SummaryTumor-educated blood platelets (TEPs) are implicated as central players in the systemic and local responses to tumor growth, thereby altering their RNA profile. We determined the diagnostic potential of TEPs by mRNA sequencing of 283 platelet samples. We distinguished 228 patients with localized and metastasized tumors from 55 healthy individuals with 96% accuracy. Across six different tumor types, the location of the primary tumor was correctly identified with 71% accuracy. Also, MET or HER2-positive, and mutant KRAS, EGFR, or PIK3CA tumors were accurately distinguished using surrogate TEP mRNA profiles. Our results indicate that blood platelets provide a valuable platform for pan-cancer, multiclass cancer, and companion diagnostics, possibly enabling clinical advances in blood-based “liquid biopsies”.

Blood-based liquid biopsies, including tumor-educated blood platelets (TEPs), have emerged as promising biomarker sources for non-invasive detection of cancer. Here we demonstrate that particle-swarm optimization (PSO)-enhanced algorithms enable efficient selection of RNA biomarker panels from platelet RNA-sequencing libraries (n = 779). This resulted in accurate TEP-based detection of early- and late-stage non-small-cell lung cancer (n = 518 late-stage validation cohort, accuracy, 88%; AUC, 0.94; 95% CI, 0.92-0.96; p < 0.001; n = 106 early-stage validation cohort, accuracy, 81%; AUC, 0.89; 95% CI, 0.83-0.95; p < 0.001), independent of age of the individuals, smoking habits, whole-blood storage time, and various inflammatory conditions. PSO enabled selection of gene panels to diagnose cancer from TEPs, suggesting that swarm intelligence may also benefit the optimization of diagnostics readout of other liquid biopsy biosources.

In feline coronavirus (FCoV) pathogenesis, the ability to infect macrophages is an essential virulence factor. Whereas the low-virulence feline enteric coronavirus (FECV) isolates primarily replicate in the epithelial cells of the enteric tract, highly virulent feline infectious peritonitis virus (FIPV) isolates have acquired the ability to replicate efficiently in macrophages, which allows rapid dissemination of the virulent virus throughout the body.

Purpose: Non-small-cell lung cancers harboring EML4-ALK rearrangements are sensitive to crizotinib. However, despite initial response, most patients will eventually relapse, and monitoring EML4-ALK rearrangements over the course of treatment may help identify these patients. However, challenges associated with serial tumor biopsies have highlighted the need for blood-based assays for the monitoring of biomarkers. Platelets can sequester RNA released by tumor cells and are thus an attractive source for the non-invasive assessment of biomarkers. Methods: EML4-ALK rearrangements were analyzed by RT-PCR in platelets and plasma isolated from blood obtained from 77 patients with non-small-cell lung cancer, 38 of whom had EML4-ALK-rearranged tumors. In a subset of 29 patients with EML4-ALK-rearranged tumors who were treated with crizotinib, EML4-ALK rearrangements in platelets were correlated with progression-free and overall survival. Results: RT-PCR demonstrated 65% sensitivity and 100% specificity for the detection of EML4-ALK rearrangements in platelets. In the subset of 29 patients treated with crizotinib, progression-free survival was 3.7 months for patients with EML4-ALK+ platelets and 16 months for those with EML4-ALK− platelets (hazard ratio, 3.5; P = 0.02). Monitoring of EML4-ALK rearrangements in the platelets of one patient over a period of 30 months revealed crizotinib resistance two months prior to radiographic disease progression. Conclusions: Platelets are a valuable source for the non-invasive detection of EML4-ALK rearrangements and may prove useful for predicting and monitoring outcome to crizotinib, thereby improving clinical decisions based on radiographic imaging alone.

A longstanding enigmatic feature of the group 1 coronaviruses is the uncleaved phenotype of their spike protein, an exceptional property among class I fusion proteins. Here, however, we show that some group 1 coronavirus spike proteins carry a furin enzyme recognition motif and can actually be cleaved, as demonstrated for a feline coronavirus. Interestingly, this feature can be lost during cell culture adaptation by a single mutation in the cleavage motif; this, however, preserves a heparan sulfate binding motif and renders infection by the virus heparan sulfate dependent. We identified a similar cell culture adaptation for the human coronavirus OC43.Enveloped viruses use different types of fusion proteins to realize the membrane fusion by which they initiate their infection. For coronaviruses, it is the spike (S) protein that is responsible for cell entry, and this S protein has been shown to belong to the class I fusion proteins (4). These proteins typically occur in virions as homotrimeric complexes primed for fusion through cleavage by furin-like enzymes. Membrane fusion by these activated proteins can then be triggered upon receptor binding (e.g., human immunodeficiency virus type 1) or by conditions such as low pH after endosomal uptake (e.g., influenza A virus) (for a recent review, see reference 50).One of the puzzling questions about coronavirus S proteinmediated membrane fusion regards the cleavage requirement of the S protein. Coronaviruses have been assigned to different groups based on antigenic and genetic criteria (41). Interestingly, while the group 1 coronaviruses carry uncleaved S proteins, the S proteins of almost all viruses from groups 2 and 3 are furin activated (10) by processing at a characteristic multibasic motif (often RRXRR) present in these proteins. The importance of cleavage for infectivity was underscored recently by the revelation that the two prominent group 2 viruses lacking such a furin recognition site, and hence carrying uncleaved spikes, appeared to depend on a different, new processing mechanism. Thus, the severe acute respiratory syndrome coronavirus (SARS-CoV) and the murine hepatitis virus strain 2 (MHV-2) were both shown to require proteolytic cleavage in their target cell, which is mediated by cathepsin enzymes (23,36,42). The cathepsin cleavage site of the SARS-CoV spike protein was mapped to the same region as that in which, in other viruses, the S protein is activated by furin (B. J. Bosch and P. J. M. Rottier, unpublished observations), hence similarly generating an amino-terminal, receptor binding domain (S1) and a membrane-anchored carboxy-terminal domain (S2) responsible for membrane fusion (for reviews, see references 3 and 9).When looking closer into the enigmatic lack of cleavage of the group 1 coronavirus spike proteins, we established that the infection of cells by two of those viruses, human coronavirus (HCoV) NL63 (23) and feline infectious peritonitis virus strain 79-1146 (our unpublished observations), is insensitive to cathepsin inhibitors. However, w...

Coronaviruses are positive-strand RNA viruses of extraordinary genetic complexity and diversity. In addition to a common set of genes for replicase and structural proteins, each coronavirus may carry multiple group-specific genes apparently acquired through relatively recent heterologous recombination events. Here we describe an accessory gene, ORF3, unique to canine coronavirus type I (CCoV-I) and characterize its product, glycoprotein gp3. Whereas ORF3 is conserved in CCoV-I, only remnants remain in CCoV-II and CCoV-II-derived porcine and feline coronaviruses. Our findings provide insight into the evolutionary history of coronavirus group 1a and into the dynamics of gain and loss of accessory genes.

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We sought to investigate the clinical response to MET inhibition in patients diagnosed with structural MET alterations and to characterize their functional relevance in cellular models. Patients were selected for treatment with crizotinib upon results of hybrid capture-based next-generation sequencing. To confirm the clinical observations, we analyzed cellular models that express these MET kinase alterations. Three individual patients were identified to harbor alterations within the MET receptor. Two patients showed genomic rearrangements, leading to a gene fusion of or and One patient diagnosed with an EML4-ALK rearrangement developed a MET kinase domain duplication as a resistance mechanism to ceritinib. All 3 patients showed a partial response to crizotinib that effectively inhibits MET and ALK among other kinases. The results were further confirmed using orthogonal cellular models. Crizotinib leads to a clinical response in patients with MET rearrangements. Our functional analyses together with the clinical data suggest that these structural alterations may represent actionable targets in lung cancer patients. .