Immune-related adverse events, particularly severe toxicities such as myocarditis, are major challenges to the utility of immune checkpoint inhibitors (ICIs) in anticancer therapy
. The pathogenesis ...of ICI-associated myocarditis (ICI-MC) is poorly understood. Pdcd1
Ctla4
mice recapitulate clinicopathological features of ICI-MC, including myocardial T cell infiltration
. Here, using single-cell RNA and T cell receptor (TCR) sequencing of cardiac immune infiltrates from Pdcd1
Ctla4
mice, we identify clonal effector CD8
T cells as the dominant cell population. Treatment with anti-CD8-depleting, but not anti-CD4-depleting, antibodies improved the survival of Pdcd1
Ctla4
mice. Adoptive transfer of immune cells from mice with myocarditis induced fatal myocarditis in recipients, which required CD8
T cells. The cardiac-specific protein α-myosin, which is absent from the thymus
, was identified as the cognate antigen source for three major histocompatibility complex class I-restricted TCRs derived from mice with fulminant myocarditis. Peripheral blood T cells from three patients with ICI-MC were expanded by α-myosin peptides. Moreover, these α-myosin-expanded T cells shared TCR clonotypes with diseased heart and skeletal muscle, which indicates that α-myosin may be a clinically important autoantigen in ICI-MC. These studies underscore the crucial role for cytotoxic CD8
T cells, identify a candidate autoantigen in ICI-MC and yield new insights into the pathogenesis of ICI toxicity.
The simple method for amplifying RNA targets (SMART) was used to detect K103N, a common HIV-1 reverse transcriptase drug-resistance mutation. Novel amplifiable SMART probes served as reporter ...molecules for RNA sequences that are captured and separated on a microfluidic platform under zero-flow conditions. Assays were performed both off chip and in a microchip reservoir using a modified version of real-time nucleic acid sequence-based amplification, without the noncyclic phase, and 65°C preheat. A total of 6000 copies/mL of the synthetic sequences were detected within 180 minutes of amplification. Although the sensitivity of research platforms is higher, SMART has the potential to offer comparable sensitivity and speed to commercially available viral load and HIV detection kits. Furthermore, SMART uses an inexpensive, practical, and more accurate isothermal exponential amplification technique. The use of molecular beacons resulted in relatively fast real-time detection (<180 minutes); however, they were also shown to hinder the amplification process when compared with end point detection. Finally, SMART probes were used for modeling of K103N concentrations within an unknown sample. Only 1% of the SMART probes was detected within the wild-type population (6 × 108 copies/mL). These results establish the groundwork for point-of-care drug resistance and viral load monitoring in clinical samples, which can revolutionize HIV patient care globally.
This work presents a novel method for detecting nucleic acid targets using a ligation step along with an isothermal, exponential amplification step. We use an engineered ssDNA with two variable ...regions on the ends, allowing us to design the probe for optimal reaction kinetics and primer binding. This two-part probe is ligated by T4 DNA Ligase only when both parts bind adjacently to the target. The assay demonstrates that the expected 72-nt RNA product appears only when the synthetic target, T4 ligase, and both probe fragments are present during the ligation step. An extraneous 38-nt RNA product also appears due to linear amplification of unligated probe (P3), but its presence does not cause a false-positive result. In addition, 40 mmol/L KCl in the final amplification mix was found to be optimal. It was also found that increasing P5 in excess of P3 helped with ligation and reduced the extraneous 38-nt RNA product. The assay was also tested with a single nucleotide polymorphism target, changing one base at the ligation site. The assay was able to yield a negative signal despite only a single-base change. Finally, using P3 and P5 with longer binding sites results in increased overall sensitivity of the reaction, showing that increasing ligation efficiency can improve the assay overall. We believe that this method can be used effectively for a number of diagnostic assays.
Fyodorov, Hiary & Keating established an intriguing connection between the maxima of log-correlated processes and the ones of the Riemann zeta function on a short interval of the critical line. In ...particular, they suggest that the analogue of the free energy of the Riemann zeta function is identical to the one of the Random Energy Model in spin glasses. In this paper, the connection between spin glasses and the Riemann zeta function is explored further. We study a random model of the Riemann zeta function and show that its two-overlap distribution corresponds to the one of a one-step replica symmetry breaking (1-RSB) spin glass. This provides evidence that the local maxima of the zeta function are strongly clustered.
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