Roland Goldbrunner

Long-term Cultures of Bone Marrow-Derived Human Mesenchymal Stem Cells Frequently Undergo Spontaneous Malignant Transformation

Human mesenchymal stem cells (hMSC) aid in tissue maintenance and repair by differentiating into specialized cell types. Due to this ability, hMSC are currently being evaluated for cell-based therapies of tissue injury and degenerative diseases. However, extensive expansion ex vivo is a prerequisite to obtain the cell numbers required for human cell-based therapy protocols. Recent studies indicate that hMSC may contribute to cancer development and progression either by acting as cancer-initiating cells or through interactions with stromal elements. If spontaneous transformation ex vivo occurs, this may jeopardize the use of hMSC as therapeutic tools. Whereas murine MSC readily undergo spontaneous transformation, there are conflicting reports about spontaneous transformation of hMSC. We have addressed this controversy in a two-center study by growing bone marrow-derived hMSC in long-term cultures (5-106 weeks). We report for the first time spontaneous malignant transformation to occur in 45.8% (11 of 24) of these cultures. In comparison with hMSC, the transformed mesenchymal cells (TMC) showed a significantly increased proliferation rate and altered morphology and phenotype. In contrast to hMSC, TMC grew well in soft agar assays and were unable to undergo complete differentiation. Importantly, TMC were highly tumorigenic, causing multiple fast-growing lung deposits when injected into immunodeficient mice. We conclude that spontaneous malignant transformation may represent a biohazard in long-term ex vivo expansion of hMSC. On the other hand, this spontaneous transformation process may represent a unique model for studying molecular pathways initiating malignant transformation of hMSC.

Phase I/IIa Study of Cilengitide and Temozolomide With Concomitant Radiotherapy Followed by Cilengitide and Temozolomide Maintenance Therapy in Patients With Newly Diagnosed Glioblastoma

PURPOSE Invasion and migration are key processes of glioblastoma and are tightly linked to tumor recurrence. Integrin inhibition using cilengitide has shown synergy with chemotherapy and radiotherapy in vitro and promising activity in recurrent glioblastoma. This multicenter, phase I/IIa study investigated the efficacy and safety of cilengitide in combination with standard chemoradiotherapy in newly diagnosed glioblastoma. PATIENTS AND METHODS Patients (age > or = 18 to < or = 70 years) were treated with cilengitide (500 mg) administered twice weekly intravenously in addition to standard radiotherapy with concomitant and adjuvant temozolomide. Treatment was continued until disease progression or for up to 35 weeks. The primary end point was progression-free survival (PFS) at 6 months. RESULTS Fifty-two patients (median age, 57 years; 62% male) were included. Six- and 12-month PFS rates were 69% (95% CI, 54% to 80%) and 33% (95% CI, 21% to 46%). Median PFS was 8 months (95% CI, 6.0 to 10.7 months). Twelve- and 24-month overall survival (OS) rates were 68% (95% CI, 53% to 79%) and 35% (95% CI, 22% to 48%). Median OS was 16.1 months (95% CI, 13.1 to 23.2 months). PFS and OS were longer in patients with tumors with O(6)-methylguanine-DNA methyltransferase (MGMT) promoter methylation (13.4 and 23.2 months) versus those without MGMT promoter methylation (3.4 and 13.1 months). The combination of cilengitide with temozolomide and radiotherapy was well tolerated, with no additional toxicity. No pharmacokinetic interactions between temozolomide and cilengitide were identified. CONCLUSION Compared with historical controls, the addition of concomitant and adjuvant cilengitide to standard chemoradiotherapy demonstrated promising activity in patients with glioblastoma with MGMT promoter methylation.

Spontaneous Malignant Transformation of Human Mesenchymal Stem Cells Reflects Cross-Contamination: Putting the Research Field on Track - Letter

Several groups, including ours, have published results showing spontaneous transformation of human mesenchymal stem cells (MSC). Recently, we reported in this journal spontaneous transformation of bone marrow-derived human MSC (hMSC), isolated and expanded independently in two laboratories ([1][1

Imaging of integrin αvβ3 expression in patients with malignant glioma by [18F] Galacto-RGD positron emission tomography

Inhibitors targeting the integrin alpha(v)beta(3) are promising new agents currently tested in clinical trials for supplemental therapy of glioblastoma multiforme (GBM). The aim of our study was to evaluate (18)F-labeled glycosylated Arg-Gly-Asp peptide ([(18)F]Galacto-RGD) PET for noninvasive imaging of alpha(v)beta(3) expression in patients with GBM, suggesting eligibility for this kind of additional treatment. Patients with suspected or recurrent GBM were examined with [(18)F]Galacto-RGD PET. Standardized uptake values (SUVs) of tumor hotspots, galea, and blood pool were derived by region-of-interest analysis. [(18)F]Galacto-RGD PET images were fused with cranial MR images for image-guided surgery. Tumor samples taken from areas with intense tracer accumulation in the [(18)F]Galacto-RGD PET images and were analyzed histologically and immunohistochemically for alpha(v)beta(3) integrin expression. While normal brain tissue did not show significant tracer accumulation (mean SUV, 0.09 +/- 0.04), GBMs demonstrated significant but heterogeneous tracer uptake, with a maximum in the highly proliferating and infiltrating areas of tumors (mean SUV, 1.6 +/- 0.5). Immunohistochemical staining was prominent in tumor microvessels as well as glial tumor cells. In areas of highly proliferating glial tumor cells, tracer uptake (SUVs) in the [(18)F]Galacto-RGD PET images correlated with immunohistochemical alpha(v)beta(3) integrin expression of corresponding tumor samples. These data suggest that [(18)F] Galacto-RGD PET successfully identifies alpha(v)beta(3) expression in patients with GBM and might be a promising tool for planning and monitoring individualized cancer therapies targeting this integrin.

PTK787/ZK222584, an inhibitor of vascular endothelial growth factor receptor tyrosine kinases, decreases glioma growth and vascularization.

OBJECTIVE The aim of this study was to test the efficacy of PTK787/ZK222584, an inhibitor of vascular endothelial growth factor (VEGF) receptor tyrosine kinases, on VEGF-dependent glioma vascularization and growth. METHODS C6 rat glioma cells were transfected with VEGF(164) in a sense (V(+)) or antisense (V(-)) direction. Spheroids generated from V(+) or V(-) cells were implanted orthotopically into 60 rat brains. Expression of VEGF and fetal liver kinase-1 (VEGF receptor 2) was assessed immunohistochemically. Animals with V(+) gliomas received orally administered PTK787/ZK222584 on postoperative Day (POD) 1 to 12 or POD 7 to 12. Untreated animals served as negative controls, and animals with V(-) gliomas served as positive controls. Growth and vascularization were evaluated by magnetic resonance imaging and immunohistochemistry. RESULTS Flk-1 expression was positive within tumor vessels in V(+) gliomas, whereas all C6 clones were negative for fetal liver kinase-1 in vitro. Early (POD 1-12) and delayed (POD 7-12) application of PTK787/ZK222584 in V(+) glioma-bearing animals resulted in a significant reduction of tumor size (71% and 36%, P < 0.05) as measured by magnetic resonance imaging volumetry. Early treated V(+) gliomas reached similar volumes compared with V(-) gliomas. Vessel density was significantly reduced (42.3% and 25.7%, P < 0.05), and areas of intratumoral necrosis were enlarged (by 1.7-fold after early treatment). Additionally, proliferation was decreased by 89% and 72% (P < 0.05). There was no growth-inhibiting effect of PTK787/ZK222584 on V(-) cells observed. CONCLUSION PTK787/ZK222584 significantly halted VEGF-mediated glioma growth by inhibition of neovascularization and proliferation, providing a promising new tool in malignant glioma therapy.S OF OPEN PAPERS 800 Difference in Risk of Cerebral Aneurysm Recurrence after Surgical Treatment between Ruptured and Unruptured Patients Motoshi Sawada, M.D., Hiromu Hadeishi, M.D., Akifumi Suzuki, M.D., Nobuyuki Yasui, M.D. INTRODUCTION: It is unclear how the risk of aneurysm recurrence varies over 10 years between ruptured and unruptured patients with a successfully treated aneurysm. This study was undertaken to compare the long-term angiographic outcome of surgically treated aneurysms between ruptured and unruptured patients. METHODS: We analyzed 502 patients who were independent in daily life and younger than 75 years in age and who had undergone surgery to treat a ruptured or unruptured cerebral aneurysm at our institution more than 10 years before the study. A letter explaining the objective and method of follow-up angiography was sent to each of the patients, and cerebral angiography was scheduled for those patients who showed an interest in undergoing angiography. A reply was obtained from 252 out of the 502 patients, with 99 patients expressing their willingness to undergo follow-up angiography. RESULTS: Ninety-nine patients (85 ruptured and 14 unruptured patients) underwent cerebral angiography. The mean interval from surgery was 15.6 years for all patients (range, 11 to 27 years). De novo aneurysms were detected in 28 out of 85 (32.9%) ruptured and in two out of 14 (14.3%) unruptured patients. The annual rate of de novo aneurysm formation was 2.8% for ruptured patients and 1.2% for unruptured patients. Aneurysm regrowth occurred only in patients with ruptured aneurysms, but not unruptured aneurysms. CONCLUSION: The annual rate of de novo aneurysm formation significantly increased in patients with a previous subarachnoid hemorrhage compared with those with surgically treated unruptured aneurysms. This rate was considerably higher than the previously reported risk, especially in female patients surgically treated for ruptured aneurysms. It is suggested that the cumulative recurrence rate becomes significant more than 10 years after the original treatment, and that follow-up angiography to detect recurrent aneurysms may be indicated for patients with clipped aneurysms 10 to 20 years after surgery. 801 Prevention of Cerebral Vasospasm after Aneurysmal Subarachnoid Hemorrhage with Clazosentan, an Endothelin Receptor Antagonist Robert L. MacDonald, M.D., A. Kakarieka, M.D., Stephan A. Mayer, M.D., Alberto Pasqualin, M.D., D. Ruefenacht, M.D., Peter Schmiedek, M.D., Neal F. Kassell, M.D. INTRODUCTION: To assess the efficacy, safety and tolerability of 1, 5, and 15 mg/hour intravenous clazosentan in preventing cerebral vasospasm after aneurysmal subarachnoid hemorrhage (SAH). METHODS: This was a Phase 2b, international, multicenter, double-blind, placebo-controlled dose-finding study. Male and female patients (age, 18–70 yr) with SAH and confirmed ruptured saccular aneurysms were screened within 48 hours after SAH and were randomized 1:1:1:1 to one of three dose groups of the study drug or to placebo. Drug or placebo infusion commenced within 56 hours of aneurysm rupture and was stopped on Day 14 after bleeding. Clipping or coiling started before or within 12 hours of the start of drug infusion. The occurrence of moderate ( 34% arterial narrowing) or severe ( 67% narrowing) vasospasm was measured quantitatively by comparison of digital subtraction angiography at baseline and on Day 9 2. The significance of test-drug treatment effects versus placebo was determined by Fisher’s exact test. Vasospasm-related morbidity and clinical outcome were assessed by comparison of computed tomographic scans obtained at baseline, 24 to 48 hours after aneurysm treatment, and at 6 weeks and clinical assessment at 12 weeks after the SAH, respectively. Safety and tolerability assessments were also conducted. RESULTS: Four hundred thirteen patients were randomized. Seventy-one percent were female and 29% were male. The mean age was 51 years. Coiling or clipping was performed in 54 and 46% of subjects, respectively. The occurrence and severity of vasospasm will be reported per treatment group. Secondary end points that will be reported for each treatment group are cerebral infarcts as determined by follow-up computed tomographic scans, vasospasm-related morbidity, and clinical outcome. The incidence of adverse events will also be presented. CONCLUSION: Results of this randomized, blinded trial with complete quantitative angiographic and computed tomographic imaging documentation will demonstrate whether clazosentan, an endothelin ET receptor antagonist, reduces angiographic vasospasm and alters clinical outcome in this patient population. 802 Experience in the Endovascular Management of 76 Patients with Middle Cerebral Artery Aneurysms Ketan R. Bulsara, M.D., Marshall E. Tolbert, M.D., Michael J. Alexander, M.D., F.A.C.S. INTRODUCTION: Endovascular management of middle cerebral artery (MCA) aneurysms has historically been associated with high morbidity. We review a single endovascular neurosurgeon’s experience in the management of these aneurysms. METHODS: All patients with MCA aneurysms evaluated between 2000 and 2006 were evaluated for endovascular management. Absolute contraindication for endovascular management were aneurysms associated with a hematoma causing mass effect and obvious origin of MCA branches from the dome of the aneurysm. RESULTS: Seventy-six patients were considered for endovascular management. In five patients, the endovascular procedure was aborted after selective microcatherization of the MCA owing to unfavorable morphology. Twenty men and 51 women underwent endovascular treatment of their aneurysm. The average age of the patients was 62 years (range, 20–84 yr). Forty-seven patients had incidentally discovered aneurysms, whereas 24 patients presented with subarachnoid hemorrhage. Twelve patients underwent stent-assisted coiling. Fifty-one percent of patients (n 36) had no immediate residual aneurysm after treatment. Thirty-six patients had follow-up angiography at an average of 9 months. Of these, 27 patients (75%) had no residual aneurysm. Four patients (11%) had some recanalization of their aneurysm, with one requiring retreatment. Five patients (14%) had small stable residual neck remnants. Two patients (2.8%) had transient thromboembolic complications and two (2.8%) had significant persistent neurological deficits. CONCLUSION: Careful pre-procedure selection can help identify NEUROSURGERY VOLUME 59 | NUMBER 2 | AUGUST 2006 | 453 MCA aneurysms that are amenable to endovascular management. Long-term data is needed on the durability of this treatment option. 803 Hyperglycemia Independently Increases the Risk of Perioperative Stroke, Myocardial Infarction, and Death after Carotid Endarterectomy Matthew McGirt, M.D., Graeme F. Woodworth, B.S., Alexander L. Coon, M.D., Donald Buck, B.S., Judy Huang, M.D., Richard E. Clatterbuck, M.D., Bruce Perler, M.D., Rafael J. Tamargo, M.D. INTRODUCTION: Clinical and experimental evidence suggests that hyperglycemia lowers the neuronal ischemic threshold, potentiates stroke volume in focal ischemia, and is associated with morbidity and mortality in the surgical critical care setting. It remains unknown whether or not hyperglycemia during carotid endarterectomy (CEA) predisposes to perioperative stroke and operative related morbidity and mortality. METHODS: The clinical and radiological records of all patients undergoing CEA and operative day glucose measurement at an academic institution from 1994 to 2004 were reviewed and 30-day outcomes were assessed. The independent association of operative day glucose before CEA and perioperative morbidity and mortality was assessed via multivariate logistic regression analysis. RESULTS: One thousand two-hundred one patients with a mean age of 72 10 years (748 men, 453 women) underwent CEA (676 asymptomatic, 525 symptomatic). Overall, stroke occurred in 46 (3.8%) patients, transient ischemic attack (TIA) in 19 (1.6%), myocardial infarction (MI) in 19 (1.6%), and death in 17 (1.4%). Increasing operative day glucose was independently associated with perioperative stroke or TIA (odds ratio [OR], 1.005; 95% confidence interval [CI], 1.00–1.01; P 0.03), MI (OR, 1.01; 95% CI, 1.004–1.016; P 0.017), and death (OR, 1.007; 95% CI, 1.00–1.015; P 0.04)]. Patients with operative day glucose greater than 200 mg/dl were 2.8-fold, 4.3-fold, and 3.3-fold more likely to experience perioperative stroke or TIA (OR, 2.78; 95% CI, 1.37–5.67; P 0.005), MI (OR, 4.29; 95% CI, 1.28–14.4; P 0.018), or death (OR, 3.29; 95% CI, 1.07–10.1; P 0.037), respectively. Median (interquartile range) length of hospitalization was greater for patients with operative day glucose greater than 200 mg/dl (4 [2–15] versus 3 [2–7] d, P 0.05). CONCLUSION: Independent of previous cardiac disease, diabetes, or other comorbidities, hyperglycemia at the time of CEA was associated with an increased risk of perioperative stroke or TIA, MI, and death. Strict glucose control should be attempted before surgery to minimize the risk of morbidity and mortality after CEA. 804 Deployment of Self-expanding Stents for Recanalization of Acute Cerebrovascular Occlusions Elad I. Levy, M.D., Ricky Mehta, B.S., Rishi Gupta, M.D., Ricardo A. Hanel, M.D., Andrea J. Chamczuk, M.D., B.Sc., M.Sc., David Fiorella, M.D., Ph.D., Henry H. Woo, M.D., Felipe Albuquerque, M.D., L. Nelson Hopkins, M.D. INTRODUCTION: Stent-assisted revascularization has been reported to increase prevailing recanalization rates (approximately 50– 60%) for occlusive vessels recalcitrant to thrombolytics. Although balloon-mounted coronary stents can displace thrombus (via angioplasty) and retain clot along vessel walls, they are difficult to track through tortuous cerebrovasculature. Intracranial self-expanding stents (SES) are more flexible and exert less radial outward force during deployment, increasing deliverability and safety

Expression of Integrin αvβ3 in Gliomas Correlates with Tumor Grade and Is not Restricted to Tumor Vasculature

In malignant gliomas, the integrin adhesion receptors seem to play a key role for invasive growth and angiogenesis. However, there is still a controversy about the expression and the distribution of αvβ3 integrin caused by malignancy. The aim of our study was to assess the extent and pattern of αvβ3 integrin expression within primary glioblastomas (GBMs) compared with low‐grade gliomas (LGGs). Tumor samples were immunostained for the detection of αvβ3 integrin and quantified by an imaging software. The expression of αvβ3 was found to be significantly higher in GBMs than in LGGs, whereby focal strong reactivity was restricted to GBMs only. Subsequent analysis revealed that not only endothelial cells but also, to a large extent, glial tumor cells contribute to the overall amount of αvβ3 integrin in the tumors. To further analyze the integrin subunits, Western blots from histologic sections were performed, which demonstrated a significant difference in the expression of the β3 integrin subunit between GBMs and LGGs. The presented data lead to new insights in the pattern of αvβ3 integrin in gliomas and are of relevance for the inhibition of αvβ3 integrin with specific RGD peptides and interfering drugs to reduce angiogenesis and tumor growth.

Cell-extracellular matrix interaction in glioma invasion.

 The complex receptor-ECM interaction in glioma cell invasion is discussed focussing upon the role of integrin receptors and matrix-metalloproteinases. Influencing these molecules or their regulation may lead to novel therapeutic approaches in the treatment of malignant glioma.

Acoustic neuroma surgery as an interdisciplinary approach: a neurosurgical series of 508 patients

OBJECTIVES To evaluate an interdisciplinary concept (neurosurgery/ear, nose, and throat (ENT)) of treating acoustic neuromas with extrameatal extension via the retromastoidal approach. To analyse whether monitoring both facial nerve EMG and BAEP improved the functional outcome in acoustic neuroma surgery. METHODS In a series of 508 patients consecutively operated on over a period of 7 years, functional outcome of the facial nerve was evaluated according to the House/Brackmann scale and hearing preservation was classified using the Gardner/Robertson system. RESULTS Facial monitoring (396 of 508 operations) and continuous BAEP recording (229 of 399 cases with preserved hearing preoperatively) were performed routinely. With intraoperative monitoring, the rate of excellent/good facial nerve function (House/Brackmann I-II) was 88.7%. Good functional hearing (Gardner/Robertson 1–3) was preserved in 39.8%. CONCLUSION Acoustic neuroma surgery via a retrosigmoidal approach is a safe and effective treatment for tumours with extrameatal extension. Functional results can be substantially improved by intraoperative monitoring. The interdisciplinary concept of surgery performed by ENT and neurosurgeons was particularly convincing as each pathoanatomical phase of the operation is performed by a surgeon best acquainted with the regional specialties.

MGMT Promoter Methylation Is a Strong Prognostic Biomarker for Benefit from Dose-Intensified Temozolomide Rechallenge in Progressive Glioblastoma: The DIRECTOR Trial.

Purpose: Rechallenge with temozolomide (TMZ) at first progression of glioblastoma after temozolomide chemoradiotherapy (TMZ/RT→TMZ) has been studied in retrospective and single-arm prospective studies, applying temozolomide continuously or using 7/14 or 21/28 days schedules. The DIRECTOR trial sought to show superiority of the 7/14 regimen. Experimental Design: Patients with glioblastoma at first progression after TMZ/RT→TMZ and at least two maintenance temozolomide cycles were randomized to Arm A [one week on (120 mg/m2 per day)/one week off] or Arm B [3 weeks on (80 mg/m2 per day)/one week off]. The primary endpoint was median time-to-treatment failure (TTF) defined as progression, premature temozolomide discontinuation for toxicity, or death from any cause. O6-methylguanine DNA methyltransferase (MGMT) promoter methylation was prospectively assessed by methylation-specific PCR. Results: Because of withdrawal of support, the trial was prematurely closed to accrual after 105 patients. There was a similar outcome in both arms for median TTF [A: 1.8 months; 95% confidence intervals (CI), 1.8–3.2 vs. B: 2.0 months; 95% CI, 1.8–3.5] and overall survival [A: 9.8 months (95% CI, 6.7–13.0) vs. B: 10.6 months (95% CI, 8.1–11.6)]. Median TTF in patients with MGMT-methylated tumors was 3.2 months (95% CI, 1.8–7.4) versus 1.8 months (95% CI, 1.8–2) in MGMT-unmethylated glioblastoma. Progression-free survival rates at 6 months (PFS-6) were 39.7% with versus 6.9% without MGMT promoter methylation. Conclusions: Temozolomide rechallenge is a treatment option for MGMT promoter-methylated recurrent glioblastoma. Alternative strategies need to be considered for patients with progressive glioblastoma without MGMT promoter methylation. Clin Cancer Res; 21(9); 2057–64. ©2015 AACR.

Heparin and Air Filters Reduce Embolic Events Caused by Intra-Arterial Cerebral Angiography A Prospective, Randomized Trial

Background—Intra-arterial cerebral angiography is associated with a low risk for neurological complications, but clinically silent ischemic events after angiography have been seen in a substantial number of patients. Methods and Results—In a prospective study, diffusion-weighted magnetic resonance imaging (DW-MRI) before and after intra-arterial cerebral angiography and transcranial Doppler sonography during angiography were used to evaluate the frequency of cerebral embolism. One hundred fifty diagnostic cerebral angiographies were randomized into 50 procedures, each using conventional angiographic technique, or systemic heparin treatment throughout the procedure, or air filters between the catheter and both the contrast medium syringe and the catheter flushing. There was no neurological complication during or after angiography. Overall, DW-MRI revealed 26 new ischemic lesions in 17 patients (11%). In the control group, 11 patients showed a total of 18 lesions. In the heparin group, 3 patients showed a total of 4 lesions. In the air filter group, 3 patients exhibited a total of 4 lesions. The reduced incidence of ischemic events in the heparin and air filter groups compared with the control group was significantly different (P=0.002). Transcranial Doppler sonography demonstrated a large number of microembolic signals that was significantly lower in the air filter group compared with the heparin and control groups (P<0.01), which did not differ from each other. Conclusions—Air filters and heparin both reduce the incidence of silent ischemic events detected by DW-MRI after intra-arterial cerebral angiography and can potentially lower clinically overt ischemic complications. This may apply to any intra-arterial angiographic procedure.