Adapting NGS for coronavirus surveillance

In work that is proving prescient, researchers at the Wuhan Institute of Virology and Duke-National University of Singapore have adapted next-generation sequencing (NGS) for coronavirus surveillance. The emergence of 2019-nCoV as a pandemic threat is the third serious public health threat by coronaviruses, pointing to a need for surveillance. However, coronaviruses are a large and genetically very diverse family, and it is likely that many of its members remain undiscovered. In such a situation, neither targeted PCR nor NGS are good options for surveillance because PCR will miss genomic diversity and NGS will miss low-abundance species, and be very costly. In their work, which was conducted prior to the emergence of 2019-nCoV, the team combined NGS on coronavirus-enriched samples based on eight strains of coronavirus. They demonstrated that “within these data sets, the ratios of viral reads to total number of reads increased by almost 100% for captured samples, in contrast to the ratios of less than 1% for most of the unbiased NGS. The high ratio of viral to total reads in conjunction with decreased data size reduces the sequencing cost and data analysis burden.” The team acknowledged that the method is in need of further improvements, including better capture of diverse spike regions, but concluded that “this methodology could thus greatly facilitate large-scale surveillance studies.” Their work appeared in the Jan. 29, 2020, online issue of mSphere.

Long QT genes mostly short on evidence

Long QT syndrome (LQTS) is an arrhythmia of the heart that can cause sudden death in otherwise healthy and often young people, without any structural abnormalities of the heart. Since it was first described 25 years ago, 17 genes have been linked to LQTS, and are tested for in patients. Now, researchers at the University of Toronto have reappraised those genes, and come to the conclusion that, using current criteria, variants in only three of those genes – KCNQ1, KCNH2 and SCN5A – had definitive evidence for causing LQTS, and 10 genes had moderate, limited or disputed evidence. The authors concluded that genes with weak evidence should not be part of clinical testing and that, more generally, “the findings of insufficient evidence to support gene-disease associations may extend to other disciplines of medicine and warrants a contemporary evidence-based evaluation for previously reported disease-causing genes to ensure their appropriate use in precision medicine.” Their work appeared in the Jan. 27, 2020, online issue of Circulation.

Reservoir dogs don’t hunt

Researchers at the Johns Hopkins University School of Medicine have uncovered sobering evidence about the HIV reservoir’s ability to escape immune clearance. Latently infected T cells, which are not dividing and not producing virus, are a major barrier to curing HIV infections. Current efforts are focused on activating such latently infected T cells to prod HIV in such cells into replicating. However, the authors showed that, in vitro, most infected cells that were activated did not produce virus, even if they were infected with replication-competent provirus. The authors concluded that “the very high fraction (98.3%) of intact proviruses that are not induced even after four rounds of T cell stimulation demands further investigation to determine whether these proviruses contribute to the latent reservoir, residual viremia, and viral rebound. Several biological mechanisms might explain this observation.” In the meantime, “this finding complicates the vision for a targeted approach for HIV-1 cure based on T cell memory subset.” Their work appeared in the Jan. 29, 2020, issue of Science Translational Medicine.

Another reason to get a flu shot

Investigators at Rush University Medical Center have discovered that intratumoral injection of unadjuvanted seasonal influenza vaccine converted “cold” tumors, which do not respond to immunotherapy, into “hot” tumors with an inflammatory microenvironment. The authors showed that an unadjuvanted flu vaccine could generate systemic CD8+ T cell-mediated antitumor immunity, deplete regulatory B cells, and sensitize resistant tumors to checkpoint blockade. An adjuvanted vaccine was ineffective because it failed to deplete B cells, though adjuvant removal, B-cell depletion or IL-10 blockade could recover its antitumor effectiveness. The authors concluded that “our findings propose that antipathogen vaccines may be utilized for both infection prevention and repurposing as a cancer immunotherapy… Repurposing the “flu shot” may increase response rates to immunotherapy, and based on its current FDA approval and safety profile, may be quickly translated for clinical care.” They reported their work in the Jan. 16, 2020, print edition of the Proceedings of the National Academy of Sciences after earlier publication online.

Cerebrospinal fluid is early culprit in stroke edema

Scientists at the University of Rochester Medical Center and the University of Copenhagen have demonstrated that cerebrospinal fluid, rather than fluid from the blood, is responsible for the early stages of edema (swelling due to fluid buildup) after stroke. The extent of edema predicts how well individuals will recover after a stroke, but treatment options to minimize edema are very limited, in part because edema itself is not well understood. After a stroke, blood-brain barrier breakdown contributes to fluid buildup after several hours, but edema is evident almost immediately. In their study, the authors used various imaging technologies to show that cerebrospinal fluid entered brain tissue within minutes after an experimentally induced stroke “initiated by ischemic spreading depolarizations along with subsequent vasoconstriction, which in turn enlarged the perivascular spaces and doubled glymphatic inflow speeds.” The authors wrote that “our understanding of post-stroke edema needs to be revised and these findings could provide a conceptual basis for development of alternative treatment strategies.” Their work appeared in the Jan. 31, 2020, issue of Science.

Different drivers can turn the wheel in glioblastoma’s vicious cycle

Researchers at the Baylor College of Medicine have discovered that several different driver variants of PIK3CA drove neuronal hyperexcitability in a mouse model of glioblastoma. Neuronal hyperexcitability is an important contributor to the growth of glioblastomas, but how tumors affect their environments to induce such hyperexcitability remains largely unknown. PIK3CA is the catalytic subunit of PI3 kinase. It is mutated in roughly 10% of glioblastoma cases, and the pathway it is a part of is mutated in 90% of glioblastoma cases. In their work, the authors used in utero gene editing in mice and screening of human glioblastoma cells to identify PIK3CA driver mutations and investigate their effects on neuronal excitability at the molecular level. They demonstrated that different PIK3CA mutations led to tumors with distinct molecular features, but converged on an ability to induce neuronal hyperexcitability and seizures. The authors wrote that their findings illustrate a “vicious cycle of glioblastoma cells driving neuronal hyperexcitability and this hyperexcitability driving further tumor growth.” They reported their results in the Jan. 30, 2020, issue of Nature.

Genes of the brain synapse with damaging mutations in schizophrenia. Credit: Gulsuner et al., Science (2019)

From African genomes, big insights with small sample size

Genomic variation is greatest in African populations and those of the African diaspora, because the prehistoric migrations out of the African continent caused repeated genetic bottlenecks in other populations. Researchers at the University of Washington and South African and U.S. colleagues have taken advantage of that diversity to gain new insights into schizophrenia risk genes by studying the South African Xhosa population. By sequencing slightly more than 900 schizophrenics and age-, gender- and residence-matched controls each, the authors were able to show that individuals with schizophrenia were more likely to harbor mutations in genes that were important for synaptic function and GABA, glutamate and dopamine neural circuits. “Schizophrenia is genetically highly heterogeneous, involving severe ultrarare mutations in genes that are critical to synaptic plasticity,” the authors wrote. “The depth of genetic variation in Africa revealed this relationship with a moderate sample size and informed our understanding of the genetics of schizophrenia worldwide. Their work appeared in the Jan. 31, 2020, issue of Science.

Commercial antibodies underwhelm for studies of PP2A

Scientists at the University of Vienna have shown that several commercial antibodies for protein phosphatase 2A (PP2A) had shortcomings that severely affected their performance. PP2A is an important cell cycle regulator, and the PP2A inhibitor LB-100 is in clinical trials for recurrent gliomas. Because PP2A is structurally complex, the authors tested the details of its binding to different forms of the enzyme. PP2A antibodies from several different commercially available kits “preferentially recognized the nonmethylated form of the enzyme, and they did not coimmunoprecipitate trimeric holoenzymes containing the regulatory subunits B or B′, an issue that precludes their use to monitor PP2A holoenzyme activity,” the authors wrote. “Furthermore, some of the antibodies also recognized the phosphatase PP4, demonstrating a lack of specificity for PP2A. Together, these findings suggest that reinterpretation of the data generated by using these reagents is required.” They reported their findings in the Jan. 28, 2020, issue of Science Signaling.

Tau keeps gliomas in check

Tau protein is bad news in Alzheimer’s disease (AD), but researchers at the Autonomous University in Madrid have shown that in gliomas, the protein can prevent transition from low-grade-gliomas to the much more aggressive glioblastoma. The team was investigating why patients with mutated isocitrate dehydrogenase (IDH) have a better prognosis, and found that such patients expressed high levels of tau. Mechanistically, the team showed that tau protein kept tumor angiogenesis in check. Gliomas have a high level of angiogenesis, and preventing that angiogenesis hampered tumor growth. The team also showed that tau was more highly expressed in IDH-mutated tumors because the metabolic product of mutated IDH, 2-hydroxyglutarate, contributed to the epigenetic control of tau expression. “Our results provide a possible explanation for the better outcome of IDH mut gliomas, which could have important implications for several aspects of glioma research and clinical practice,” the authors wrote. “Moreover, understanding how tau governs the vascular component of the [tumor microenvironment] in gliomas could provide insight for the neurovascular dysfunction observed in AD and other dementias.” They reported their findings in the Jan. 22, 2020, issue of Science Translational Medicine.

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