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Tissue next-generation sequencing gene coverage (152 genes)

Plasma cell-free DNA (cfDNA) concentration (ng/mL) is correlated with (A), total radiographic tumor burden (contrast-enhancing tumor + T2/FLAIR non-enhancing tumor) and (B), contrast-enhancing tumor burden at the first post-radiation MRI scan in patients with newly diagnosed glioblastoma more...

Purpose:The clinical utility of plasma cell-free DNA (cfDNA) has not been assessed prospectively in patients with glioblastoma (GBM). We aimed to determine the prognostic impact of plasma cfDNA in GBM, as well as its role as a surrogate of tumor burden and substrate for next-generation sequencing (NGS).Experimental Design:We conducted a prospective cohort study of 42 patients with newly diagnosed GBM. Plasma cfDNA was quantified at baseline prior to initial tumor resection and longitudinally during chemoradiotherapy. Plasma cfDNA was assessed for its association with progression-free survival (PFS) and overall survival (OS), correlated with radiographic tumor burden, and subjected to a targeted NGS panel.Results:Prior to initial surgery, GBM patients had higher plasma cfDNA concentration than age-matched healthy controls (mean 13.4 vs. 6.7 ng/mL, P < 0.001). Plasma cfDNA concentration was correlated with radiographic tumor burden on patients' first post-radiation magnetic resonance imaging scan (ρ = 0.77, P = 0.003) and tended to rise prior to or concurrently with radiographic tumor progression. Preoperative plasma cfDNA concentration above the mean (>13.4 ng/mL) was associated with inferior PFS (median 4.9 vs. 9.5 months, P = 0.038). Detection of ≥1 somatic mutation in plasma cfDNA occurred in 55% of patients and was associated with nonstatistically significant decreases in PFS (median 6.0 vs. 8.7 months, P = 0.093) and OS (median 5.5 vs. 9.2 months, P = 0.053).Conclusions:Plasma cfDNA may be an effective prognostic tool and surrogate of tumor burden in newly diagnosed GBM. Detection of somatic alterations in plasma is feasible when samples are obtained prior to initial surgical resection. more...

BACKGROUND Tumor-associated macrophages (TAMs) are a key component of glioblastoma (GBM) tumor microenvironment. Considering the differential role of different TAM phenotypes in iron metabolism with the M1 phenotype storing intracellular iron, and M2 phenotype releasing iron in the tumor microenvironment, here we investigated non-invasive quantitative susceptibility mapping (QSM) and T2* MRI relaxometry to quantify iron as imaging biomarkers for TAMs in adult patients with GBM. METHODS In this prospective study, 21 adult patients with GBM were enrolled between 2016 to 2019. Patients underwent a 3D single echo gradient echo sequence in addition to standard anatomical sequences on a 3 Tesla MRI. QSM images were reconstructed using the morphology-enabled dipole inversion (MEDI) algorithm. In 3 subjects, ex vivo imaging of surgical specimens was performed on a Bruker 9.4 Tesla 8.9 cm vertical bore MR using 3D multi-echo GRE scans, and R2* (1/T2*) maps were generated by a pixel-wise monoexponential fitting. Each specimen was stained with hematoxylin and eosin, as well as with CD68, CD86, CD206, and L-Ferritin. RESULTS Significant positive correlation was observed between mean susceptibility for the tumor enhancing zone and the L-ferritin positivity percent (r =0.56, p=0.018) and the combination of tumor’s enhancing zone and necrotic core and the L-Ferritin positivity percent (r=0.72; p=0.001). Moreover, the mean susceptibility significantly correlated with positivity percent for CD68 (ρ=0.52, p=0.034) and CD86 (r=0.7 p=0.001), but not for CD206 (ρ=0.09; p=0.7). There was a positive correlation between mean R2* values and CD68 positive cell counts (r =0.6, p=0.016). Similarly, mean R2* values significantly correlated with CD86 (r=0.54, p=0.03) but not with CD206 (r=0.15, p=0.5). High mean susceptibility in the necrotic core was associated with inferior PFS (hazard ratio,4.5, p=0.016). CONCLUSION MR Susceptibility Imaging can quantify the iron content of GBM and provide a non-invasive method for TAM quantification and phenotyping. more...

Purpose Tumor-associated macrophages (TAMs) are a key component of glioblastoma (GBM) microenvironment. Considering the differential role of different TAM phenotypes in iron metabolism with the M1 phenotype storing intracellular iron, and M2 phenotype releasing iron in the tumor microenvironment, we investigated MRI to quantify iron as an imaging biomarker for TAMs in GBM patients. Methods 21 adult patients with GBM underwent a 3D single echo gradient echo MRI sequence and quantitative susceptibility maps were generated. In 3 subjects, ex vivo imaging of surgical specimens was performed on a 9.4 Tesla MRI using 3D multi-echo GRE scans, and R2* (1/T2*) maps were generated. Each specimen was stained with hematoxylin and eosin, as well as CD68, CD86, CD206, and L-Ferritin. Results Significant positive correlation was observed between mean susceptibility for the tumor enhancing zone and the L-ferritin positivity percent (r =0.56, p=0.018) and the combination of tumor’s enhancing zone and necrotic core and the L-Ferritin positivity percent (r=0.72; p=0.001). The mean susceptibility significantly correlated with positivity percent for CD68 (ρ = 0.52, p=0.034) and CD86 (r=0.7 p=0.001), but not for CD206 (ρ = 0.09; p=0.7). There was a positive correlation between mean R2* values and CD68 positive cell counts (r =0.6, p=0.016). Similarly, mean R2* values significantly correlated with CD86 (r=0.54, p=0.03) but not with CD206 (r=0.15, p=0.5). Conclusion MR quantitative susceptibility mapping can quantify the iron content of GBM and provide a non-invasive method for TAM quantification and phenotyping. more...

Glioblastoma (GBM) has high metabolic demands, which can lead to acidification of the tumor microenvironment. We hypothesize that a machine learning model built on temporal principal component analysis (PCA) of dynamic susceptibility contrast-enhanced (DSC) perfusion MRI can be used to estimate tumor acidity in GBM, as estimated by pH-sensitive amine chemical exchange saturation transfer echo-planar imaging (CEST-EPI). We analyzed 78 MRI scans in 32 treatment naïve and post-treatment GBM patients. All patients were imaged with DSC-MRI, and pH-weighting that was quantified from CEST-EPI estimation of the magnetization transfer ratio asymmetry (MTRasym) at 3 ppm. Enhancing tumor (ET), non-enhancing core (NC), and peritumoral T2 hyperintensity (namely, edema, ED) were used to extract principal components (PCs) and to build support vector machines regression (SVR) models to predict MTRasym values using PCs. Our predicted map correlated with MTRasym values with Spearman’s r equal to 0.66, 0.47, 0.67, 0.71, in NC, ET, ED, and overall, respectively (p more...

Glioblastoma (GBM) has high metabolic demands, which can lead to acidification of the tumor microenvironment. We hypothesize that a machine learning model built on temporal principal component analysis (PCA) of dynamic susceptibility contrast-enhanced (DSC) perfusion MRI can be used to estimate tumor acidity in GBM, as estimated by pH-sensitive amine chemical exchange saturation transfer echo-planar imaging (CEST-EPI). We analyzed 78 MRI scans in 32 treatment naïve and post-treatment GBM patients. All patients were imaged with DSC-MRI, and pH-weighting that was quantified from CEST-EPI estimation of the magnetization transfer ratio asymmetry (MTRasym) at 3 ppm. Enhancing tumor (ET), non-enhancing core (NC), and peritumoral T2 hyperintensity (namely, edema, ED) were used to extract principal components (PCs) and to build support vector machines regression (SVR) models to predict MTRasym values using PCs. Our predicted map correlated with MTRasym values with Spearman’s r equal to 0.66, 0.47, 0.67, 0.71, in NC, ET, ED, and overall, respectively (p). The results of this study demonstrates that PCA analysis of DSC imaging data can provide information about tumor pH in GBM patients, with the strongest association within the peritumoral regions. more...

OBJECTIVE Spasmodic dysphonia (SD) is a neurological disorder characterized by involuntary spasms in the laryngeal muscles. It is thought to selectively affect speaking; other vocal behaviors remain intact. However, the patients' own perspective on their symptoms is largely missing, leading to partial understanding of the full spectrum of voice alterations in SD. METHODS A cohort of 178 SD patients rated their symptoms on the visual analog scale based on the level of effort required for speaking, singing, shouting, whispering, crying, laughing, and yawning. Statistical differences between the effort for speaking and the effort for other vocal behaviors were assessed using nonparametric Wilcoxon rank-sum tests within the overall SD cohort as well as within different subgroups of SD. RESULTS Speech production was found to be the most impaired behavior, ranking as the most effortful type of voice production in all SD patients. In addition, singing required nearly similar effort as speaking, ranking as the second most altered vocal behavior. Shouting showed a range of variability in its alterations, being especially difficult to produce for patients with adductor form, co-occurring voice tremor, late onset of disorder, and familial history of dystonia. Other vocal behaviors, such as crying, laughing, whispering, and yawning, were within the normal ranges across all SD patients. CONCLUSION Our findings widen the symptomatology of SD, which has predominantly been focused on selective speech impairments. We suggest that a separation of SD symptoms is rooted in selective aberrations of the neural circuitry controlling learned but not innate vocal behaviors. LEVEL OF EVIDENCE 4 Laryngoscope, 129:1627-1633, 2019. more...

Background Plasma cell-free DNA (cfDNA) concentration is lower in glioblastoma (GBM) compared to other solid tumors, which can lead to low circulating tumor DNA (ctDNA) detection. In this study, we investigated the relationship between multimodality magnetic resonance imaging (MRI) and histopathologic features with plasma cfDNA concentration and ctDNA detection in patients with treatment-naive GBM. Methods We analyzed plasma cfDNA concentration, MRI scans, and tumor histopathology from 42 adult patients with newly diagnosed GBM. Linear regression analysis was used to examine the relationship of plasma cfDNA concentration before surgery to imaging and histopathologic characteristics. In a subset of patients, imaging and histopathologic metrics were also compared between patients with and without a detected tumor somatic mutation. Results Tumor volume with elevated (>1.5 times contralateral white matter) rate transfer constant (Kep, a surrogate of blood–brain barrier [BBB] permeability) was independently associated with plasma cfDNA concentration (P = .001). Histopathologic characteristics independently associated with plasma cfDNA concentration included CD68+ macrophage density (P = .01) and size of tumor vessels (P = .01). Patients with higher (grade ≥3) perivascular CD68+ macrophage density had lower volume transfer constant (Ktrans, P = .01) compared to those with lower perivascular CD68+ macrophage density. Detection of at least 1 somatic mutation in plasma cfDNA was associated with significantly lower perivascular CD68+ macrophages (P = .01). Conclusions Metrics of BBB disruption and quantity and distribution of tumor-associated macrophages are associated with plasma cfDNA concentration and ctDNA detection in GBM patients. These findings represent an important step in understanding the factors that determine plasma cfDNA concentration and ctDNA detection. more...

BACKGROUND: Over the last quarter-century, the number of publications using vessel wall MR imaging has increased. Although many narrative reviews offer insight into technique and diagnostic applications, a systematic review of publication trends and reporting quality has not been conducted to identify unmet needs and future directions. PURPOSE: We aimed to identify which intracranial vasculopathies need more data and to highlight areas of strengths and weaknesses in reporting. DATA SOURCES: PubMed, EMBASE, and MEDLINE databases were searched up to September 2018 in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. DATA ANALYSIS: Two independent reviewers screened and extracted data from 128 articles. The Strengthening the Reporting of Observational Studies in Epidemiology guidelines were used to assess the reporting quality of analytic observational studies. DATA SYNTHESIS: There has been an exponentially increasing trend in the number of vessel wall MR imaging publications during the past 24 years (P more...

BACKGROUND: Quantitative susceptibility mapping (QSM) uses prior information to reconstruct maps, but prior information may not show pathology and introduce inconsistencies with susceptibility maps, degrade image quality and inadvertently smoothing image features. PURPOSE: To develop a local field data-driven QSM reconstruction that does not depend on spatial edge prior information. STUDY TYPE: Retrospective. SUBJECTS, ANIMAL MODELS: A dataset from 2016 ISMRM QSM Challenge, 11 patients with glioblastoma, a patient with microbleeds and porcine heart. SEQUENCE/FIELD STRENGTH: 3D gradient echo sequence on 3T and 7T scanners. ASSESSMENT: Accuracy was compared to Calculation of Susceptibility through Multiple Orientation Sampling (COSMOS), and several published techniques using region of interest (ROI) measurements, root-mean-squared error (RMSE), structural similarity index metric (SSIM), and high-frequency error norm (HFEN). Numerical ranking and semiquantitative image grading was performed by three expert observers to assess overall image quality (IQ) and image sharpness (IS). STATISTICAL TESTS: Bland–Altman, Friedman test, and Conover multiple comparisons. RESULTS: Loss adaptive dipole inversion (LADI) (β = 0.82, R(2) = 0.96), morphology-enabled dipole inversion (MEDI) (β = 0.91, R(2) = 0.97), and fast nonlinear susceptibility inversion (FANSI) (β = 0.81, R(2) = 0.98) had excellent correlation with COSMOS and no bias was detected (bias = 0.006 ± 0.014, P < 0.05). In glioblastoma patients, LADI showed consistently better performance (IQ(Grade) = 2.6 ± 0.4, IS(Grade) = 2.6 ± 0.3, IQ(Rank) = 3.5 ± 0.4, IS(Rank) = 3.9 ± 0.2) compared with MEDI (IQ(Grade) = 2.1 ± 0.3, IS(Grade) = 2 ± 0.5, IQ(Rank) = 2.4 ± 0.5, IS(Rank) = 2.8 ± 0.2) and FANSI (IQ(Grade) = 2.2 ± 0.5, IS(Grade) = 2 ± 0.4, IQ(Rank) = 2.8 ± 0.3, IS(Rank) = 2.1 ± 0.2). Dark artifact visible near the infarcted region in MEDI (Inf(MEDI) = −0.27 ± 0.06 ppm) was better mitigated by FANSI (Inf(FANSI-TGV) = −0.17 ± 0.05 ppm) and LADI (Inf(LADI) = −0.18 ± 0.05 ppm). CONCLUSION: For neuroimaging applications, LADI preserved image sharpness and fine features in glioblastoma and microbleed patients. LADI performed better at mitigating artifacts in cardiac QSM. EVIDENCE LEVEL: 4 TECHNICAL EFFICACY STAGE: 1 more...

Purpose: The clinical utility of plasma cell-free DNA (cfDNA) has not been assessed prospectively in patients with glioblastoma (GBM). We aimed to determine the prognostic impact of plasma cfDNA in GBM, as well as its role as a surrogate of tumor burden and substrate for next-generation sequencing (NGS). Experimental Design: We conducted a prospective cohort study of 42 patients with newly diagnosed GBM. Plasma cfDNA was quantified at baseline prior to initial tumor resection and longitudinally during chemoradiotherapy. Plasma cfDNA was assessed for its association with progression-free survival (PFS) and overall survival (OS), correlated with radiographic tumor burden, and subjected to a targeted NGS panel. Results: Prior to initial surgery, GBM patients had higher plasma cfDNA concentration than age-matched healthy controls (mean 13.4 vs. 6.7 ng/mL, P < 0.001). Plasma cfDNA concentration was correlated with radiographic tumor burden on patients' first post-radiation magnetic resonance imaging scan (ρ = 0.77, P = 0.003) and tended to rise prior to or concurrently with radiographic tumor progression. Preoperative plasma cfDNA concentration above the mean (>13.4 ng/mL) was associated with inferior PFS (median 4.9 vs. 9.5 months, P = 0.038). Detection of ≥1 somatic mutation in plasma cfDNA occurred in 55% of patients and was associated with nonstatistically significant decreases in PFS (median 6.0 vs. 8.7 months, P = 0.093) and OS (median 5.5 vs. 9.2 months, P = 0.053). Conclusions: Plasma cfDNA may be an effective prognostic tool and surrogate of tumor burden in newly diagnosed GBM. Detection of somatic alterations in plasma is feasible when samples are obtained prior to initial surgical resection. more...

Glioblastoma (GBM) is the most common primary malignant brain tumor in adults and carries a dismal prognosis. Significant challenges in the care of patients with GBM include marked vascular heterogeneity and arteriovenous (AV) shunting, which results in tumor hypoxia and inadequate delivery of systemic treatments to reach tumor cells. In this study, we investigated the utility of different MR perfusion techniques to detect and quantify arteriovenous (AV) shunting and tumor hypoxia in patients with GBM. Macrovascular shunting was present in 33% of subjects, with the degree of shunting ranging from (37–60%) using arterial spin labeling perfusion. Among the dynamic susceptibility contrast-enhanced perfusion curve features, there were a strong negative correlation between hypoxia score, DSC perfusion curve recovery slope (r = −0.72, P = 0.018) and angle (r = −0.73, P = 0.015). The results of this study support the possibility of using arterial spin labeling and pattern analysis of dynamic susceptibility contrast-enhanced MR Imaging for evaluation of arteriovenous shunting and tumor hypoxia in glioblastoma. more...

2508 Background: Due to significant interpatient heterogeneity, survival outcomes vary widely in patients with GBM. Novel prognostic biomarkers are needed. We aimed to determine the prognostic impact of baseline plasma cfDNA concentration in patients with GBM. Methods: We analyzed 84 patients with newly diagnosed GBM and at least 7 months of follow-up time. The first 41 patients comprised a previously published derivation cohort (Bagley, Clin Cancer Res 2020). The subsequent 43 patients served as an independent validation cohort. cfDNA was extracted from plasma collected prior to initial surgical resection and quantified by qPCR for a 115 bp amplicon of the human ALU repeat element. Receiver operating characteristic (ROC) curve analysis was used in the derivation cohort to (1) assess the accuracy of plasma cfDNA concentration for predicting progression-free survival status at 7 months (PFS-7), a landmark based on the median PFS for newly diagnosed GBM (Stupp, N Engl J Med 2005), and (2) derive the optimal cutoff for dichotomizing patients into high- and low-cfDNA groups. In the validation cohort, logistic regression was used to measure the association of plasma cfDNA concentration (high vs. low) with PFS-7, adjusted for age, isocitrate dehydrogenase ( IDH) 1/2 mutational status, 0-6-methylguanine-methyltransferase ( MGMT) methylation, extent of resection, and performance status. Multivariate Cox regression was used for overall survival (OS) analysis. Results: In the derivation cohort, the optimal cutoff for plasma cfDNA was 25.0 ng/mL (area under the curve [AUC] = 0.663), with inferior PFS and OS in patients with cfDNA above this cutoff (PFS, median 4.9 vs. 9.5 months, log-rank p = 0.001; OS, median 8.5 vs. 15.5 months, log-rank p = 0.03). In the validation cohort, baseline plasma cfDNA concentration over the cutoff was independently associated with a lower likelihood of being alive and progression-free at 7 months (adjusted OR 0.13, 95% CI 0.02 – 0.75, p = 0.02). OS was also worse in in the validation cohort in patients with high plasma cfDNA (adjusted HR 3.0, 95% CI 1.1 – 8.0, p = 0.03). Conclusions: In patients with newly diagnosed GBM, high baseline plasma cfDNA concentration is associated with worse survival outcomes independent of other prognostic factors. Further validation in a larger, multicenter study is warranted. more...

BACKGROUND Liquid biopsy has been not been widely utilized in patients with glioblastoma (GBM) compared to other solid tumors. However, the clinical utility of plasma cell-free DNA (cfDNA) in GBM has not been assessed prospectively or at the time of initial diagnosis. METHODS We conducted a prospective cohort study of patients with newly diagnosed GBM. Whole blood was collected in Streck® tubes at baseline prior to initial surgical resection and longitudinally during the course of adjuvant chemoradiotherapy. Plasma cfDNA concentration (ng/mL) was quantified by qPCR for a 115 bp amplicon of the human ALU repeat element, correlated with radiographic tumor burden by volumetry at multiple time points using Spearman rank correlation, and assessed for its impact on progression-free (PFS) and overall survival (OS) by Cox regression. RESULTS Prior to initial resection, GBM patients (N=42) had higher plasma cfDNA concentration compared to age-matched healthy controls (N=42) (mean 13.43 vs. 6.70 ng/mL, p< 0.001). Plasma cfDNA concentration was correlated with radiographic tumor burden on subjects’ first post-radiation MRI scan (r=0.77, p=0.003) and tended to rise prior to or concurrently with radiographic tumor progression. Pre-operative plasma cfDNA concentration above the mean (>13.4 ng/mL) was associated with inferior PFS (median 4.9 vs. 9.5 months, p=0.038) and OS (median 8.9 vs. 14.8 months, p=0.078). The impact on PFS persisted after adjusting for age, extent of resection, performance status, MGMT promoter methylation, and IDH1/2 mutational status (HR 2.48, 95% CI 1.1–6.1, p=0.046). CONCLUSIONS Plasma cfDNA may be an effective prognostic tool and noninvasive surrogate of tumor burden in newly diagnosed GBM. Tumor tissue samples from our cohort have been subjected to targeted next generation sequencing (NGS), and baseline plasma samples have been sent to Guardant Health for NGS. Plasma NGS results and concordance with matched tissue NGS will be included at time of presentation. more...