The landscape of postsurgical recurrence patterns in diffuse low-grade gliomas.

Early and maximal safe surgical resection optionally followed by adjuvant treatment is currently recommended in diffuse low-grade glioma (DLGG). Although this management delays malignant transformation (MT), recurrence will most often occur. Because this relapse usually arises locally, reoperation can be considered, with possible further chemotherapy/radiotherapy. However, due to a prolonged overall survival, a large spectrum of unusual recurrence patterns begins to emerge during long-term follow-up, beyond the classical slow and local tumor re-growth. We review various atypical patterns of DLGG relapse, we discuss their pathophysiological mechanisms and how to adapt the treatment(s). Those patterns include very diffuse, ipsi- or bilateral gliomatosis-like progression, multicentric recurrence with emergence of remote low-grade or high-grade glioma, leptomeningeal dissemination, acute (early or delayed) local MT or bulky relapse into the operating cavity. This landscape of recurrence patterns may allow physicians to elaborate new tailored therapeutic strategies and scientists to develop original hypotheses for basic research.

Impact of 9p deletion and p16, Cyclin D1, and Myc hyperexpression on the outcome of anaplastic oligodendrogliomas.

OBJECTIVE: To study the presence of 9p deletion and p16, cyclin D1 and Myc expression and their respective diagnostic and prognostic interest in oligodendrogliomas. METHODS: We analyzed a retrospective series of 40 consecutive anaplastic oligodendrogliomas (OIII) from a single institution and compared them to a control series of 10 low grade oligodendrogliomas (OII). Automated FISH analysis of chromosome 9p status and immunohistochemistry for p16, cyclin D1 and Myc was performed for all cases and correlated with clinical and histological data, event free survival (EFS) and overall survival (OS). RESULTS: Chromosome 9p deletion was observed in 55% of OIII (22/40) but not in OII. Deletion was highly correlated to EFS (median = 29 versus 53 months, p<0.0001) and OS (median = 48 versus 83 months, p<0.0001) in both the total cohort and the OIII population. In 9p non-deleted oligodendrogliomas, p16 hyperexpression correlated with a shorter OS (p = 0.02 in OII and p = 0.0001 in OIII) whereas lack of p16 expression was correlated to a shorter EFS and OS in 9p deleted OIII (p = 0.001 and p = 0.0002 respectively). Expression of Cyclin D1 was significantly higher in OIII (median expression 45% versus 14% for OII, p = 0.0006) and was correlated with MIB-1 expression (p<0.0001), vascular proliferation (p = 0.002), tumor necrosis (p = 0.04) and a shorter EFS in the total cohort (p = 0.05). Hyperexpression of Myc was correlated to grade (median expression 27% in OII versus 35% in OIII, p = 0.03), and to a shorter EFS in 9p non-deleted OIII (p = 0.01). CONCLUSION: Chromosome 9p deletion identifies a subset of OIII with significantly worse prognosis. The combination of 9p status and p16 expression level identifies two distinct OIII populations with divergent prognosis. Hyperexpression of Bcl1 and Myc appears highly linked to anaplasia but the prognostic value is unclear and should be investigated further.

Impact of standard care on elderly glioblastoma patients.

BACKGROUND: Uncertainty persists about the survival advantage of concomitant and adjuvant temozolomide (TMZ) plus radiotherapy (RT) in elderly patients with newly diagnosed glioblastoma (GBM). We compared the clinical outcome of unselected elderly GBM patients treated with 4 adjuvant treatment modalities, including the Stupp protocol. METHODS: From 2010 to 2014, retrospective chart review was performed on 171 GBM patients aged ≥55 who received either concurrent chemoradiation therapy (CCRT) with standard 60 Gy/30 (SRT); CCRT with hypofractionated 40 Gy/15 (HRT); HRT alone; or TMZ alone. Stratification is by age (55-69, ≥70), KPS (<70, ≥70), and resection status (biopsy, resection). RESULTS: Out of 171 patients identified, 128(75%) had surgical resection, median age was 66(55-83), and median overall survival (mOS) 11.4mo. Majority (109/171) were treated according to the Stupp protocol (CCRT-SRT), and 106/171 received post-CCRT adjuvant TMZ (median of 3 cycles). In our population, age <70yo was a significant prognostic factor (mOS of patients aged 55-69 vs ≥70 yo = 13.3 vs 6.6 mo; P = .001). However, among the population receiving the Stupp regimen, there was no difference in survival between patients aged 55-69 and those ≥70 (respectively, 14.4 vs 13.2 mo; P = .798). Patients ≥70 yo had similar survival when treated with CCRT-HRT and CCRT-SRT (P = .248), although numbers were small. CONCLUSIONS: Our data suggests that, despite having a worse global prognostic than their younger counterparts, GBM patients ≥70yo with a good performance status could be treated according to the Stupp protocol with similar survival. Theses results need prospective confirmation.

Contribution of 1p, 19q, 9p and 10q Automated Analysis by FISH to the Diagnosis and Prognosis of Oligodendroglial Tumors According to WHO 2016 Guidelines.

OBJECTIVE: To study the feasibility and the diagnostic and prognostic interest of automated analysis of 1p, 19q, 9p and 10q status by FISH technique in oligodendroglial tumors. METHODS: We analyzed a retrospective series of 33 consecutive gliomas with oligodendroglial histology (originally diagnosed as 24 oligodendrogliomas and 9 oligoastrocytomas). For all cases, automated FISH analysis of 1p, 19q, 9p and 10q status were performed and compared to clinical and histological data, ATRX, IDH1R132H and alpha-internexin status (studied by immunohistochemistry) and overall survival (OS). Manual analysis of 9p and 10q status were also performed and compared to automated analysis to verify the concordance of the two methods. RESULTS: The 33 gliomas were reclassified into 13 low-grade oligodendrogliomas (OII), 10 anaplastic oligodendrogliomas (OIII), 3 diffuse astrocytomas (AII), 3 anaplastic astrocytomas (AIII) and 4 glioblastomas (GBM) according to the WHO 2016 histological criteria. The 1p and/or 19q imbalanced status were restricted to astrocytomas with no correlation to their grade or their OS. Chromosome 9p deletion was restricted to OIII (70%) and GBM (100%) and was correlated with a shorter OS in the total cohort (p = 0.0007), the oligodendroglioma cohort (p = 0.03) and the astrocytoma cohort (p = 0.001). Concordance between 9p manual and automated analysis was satisfactory (81%, κ = 0.69). Chromosome 10q deletion was restricted to GBMs (50%) and was correlated with a poor OS in both the total cohort (p = 0.003) and the astrocytoma (AS) cohort (p = 0.04). Concordance between manual and automated analysis was satisfactory (79%, κ = 0.62). CONCLUSION: Automated analysis of 1p, 19q, 9p and 10q status by FISH is a reliable technique which allows for refined classification of oligodendroglial tumors. 1p and/or 19q imbalanced status is evidence of astrocytic differentiation. 9p deletion is found in high grade oligodendrogliomas and astrocytomas with a poor OS. 10q is related to GBM status and a poor OS.

Surgery of insular and paralimbic diffuse low-grade gliomas: technical considerations.

Once considered a "no man's land" especially when invaded by a diffuse low grade glioma (DLGG), the insula remains to this day a surgical challenge. Surgery for insular DLGG involves consideration of its hidden location under the potentially eloquent operculae, the proximity to vascular tree and high density of functions not only in the insular cortex but also in the white fiber pathways passing under the insular lobe. The natural history of DLGG and the potential benefits and consequences of the surgical approach also need a close look. In the last decade, a better knowledge of the functional anatomy and connectivity of this region, as well as an improvement in surgical techniques as direct stimulation mapping, combined with an increasing literature showing a favorable impact of maximal resection for DLGG, were deciding factors in the paradigmatic shift from expectative treatment to early surgical management. Here, our goal is to discuss the structural and functional aspects of the insula, the specificities of insular and paralimbic DLGG by emphasizing the technical considerations of surgery in this region, as well as its oncological and functional outcomes. In summary, this new strategy based upon early maximal safe surgical resection showed both oncological benefit and preservation of quality of life-or even an improvement thanks to epilepsy relief.

Automated Analysis of 1p/19q Status by FISH in Oligodendroglial Tumors: Rationale and Proposal of an Algorithm.

OBJECTIVE: To propose a new algorithm facilitating automated analysis of 1p and 19q status by FISH technique in oligodendroglial tumors with software packages available in the majority of institutions using this technique. METHODS: We documented all green/red (G/R) probe signal combinations in a retrospective series of 53 oligodendroglial tumors according to literature guidelines (Algorithm 1) and selected only the most significant combinations for a new algorithm (Algorithm 2). This second algorithm was then validated on a prospective internal series of 45 oligodendroglial tumors and on an external series of 36 gliomas. RESULTS: Algorithm 2 utilizes 24 G/R combinations which represent less than 40% of combinations observed with Algorithm 1. The new algorithm excludes some common G/R combinations (1/1, 3/2) and redefines the place of others (defining 1/2 as compatible with normal and 3/3, 4/4 and 5/5 as compatible with imbalanced chromosomal status). The new algorithm uses the combination + ratio method of signal probe analysis to give the best concordance between manual and automated analysis on samples of 100 tumor cells (91% concordance for 1p and 89% concordance for 19q) and full concordance on samples of 200 tumor cells. This highlights the value of automated analysis as a means to identify cases in which a larger number of tumor cells should be studied by manual analysis. Validation of this algorithm on a second series from another institution showed a satisfactory concordance (89%, κ = 0.8). CONCLUSION: Our algorithm can be easily implemented on all existing FISH analysis software platforms and should facilitate multicentric evaluation and standardization of 1p/19q assessment in gliomas with reduction of the professional and technical time required.

ImmunoFISH is a reliable technique for the assessment of 1p and 19q status in oligodendrogliomas.

OBJECTIVE: To develop a new ImmunoFISH technique for the study of oligodendrogliomas by combining a standard immunohistochemical stain using MIB-1 antibody with a standard FISH technique using commercial 1p36 and 19q13 chromosomal probes. METHODS: Validation was performed by two observers on a series of 36 pre-selected oligodendrogliomas and compared to the results previously determined by FISH alone. RESULTS: The ImFISH technique is easy to perform and to analyze and is no more time-consuming than the usual FISH technique. Our results show that the inter-observer reliability of ImFISH is high (κ = 0.86 and 0.95 respectively for 1p and 19q). Compared to FISH, the ImFISH exhibits a very high sensitivity (∼100%) and specificity (∼90%) for 1p and/or 19q deleted cases. The sensitivity is high for normal cases (∼85%) and imbalanced cases (∼90%) with a specificity ranging between 50 and 85%. Finally, there were no significant differences between FISH and ImFISH results calculated on 60, 40 or 20 cells. CONCLUSION: Our study demonstrates the reliability of the ImFISH technique in oligodendrogliomas and emphasizes its advantage in poorly cellular tumoral specimen.

A new method of quantitatively assessing the opening of the blood-brain barrier in murine animal models.

The blood-brain barrier (BBB) restricts the delivery of drugs into the brain. Different strategies have been developed to circumvent this obstacle. One such approach, the osmotic BBB disruption (BBBD), has been under pre-clinical study since the 70's. Typically, qualitative ex vivo assessment of the extent of BBBD has been performed using Evan's blue staining technique. In this study, we describe a simple quantitative technique based on albumin indirect immunohistochemistry to measure the extent of BBB breach. Thirty Fischer rats were assigned to one of 6 groups: a control group, and BBBD groups with escalation in IA mannitol infusion rate: 0.06, 0.08, 0.10, 0.12 and 0.15 cc/s. Fifteen minutes after the BBBD procedure, the animals were sacrificed, brain harvested and sections stained for albumin. Using an image analysis software, isolated albumin staining pixels were expressed as a fraction of the treated hemisphere. This ratio was used as a percentage value in the intensity of the BBB permeabilization. All sections studied harbored staining, averaging 0.37% for the controls (group 1), 5.69% for group 2 (0.06 cc/s), 10.44% for group 3 (0.08 cc/s), 6.99% for group 4 (0.1 cc/s), 18.50% for group 5 (0.12 cc/s) and reaching 61.70% for group 6 (0.15 cc/s). Important variations were observed between animals. A threshold effect was observed, and animals in group 6 presented a significant increase in BBB permeabilization compared to the other groups. We hereby detail a simple technique that can be applied to quantitatively measure the extent of the BBB breach notwithstanding the pathological process.

Investigation of intravenous delivery of nanoliposomal topotecan for activity against orthotopic glioblastoma xenografts.

Achieving effective treatment outcomes for patients with glioblastoma (GBM) has been impeded by many obstacles, including the pharmacokinetic limitations of antitumor agents, such as topotecan (TPT). Here, we demonstrate that intravenous administration of a novel nanoliposomal formulation of TPT (nLS-TPT) extends the survival of mice with intracranial GBM xenografts, relative to administration of free TPT, because of improved biodistribution and pharmacokinetics of the liposome-formulated drug. In 3 distinct orthotopic GBM models, 3 weeks of biweekly intravenous therapy with nLS-TPT was sufficient to delay tumor growth and significantly extend animal survival, compared with treatment with free TPT (P ≤ .03 for each tumor tested). Analysis of intracranial tumors showed increased activation of cleaved caspase-3 and increased DNA fragmentation, both indicators of apoptotic response to treatment with nLS-TPT. These results demonstrate that intravenous delivery of nLS-TPT is a promising strategy in the treatment of GBM and support clinical investigation of this therapeutic approach.

Sex differences in perceived pain are affected by an anxious brain.

Decades of research confirm that women have greater pain sensitivity than men. Women also show greater overall anxiety sensitivity than men. Given these differences, we hypothesized that sex differences in anxiety would explain sex differences in experienced pain and physiological responses to pain (at both spinal and cortical levels). By measuring subjective pain, state/trait anxiety, nociceptive flexion reflexes, and somatosensory evoked potentials (SEPs), it was possible to test the effects of anxiety on the processing of painful drives at different levels of the neuraxis while also documenting the role played by anxiety on sex differences in experienced pain. Results confirm that women are indeed more sensitive to pain than men. Importantly, this difference was accompanied by a significant sex difference in cortical activity (SEP amplitude) but not spinal nociceptive activity, suggesting that much of the sex difference in experienced pain is attributable to variations in thalamocortical processing and to ensuing changes in the appraisal of and/or emotional response to noxious insult. In support of this claim, we found that sex differences in cortical activity and subjective pain disappeared when trait anxiety was controlled for. This means that stable predispositions to respond with heightened apprehension contribute to baseline pain sensitivity differences between the sexes. These results indicate that the modulatory effect of affect on pain-related brain processes may explain why men and women experience painful shocks so differently. In our study, the mediating role of anxiety on sex differences in pain was tested and confirmed using path analysis.

Pharmacologic inhibition of cyclin-dependent kinases 4 and 6 arrests the growth of glioblastoma multiforme intracranial xenografts.

Activation of cyclin-dependent kinases 4 and 6 (cdk4/6) occurs in the majority of glioblastoma multiforme (GBM) tumors, and represents a promising molecular target for the development of small molecule inhibitors. In the current study, we investigated the molecular determinants and in vivo response of diverse GBM cell lines and xenografts to PD-0332991, a cdk4/6-specific inhibitor. In vitro testing of PD-0332991 against a panel of GBM cell lines revealed a potent G(1) cell cycle arrest and induction of senescence in each of 16 retinoblastoma protein (Rb)-proficient cell lines regardless of other genetic lesions, whereas 5 cell lines with homozygous inactivation of Rb were completely resistant to treatment. Short hairpin RNA depletion of Rb expression conferred resistance of GBM cells to PD-0332991, further demonstrating a requirement of Rb for sensitivity to cdk4/6 inhibition. PD-0332991 was found to efficiently cross the blood-brain barrier and proved highly effective in suppressing the growth of intracranial GBM xenograft tumors, including those that had recurred after initial therapy with temozolomide. Remarkably, no mice receiving PD-0332991 died as a result of disease progression while on therapy. Additionally, the combination of PD-0332991 and radiation therapy resulted in significantly increased survival benefit compared with either therapy alone. In total, our results support clinical trial evaluation of PD-0332991 against newly diagnosed as well as recurrent GBM, and indicate that Rb status is the primary determinant of potential benefit from this therapy.

Tumor bed radiosurgery after resection of cerebral metastases.

OBJECTIVE: Adjuvant irradiation after resection of brain metastases reduces the risk of local recurrence. Whole-brain radiation therapy can be associated with significant neurotoxicity in long-term survivors of brain metastases. This retrospective study evaluates the role of tumor bed stereotactic radiosurgery as an alternative method of irradiation after initial resection of brain metastases to prevent local recurrence. METHODS: Forty patients underwent tumor bed radiosurgery after resection of brain metastases at two separate academic medical centers. The median age was 59.5 years. Twenty patients (67.5%) had single metastases. Resection was complete in 80% and partial in 20% of the patients. At the time of radiosurgery, systemic disease was active in 57.5%, inactive in 32.5%, and in remission in 10% of the patients. The median Karnofsky Performance Scale score was 80% (range, 60-100%). Radiosurgery was performed a median of 4 weeks after tumor resection. The median cavity radiosurgery volume was 9.1 ml (range, 0.6-39.9 ml). The median margin and maximum radiation dose were 16 and 32 Gy, respectively. RESULTS: Local control at the resection site was achieved in 73% of patients at a median follow-up period of 13 months. No variable significantly affected local control. New remote brain metastases occurred in 54% of the patients. Symptomatic radiation effect was seen in 5.4% of the patients. The median survival was 13 months after radiosurgery (range, 2-56 mo). CONCLUSION: Tumor bed radiosurgery provides effective local control of the tumor after resection in most patients. These preliminary data support radiosurgery after resection rather than traditional radiation therapy.