5-Aminolevulinic Acid Fluorescence-Guided Resection in High-Grade Glioma

J.Sales-Llopis

Neurosurgery Department, General University Hospital Alicante, Spain.

• 5-Aminolevulinic acid as an emerging radiosensitizer for radiodynamic therapy in solid tumors: a systematic review of available data and clinical potential
• Fluorescence-Guided Surgery for Gliomas: Past, Present, and Future
• Double fluorescence-guided surgery with 5-ALA and fluorescein sodium in grade 2 and grade 3 adult-type diffuse gliomas: retrospective analysis of 112 cases
• Comparative Analysis of 5-ALA and Fluorescent Techniques in High-Grade Glioma Treatment
• Integration of 5-ALA fluorescence and intraoperative MRI in awake craniotomy for glioma resection: a six-year retrospective analysis
• Current and Future Applications of 5-Aminolevulinic Acid in Neurosurgical Oncology
• Real-time fluorescence-guided glioblastoma resection with 5-aminolevulinic acid using ORBEYE
• Fluorescence-Guided Cord Transection for Supramaximal Resection of Spinal High-Grade Glioma: 2-Dimensional Operative Video

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1. Improved Tumor Visualization:

1. 5-ALA fluorescence distinguishes tumor margins and infiltrative regions, providing a clear visual aid to neurosurgeons.
2. It is particularly effective in areas where MRI is less reliable, such as brain regions near the ventricles or eloquent cortex.

2. Extent of Resection (EOR):

1. Studies have demonstrated that 5-ALA-guided resection significantly increases the rate of gross total resection (GTR), achieving higher volumes of tumor removal compared to conventional white-light surgery.
2. Achieving GTR is associated with prolonged progression-free survival (PFS) and overall survival (OS) in HGG patients.

3. Residual Tumor Detection:

1. Residual fluorescence during surgery indicates tumor tissue that might be missed under standard visualization. This allows for more thorough resection without relying solely on preoperative imaging.

4. Safety and Efficacy:

1. The use of 5-ALA is associated with reduced risk of neurological deficits when used alongside intraoperative neurophysiological monitoring, as it helps delineate tumor margins near functional areas.

1. Preoperative Preparation:

1. 5-ALA is administered orally (20 mg/kg) approximately 3–4 hours before anesthesia induction.

2. Intraoperative Imaging:

1. During surgery, a special operating microscope equipped with a blue light source is used to visualize PpIX fluorescence.
2. Fluorescence intensity correlates with tumor density:
3. Strong fluorescence: Solid tumor areas.
4. Faint fluorescence: Tumor infiltration zones.

3. Postoperative Assessment:

1. MRI is performed within 48 hours post-surgery to confirm the extent of resection and assess residual disease.

- False Negatives: Some tumor regions may not fluoresce due to low metabolic activity or heterogeneity in 5-ALA uptake. - False Positives: Inflammatory or reactive tissues may occasionally fluoresce, requiring correlation with other intraoperative findings. - Restricted Use: Not effective for low-grade gliomas due to insufficient fluorescence signal.

- The pivotal Phase III trial by Stummer et al. (2006) demonstrated that 5-ALA significantly improves the rate of complete tumor resection (65% vs. 36% with conventional surgery) and enhances progression-free survival. - Subsequent studies have confirmed its utility and safety, establishing 5-ALA as a standard adjunct in HGG resection.

- Combination with advanced technologies like intraoperative MRI, ultrasound, and neuronavigation. - Exploration of fluorescence in low-grade gliomas and non-glioma tumors. - Development of next-generation fluorescent agents for multimodal imaging.

5-aminolevulinic-acid (5-ALA) guided resection. In addition to stereotactic localization, as well as intraoperative brain mapping, techniques to enhance visual identification of tumor, intraoperatively may be used and include 5-aminolevulinic acid (5-ALA). 5-ALA is metabolized into fluorescent porphyrins, which accumulate in malignant glioma cells. This property permits the use of ultraviolet illumination during surgery as an adjunct to map out the tumor. This has been proven with RCT where the use of 5-ALA leads to more complete resection (65% vs. 36%, p < 0.0001), which translates into a higher 6-month progression-free survival (41% vs. 21.1%, p = 0.0003) but no effect on OS ¹⁾.

Naik et al. compared 5-aminolevulinic acid fluorescence guided resection of high-grade glioma, Fluorescein sodium guided resection of high-grade glioma. (FS), and Intraoperative magnetic resonance imaging-guided resection of high-grade glioma (IMRI) with no image guidance to determine the best intraoperative navigation method to maximize rates of gross total resection (GTR) and outcomes. A frequentist network meta-analysis was performed following standard PRISMA guidelines (PROSPERO registration CRD42021268659). Surface-under-the-cumulative ranking (SUCRA) analysis was executed to hierarchically rank modalities by the outcomes of interest. Heterogeneity was measured by the I2 statistic. Publication bias was assessed by funnel plots and the use of Egger's test. Statistical significance was determined by p < 0.05. Twenty-three studies were included for analysis with a total of 2,643 patients. Network meta-analysis comparing 5-ALA, IMRI, and FS was performed. The primary outcome assessed was the rate of GTR. Analysis revealed the superiority of all intraoperative navigation to control (no navigation). SUCRA analysis revealed the superiority of IMRI + 5-ALA, IMRI alone, followed by FS, and 5-ALA. Overall survival (OS) and progression-free survival (PFS) were also examined. FS (vs. control) was associated with improved OS, while IMRI was associated with improved PFS (vs. control, FS, and 5-ALA). Intraoperative navigation using IMRI, FS, and 5-ALA lead to greater rates of GTR in HGGs. FS and 5-ALA also yielded improvement in OS and PFS. Further studies are needed to evaluate differences in survival benefit, operative duration, and cost ²⁾.

The introduction of 5-aminolevulinic acid (5-ALA) fluorescence-guided surgery has allowed surgeons to better differentiate between neoplastic tissue and normal tissue, thus achieving a greater extent of resection. The development of new intraoperative imaging modalities in combination with 5-ALA may provide additional benefits for glioma patients ³⁾.

Wei et al., describe some of the main factors that limit the sensitivity and accuracy of conventional WF surgical microscopy, and then provide a survey of commercial and research prototypes being developed to address these challenges, along with their principles, advantages and disadvantages, as well as the current status of clinical translation for each technology. They also provide a neurosurgical perspective on how these visualization technologies might best be implemented for guiding glioma surgeries in the future

Detection of PpIX expression in low-grade gliomas and at the infiltrative margins of all gliomas has been achieved with high-sensitivity probe-based visualization techniques. Deep-tissue PpIX imaging of up to 5 mm has also been achieved using red-light illumination techniques. Spectroscopic approaches have enabled more accurate quantification of PpIX expression.

Advancements in visualization technologies have extended the sensitivity and accuracy of conventional WF surgical microscopy. These technologies will continue to be refined to further improve the extent of resection in glioma patients using 5-ALA-induced fluorescence ⁴⁾.

The 5-aminolevulinic acid has been used in glioma surgery and recent studies applied in Sylvian and spinal meningiomas ^{5) 6)}.

Schebesch et al., from the University Medical Center Regensburg, Germany published five patients (3 female, 2 male; mean age 45.4 years) who underwent fluorescence-guided surgery for supratentorial, intracerebral lesions which showed no contrast-enhancement in the preoperative MRI but were, however, strongly suspicious for gliomas. Accordingly, all patients received a preoperative FET-PET scan and detailed histopathological workup was performed. After giving written informed consent, all patients received 5 mg/kg of FL at the induction of anesthesia. Surgery was conducted under white light and under the YELLOW 560 nm filter. We reviewed the surgical protocols, navigational storage and the image databases of our surgical microscopes for evidence of intraoperative fluorescence that corresponded to the FET-PET positive area.

In all patients they found distinct accordances between the FET-PET positive areas and the fluorescing regions within the targeted lesions. Histopathological workup of the fluorescent tissue revealed anaplastic oligodendroglioma, IDH-mutant and 1p/19-codeleted (WHO grade III) (n = 2), anaplastic astrocytoma, IDH-mutant (WHO grade III) (n = 1), oligodendroglioma, IDH-mutant and 1p/19q-codeleted (WHO grade II) (n = 1) and pilocytic astrocytoma (WHO grade I) (n = 1). No adverse events were noted.

Despite the lack of gadolinium-enhancement in the preoperative MRI, all patients intravenously received FL to guide resection. Irrespective of the final grading, FL was extremely helpful in detecting the lesions and in identifying their border zones. In selected patients with NEG, but strong metabolic activity according to the FET-PET, FL may significantly increase the accuracy of surgery ⁷⁾.

5-aminolevulinic acid has initiated a radical change. Over the past years, various groups have published rates of complete resection of the enhancing tumor that exceed 80%. In the coming years, as the use of the technology expands, complete resection should become a common, predictable result at many centers. Consequently, adjuvant therapies that benefit from resection could play a bigger role, resection could be incorporated as a variable in randomized trials and distant recurrence might become a more common problem ⁸⁾.

Fluorescence guided resection (FGR) in high-grade glioma surgery aims at increasing complete resections and, thus, local control. This technique uses 5-aminolevulinic acid (5-ALA), a natural intermediate substance in the heme-porphyrin biosynthesis pathway, and a protoporphyrin IX (PpIX) precursor. PpIX is fluorescent under blue light exposure ⁹⁾.

The highest visible and measurable fluorescence was yielded by 20 mg/kg. No fluorescence was elicited at 0.2 mg/kg. Increasing 5-ALA doses did not result in proportional increases in tissue fluorescence or PPIX accumulation in plasma, indicating that doses higher than 20 mg/kg will not elicit useful increases in fluorescence ¹⁰⁾.

In 2000 Stummer et al published that (5-ALA)-derived fluorescence was approved for fluorescence-guided resections of malignant gliomas, relying on selective synthesis and accumulation of protoporphyrin IX (PPIX) within malignant glioma cells ¹¹⁾.

In 2006 Stummer et al., published that tumor fluorescence derived from 5-aminolevulinic acid enables more complete resections of the contrast-enhancing tumour, leading to improved progression-free survival in patients with malignant glioma ¹²⁾.

The positive predictive value (PPV) of utilizing the most robust ALA fluorescence intensity (lava-like orange) as a predictor of tumor presence is high. However, the negative predictive value (NPV) of utilizing the absence of fluorescence as an indicator of no tumor is poor. ALA intensity is a strong predictor for degree of tumor cellularity for the most fluorescent areas but less so for lower ALA intensities. Even in the absence of tumor cells, reactive changes may lead to ALA fluorescence ¹³⁾.

Many studies have shown that the ratio of gross total resections was higher if the fluorescence technique was used. The fluorescence signal intensity is correlated to the cell density and the PpIX concentration. The current method has a very high specificity but still lower sensitivity, particularly regarding the zones with poor tumoral infiltration ¹⁴⁾.

PpIX fluorescence was also identified as a novel marker for intraoperative detection of anaplastic foci in nonenhancing gliomas that ensures a precise histopathological diagnosis and optimal patient treatment. Furthermore, 5 prospective studies have confirmed that PpIX fluorescence is able to identify residual tumor tissue after assumed maximal resection of malignant gliomas with conventional white-light microscopy. Because intraoperative detection of malignant glioma tissue is significantly improved by 5-ALA FGS, high rates of complete tumor resections are achieved, especially in combination with intraoperative monitoring and mapping ¹⁵⁾.

In the future, chemotherapy with new anticancer agents, immunotherapy, and new methods of radiotherapy and gene therapy will be developed; however, ALA will play a key role in malignant glioma treatment before the development of these new treatments ¹⁶⁾.

Visual assessment of PpIX fluorescence is subjective and limited by the distorting effects of light attenuation and tissue autofluorescence ¹⁷⁾.

Resection of reactive tissue without active recurrent tumor after multimodal treatment for glioblastoma is frequently associated with solid or vague 5-AIF. Therefore, neurosurgeons should remain cautious when attempting to employ intraoperative 5-ALA induced fluorescence (5-AIF) to discriminate radiation- and chemotherapy-induced tissue changes from true disease progression. Nevertheless, 5-AIF-guided resection remains a valid tool in the neurosurgical treatment of recurrent gliomas ¹⁸⁾.

The resection cavity underestimates the volume of resected tissue and 5-ALA complete resections go significantly beyond the volume of pre-operative contrast-enhancing tumor bulk on MRI, indicating that 5-ALA also stains MRI non-enhancing tumor tissue. Use of 5-ALA may thus enable extension of coalescent tumor resection beyond radiologically evident tumor. The impact of this more extended resection method on time to progression and overall survival has not been determined, and potentially puts adjacent and functionally intact tissue at risk ¹⁹⁾.

In 2013 based on available literature, there was level of evidence 2 that 5-ALA-guided surgery is more effective than conventional neuronavigation-guided surgery in increasing diagnostic accuracy and extent of tumor resection, enhancing quality of life, or prolonging survival in patients with high-grade malignant gliomas ²⁰⁾.

Barone et al., in a Cochrane Database Systematic Review published in 2014, that there is low to very low quality evidence (according to GRADE criteria) that image guided surgery using iMRI, 5-ALA or DTI-neuronavigation increases the proportion of patients with high-grade glioma that have a complete tumour resection on post-operative MRI. There is a theoretical concern that maximising the extent of resection may lead to more frequent adverse events but this was poorly reported in the included studies. Effects of image guided surgery on survival quality of life (QoL) are unclear. Further research, including studies of ultrasound guided surgery, is needed ²¹⁾.

In 2015, a literature review produced 503 potential publications; only 20 of these fulfilled the inclusion criteria of this analysis, including a total of 565 patients treated with 5-ALA-FIGR reporting on its outcomes and 800 histological samples reporting 5-ALA-FIGR sensitivity and specificity.

The mean gross total resection (GTR) rate was 75.4% (95% CI: 67.4-83.5, p<0.001). The mean time to tumor progression (TTP) was 8.1 months (95% CI: 4.7-12, p<0.001). The mean overall survival gain reported was 6.2 months (95% CI: -1-13, p<0.001). The specificity was 88.9% (95% CI: 83.9-93.9, p<0.001) and the sensitivity was 82.6% (95% CI: 73.9-91.9, p<0.001).

5-ALA-FIGR in Glioblastoma is highly sensitive and specific, and imparts significant benefits to patients in terms of improved GTR and TTP ²²⁾.

Within the limitations of a preliminary analysis, the use of the exoscope in fluorescence-guided surgery for high-grade gliomas provided significant advantages in terms of visualization of the surgical field under a blue filter and linearity of surgical flow ²³⁾.

CLR1501 (green) and CLR1502 (near infrared) are novel tumor-selective fluorescent agents for discriminating tumor from normal brain ²⁴⁾.

5-aminolevulinic-acid fluorescence-guided resection of glioma has different intensities within tumors and among other patients, some molecular and external factors have been implicated, but there is no clear evidence analyzing the difference of fluorescence according to glioma molecular characteristics.

Garfias-Arjona et al. in an observational comparative and exploratory study aimed to compare molecular factors of glioma samples with fluorescence intensity to identify potential confounders and associations with clinically relevant tumor features.

Tumor samples of high-grade glioma patients operated using 5-ALA for guided resection were included for comparative analysis of fluorescence intensity and molecular features. All the samples were processed under the same conditions. The power for fluorescent stimulation and acquisition time was the same between samples. An inverted fluorescence microscope compared the mean fluorescence for each molecular variation. p53, ATRX and Ki67 expression and IDH1 mutation were assessed by immunohistochemistry. Follow-up was made with the patients for progression-free survival and overall survival.

They found that the fluorescence intensity for each specific tumor was independent of the methylation of the methylguanine-DNA-methyltransferase (MGMT) promoter region assessed by pyrosequencing, there was no association of fluorescence with p53, ATRX, IDH1 mutation as assessed by immunochemistry. Also, fluorescence intensity has no relation to the time of tumor recurrence or overall survival.

With the results, they argue that many factors are involved in fluorescence intensity that may be related to the specific metabolic status of the glioma cells analyzed, which is more likely to be responsible for the variation of fluorescence ²⁵⁾

This study highlights the complexity of 5-ALA fluorescence in high-grade gliomas and raises questions about its underlying mechanisms. While methodologically sound, the findings are limited by the lack of novel insights and an incomplete exploration of proposed mechanisms. Future research should delve deeper into metabolic influences, broaden the molecular scope, and incorporate clinical correlates to unlock the full potential of 5-ALA-guided surgery.

Data of 47 consecutive patients with HGG have been collected in our study (25 males, 22 females; mean age: 60.3 years, range: 27-86 years). Fluorescein (5 mg/kg of body weight) was injected intravenously right after the induction of general anesthesia. A YELLOW 560 filter was used on an OPMI Pentero 900 microscope (Carl Zeiss Meditec, Oberkochen, Germany) to complete a microsurgical tumor removal. Glioma resection and quality of life were evaluated preoperative and postoperatively.

Gross total resection (GTR) was achieved in 53.2% (n = 25) of patients. A subtotal resection (STR) (>95%) was achieved in 29.8% (n = 14), while a partial resection (PR) (<95%) was obtained in 17% (n = 8) of patients. Overall, in 83% (n = 39) of patients who underwent fluorescence-guided surgery the resection rate achieved was >95%. No adverse effects correlated to fluorescein have been recorded.

Fluorescein seems to be safe and effective in the resection of HGGs, allowing a high rate of gross total removal of contrast enhanced areas ²⁶⁾.

Thirty-two patients received fluorescein sodium (3 mg/kg) intravenously prior to resection. Fluorescence was intraoperatively visualized using a Zeiss Pentero surgical microscope equipped with a YELLOW 560 filter. Stereotactically localized biopsy specimens were acquired from CE and NCE regions based on preoperative MRI in conjunction with neuronavigation. The fluorescence intensity of these specimens was subjectively classified in real-time with subsequent quantitative image analysis, histopathological evaluation of localized biopsy specimens, and radiological volumetric assessment of the extent of resection. RESULTS Bright fluorescence was observed in all Glioblastomas and localized to the CE regions and portions of the NCE margins of the tumors, thus serving as a visual guide during resection. Gross-total resection (GTR) was achieved in 84% of the patients with an average resected volume of 95%, and this rate was higher among patients for whom GTR was the surgical goal (GTR achieved in 93.1% of patients, average resected volume of 99.7%). Intraoperative fluorescein staining correlated with histopathological alteration in both CE and NCE regions, with positive predictive values by subjective fluorescence evaluation greater than 96% in NCE regions.

Intraoperative administration of fluorescein provides an easily visualized marker for glioma pathology in both CE and NCE regions of Glioblastoma. These findings support the use of fluorescein as a microsurgical adjunct for guiding Glioblastoma resection to facilitate safe maximal removal ²⁷⁾.

A single-center, prospective, single-arm, open-label Phase II clinical trial of ALA fluorescence-guided resection of high-grade gliomas (Grade III and IV) was held over a 43-month period (August 2010 to February 2014). ALA was administered at a dose of 20 mg/kg body weight. Intraoperative biopsies from resection cavities were collected. The biopsies were graded on a 4-point scale (0 to 3) based on ALA fluorescence intensity by the surgeon and independently based on tumor cellularity by a neuropathologist. The primary outcome of interest was the correlation of ALA fluorescence intensity to tumor cellularity. The secondary outcome of interest was ALA adverse events. Sensitivities, specificities, positive predictive values (PPVs), negative predictive values (NPVs), and Spearman correlation coefficients were calculated. RESULTS A total of 211 biopsies from 59 patients were included. Mean age was 53.3 years and 59.5% were male. The majority of biopsies were glioblastoma (Glioblastoma) (79.7%). Slightly more than half (52.5%) of all tumors were recurrent. ALA intensity of 3 correlated with presence of tumor 97.4% (PPV) of the time. However, absence of ALA fluorescence (intensity 0) correlated with the absence of tumor only 37.7% (NPV) of the time. For all tumor types, Glioblastoma, Grade III gliomas, and recurrent tumors, ALA intensity 3 correlated strongly with cellularity Grade 3; Spearman correlation coefficients ® were 0.65, 0.66, 0.65, and 0.62, respectively. The specificity and PPV of ALA intensity 3 correlating with cellularity Grade 3 ranged from 95% to 100% and 86% to 100%, respectively. In biopsies without tumor (cellularity Grade 0), 35.4% still demonstrated ALA fluorescence. Of those biopsies, 90.9% contained abnormal brain tissue, characterized by reactive astrocytes, scattered atypical cells, or inflammation, and 8.1% had normal brain. In nonfluorescent (ALA intensity 0) biopsies, 62.3% had tumor cells present. The ALA-associated complication rate among the study cohort was 3.4%.

The PPV of utilizing the most robust ALA fluorescence intensity (lava-like orange) as a predictor of tumor presence is high. However, the NPV of utilizing the absence of fluorescence as an indicator of no tumor is poor. ALA intensity is a strong predictor for degree of tumor cellularity for the most fluorescent areas but less so for lower ALA intensities. Even in the absence of tumor cells, reactive changes may lead to ALA fluorescence ²⁸⁾.

58 patients with high-grade glioma s (°III and °IV) were included. 10 of 63 tumors (15.9 %) failed to fluoresce intraoperatively of which nine (90 %) had been adjuvantly treated prior to recurrence, as were 46 of the 53 fluorescing tumors (86.8 %). Non-fluorescing tumors were IDH mutated significantly more often (p = 0.005). 30 tumors (47.6 %) were located eloquently. 51 (80.9 %) patients showed no new neurologic deficits postoperatively. 13 patients (20.6 %) showed no signs of recurrence at their latest follow up. Eight patients were lost to follow up. Overall survival was significantly longer in the 5-ALA group (p = 0.025). Fluorescence-guided surgery in recurrent gliomas is safe and allows for a good surgical and neurological outcome in a difficult surgical environment, especially when used in combination with neuronavigation and intraoperative ultrasound to prevent over-resection. Adjuvant therapy did not significantly influence fluorescing properties ²⁹⁾.

Clinical and surgical data from patients affected by HGG who underwent surgery guided by 5-ALA fluorescence between June 2011 and February 2014 were retrospectively evaluated. Surgical outcome was evaluated by assessing the resection rate as gross total resection (GTR) > 98% and GTR > 90%. We finally stratified data for recurrent surgery, tumor location, tumor size, and tumor grade (IV versus III grade sec. WHO).

94 patients were finally enrolled. Overall GTR > 98% and GTR > 90% was achieved in 93% and 100% of patients. Extent of resection (GTR > 98%) was dependent on tumor location, tumor grade (P < 0.05), and tumor size (P < 0.05). In 43% of patients the boundaries of fluorescent tissue exceeded those of tumoral tissue detected by neuronavigation, more frequently in larger (57%) (P < 0.01) and recurrent (60%) tumors.

5-ALA fluorescence in HGG surgery enables a GTR in 100% of cases even if selection of patients remains a main bias. Recurrent surgery, and location, size, and tumor grade can predict both the surgical outcome and the intraoperative findings ³⁰⁾.

Schucht et al., prospectively studied 72 consecutive patients who underwent 5-ALA-guided surgery for a glioblastoma adjacent to the corticospinal tract (CST; < 10 mm) with continuous dynamic monopolar motor mapping (short-train interstimulus interval 4.0 msec, pulse duration 500 μsec) coupled to an acoustic motor evoked potential (MEP) alarm. The extent of resection was determined based on early (< 48 hours) postoperative MRI findings. Motor function was assessed 1 day after surgery, at discharge, and at 3 months.

Five patients were excluded because of nonadherence to protocol; thus, 67 patients were evaluated. The lowest motor threshold reached during individual surgery was as follows (motor threshold, number of patients): > 20 mA, n = 8; 11-20 mA, n = 13; 6-10 mA, n = 10; 4-5 mA, n = 13; and 1-3 mA, n = 23. Motor deterioration at postsurgical Day 1 and at discharge occurred in 30% (n = 20) and 10% (n = 7) of patients, respectively. At 3 months, 3 patients (4%) had a persisting postoperative motor deficit, 2 caused by vascular injury and 1 by mechanical injury. The rates of intra- and postoperative seizures were 1% and 0%, respectively. Complete resection of enhancing tumor was achieved in 73% of patients (49/67) despite proximity to the CST.

A rather high rate of CRET can be achieved in glioblastomas in motor eloquent areas via a combination of 5-ALA for tumor identification and intraoperative mapping for distinguishing between presumed and actual motor eloquent tissues. Continuous dynamic mapping was found to be a very ergonomic technique that localizes the motor tissue early and reliably ³¹⁾.

Schucht et al., selected 13 patients who had received a complete resection according to intraoperative 5-ALA induced fluorescence and CRET according to post-operative T1 contrast-enhanced MRI. The volumes of pre-operative contrast enhancing tissue, post-operative resection cavity and resected tissue were determined through shift-corrected volumetric analysis.

The mean resection cavity (29 cm(3)) was marginally smaller than the pre-operative contrast-enhancing tumor (39 cm(3), p = 0.32). However, the mean overall resection volume (84 cm(3)) was significantly larger than the pre-operative contrast-enhancing tumor (39 cm(3), p = 0.0087). This yields a mean volume of resected 5-ALA positive, but radiological non-enhancing tissue of 45 cm(3). The mean calculated rim of resected tissue surpassed pre-operative tumor diameter by 6 mm (range 0-10 mm).

Results of the current study imply that (i) the resection cavity underestimates the volume of resected tissue and (ii) 5-ALA complete resections go significantly beyond the volume of pre-operative contrast-enhancing tumor bulk on MRI, indicating that 5-ALA also stains MRI non-enhancing tumor tissue. Use of 5-ALA may thus enable extension of coalescent tumor resection beyond radiologically evident tumor. The impact of this more extended resection method on time to progression and overall survival has not been determined, and potentially puts adjacent and functionally intact tissue at risk ³²⁾.

A retrospective review found 118 consecutive patients with high-grade gliomas operated on with the use of fluorescence-guided surgery with 5-aminolevulinic acid. Within that series, the 52 patients with newly diagnosed Glioblastoma and complete resection of enhancing tumor (CRET) in early MRI were selected for analysis. They studied the influence of residual fluorescence in the surgical field on overall survival and neurological complication rate. Multivariate analysis included potential relevant factors: age, Karnofsky Performance Scale, O-methylguanine methyltransferase methylation promoter status, tumor eloquent location, preoperative tumor volume, and adjuvant therapy.

The median overall survival was 27.0 months (confidence interval = 22.4-31.6) in patients with nonresidual fluorescence (n = 25) and 17.5 months (confidence interval = 12.5-22.5) for the group with residual fluorescence (n = 27) (P = .015). The influence of residual fluorescence was maintained in the multivariate analysis with all covariables, hazard ratio = 2.5 (P = .041). The neurological complication rate was 18.5% in patients with nonresidual fluorescence and 8% for the group with residual fluorescence (P = .267).

Glioblastoma patients with CRET in early MRI and no fluorescent residual tissue had longer overall survival than patients with CRET and residual fluorescent tissue ³³⁾.

One hundred three consecutive patients underwent resection of glioblastoma from August 2008 to November 2010. Eligibility for CRET was based on the initial magnetic resonance imaging assessed by 2 reviewers. The primary end point was the number of patients with CRET and GTR. Secondary end points were volume of residual contrast-enhancing tissue and new postoperative neurological deficits.

Fifty-three patients were eligible for GTR/CRET (n = 43 newly diagnosed glioblastoma, n = 10 recurrent); 13 additional patients received surgery for GTR/CRET-ineligible glioblastoma. GTR was achieved in 96% of patients (n = 51, no residual enhancement >0.175 cm); CRET was achieved in 89% (n = 47, no residual enhancement). Postoperatively, 2 patients experienced worsening of preoperative hemianopia, 1 patient had a new mild hemiparesis, and another patient sustained sensory deficits.

Using 5-aminolevulinic acid imaging and intraoperative mapping/monitoring together leads to a high rate of CRET and an increased rate of GTR compared with the literature without increasing the rate of permanent morbidity. The combination of safety and resection-enhancing intraoperative technologies was likely to be the major drivers for this high rate of CRET/GTR ³⁴⁾.

322 patients aged 23-73 years with suspected malignant glioma amenable to complete resection of contrast-enhancing tumour were randomly assigned to 20 mg/kg bodyweight 5-aminolevulinic acid for fluorescence-guided resection (n=161) or to conventional microsurgery with white light (n=161). The primary endpoints were the number of patients without contrast-enhancing tumour on early MRI (ie, that obtained within 72 h after surgery) and 6-month progression-free survival as assessed by MRI. Secondary endpoints were volume of residual tumour on postoperative MRI, overall survival, neurological deficit, and toxic effects. We report the results of an interim analysis with 270 patients in the full-analysis population (139 assigned 5-aminolevulinic acid, 131 assigned white light), which excluded patients with ineligible histological and radiological findings as assessed by central reviewers who were masked as to treatment allocation; the interim analysis resulted in termination of the study as defined by the protocol. Primary and secondary endpoints were analysed by intention to treat in the full-analysis population. The study is registered at http://www.clinicaltrials.gov as NCT00241670.

Median follow-up was 35.4 months (95% CI 1.0-56.7). Contrast-enhancing tumour was resected completely in 90 (65%) of 139 patients assigned 5-aminolevulinic acid compared with 47 (36%) of 131 assigned white light (difference between groups 29% [95% CI 17-40], p<0.0001). Patients allocated 5-aminolevulinic acid had higher 6-month progression free survival than did those allocated white light (41.0% [32.8-49.2] vs 21.1% [14.0-28.2]; difference between groups 19.9% [9.1-30.7], p=0.0003, Z test). Groups did not differ in the frequency of severe adverse events or adverse events in any organ system class reported within 7 days after surgery.

Tumour fluorescence derived from 5-aminolevulinic acid enables more complete resections of contrast-enhancing tumour, leading to improved progression-free survival in patients with malignant glioma ³⁵⁾.

Fifty-two consecutive patients with Glioblastoma received oral doses of 5-ALA (20 mg/kg body weight) 3 hours before induction of anesthesia. Intraoperatively, tumor fluorescence was visualized using a modified operating microscope. Fluorescing tissue was removed whenever it was considered safely possible. Residual enhancement on early postoperative MR imaging was quantified and related to each patient's characteristics to determine which factors influenced resection. Survival was analyzed using the Kaplan-Meier method and multivariate analysis was performed in which the Karnofsky Performance Scale (KPS) score, residual fluorescence, patient age, and residual enhancement on MR images were considered. Intraoperatively, two fluorescence qualities were perceived: solid fluorescence generally reflected coalescent tumor, whereas vague fluorescence mostly corresponded to infiltrative tumor. Complete resection of contrast-enhancing tumor was accomplished in 33 patients (63%). Residual intraoperative tissue fluorescence left unresected for safety reasons predicted residual enhancement on MR images in 18 of the 19 remaining patients. Age, residual solid fluorescence, and absence of contrast enhancement in MR imaging were independent explanatory factors for survival, whereas the KPS score was significant only in univariate analysis. No perioperative deaths and one case of permanent morbidity were encountered.

The observations in this study indicate the usefulness of 5-ALA-induced tumor fluorescence for guiding tumor resection. The completeness of resection, as determined intraoperatively from residual tissue fluorescence, was related to postoperative MR imaging findings and to survival in patients suffering from Glioblastoma ³⁶⁾.