic value of IDH1, MGMT, ATRX, EGFR, and TP53 gene mutations, and of Ki67.
Mutation of IDH1 gene was found in less than 10% of the sample and was associated, in line with current literature, with a significant increase in OS and PFS (without statistically significantly affecting postoperative KPS).
A recent metanalysis pointed out the scarcity of evidence in terms of a direct relationship between methylated MGMT promoter and PFS [20
]. The results of our study showed that MGMT promoter methylation was an independent positive prognostic factor for both OS and PFS, but—as for IDH1—not a predictive factor for postoperative KPS. This was, however, the molecular marker that correlated the most with survival.
A similar phenotype was also induced by ATRX loss and low-ATRX mRNA expression which, in our study, were associated with an increase in survival, though without statistical significance. This finding is in agreement with the Jiao report [21
In their study, Ramamoorthy et al. proved that in the absence of ATRX, the histone variant macroH2A1.1 binds to the polymerase tankyrase 1, preventing it from localizing to telomeres and resolving cohesion, thus promoting recombination between sister telomeres. Forced resolution of this event induces genomic instability, thereby impeding cell growth [22
Liu et al. [23
] highlighted the absence of ATRX within secondary glioblastomas, and more particularly in younger patients, whereas Cai et al. [24
] observed a higher rate of lower ATRX expression in primary GBM and grade III gliomas than in grade II gliomas, and suggested this as a malignancy marker.
Our study showed that ATRX loss and low-ATRX mRNA expression play an important role, not only in the survival of patients affected by LGG, but also in case of HGG. These findings may be a potential therapeutic target for high grade glial tumors, hence the need for further investigations.
EGFR amplification/EGFRvIII mutation was—in our population—a negative independent prognostic factor in terms of OS, presenting a trend towards statistical significance for PFS too. Nonetheless, no improvement in survival was found with the EGFRvIII Rindopepimut®
]. The addition to the standard therapy of Nimotuzumab®
, a humanized therapeutic monoclonal antibody against EGFR, yielded, on the other hand, positive results only in a post hoc analysis where it revealed an improvement in survival in patients with residual tumor and nonmethylated MGMT (PFS 6.2 vs. 4 months; OS 19 vs. 13.8 months). This could be due to receptor interference and associations with multiple transduction pathways and proteins of invasion and angiogenesis regulation and the development of resistance mechanisms. Therefore, new therapies are focused on a combination of targeted gene therapy against EGFR and EGFRvIII and transduction pathways and proteins related to this pathway [26
EGFR amplification/EGFRvIII mutation, like the previous markers, did not appear to influence postoperative KPS.
Our study, with the advantage of investigating a large population through the elimination of the main confounding factor represented by multioperator treatment, showed that—in spite of the negative results of the clinical trials over Rindopepimut®
vaccine and Nimotuzumab®
—there is solid clinical evidence of the role of EGFR amplification/EGFRvIII mutation in OS and PFS. Therefore, new clinical trials trying to block the EGFR signal transduction pathway at different levels in order to reduce resistance to therapy may be fundamental, and increased survival with anti-EGFR drugs in patients with nonmethylated MGMT should be particularly investigated. Recently, new EGFR-targeted therapies have been proposed, e.g., depatuxizumab mafodotin, which completed Phase I in a study with recurrent GBM patients with EGFR amplification and entered Phase III in the RTOG 3508 trial [27
] as an adjunct therapy to standard therapy. In addition, a phase I study with T cells activated with a chimerical antigen against EGFRvIII shows good treatment tolerance and encouraging results [27
TP53 is one of the most commonly deregulated genes in cancer. Deregulated p53 pathway components have been implicated in GBM cell invasion, migration, proliferation, evasion of apoptosis, and cancer cell stemness. Recent studies show that mutant TP53 is also strongly associated with a poor prognosis in terms of overall survival and with a decrease in chemosensitivity of GBM to TMZ by increasing MGMT expression [29
]. In our study, TP53 mutation was associated with shorter OS and PFS, even though without statistical significance.
Ki-67 is a nonhistone nuclear protein, which is expressed throughout all active cell cycle phases, but not in the resting cell phase, G0. We evaluated the relationship between Ki-67 index and the outcome of patients. The analysis revealed that OS and PFS were inversely related with Ki-67 index. We divided our patients into three groups based on this parameter (Figure 6
): <10%, between 10% and 20% and >20%. We found a statistically significant difference in terms of survival in the group with Ki-67 < 10% compared to the group with Ki-67 > 20% (p
-value = 0.021).
In a recent report, Alkhaibary et al. [30
] reported similar results, but in a series of 44 multioperator patients. Our investigation highlighted the strong role of Ki-67 as an unfavorable prognostic factor in GBM.
Postoperative KPS was not related to any of the independent variables examined. The only variable tending to statistical significance was, as previously reported, the number of months between tumor removal and recurrence, which was also the variable related the most to OS in the multivariate analysis [31