The role of AKT isoforms in glioblastoma: AKT3 delays tumor progression.

The growth factor receptor/PI3K/AKT pathway is an important drug target in many cancers including Glioblastoma. AKT, a key node in the pathway, has 3 isoforms, AKT1, AKT2 and AKT3. Here we investigate their role in GBM. We find each activated, ser473 phosphorylated isoform is present in some GBMs but expression patterns vary. There is a direct relationship between human GBM patient outcome and both AKT1 and AKT2 mRNA levels, but an inverse relationship with AKT3 mRNA. Furthermore, AKT3 mRNA levels were high in a less aggressive GBM subtype. Overexpressing AKT3 improves survival in a rodent model of GBM and decreases colony forming efficiency, but not growth rate, in glioma cells. Silencing AKT3 slows cell cycle progression in one cell line and increases apoptosis in another. Our studies of AKT3 substrates indicate (1) silencing both AKT2 and AKT3 reduces GSK3 phosphorylation (2) only AKT2 silencing reduces S6 phosphorylation. Since S6 phosphorylation is a marker of mTORC1 activity this indicates that AKT2 activates mTORC1, but AKT3 does not. Our results indicate AKT isoforms have different roles and downstream substrates in GBM. Unexpectedly, they indicate AKT3 delays tumor progression. Therefore strategies that inhibit AKT3 may be unhelpful in some GBM patients.

Molecular subclasses of high-grade glioma predict prognosis, delineate a pattern of disease progression, and resemble stages in neurogenesis.

Previously undescribed prognostic subclasses of high-grade astrocytoma are identified and discovered to resemble stages in neurogenesis. One tumor class displaying neuronal lineage markers shows longer survival, while two tumor classes enriched for neural stem cell markers display equally short survival. Poor prognosis subclasses exhibit markers either of proliferation or of angiogenesis and mesenchyme. Upon recurrence, tumors frequently shift toward the mesenchymal subclass. Chromosomal locations of genes distinguishing tumor subclass parallel DNA copy number differences between subclasses. Functional relevance of tumor subtype molecular signatures is suggested by the ability of cell line signatures to predict neurosphere growth. A robust two-gene prognostic model utilizing PTEN and DLL3 expression suggests that Akt and Notch signaling are hallmarks of poor prognosis versus better prognosis gliomas, respectively.

Integrated genomic and epigenomic analyses pinpoint biallelic gene inactivation in tumors.

Aberrant methylation of CpG islands and genomic deletion are two predominant mechanisms of gene inactivation in tumorigenesis, but the extent to which they interact is largely unknown. The lack of an integrated approach to study these mechanisms has limited the understanding of tumor genomes and cancer genes. Restriction landmark genomic scanning (RLGS; ref. 1) is useful for global analysis of aberrant methylation of CpG islands, but has not been amenable to alignment with deletion maps because the identity of most RLGS fragments is unknown. Here, we determined the nucleotide sequence and exact chromosomal position of RLGS fragments throughout the genome using the whole chromosome of origin of the fragments and in silico restriction digestion of the human genome sequence. To study the interaction of these gene-inactivation mechanisms in primary brain tumors, we integrated RLGS-based methylation analysis with high-resolution deletion maps from microarray-based comparative genomic hybridization (array CGH; ref. 3). Certain subsets of gene-associated CpG islands were preferentially affected by convergent methylation and deletion, including genes that exhibit tumor-suppressor activity, such as CISH1 (encoding SOCS1; ref. 4), as well as genes such as COE3 that have been missed by traditional non-integrated approaches. Our results show that most aberrant methylation events are focal and independent of deletions, and the rare convergence of these mechanisms can pinpoint biallelic gene inactivation without the use of positional cloning.

DNA copy number alterations in central primitive neuroectodermal tumors and tumors of the pineal region: an international individual patient data meta-analysis.

Little is known about frequency, association with clinical characteristics, and prognostic impact of DNA copy number alterations (CNA) on survival in central primitive neuroectodermal tumors (CNS-PNET) and tumors of the pineal region. Searches of MEDLINE, Pubmed, and EMBASE--after the original description of comparative genomic hybridization in 1992 and July 2010--identified 15 case series of patients with CNS-PNET and tumors of the pineal region whose tumors were investigated for genome-wide CNA. One additional case study was identified from contact with experts. Individual patient data were extracted from publications or obtained from investigators, and CNAs were converted to a digitized format suitable for data mining and subgroup identification. Summary profiles for genomic imbalances were generated from case-specific data. Overall survival (OS) was estimated using the Kaplan-Meier method, and by univariable and multivariable Cox regression models. In their overall CNA profiles, low grade tumors of the pineal region clearly diverged from CNS-PNET and pineoblastoma. At a median follow-up of 89 months, 7-year OS rates of CNS-PNET, pineoblastoma, and low grade tumors of the pineal region were 22.9 ± 6, 0 ± 0, and 87.5 ± 12 %, respectively. Multivariable analysis revealed that histology (CNS-PNET), age (≤2.5 years), and possibly recurrent CNAs were associated with unfavorable OS. DNA copy number profiling suggests a close relationship between CNS-PNET and pineoblastoma. Low grade tumors of the pineal region differed from CNS-PNET and pineoblastoma. Due to their high biological and clinical variability, a coordinated prospective validation in future studies is necessary to establish robust risk factors.

Provision of rapid and specific ex vivo diagnosis of central nervous system lymphoma from rodent xenograft biopsies by a fluorescent aptamer.

OBJECTIVE: Differentiating central nervous system (CNS) lymphoma from other intracranial malignancies remains a clinical challenge in surgical neuro-oncology. Advances in clinical fluorescence imaging contrast agents and devices may mitigate this challenge. Aptamers are a class of nanomolecules engineered to bind cellular targets with antibody-like specificity in a fraction of the staining time. Here, the authors determine if immediate ex vivo fluorescence imaging with a lymphoma-specific aptamer can rapidly and specifically diagnose xenografted orthotopic human CNS lymphoma at the time of biopsy. METHODS: The authors synthesized a fluorescent CNS lymphoma-specific aptamer by conjugating a lymphoma-specific aptamer with Alexa Fluor 488 (TD05-488). They modified human U251 glioma cells and Ramos lymphoma cells with a lentivirus for constitutive expression of red fluorescent protein and implanted them intracranially into athymic nude mice. Three to 4 weeks postimplantation, acute slices (biopsies, n = 28) from the xenografts were collected, placed in aptamer solution, and imaged with a Zeiss fluorescence microscope. Three aptamer staining concentrations (0.3, 1.0, and 3.0 µM) and three staining times (5, 10, and 20 minutes) followed by a 1-minute wash were tested. A file of randomly selected images was distributed to neurosurgeons and neuropathologists, and their ability to distinguish CNS lymphoma from negative controls was assessed. RESULTS: The three staining times and concentrations of TD05-488 were tested to determine the diagnostic accuracy of CNS lymphoma within a frozen section time frame. An 11-minute staining protocol with 1.0-µM TD05-488 was most efficient, labeling 77% of positive control lymphoma cells and less than 1% of negative control glioma cells (p < 0.001). This protocol permitted clinicians to positively identify all positive control lymphoma images without misdiagnosing negative control images from astrocytoma and normal brain. CONCLUSIONS: Ex vivo fluorescence imaging is an emerging technique for generating rapid histopathological diagnoses. Ex vivo imaging with a novel aptamer-based fluorescent nanomolecule could provide an intraoperative tumor-specific diagnosis of CNS lymphoma within 11 minutes of biopsy. Neurosurgeons and neuropathologists interpreted images generated with this molecular probe with high sensitivity and specificity. Clinical application of TD05-488 may permit specific intraoperative diagnosis of CNS lymphoma in a fraction of the time required for antibody staining.

Amplifying small amounts of tumor DNA allows detection of DNA copy number abberations with array-CGH.

Array comparative genomic hybridization (aCGH) is a powerful tool to detect relative DNA copy number at a resolution limited only by the coverage of bacterial artificial chromosomes (BACs) used to print the genomic array. The amount of DNA needed to perform a reliable aCGH analysis has been a limiting factor, especially on minute tissue samples where limited DNA is available. Here we report a simple, highly sensitive and reliable aCGH method to analyze samples of no more than 1 ng genomic DNA. The speed and simplicity of the technique are ideal for studies on small clinical samples such as needle biopsies.

Contribution of Notch signaling activation to human glioblastoma multiforme.

OBJECT: Because activation of Notch receptors has been suggested to be critical for Ras-mediated transformation, and because many gliomas exhibit deregulated Ras signaling, the authors measured Notch levels and activation in primary samples and cell lines derived from glioblastoma multiforme (GBM) as well as the contribution of Notch pathway activation to astrocytic transformation and growth. METHODS: Western blot analysis of Notch 1 expression and activation showed that Notch 1 protein was overexpressed and/or activated in Ras-transformed astrocytes, in three of four GBM cell lines, and in four of five primary GBM samples. Expansion of these studies to assess mRNA expression of components of the Notch signaling pathway by cDNA expression array showed that cDNAs encoding components of the Notch signaling pathway, including the Notch ligand Jagged-1, Notch 3, and the downstream targets of Notch (HES1 and HES2), were also overexpressed relative to non-neoplastic brain controls in 23, 71, and 51% of 35 primary GBMs, respectively. Furthermore, inhibition of Notch signaling by genetic or pharmacological means led to selective suppression of the growth and expression of markers of differentiation in cells exhibiting Notch pathway deregulation. CONCLUSIONS: Notch activation contributes to Ras-induced transformation of glial cells and to glioma growth, survival, or both and as such may represent a new target for GBM therapy.

Integrated array-comparative genomic hybridization and expression array profiles identify clinically relevant molecular subtypes of glioblastoma.

Glioblastoma, the most aggressive primary brain tumor in humans, exhibits a large degree of molecular heterogeneity. Understanding the molecular pathology of a tumor and its linkage to behavior is an important foundation for developing and evaluating approaches to clinical management. Here we integrate array-comparative genomic hybridization and array-based gene expression profiles to identify relationships between DNA copy number aberrations, gene expression alterations, and survival in 34 patients with glioblastoma. Unsupervised clustering on either profile resulted in similar groups of patients, and groups defined by either method were associated with survival. The high concordance between these separate molecular classifications suggested a strong association between alterations on the DNA and RNA levels. We therefore investigated relationships between DNA copy number and gene expression changes. Loss of chromosome 10, a predominant genetic change, was associated not only with changes in the expression of genes located on chromosome 10 but also with genome-wide differences in gene expression. We found that CHI3L1/YKL-40 was significantly associated with both chromosome 10 copy number loss and poorer survival. Immortalized human astrocytes stably transfected with CHI3L1/YKL-40 exhibited changes in gene expression similar to patterns observed in human tumors and conferred radioresistance and increased invasion in vitro. Taken together, the results indicate that integrating DNA and mRNA-based tumor profiles offers the potential for a clinically relevant classification more robust than either method alone and provides a basis for identifying genes important in glioma pathogenesis.

Isochromosome 17q is a negative prognostic factor in poor-risk childhood medulloblastoma patients.

BACKGROUND: Medulloblastomas are the most common primary malignant childhood intracranial neoplasms. Patients are currently sorted into three risk groups based on clinical criteria: standard, poor, and infant (<18 months old). We hypothesized that genetic copy number aberrations (CNA) predict prognosis and would provide improved criteria for predicting outcome. METHODS: DNA from 35 medulloblastoma patients from four Children's Cancer Group trials was analyzed by comparative genomic hybridization to determine CNAs. The genetic alterations were evaluated using statistical and cluster analyses. RESULTS: The most frequent CNAs were gains on 17q, 7, 1q, and 7q and losses on 17p, 10q, X, 16q, and 11q. Amplification at 5p15.1-p15.3 was also detected. Isochromosome 17q (i(17)(q10)) was associated with poor overall survival (P = 0.03) and event-free survival (P = 0.04) independent of poor risk group classification. Age <3 tended to be associated with <3 CNAs (P = 0.06). Unsupervised cluster analysis sorted the study patients into four subgroups based on CNAs. Supervised analysis using the program Significance Analysis of Microarrays (SAM) quantitatively validated those CNAs identified by unsupervised clustering that significantly distinguished among the four subgroups. CONCLUSIONS: Medulloblastomas are genetically heterogeneous and can be categorized into separate genetic subgroups by their CNAs using unsupervised cluster analysis and SAM. i(17)(q10) was a significant independent negative prognostic factor. Infant medulloblastomas may be a distinct genetic subset from those of older patients.

Array comparative genomic hybridization identifies genetic subgroups in grade 4 human astrocytoma.

Alterations of DNA copy number are believed to be important indicators of tumor progression in human astrocytoma. We used an array of bacterial artificial chromosomes to map relative DNA copy number in 50 primary glioblastoma multiforme tumors at approximately 1.4-Mb resolution. We identified 33 candidate sites for amplification and homozygous deletion in these tumors. We identified three major genetic subgroups within these glioblastoma multiforme tumors: tumors with chromosome 7 gain and chromosome 10 loss, tumors with only chromosome 10 loss in the absence of chromosome 7 gain, and tumors without copy number change in chromosomes 7 or 10. The significance of these genetic groups to therapeutics needs further study.

Genetic aberrations defined by comparative genomic hybridization distinguish long-term from typical survivors of glioblastoma.

Glioblastoma (GBM) remains a highly lethal neoplasm, refractory to current therapies. The molecular genetic aberrations most closely related to clinical aggressiveness in GBM have been difficult to identify, perhaps due in part to the short survival range observed in cohorts of GBM patients. To address this, we characterized 39 tumors from rare patients (2-5% of all GBM cases) who experienced long-term survival (>3 years) using comparative genomic hybridization as a genome-wide screen. We then compared the frequency and type of aberrations with those in tumors from 24 typical or short-term survivors [STSs (<1.5 years)]. Losses of 9p and 10 and simple gains of chromosome 7 showed at least trends toward increased frequency in the STS group. Additional aberrations, including loss of 6q and gains of 19q and 20q, were significantly more frequent in the STS group. The presence of 19q loss was exclusive to the long-term survivor (LTS) group. Multivariate analyses indicated that 6q loss, 10q loss, and 19q gain were associated with short-term survival (all P < 0.01). The combination of any two of these three aberrations was seen in 16 of 24 STSs but only 1 of 39 LTSs. This comparison of rare LTSs with STSs (typical GBM survivors) identified 6q loss, 10q loss, and 19q gain, particularly when two or more of these were present, as most closely associated with aggressive clinical behavior in GBM. Loss of 19q may be a marker of long-term survival.

Handheld confocal laser endomicroscopic imaging utilizing tumor-specific fluorescent labeling to identify experimental glioma cells in vivo.

BACKGROUND: We have reported that handheld confocal laser endomicroscopy (CLE) can be used with various nonspecific fluorescent dyes to improve the microscopic identification of brain tumor and its boundaries. Here, we show that CLE can be used experimentally with tumor-specific fluorescent labeling to define glioma margins in vivo. METHODS: Thirteen rats underwent craniectomy and in vivo imaging 21 days after implantation with green fluorescent protein (GFP)-labeled U251 (n = 7) cells or epidermal growth factor receptor (EGFR) overexpressing F98 cells (n = 6). Fluorescein isothiocyanate (FITC) conjugated EGFR fluorescent antibody (FITC-EGFR) was applied for contrast in F98 tumors. Confocal images of normal brain, obvious tumor, and peritumoral zones were collected using the CLE system. Bench-top confocal microscopy and hematoxylin and eosin-stained sections were correlated with CLE images. RESULTS: GFP and FITC-EGFR fluorescence of glioma cells were detected by in vivo visible-wavelength fluorescence CLE. CLE of GFP-labeled tumors revealed bright individual satellite tumor cells within peritumoral tissue, a definitive tumor border, and subcellular structures. Imaging with FITC-EGFR labeling provided weaker contrast in F98-EGFR tumors but was able to delineate tumor cells. Imaging with both methods in various tumor regions correlated with standard confocal imaging and clinical histology. CONCLUSIONS: These data suggest that in vivo CLE of selectively tagged neoplasms could allow specific interactive identification of tumoral areas. Imaging of GFP and FITC-EGFR provides real-time histologic information precisely related to the site of microscopic imaging of tumor.

Molecular cytogenetic analysis of chromosomes 1 and 19 in glioma cell lines.

Deletions of chromosome 1p and 19q arms are frequent genetic abnormalities in primary human gliomas and are especially common in oligodendrogliomas. However, the chromosome 1p and 19q status of many glioma cell lines has not been established. Using homozygosity mapping, fluorescence in situ hybridization (FISH), and comparative genomic hybridization to arrayed BAC (CGHa), we screened 17 glioma cell lines for chromosome 1 and 19 deletions. Sequence tagged site polymorphisms were used to evaluate the cell lines for regions of chromosome 1p and 19q homozygosity. Cell lines A172, U251, TP265, U118, SW1088, U87, SW1783, and D32 contained significant regions of 19q homozygosity. In addition, A172, U87, TP483, D37, U118, MO67, and TP265 contained significant regions of 1p homozygosity. FISH probes localized to 1p36.32 and 19q13.33 as well as CGHa were used to determine which cell lines had deletions of 1p and/or 19q. Cell lines A172, U87, TP483, TP265, H4, U251, and D37 were deleted for portions of 1p. CGHa and homozygosity mapping of these cell lines define a 700-kilobase (Kb) common deletion region that is encompassed by a larger deletion region previously mapped in sporadic gliomas. This common deletion region is localized at 1p36.31 and includes CHD5, a putative tumor suppressor gene. Cell line A172 was observed to have a deletion between 19q13.33 and 19q13.41, while U87 was observed to have a smaller deletion of 19q13.33. Cell lines A172 and U87 contain 1p and 19q deletions similar to those found in sporadic gliomas and will be useful cellular reagents for evaluating the function of putative 1p and 19q glioma tumor suppressor genes.

Use of a conformational switching aptamer for rapid and specific ex vivo identification of central nervous system lymphoma in a xenograft model.

Improved tools for providing specific intraoperative diagnoses could improve patient care. In neurosurgery, intraoperatively differentiating non-operative lesions such as CNS B-cell lymphoma from operative lesions can be challenging, often necessitating immunohistochemical (IHC) procedures which require up to 24-48 hours. Here, we evaluate the feasibility of generating rapid ex vivo specific labeling using a novel lymphoma-specific fluorescent switchable aptamer. Our B-cell lymphoma-specific switchable aptamer produced only low-level fluorescence in its unbound conformation and generated an 8-fold increase in fluorescence once bound to its target on CD20-positive lymphoma cells. The aptamer demonstrated strong binding to B-cell lymphoma cells within 15 minutes of incubation as observed by flow cytometry. We applied the switchable aptamer to ex vivo xenograft tissue harboring B-cell lymphoma and astrocytoma, and within one hour specific visual identification of lymphoma was routinely possible. In this proof-of-concept study in human cell culture and orthotopic xenografts, we conclude that a fluorescent switchable aptamer can provide rapid and specific labeling of B-cell lymphoma, and that developing aptamer-based labeling approaches could simplify tissue staining and drastically reduce time to histopathological diagnoses compared with IHC-based methods. We propose that switchable aptamers could enhance expeditious, accurate intraoperative decision-making.

A complex rearrangement of chromosome 7 in human astrocytoma.

Chromosome 7 is a frequent site of cytogenetic aberrations in human astrocytomas. One region that is often targeted in human astrocytomas is on 7p. The U251 human glioblastoma cell line has a region of gain of genetic material on 7p similar to that seen in human astrocytomas. We used several cytogenetic techniques to study chromosome 7 in U251 cells and identified a complex rearrangement that accounts for gain of chromosome 7 genetic material in the cell line. The characteristic rearrangement suggests a mechanism leading to 7p gain in primary grade IV astrocytomas.

Grade II astrocytomas are subgrouped by chromosome aberrations.

Grade II astrocytoma is defined as a low-grade tumor, yet patients have a wide range of survival and tumors can quickly progress to high-grade astrocytoma/glioblastoma. Previous studies using comparative genomic hybridization (CGH) failed to demonstrate frequent copy number aberrations (CNA) in these tumors. This may be related to technical difficulties because infiltrating astrocytic tumors are often intermixed with normal brain tissue. We developed methods to exclude most normal tissue and use small amounts of DNA for CGH by microdissecting small regions of tumor from paraffin sections and amplifying extracted DNA using degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR). Using this method, we examined 30 grade II astrocytoma cases. We found CNA in 25 cases (83%), with a mean of two CNA per case. The most frequent CNA were gains on 7q (12 cases), 5p (5 cases), 9 (5 cases), and 19p (3 cases), and losses on 19q (7 cases), 1p (6 cases), and Xp (3 cases). Gain on 7q and losses on 1p/19q were mutually exclusive. This is the first report on the genetic characterization of low-grade astrocytomas using CGH from microdissected and formalin-fixed tissue. The comparatively large number of cases in this study allows us to suggest that these tumors are genetically subgrouped.

AKT pathway genes define 5 prognostic subgroups in glioblastoma.

Activity of GFR/PI3K/AKT pathway inhibitors in glioblastoma clinical trials has not been robust. We hypothesized variations in the pathway between tumors contribute to poor response. We clustered GBM based on AKT pathway genes and discovered new subtypes then characterized their clinical and molecular features. There are at least 5 GBM AKT subtypes having distinct DNA copy number alterations, enrichment in oncogenes and tumor suppressor genes and patterns of expression for PI3K/AKT/mTOR signaling components. Gene Ontology terms indicate a different cell of origin or dominant phenotype for each subgroup. Evidence suggests one subtype is very sensitive to BCNU or CCNU (median survival 5.8 vs. 1.5 years; BCNU/CCNU vs other treatments; respectively). AKT subtyping advances previous approaches by revealing additional subgroups with unique clinical and molecular features. Evidence indicates it is a predictive marker for response to BCNU or CCNU and PI3K/AKT/mTOR pathway inhibitors. We anticipate Akt subtyping may help stratify patients for clinical trials and augment discovery of class-specific therapeutic targets.

Molecular interactions of ErbB1 (EGFR) and integrin-ß1 in astrocytoma frozen sections predict clinical outcome and correlate with Akt-mediated in vitro radioresistance.

INTRODUCTION: Treatment of astrocytoma is frequently hampered by radioresistance of the tumor. In addition to overexpression of ErbB1/EGFR, functional crosstalk between receptor tyrosine kinases and cell adhesion molecules may also contribute to therapy resistance. METHODS: Acceptor photobleaching FRET was implemented on frozen sections of clinical astrocytoma to check the role of ErbB1-integrin-ß1 interaction. U251 glioma subclones were obtained by introducing extra CHR7 material or the ErbB1 gene to test the relevance and mechanism of this interaction in vitro. RESULTS: Grade IV tumors showed higher ErbB1 and integrin-ß1 expression and greater ErbB1-integrin-ß1 heteroassociation than did grade II tumors. Of these, the extent of molecular association was a single determinant of tumor grade and prognosis in stepwise logistic regression. In vitro, integrin-ß1 was upregulated, and radiosensitivity was diminished by ectopic ErbB1 expression. Great excess of ErbB1 provided colony forming advantage over medium excess but did not yield better radiation resistance or faster proliferation and decreased to medium level over time, whereas integrin-ß1 levels remained elevated and defined the extent of radioresistance. Increased expression of ErbB1 and integrin-ß1 was paralleled by decreasing ErbB1 homoassociation and increasing ErbB1-integrin-ß1 heteroassociation. Microscopic two-sided FRET revealed that pixels with higher ErbB1-integrin-ß1 heteroassociation exhibited lowed ErbB1 homoassociation, indicating competition for association partners among these molecules. Boosted Akt phosphorylation response to EGF accompanied this shift toward heteroassociation, and the consequentially increased radioresistance could be reverted by inhibiting PI3K. CONCLUSION: The clinically relevant ErbB1-integrin-ß1 heteroassociation may be used as a target of both predictive diagnostics and molecular therapy.

Intraoperative fluorescent imaging of intracranial tumors: a review.

A review of fluorescent imaging for intracranial neoplasms is presented. Complete resection of brain cancer is seldom possible because of the goal to preserve brain tissue and the inability to visualize individual infiltrative tumor cells. Verification of histology and identification of tumor invasion in macroscopically normal-appearing brain tissue determine prognosis after resection of malignant gliomas. Therefore, imaging modalities aim to facilitate intraoperative decision-making. Intraoperative fluorescent imaging techniques have the potential to enable precise histopathologic diagnosis and to detect tumor remnants in the operative field. Macroscopic fluorescence imaging is effective for gross tumor detection. Microscopic imaging techniques enhance the sensitivity of the macroscopic observations and provide real-time histological information. Further development of clinical grade fluorescent agents specifically targeting tumor cells could improve the diagnostic and prognostic yield of intraoperative imaging.

Reevaluating the imaging definition of tumor progression: perfusion MRI quantifies recurrent glioblastoma tumor fraction, pseudoprogression, and radiation necrosis to predict survival.

INTRODUCTION: Contrast-enhanced MRI (CE-MRI) represents the current mainstay for monitoring treatment response in glioblastoma multiforme (GBM), based on the premise that enlarging lesions reflect increasing tumor burden, treatment failure, and poor prognosis. Unfortunately, irradiating such tumors can induce changes in CE-MRI that mimic tumor recurrence, so called post treatment radiation effect (PTRE), and in fact, both PTRE and tumor re-growth can occur together. Because PTRE represents treatment success, the relative histologic fraction of tumor growth versus PTRE affects survival. Studies suggest that Perfusion MRI (pMRI)-based measures of relative cerebral blood volume (rCBV) can noninvasively estimate histologic tumor fraction to predict clinical outcome. There are several proposed pMRI-based analytic methods, although none have been correlated with overall survival (OS). This study compares how well histologic tumor fraction and OS correlate with several pMRI-based metrics. METHODS: We recruited previously treated patients with GBM undergoing surgical re-resection for suspected tumor recurrence and calculated preoperative pMRI-based metrics within CE-MRI enhancing lesions: rCBV mean, mode, maximum, width, and a new thresholding metric called pMRI-fractional tumor burden (pMRI-FTB). We correlated all pMRI-based metrics with histologic tumor fraction and OS. RESULTS: Among 25 recurrent patients with GBM, histologic tumor fraction correlated most strongly with pMRI-FTB (r = 0.82; P < .0001), which was the only imaging metric that correlated with OS (P<.02). CONCLUSION: The pMRI-FTB metric reliably estimates histologic tumor fraction (i.e., tumor burden) and correlates with OS in the context of recurrent GBM. This technique may offer a promising biomarker of tumor progression and clinical outcome for future clinical trials.