Faculty Bibliography

BACKGROUND:The goal of precision oncology is to find effective therapeutics for every patient. Through the inclusion of emerging therapeutics in a high-throughput drug screening platform, our functional genomics pipeline inverts the common paradigm to identify patient populations that are likely to benefit from novel therapeutic strategies. APPROACH/METHODS:Utilizing drug screening data across a panel of 46 cancer cell lines from 11 tumor lineages, we identified an ovarian cancer-specific sensitivity to the first-in-class CRL4 inhibitors KH-4-43 and 33-11. CRL4 (i.e., Cullin-4 RING E3 ubiquitin ligase) is known to be dysregulated in a variety of cancer contexts, making it an attractive therapeutic target. Unlike proteasome inhibitors that are associated with broad toxicity, CRL4 inhibition offers the potential for tumor-specific effects. RESULTS:We observed that CRL4 inhibition negatively regulates core gene signatures that are upregulated in ovarian tumors and significantly slowed tumor growth as compared to the standard of care, cisplatin, in OVCAR8 xenografts. Building on this, we performed combination drug screening in conjunction with proteomic and transcriptomic profiling to identify ways to improve the antitumor effects of CRL4 inhibition in ovarian cancer models. CRL4 inhibition consistently resulted in activation of the mitogen-activated protein kinase (MAPK) signaling cascade at both the transcriptomic and protein levels, suggesting that survival signaling is induced in response to CRL4 inhibition. These observations were concordant with the results of the combination drug screens in seven ovarian cancer cell lines that showed CRL4 inhibition cooperates with MEK inhibition. Preclinical studies in OVCAR8 and A2780 xenografts confirmed the therapeutic potential of the combination of KH-4-43 and trametinib, which extended overall survival and slowed tumor progression relative to either single agent or the standard of care. CONCLUSIONS:Together, these data demonstrate the prospective utility of functional modeling pipelines for therapeutic development and underscore the clinical potential of CRL4 inhibition in the ovarian cancer context. HIGHLIGHTS/CONCLUSIONS:A precision medicine pipeline identifies ovarian cancer sensitivity to CRL4 inhibitors. CRL4 inhibition induces activation of MAPK signalling as identified by RNA sequencing, proteomics, and phosphoproteomics. Inhibitor combinations that target both CRL4 and this CRL4 inhibitor-induced survival signalling enhance ovarian cancer sensitivity to treatment.

T cell receptor (TCR) gene therapy is a potent form of cellular immunotherapy in which patient T cells are genetically engineered to express TCRs with defined tumor reactivity. However, the isolation of therapeutic TCRs is complicated by both the general scarcity of tumor-specific T cells among patient T cell repertoires and the patient-specific nature of T cell epitopes expressed on tumors. Here we describe a high-throughput, personalized TCR discovery pipeline that enables the assembly of complex synthetic TCR libraries in a one-pot reaction, followed by pooled expression in reporter T cells and functional genetic screening against patient-derived tumor or antigen-presenting cells. We applied the method to screen thousands of tumor-infiltrating lymphocyte (TIL)-derived TCRs from multiple patients and identified dozens of CD4⁺ and CD8⁺ T-cell-derived TCRs with potent tumor reactivity, including TCRs that recognized patient-specific neoantigens.

BACKGROUND:Life-threatening, space-occupying mass effect due to cerebral edema and/or hemorrhagic transformation is an early complication of patients with middle cerebral artery stroke. Little is known about longitudinal trajectories of laboratory and vital signs leading up to radiographic and clinical deterioration related to this mass effect. METHODS:We curated a retrospective data set of 635 patients with large middle cerebral artery stroke totaling 95,463 data points for 10 longitudinal covariates and 40 time-independent covariates. We assessed trajectories of the 10 longitudinal variables during the 72 h preceding three outcomes representative of life-threatening mass effect: midline shift ≥ 5 mm, pineal gland shift (PGS) > 4 mm, and decompressive hemicraniectomy (DHC). We used a "backward-looking" trajectory approach. Patients were aligned based on outcome occurrence time and the trajectory of each variable was assessed before that outcome by accounting for cases and noncases, adjusting for confounders. We evaluated longitudinal trajectories with Cox proportional time-dependent regression. RESULTS:Of 635 patients, 49.0% were female, and the mean age was 69 years. Thirty five percent of patients had midline shift ≥ 5 mm, 24.3% of patients had PGS > 4 mm, and 10.7% of patients underwent DHC. Backward-looking trajectories showed mild increases in white blood cell count (10-11 K/UL within 72 h), temperature (up to half a degree within 24 h), and sodium levels (1-3 mEq/L within 24 h) before the three outcomes of interest. We also observed a decrease in heart rate (75-65 beats per minute) 24 h before DHC. We found a significant association between increased white blood cell count with PGS > 4 mm (hazard ratio 1.05, p value 0.007). CONCLUSIONS:Longitudinal profiling adjusted for confounders demonstrated that white blood cell count, temperature, and sodium levels appear to increase before radiographic and clinical indicators of space-occupying mass effect. These findings will inform the development of multivariable dynamic risk models to aid prediction of life-threatening, space-occupying mass effect.

The α-synuclein seed amplification assay (αSyn-SAA) sensitively detects Lewy pathology, the amyloid state of α-synuclein, in the cerebrospinal fluid (CSF) of patients with Parkinson's disease (PD). The αSyn-SAA harnesses the physics of seeding, whereby a superconcentrated solution of recombinant α-synuclein lowers the thermodynamic threshold (nucleation barrier) for aggregated α-synuclein to act as a nucleation catalyst ("seed") to trigger the precipitation (nucleation) of monomeric α-synuclein into pathology. This laboratory setup increases the signal for identifying a catalyst if one is present in the tissue examined. The result is binary: positive, meaning precipitation occurred, and a catalyst is present, or negative, meaning no precipitation, therefore no catalyst. Since protein precipitation via seeding can only occur at a concentration many-fold higher than the human brain, laboratory-elicited seeding does not mean human brain seeding. We suggest that a positive αSyn-SAA reveals the presence of pathological α-synuclein but not the underlying etiology for the precipitation of monomeric α-synuclein into its pathological form. Thus, a positive αSyn-SAA supports a clinical diagnosis of PD but cannot inform disease pathogenesis, ascertain severity, predict the rate of progression, define biology or biological subtypes, or monitor treatment response.

BACKGROUND:α-Synuclein seed amplification assay on cerebrospinal fluid (CSF-αSyn-SAA) has shown high accuracy for Parkinson's disease (PD) diagnosis. The analysis of CSF-αSyn-SAA parameters may provide useful insight to dissect the heterogeneity of synucleinopathies. OBJECTIVE:To assess differences in CSF-αSyn-SAA amplification parameters in participants with PD stratified by rapid eye movement (REM) sleep behavior disorder (RBD), dysautonomia, GBA, and LRRK2 variants. METHODS:(T50), time to threshold (TTT), slope, and area under the curve (AUC). Sporadic PD (n = 371) was stratified according to RBD and dysautonomia (DysA) symptoms. Genetic PD included carriers of pathogenic variants of GBA (GBA-PD, n = 52) and LRRK2 (LRRK2-PD, n = 124) gene. RESULTS:CSF-αSyn-SAA was positive in 77% of LRRK2-PD, 92.3% of GBA-PD, and 93.8% of sporadic PD. The LRRK2-PD cohort showed longer T50 and TTT, and smaller AUC than GBA-PD (P = 0.029, P = 0.029, P = 0.016, respectively) and sporadic PD (P = 0.034, P = 0.033, P = 0.014, respectively). In the sporadic cohort, CSF-αSyn-SAA parameters were similar between PD with (n = 157) and without (n = 190) RBD, whereas participants with DysA (n = 193) presented shorter T50 (P = 0.026) and larger AUC (P = 0.029) than those without (n = 150). CONCLUSION/CONCLUSIONS:CSF-αSyn-SAA parameters vary across genetic and non-genetic PD subtypes at the group level. These differences are mostly driven by the presence of LRRK2 variants and DysA. Significant overlaps in the amplification parameter values exist between groups and limit their use at the individual level. Further studies are necessary to understand the mechanisms of CSF-αSyn-SAA parameter differences. © 2024 International Parkinson and Movement Disorder Society.

Cerebral accumulation of alpha-synuclein (αSyn) aggregates is the hallmark event in a group of neurodegenerative diseases-collectively called synucleinopathies-which include Parkinson's disease, dementia with Lewy bodies and multiple system atrophy. Currently, these are diagnosed by their clinical symptoms and definitively confirmed postmortem by the presence of αSyn deposits in the brain. Here, we summarize the drawbacks of the current clinical definition of synucleinopathies and outline the rationale for moving toward an earlier, biology-anchored definition of these disorders, with or without the presence of clinical symptoms. We underscore the utility of the αSyn seed amplification assay to detect aggregated αSyn in living patients and to differentiate between neuronal or glial αSyn pathology. We anticipate that a biological definition of synucleinopathies, if well-integrated with the current clinical classifications, will enable further understanding of the disease pathogenesis and contribute to the development of effective, disease-modifying therapies.

The adoption of large language models (LLMs) in healthcare demands a careful analysis of their potential to spread false medical knowledge. Because LLMs ingest massive volumes of data from the open Internet during training, they are potentially exposed to unverified medical knowledge that may include deliberately planted misinformation. Here, we perform a threat assessment that simulates a data-poisoning attack against The Pile, a popular dataset used for LLM development. We find that replacement of just 0.001% of training tokens with medical misinformation results in harmful models more likely to propagate medical errors. Furthermore, we discover that corrupted models match the performance of their corruption-free counterparts on open-source benchmarks routinely used to evaluate medical LLMs. Using biomedical knowledge graphs to screen medical LLM outputs, we propose a harm mitigation strategy that captures 91.9% of harmful content (F1 = 85.7%). Our algorithm provides a unique method to validate stochastically generated LLM outputs against hard-coded relationships in knowledge graphs. In view of current calls for improved data provenance and transparent LLM development, we hope to raise awareness of emergent risks from LLMs trained indiscriminately on web-scraped data, particularly in healthcare where misinformation can potentially compromise patient safety.

BACKGROUND:Underlying intracranial stenosis is the most common cause of failed mechanical thrombectomy in acute ischemic stroke patients with large vessel occlusion. Adjunct emergent stenting is sometimes performed to improve or maintain reperfusion, despite limited data regarding its safety or efficacy. METHODS:We conducted a prospective multicenter observational international cohort study. Patients were enrolled between January 2022 and December 2023 at 25 thrombectomy capable centers in North America, Europe, and Asia. Consecutive patients treated with mechanical thrombectomy were included if they were identified as having underlying intracranial stenosis, defined as 50-99% residual stenosis of the target vessel or intra-procedural re-occlusion. The primary outcome was functional independence, defined as modified Rankin Scale of 0-2 at 90 days. After applying inverse probability of treatment weighting (IPTW) based on propensity scores, we compared outcomes among patients who underwent adjunct emergent intracranial stenting (stenting) versus those who received mechanical thrombectomy alone. RESULTS:A total of 417 patients were included; 218 patients treated with mechanical thrombectomy alone (168 anterior circulation) and 199 with mechanical thrombectomy plus stenting (144 anterior circulation). Patients in the stenting group were less likely to be non-Hispanic White (51.8% vs 62.4%, p=0.03), and less likely to have diabetes (33.2% vs 43.1%, p=0.037) or hyperlipidemia (43.2% vs 56%, p= 0.009). In addition, there was a lower rate of IV thrombolysis use in the stenting group (18.6% vs 27.5%, p=0.03). There was a higher rate of successful reperfusion (modified Treatment In Cerebral Infarction score ≥ 2B) in the stenting versus mechanical thrombectomy alone group (90.9% vs 77.9%, p<0.001) and a higher rate of a 24-hour infarct volume of <30 mL (n=260, 67.9% vs 50.3%, p=0.005). The overall complication rate was higher in the stenting group (12.6% vs 5%, p=0.006), but there was not a significant difference in the rate of symptomatic hemorrhage (9% vs 5.5%, p=0.162). Functional independence at 90 days was significantly higher in the stenting group (42.2% vs. 28.4%, adjusted odds ratio 2.67; 95% CI, 1.66-4.32). CONCLUSIONS:In patients with underlying stenosis who achieved reperfusion with mechanical thrombectomy, adjunct emergent stenting was associated with better functional outcome without a significantly increased risk of symptomatic hemorrhage. REGISTRATION/BACKGROUND:https://clinicaltrials.gov/study/NCT05403593.

OBJECTIVE:This study aimed to externally validate the Memory Assessment Clinics Scale for Epilepsy (MAC-E), a brief self-report measure of subjective memory complaints in adults with epilepsy. METHODS:A cross-sectional study was conducted including adults with focal pharmacoresistant epilepsy from three Level 4 epilepsy centers in the U.S., who completed the MAC-E as part of a clinical neuropsychological evaluation. Confirmatory factor analysis was conducted, and goodness-of-fit criteria were calculated to assess model fit: comparative fit index (CFI), root mean square error of approximation (RMSEA), and standardized root mean residual (SRMR). Item response theory models were constructed, and Mokken analysis was used to assess discrimination and unidimensionality. Internal consistency was evaluated with McDonald's Omega. RESULTS:values for each of the 5 factors (0.58-0.91 and 0.34-0.82, respectively). MAC-E items demonstrated high levels of discrimination as well as the ability to evaluate across the entirety of each latent trait. Score responses were uniformly distributed across latent traits, and unidimensionality was established by factor (all H coefficients > 0.4). Internal consistency was high across factors (omega range: 0.77-0.88). CONCLUSIONS:Results of this study demonstrate good external validation of the MAC-E in an independent, multicenter cohort of adults with epilepsy. These findings provide further support that the MAC-E is a psychometrically valid, self-report instrument to assess every-day memory abilities in adults with epilepsy in both clinical and research settings.

BACKGROUND/UNASSIGNED:In the DISCOMS (DISCOntinuation of disease-modifying therapies (DMTs) in multiple sclerosis (MS)) randomized clinical trial, we could not demonstrate that discontinuing MS DMTs in older, stable adults was not inferior to continuing DMTs. Relapses were rare in both groups, and most new disease activity was one to two new brain magnetic resonance imaging (MRI) lesions unassociated with clinical changes. OBJECTIVE/AIMS/UNASSIGNED:Describe results of the DISCOMS extension study. METHODS/UNASSIGNED:Among 10/19 of the original sites, we enrolled patients who completed DISCOMS; did not reach the primary endpoint during the original trial; and retained original randomized assignment. Participants completed one study visit and brain MRI at least 30 months after original enrollment in DISCOMS. Primary endpoint was time from entry into the primary study to relapse or new brain MRI activity. RESULTS/UNASSIGNED:= 0.043 from log-rank test). CONCLUSIONS/UNASSIGNED:From entry into DISCOMS extension study, time to new MS activity remained shorter in discontinuers, but relapses were absent and new brain MRI lesions were rare.