Citations

Abstract BACKGROUND Few methods exist via which to assess treatment response with high fidelity in gliomas. Cell-free DNA (cfDNA) is of growing interest as a liquid biomarker in plasma and CSF and has been used in other cancer types to optimize therapeutic regimens for individual patients. While CSF has previously been used as a source of cfDNA for gliomas, it remains unknown how cfDNA abundance and sequencing change throughout treatment and inevitable recurrence. To that end, we leveraged our growing biobank of longitudinal CSF specimens, acquired via CSF access devices, to determine the potential utility of CSF cfDNA as a monitoring tool for treatment response in patients with gliomas. MATERIAL AND METHODS Longitudinal CSF specimens were acquired via Ommaya reservoirs (NCT04692337) or ventriculoperitoneal shunts (NCT04692324) in patients with brain tumors, including gliomas. cfDNA was extracted from 1-5 mL of CSF based on available sample volumes and quantified via Qubit. Next-Generation sequencing (PredicineCARE) or low-pass whole genome sequencing (LPWGS) was performed by Predicine, Inc based on the amount of cfDNA available. RESULTS CSF acquired prior to versus after resection demonstrated an increase in quantified cfDNA (2.97x, range: 1.58-5.26x), consistent with the impact of increased parenchymal disruption and closer contact with CSF. Thereafter, CSF cfDNA continued to decrease throughout chemoradiation and adjuvant treatment and increased at recurrence, including in patients co-enrolled in immunotherapy trials for IL-7 agonists and pembrolizumab. Interestingly, copy number burden (CNB) increased by the time of recurrence in three patients with known disease progression, and continued to decrease throughout immunotherapy treatment in one patient despite concerns of pseudoprogression. In one patient with a hypermutated glioblastoma, CSF cfDNA revealed over 59 genomic alterations, including MAP2K1, KIT, and PDGFRA, and over 100 more variants of unknown significance, each of which had been detected in sequencing of the tissue acquired at resection and subsequently decreased with chemoradiation. In another patient with a known progressing EGFR-amplified GBM, over 200 new variants were identified by the patient’s final sample and EGFR copy number increased from 2 to over 30. CONCLUSION Analysis of cfDNA over a patient’s disease course, including resection and treatment with standard-of-care and experimental therapies, may be of use for disease monitoring and treatment response. Further work is needed to evaluate these findings in a larger cohort of patients and to determine the sensitivity of changes in mutations, CNB, and cfDNA quantity for glioma disease burden.

Abstract BACKGROUND The microenvironment of glioblastoma is complex and defined in part by protumorigenic metabolic changes. Within this microenvironment, blood-brain barrier disruption is often associated with the most aggressive portions of the tumor. To date, the metabolic tumor microenvironments of enhancing, blood-brain barrier (BBB) disrupted portions of gliomas, as compared to BBB-intact tumor, remain poorly understood. Specifically, it is unclear if the associated blood-brain barrier disruption merely reflects or functionally supports tumor aggressiveness. METHODS Intra-operative high-molecular weight microdialysis was utilized under an investigational device exemption (NCT NCT04047264) to sample the extracellular metabolome of radiographically diverse regions during fifteen neurosurgical resections. The global extracellular metabolome of recovered microdialysate from contrast-enhancing and non-enhancing tumor, as well as brain adjacent to tumor, was evaluated via ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) on the Metabolon platform and assessed using enrichment and correlation analyses. RESULTS Among 162 named metabolites identified via ultra-performance liquid chromatography tandem mass spectrometry, a novel glioma-associated metabolite, guanidinoacetate (GAA), was 126.32x higher in enhancing tumor than in adjacent brain and 13.63x higher in non-enhancing tumor than brain. Forty-eight additional metabolites were 2.05-10.18x more abundant in enhancing tumor than brain, including carnitine family members and all nine large neutral amino acids. Except for GAA and 2-hydroxyglutarate in IDH-mutant gliomas, differences between non-enhancing tumor and brain microdialysate were comparatively modest and less consistent, suggesting heterogeneity in the glioma metabolic microenvironment of blood-brain barrier (BBB) intact areas as compared to BBB-disrupted portions of gliomas. Interestingly, the enhancing but not the non-enhancing glioma metabolome was significantly enriched for plasma-associated metabolites, largely comprising amino acids and carnitines. CONCLUSION Our patient-derived data suggest that the BBB-disrupted metabolome of high-grade gliomas is characterized by metabolite diffusion from circulation into the tumor microenvironment. Further studies are ongoing to determine how this alteration in the glioma extracellular metabolome impacts glioma behavior and how it could be therapeutically targeted via BBB-modulating drugs.

Abstract BACKGROUND Liquid biopsy in glioblastoma (GBM) is hindered by a lack of requisite circulating-free DNA (cfDNA) levels in blood due to the blood-brain barrier (BBB). This results in challenges to the identification of blood-based biomarkers and the development of novel biomarker-driven systemic therapies. Real-time image-guided low intensity focused ultrasound (LIFU) combined with intravenously administered microbubble oscillators non-invasively causes BBB disruption (BBBD). This clinical trial aimed to evaluate the utility of LIFU for increasing cfDNA in blood for liquid biopsy in GBM. METHODS LIBERATE is a prospective, multi-center, self-controlled, ongoing, pivotal trial evaluating the safety and technical efficacy of LIFU for BBBD to increase cfDNA in blood for GBM. Patients aged 18-80 years with suspected GBM planned for tumor biopsy or resection at ten centers in the US are being included. Patients with multifocal tumors or tumors arising from deep midline, thalamus, cerebellum, or brainstem are excluded. Patients are administered intravenous oscillating microbubbles for enhancing sonication, after which MR-guided BBBD using a 220 kHz LIFU device is performed with real-time acoustic feedback for effective cavitation. Before and after the procedure, phlebotomies and MRI brain are performed to evaluate outcomes. The primary study endpoint is defined, per subject, as the ratio between their cfDNA level in blood 1-hour post-LIFU procedure compared to cfDNA level in blood pre-procedure. The primary study hypothesis is that BBBD with LIFU leads to a ≥2-fold increase in cfDNA in blood. The secondary hypothesis is that there exists ≥75% agreement between biomarker pattern in cfDNA sample from 1-hour post-LIFU sample and biomarker pattern in tumor tissue obtained later. The trial has been powered to evaluate both primary and secondary hypotheses. Based on an assumed true agreement rate of 91% and a one-sided alpha of 0.025, an exact test for binomial proportions provides a sample of N=50 with 84% power for the secondary hypothesis. Exploratory endpoints include (1) sensitivity of detection of known somatic mutations in cfDNA from blood samples collected before and after LIFU, (2) estimation of cfDNA levels post-LIFU in samples collected at 30 minutes, 1 hour, 2 hours, and 3 hours to determine the time of greatest yield, (3) correlation of MRI parameters related to grading of BBBD and biomarkers positive in cfDNA from post-LIFU blood samples. RESULTS Patient enrollment commenced in 2022, and 8 patients have been recruited by 04/11/2023. CONCLUSION Findings from this trial will be critical to liquid biopsy in GBM (Clinical trial information: NCT05383872).

Abstract Background Bevacizumab is commonly used to manage cerebral edema associated with brain tumors. However, its long half-life poses challenges for patients requiring urgent surgery due to wound complications. We present a case of utilizing therapeutic plasma exchange (TPE) to remove bevacizumab in a patient with recurrent glioblastoma requiring urgent surgery. Methods A 58-year-old male with recurrent glioblastoma, IDH-wildtype, presented with clinical and radiographic concern for ventriculitis requiring urgent wound washout only 4 days after his last bevacizumab infusion. TPE was performed for 3 sessions after surgery using a centrifugation-based cell separator. Replacement fluids included normal serum albumin, normal saline, and fresh frozen plasma. Bevacizumab levels were quantified using an enzyme-linked immunoabsorbent assay before and after each TPE session. Results TPE effectively removed bevacizumab, enabling safe surgery without new complications. Plasma bevacizumab levels decreased from 1087.63 to 145.35 ng/mL (13.4% of original) by the end of the last TPE session. This decline is consistent with nearly 3 half-lives, which compares favorably to the expected timeline of natural decline given the 21-day half-life. Conclusions We report a complex clinical scenario of a patient requiring urgent wound washout 4 days after last bevacizumab infusion for CNS infection. Surgery was successfully performed without new complications with use of TPE to remove bevacizumab immediately following surgery. This case highlights the feasibility of this approach, which may be utilized effectively in patients requiring surgery after having recently received bevacizumab.

Background: The prevalence of non-alcoholic fatty liver disease (NAFLD) in U.S. adults is over 30%, yet the role of lifestyle factors in the etiology of NAFLD remains understudied. We examined the associations of physical activity, by intensity and type, and television viewing with prevalent NAFLD. Methods: Cross-sectional analysis of a population-based sample of 2726 Black (49%) and White (51%) adults (Mean (SD) age, 50 (3.6) years; 57.3% female) from the CARDIA study. Exposures were aerobic activity by intensity (moderate, vigorous; hours/week); activity type (aerobic, muscle-strengthening; hours/week); and television viewing (hours/week), examined concurrently in all models and assessed by validated questionnaires. Our outcome was NAFLD (liver attenuation < 51 Hounsfield Units), measured by non-contrast computed tomography, after exclusions for other causes of liver fat. Covariates were sex, age, race, study center, education, diet quality, smoking status, alcohol consumption, and body mass index or waist circumference. Results: 648 participants had NAFLD. In the fully adjusted modified Poisson regression model, the risk ratios per interquartile range of each exposure were moderate-intensity aerobic activity, 1.10 (95% CI, 0.97–1.26); vigorous-intensity aerobic activity, 0.72 (0.63–0.82); muscle-strengthening activity, 0.89 (0.80–1.01); and television viewing, 1.20 (1.10–1.32). Relative to less active participants with higher levels of television viewing, those who participated in ≥2 h/week of both vigorous-intensity aerobic and muscle-strengthening activity and <7 h/week of television viewing had 65% lower risk of NAFLD (risk ratio = 0.35, 95% CI = 0.23–0.51). Conclusion: Adults who follow public health recommendations for vigorous-aerobic and muscle-strengthening activity, as well as minimize television viewing, are considerably less likely to have NAFLD than those who do not follow the recommendations and who have relatively high levels of television viewing.

Abstract Glioblastoma (GBM) is a lethal brain tumor with limited treatment options, due in part to the low blood-brain-barrier (BBB) penetrance of many anti-cancer compounds. Development of a non-invasive test that can measure drug delivery and target engagement in GBM represents an opportunity to enhance knowledge during early phase clinical trials of a new compound’s pharmacological suitability and potential efficacy. We present here data supporting that the cargo within extracellular vesicles (EV) can potentially serve as a liquid biopsy that reports on the pharmacodynamics of GBM therapeutics. We treated short-term cell cultures established from four patient-derived xenografts with vehicle, temozolomide, MLN4924, or arsenic trioxide. EVs were isolated from culture supernatants using size exclusion chromatography and then analyzed by RNA sequencing and mass spectrometry (proteomics) to characterize and compare cargo profiles for each condition. We found that depending on treatment, there were shared alterations in EV cargo between models in the expression of long non-coding RNAs, genes that are overrepresented in biological processes such as cytoplasmic translation and cellular macromolecule biosynthesis, and circular RNAs. Mass spectrometry of protein cargo further demonstrated both upregulated and downregulated peptide abundances with treatment compared to vehicle. Thus, our preliminary results indicate that EV cargo post-treatment may be a novel pharmacodynamic reporter in GBM. To allow for the selective interrogation of patient GBM-derived EVs, we have simultaneously pursued the identification of biomarkers to allow for immunoprecipitation of EVs from patient biofluids. Potential biomarkers, including PTPRZ1, B7H3, and IL13RA2, were screened across isolated EVs from both pre-clinical and clinical patient samples. Future efforts are focused on evaluating immunoprecipitated GBM-EVs from longitudinally collected patient biofluids. Overall, we anticipate that the results of this study will lead to development of a clinical test that reflects BBB penetration and tumor response, which will likely aid in novel drug development efforts.

Abstract WSD0922-Fu is an oral, central nervous system (CNS)-penetrant, small molecule, ATP non-competitive, reversible EGFR inhibitor which potently inhibits EGFR aberrations specific to non-small cell lung cancer (NSCLC) and high-grade astrocytoma (HGA). MC1914 (NCT04197934, sponsored by Wayshine Biopharma and FDA OOPD) is a first-in-human phase 1 dose-escalation trial investigating the safety and tolerability of WSD0922-Fu in patients with EGFR aberrant recurrent HGA and NSCLC. Adult patients with either recurrent EGFR mutant NSCLC with CNS metastases or recurrent EGFR mutant and/or EGFR amplified HGA were initially treated with oral WSD0922-FU, with doses escalated in cohorts based on a standard 3 + 3 design to determine the maximum tolerated dose (MTD). After establishing the MTD, this trial has now activated three dose expansion cohorts. Two of these cohorts evaluate different dose/schedule strategies in independent populations. The NSCLC cohort includes patients with EGFR mutant NSCLC and CNS metastases, and examines the safety, tolerability, plasma pharmacokinetics and preliminary efficacy of continuous WSD0922-Fu dosing (twice daily, seven days per week). The HGA cohort includes patients with EGFRvIII mutant HGA at first recurrence after prior radiation/temozolomide and examines the impact of higher doses of WSD0922-Fu administered intermittently (twice daily, five days per week). The HGA cohort also includes a food-effect study which evaluates the effect of the fed and fasted states on plasma PK. The third cohort (Brain tumor penetration cohort) includes patients with HGA (any EGFR mutation and/or EGFR amplification) who need a surgical resection for recurrent disease. Patients enrolled will be treated pre-operatively with WSD0922-Fu for five days, and tumor PK/PD will be examined in samples obtained from image-registered brain tumor biopsies. WSD0922-Fu therapy will be resumed after post-operative recovery. Patient enrollment to all three dose expansion cohorts is ongoing.

Abstract Rapid feedback is needed to understand the individualized biological impacts of novel glioma therapies. We are performing glioma biomarker discovery by serial cerebrospinal fluid (CSF) sampling from Ommaya reservoirs or lumbar punctures to determine how the CSF proteome can reveal early longitudinal intelligence regarding glioma status, biology, and therapeutic response. We aimed to leverage access to each patient’s glioma via CSF to identify candidate biomarkers of tumor burden, biology, and therapeutic challenge. Global proteomic CSF analysis was performed via Somalogic– an aptamer-based technology for highly sensitive and specific analysis of over 7,000 proteins. Discovery analysis comprised of the top 500 ranked proteins in the CSF from seven patients with high-grade gliomas versus non-glioma (normal pressure hydrocephalus) controls. The top HGG proteins were then filtered to include only proteins that met two additional criteria of 1) decrease with resection, and 2) increase with recurrence in individual paired patient samples. Additionally, in individual patients for whom serial CSF samples were available, protein fold change lists were created and ranked to determine the most or least differentially abundant proteins between various time points. Proteomic enrichment analysis revealed a conserved HGG CSF proteomic signature defined by 79 proteins, including ones known to be over-expressed in solid tumor malignancies, such as retinoblastoma binding protein 4, heat shock protein 90, and sorcin. The HGG proteomic signature was consistently enriched in an independent validation cohort consisting of 13 gliomas diverse in primary versus recurrent status, subtype, and grade, when compared to independent control CSF samples. Encouragingly, proteins in the HGG signature decreased in two patients following decreased tumor burden from surgical resection. Our preliminary data demonstrate the ability to gain detailed, individualized insights regarding glioma biology, tumor burden, and evolution through global CSF proteomics acquired from longitudinal neurosurgical access to each patient’s unique glioma.

Background and Aims: Fatty liver disease is a major public health threat due to its very high prevalence and related morbidity and mortality. Focused and dedicated interventions are urgently needed to target disease prevention, treatment, and care. Approach and Results: We developed an aligned, prioritized action agenda for the global fatty liver disease community of practice. Following a Delphi methodology over 2 rounds, a large panel (R1 n = 344, R2 n = 288) reviewed the action priorities using Qualtrics XM, indicating agreement using a 4-point Likert-scale and providing written feedback. Priorities were revised between rounds, and in R2, panelists also ranked the priorities within 6 domains: epidemiology, treatment and care, models of care, education and awareness, patient and community perspectives, and leadership and public health policy. The consensus fatty liver disease action agenda encompasses 29 priorities. In R2, the mean percentage of “agree” responses was 82.4%, with all individual priorities having at least a super-majority of agreement (> 66.7% “agree”). The highest-ranked action priorities included collaboration between liver specialists and primary care doctors on early diagnosis, action to address the needs of people living with multiple morbidities, and the incorporation of fatty liver disease into relevant non-communicable disease strategies and guidance. Conclusions: This consensus-driven multidisciplinary fatty liver disease action agenda developed by care providers, clinical researchers, and public health and policy experts provides a path to reduce the prevalence of fatty liver disease and improve health outcomes. To implement this agenda, concerted efforts will be needed at the global, regional, and national levels.

ABSTRACT Pancreatic cysts with high-risk characteristics are at increased risk of harboring high-grade dysplasia or pancreatic cancer. Endoscopic ultrasound may clarify the nature of the cystic lesion and its malignant potential. A mural nodule found through endoscopic ultrasound within a cyst may represent malignancy and require fine-needle aspiration. Pancreatic pseudocysts are benign walled-off fluid collections that form in the setting of pancreatitis and may be difficult to differentiate from neoplastic cysts. Pseudoaneurysms form when pancreatitis inflammation damages vessel walls and can cause fatal hemorrhage. We present a pancreatic pseudocyst with pseudoaneurysm mimicking a neoplastic cyst with a mural nodule.