Paraneoplastic neurological syndromes: clinical presentations and management.

We provide an overview of the varied presentations of paraneoplastic neurological syndromes. We also review the onconeural antibodies and their particular oncological and neurological associations. Recognition of these syndromes and their oncological associations is crucial, as early diagnosis and management has been associated with better patient outcomes. Specific management strategies and prognosis vary widely depending on the underlying etiology. An understanding of the relevant clinical details, imaging findings, and other diagnostic information can help tailor treatment approaches. We provide an outline of the diagnostic evaluation and treatment of various paraneoplastic neurological disorders, presenting with central and/or peripheral nervous system involvement. We briefly discuss neurologic immune checkpoint inhibitor-related adverse events, which can occasionally present with paraneoplastic neurological syndrome phenotypes.

Single Cell Mass Spectrometry Quantification of Anticancer Drugs: Proof of Concept in Cancer Patients.

In clinical cancer medicine, the current inability to quantify intracellular chemotherapy drug concentrations in individual human cells limits the personalization and overall effectiveness of drug administration. New bioanalytical methods capable of real-time measurement of drug levels in live single cancer cells would allow for more adaptive and personalized administration of chemotherapy drugs, potentially leading to better clinical outcomes with fewer side effects. In this study, we report the development of a new quantitative single cell mass spectrometry (qSCMS) method capable of providing absolute drug amounts and concentrations in single cancer cells. Using this qSCMS system, quantitative analysis of the intracellular drug gemcitabine present in individual bladder cancer cells is reported, including in bladder cancer cells isolated from patients undergoing standard-of-care gemcitabine chemotherapy. The development of single cell pharmacology bioanalytical methods can potentially lead to more effective and safely administered drug medications in patients, especially in the treatment of cancer.

Quantification of Drug Molecules in Live Single Cells Using the Single-Probe Mass Spectrometry Technique.

Analyzing cellular constituents on the single-cell level through mass spectrometry (MS) allows for a wide range of compounds to be studied simultaneously. However, there is a need for quantitative single-cell mass spectrometry (qSCMS) methods to fully characterize drug efficacy from individual cells within cell populations. In this study, qSCMS experiments were carried out using the Single-probe MS technique. The method was successfully used to perform rapid absolute quantifications of the anticancer drug irinotecan in individual mammalian cancer cells under ambient conditions in real time. Traditional liquid chromatography/mass spectrometry (LC/MS) quantifications of irinotecan in cell lysate samples were used to compare the results from Single-probe qSCMS. This technique showcases heterogeneity of drug efficacy on the single-cell level.

Transient Compound Treatment Induces a Multigenerational Reduction of Oxysterol-Binding Protein (OSBP) Levels and Prophylactic Antiviral Activity.

Oxysterol-binding protein (OSBP) is a lipid transport and regulatory protein required for the replication of Enterovirus genus viruses, which includes many significant human pathogens. Short-term exposure (i.e., 1-6 h) to a low dose (i.e., 1 nM) of the natural product compound OSW-1 induces a reduction of cellular OSBP levels by ∼90% in multiple different cell lines with no measurable cytotoxicity, defect in cellular proliferation, or global proteome reduction. Interestingly, the reduction of OSBP levels persists multiple days after the low-dose, transient OSW-1 compound treatment is ended and the intracellular OSW-1 compound levels drop to undetectable levels. The reduction in OSBP levels is inherited in multiple generations of cells that are propagated after the OSW-1 compound treatment is stopped. The enduring multiday, multigenerational reduction of OSBP levels triggered by the OSW-1 compound is not due to proteasome degradation of OSBP or due to a reduction in OSBP mRNA levels. OSW-1 compound treatment induces transient autophagy in cells, but blocking autophagy does not rescue OSBP levels. Although the specific cellular mechanism of long-term OSBP repression is not yet identified, these results clearly show the existence of an OSBP specific cellular regulation process that is triggered upon treatment with an OSBP-binding compound. The stable reduction of OSBP levels upon short-term, transient OSW-1 compound treatment will be a powerful tool to understand OSBP regulation and cellular function. Additionally, the persistent reduction in OSBP levels triggered by the transient OSW-1 compound treatment substantially reduces viral replication in treated cells. Therefore, the long-term, compound-induced reduction of OSBP in cells presents a new route to broad spectrum anti- Enterovirus activity, including as a novel route to antiviral prophylactic treatment through small molecule targeting a human host protein.

Predictors of neural-specific autoantibodies and immunotherapy response in patients with cognitive dysfunction.

Recognition of autoimmunity as a cause of encephalopathy has increased. Recent studies have validated the use of Antibody-Prevalence-in-Epilepsy (APE) and Responsive-to-immunotherapy-in-Epilepsy (RITE) scores in the evaluation and management of autoimmune-epilepsy. We aim to assess the utility of these models for patients with cognitive dysfunction. Among the evaluated patients, 17% had antibodies universally associated with autoimmune-encephalopathy. NMDA-R-IgG and LGI1-IgG were the most common antibody specificities. Antibody-Prevalence-in-Epilepsy-and-Encephalopathy (APE2) score ≥ 4 was 99% sensitive and 93% specific for neural-specific-antibodies. Responsive-to-immunotherapy-in-Epilepsy-and-Encephalopathy (RITE2) score ≥ 7 had 96% sensitivity and 86% specificity for favorable initial immunotherapy response. Application of these models may optimize autoantibody evaluations and immunotherapeutic trials.

Association of MOG-IgG Serostatus With Relapse After Acute Disseminated Encephalomyelitis and Proposed Diagnostic Criteria for MOG-IgG-Associated Disorders.

Importance: Recent studies have reported a higher relapse rate following an initial inflammatory demyelinating disorder in pediatric patients with persistent seropositivity of antibodies targeting myelin oligodendrocyte glycoprotein (MOG-IgG1). To date, the clinical implications of longitudinal MOG-IgG1 seropositivity using live cell assays with IgG1 secondary antibodies in adults after acute disseminated encephalomyelitis (ADEM) are unknown. Objective: To determine whether MOG-IgG1 serostatus (transient vs persistent) and titer change over time provide clinical utility in predicting the likelihood of relapse after ADEM. Design, Setting, and Participants: This cohort study identified patients with an initial diagnosis of ADEM evaluated at a single referral center between January 1, 1990, and October 1, 2017. Fifty-one patients were included, including 31 children and 20 adults. Longitudinal serologic testing was performed detecting autoantibodies targeting aquaporin 4 (AQP4-IgG) and MOG-IgG1 with clinically validated fluorescence-activated cell sorting assays. Patients were divided into 3 cohorts: persistent seropositivity, transient seropositivity, and seronegativity. Main Outcomes and Measures: Clinical demographic characteristics, longitudinal AQP4-IgG and MOG-IgG1 serostatus, titers, relapses, use of immunotherapy, and Expanded Disability Status Scale score at follow-up. Results: Of 51 patients presenting with an initial diagnosis of ADEM, 20 (39%) were adult, 24 (47%) were female, and ages ranged from 12 months to 57 years. Seventeen patients fulfilled criteria for persistent seropositivity; of those, 8 of 9 children (89%) and 7 of 8 adults (88%) had at least 1 relapse after median (range) follow-up periods of 75 (15-236) months and 39 (9-161) months, respectively. Eight patients (16%), including 4 adults, fulfilled criteria for transient seropositivity; of those, no children and 1 of 4 adults (25%) relapsed after median (range) follow-up periods of 32 (24-114) months and 16 (13-27) months, respectively. Of 24 patients with AQP4-IgG and MOG-IgG seronegativity, 6 of 17 children (35%) and 2 of 7 adults (29%) had at least 1 relapse after median (range) follow-up periods of 36 (3-203) months and 34 (15-217) months, respectively. There were only 2 patients, including 1 adult, with AQP4-IgG seropositivity, and both relapsed. The hazard ratio for relapses in those with persistent MOG-IgG1 positivity compared with AQP4-IgG and MOG-IgG1 seronegativity was 3.1 (95% CI, 1.1-8.9; P = .04) in children and 5.5 (95% CI, 1.4-22.5; P = .02) in adults. Immunotherapy was used in 5 of 9 children (56%) and 6 of 8 adults (75%) with persistent seropositivity and in 3 of 17 children (18%) and 1 of 7 adults (14%) with AQP4-IgG and MOG-IgG seronegativity. Conclusions and Relevance: Relapse occurred in 15 of 17 patients (88%) with persistent MOG-IgG1 seropositivity after ADEM; only 1 patient with transient seropositivity experienced relapse. Our data extend the clinical utility of MOG-IgG1 serological testing to adult patients and highlights that longitudinal serologic evaluation of MOG-IgG1 could help predict disease course and consideration of immunotherapy.

The single-probe: a miniaturized multifunctional device for single cell mass spectrometry analysis.

We have developed a new mass spectrometry (MS) technology, the Single-probe MS, capable of real-time, in situ metabolomic analysis of individual living cells. The Single-probe is a miniaturized multifunctional sampling and ionization device that is directly coupled to the mass spectrometer. With a sampling tip smaller than individual eukaryotic cells (<10 µm), the Single-probe can be inserted into single cells to sample the intracellular compounds for real-time MS analysis. We have used the Single-probe to detect several cellular metabolites and the anticancer small molecules paclitaxel, doxorubicin, and OSW-1 in individual cervical cancer cells (HeLa). Single cell mass spectrometry (SCMS) is an emerging scientific technology that could reshape the analytical science of many research disciplines, and the Single-probe MS technology is a novel method for SCMS that, through its accessible fabrication protocols, can be broadly applied to different research areas.

Separation of mutational and transcriptional enhancers in Ig genes.

Secondary Ig gene diversification relies on activation-induced cytidine deaminase (AID) to create U:G mismatches that are subsequently fixed by mutagenic repair pathways. AID activity is focused to Ig loci by cis-regulatory DNA sequences named targeting elements. In this study, we show that in contrast to prevailing thought in the field, the targeting elements in the chicken IGL locus are distinct from classical transcriptional enhancers. These mutational enhancer elements (MEEs) are required over and above transcription to recruit AID-mediated mutagenesis to Ig loci. We identified a small 222-bp fragment in the chicken IGL locus that enhances mutagenesis without boosting transcription, and this sequence represents a key component of an MEE. Lastly, MEEs are evolutionarily conserved among birds, both in sequence and function, and contain several highly conserved sequence modules that are likely involved in recruiting trans-acting targeting factors. We propose that MEEs represent a novel class of cis-regulatory elements for which the function is to control genomic integrity.

Classical Mus musculus Igκ enhancers support transcription but not high level somatic hypermutation from a V-lambda promoter in chicken DT40 cells.

Somatic hypermutation (SHM) of immunoglobulin genes is initiated by activation-induced cytidine deaminase (AID) in activated B cells. This process is strictly dependent on transcription. Hence, cis-acting transcriptional control elements have been proposed to target SHM to immunoglobulin loci. The Mus musculus Igκ locus is regulated by the intronic enhancer (iE/MAR) and the 3' enhancer (3'E), and multiple studies using transgenic and knock-out approaches in mice and cell lines have reported somewhat contradictory results about the function of these enhancers in AID-mediated sequence diversification. Here we show that the M. musculus iE/MAR and 3'E elements are active solely as transcriptional enhancer when placed in the context of the IGL locus in Gallus gallus DT40 cells, but they are very inefficient in targeting AID-mediated mutation events to this locus. This suggests that either key components of the cis-regulatory targeting elements reside outside the murine Igκ transcriptional enhancer sequences, or that the targeting of AID activity to Ig loci occurs by largely species-specific mechanisms.

Targeting of AID-mediated sequence diversification to immunoglobulin genes.

Activation-induced cytidine deaminase (AID) is a key enzyme for antibody-mediated immune responses. Antibodies are encoded by the immunoglobulin genes and AID acts as a transcription-dependent DNA mutator on these genes to improve antibody affinity and effector functions. An emerging theme in field is that many transcribed genes are potential targets of AID, presenting an obvious danger to genomic integrity. Thus there are mechanisms in place to ensure that mutagenic outcomes of AID activity are specifically restricted to the immunoglobulin loci. Cis-regulatory targeting elements mediate this effect and their mode of action is probably a combination of immunoglobulin gene specific activation of AID and a perversion of faithful DNA repair towards error-prone outcomes.