No abstract

The immune response of macrophage cells to internalized polyvalent nucleic acid-functionalized gold nanoparticles has been studied. This study finds that the innate immune response (as measured by interferon-β levels) to densely functionalized, oligonucleotide-modified nanoparticles is significantly less (up to a 25-fold decrease) when compared to a lipoplex carrying the same DNA sequence. The magnitude of this effect is inversely proportional to oligonucleotide density. It is proposed that the enzymes involved in recognizing foreign nucleic acids and triggering the immune response are impeded due to the local surface environment of the particle, in particular high charge density. The net effect is an intracelluar gene regulation agent that elicits a significantly lower cellular immune response than conventional DNA transfection materials.

Acute lymphoblastic leukemia (ALL) is the most common childhood malignancy, and implementation of risk-adapted therapy has been instrumental in the dramatic improvements in clinical outcomes. A key to risk-adapted therapies includes the identification of genomic features of individual tumors, including chromosome number (for hyper- and hypodiploidy) and gene fusions, notably ETV6-RUNX1, TCF3-PBX1, and BCR-ABL1 in B-cell ALL (B-ALL). RNA-sequencing (RNA-seq) of large ALL cohorts has expanded the number of recurrent gene fusions recognized as drivers in ALL, and identification of these new entities will contribute to refining ALL risk stratification. We used RNA-seq on 126 ALL patients from our clinical service to test the utility of including RNA-seq in standard-of-care diagnostic pipelines to detect gene rearrangements and IKZF1 deletions. RNA-seq identified 86% of rearrangements detected by standard-of-care diagnostics. KMT2A (MLL) rearrangements, although usually identified, were the most commonly missed by RNA-seq as a result of low expression. RNA-seq identified rearrangements that were not detected by standard-of-care testing in 9 patients. These were found in patients who were not classifiable using standard molecular assessment. We developed an approach to detect the most common IKZF1 deletion from RNA-seq data and validated this using an RQ-PCR assay. We applied an expression classifier to identify Philadelphia chromosome–like B-ALL patients. T-ALL proved a rich source of novel gene fusions, which have clinical implications or provide insights into disease biology. Our experience shows that RNA-seq can be implemented within an individual clinical service to enhance the current molecular diagnostic risk classification of ALL.

B-cell acute lymphoblastic leukemia (B-ALL) is the most common childhood cancer. Subtypes within B-ALL are distinguished by characteristic structural variants and mutations, which in some instances strongly correlate with responses to treatment. The World Health Organisation (WHO) recognises seven distinct classifications, or subtypes, as of 2016. However, recent studies have demonstrated that B-ALL can be segmented into 23 subtypes based on a combination of genomic features and gene expression profiles. A method to identify a patient's subtype would have clear utility. Despite this, no publically available classification methods using RNA-Seq exist for this purpose. Here we present ALLSorts: a publicly available method that uses RNA-Seq data to classify B-ALL samples to 18 known subtypes and five meta-subtypes. ALLSorts is the result of a hierarchical supervised machine learning algorithm applied to a training set of 1223 B-ALL samples aggregated from multiple cohorts. Validation revealed that ALLSorts can accurately attribute samples to subtypes and can attribute multiple subtypes to a sample. Furthermore, when applied to both paediatric and adult cohorts, ALLSorts was able to classify previously undefined samples into subtypes. ALLSorts is available and documented on GitHub (https://github.com/Oshlack/AllSorts/).

Emerging viruses in the paramyxovirus genusThe genus Henipavirus within the family Paramyxoviridae was created in 2002 to accommodate the recently emerging and closely related Nipah and Hendra viruses (10,28,46,47). Nipah virus emerged in Peninsular Malaysia in 1998 as a result of human contact with infected swine, and recent outbreaks have been reported in Bangladesh and India between 2001 and 2005 (7, 17, 20, 27). Hendra virus emerged in Brisbane, Australia, in 1994 as a result of human contact with infected horses and last appeared in Queensland, Australia, in 2004 (3, 32). The viruses were responsible for zoonotic respiratory disease and severe acute encephalitis in humans and livestock and exhibit respiratory and neurological tropism (24). These human pathogens are set apart by their low homology to other paramyxoviruses, wide host range, and high level of virulence, which has limited investigations into the interaction of virally encoded Henipavirus proteins with their natural host and susceptible livestock. Furthermore, these viruses caused significant economic losses for the Malaysian swine industry (30), and their potential use in bioterrorist scenarios (23), due to a lack of therapeutic intervention, warrants further investigation.The cellular response to type I interferons (IFNs) (IFN-␣ and IFN-␤, collectively referred to as IFN) results in the establishment of a potent antiviral state in addition to regulating components of the adaptive immune response (21,22). This process is mediated by the signal transducer and activator of transcription (STAT) protein family. Following IFN exposure, STAT proteins translocate to the nucleus to directly regulate cellular gene transcription, creating a cellular antiviral state (1). Many viruses have evolved well-characterized adaptations to evade the IFN-induced antiviral responses of their hosts (15). The family Paramyxoviridae, comprised of enveloped negative-strand RNA viruses in several distinct genera, has developed unique molecular mechanisms of IFN signaling inhibition, several of which have been attributed to a virus-encoded protein derived from a polycistronic gene (10,18,19).As with other paramyxoviruses, the cotranscriptional insertion of nontemplated nucleotides generates alternative mRNAs from the Henipavirus P locus, encoding the P, V, and W proteins (45). A fourth protein, C, is generated by alternate translation initiation site selection from all these mRNAs and is unrelated to the other products. The P, V, and W proteins share 407 amino acids in their N termini but differ in their C termini as a result of alternate reading frames produced by the addition of nontemplated nucleotides at the "editing site" (Fig. 1A). The P protein contains an additional 301 amino acids to produce a 78-kDa polypeptide. The P protein is highly phosphorylated, albeit by unknown kinase(s) (43), and is an essential component of the RNA transcription and replication machinery (8). The V protein C-terminal domain (CTD) is 50 amino acids in length and encodes a zinc finger do...