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Introduction: Clinical sequencing has provided molecular and therapeutic insights into the field of clinical oncology. However, despite its significance, its clinical utility in Japanese patients remains unknown. Here, we examined the clinical utility of tissue-based clinical sequencing with FoundationOne® CDx and FoundationOne® Heme. Between August 2018 and August 2019, 130 Japanese pretreated patients with advanced solid tumors were tested with FoundationOne® CDx or FoundationOne® Heme. Results: The median age of 130 patients was 60.5 years (range: 3 to 84 years), and among them, 64 were males and 66 were females. Major cancer types were gastrointestinal cancer (23 cases) and hepatic, biliary, and pancreatic cancer (21 cases). A molecular tumor board had been completed on all 130 cases by October 31, 2019. The median number of gene alterations detected by Foundation testing, excluding variants of unknown significance (VUS) was 4 (ranged 0 to 21) per case. Of the 130 cases, one or more alterations were found in 123 cases (94.6%), and in 114 cases (87.7%), actionable alterations with candidates for therapeutic agents were found. In 29 (22.3%) of them, treatment corresponding to the gene alteration was performed. Regarding secondary findings, 13 cases (10%) had an alteration suspected of a hereditary tumor. Of the 13 cases, only one case received a definite diagnosis of hereditary tumor. Conclusions: Our study showed that clinical sequencing might be useful for detecting gene alterations in various cancer types and exploring treatment options. However, many issues still need to be improved. [ABSTRACT FROM AUTHOR]

Zika virus (ZIKV) strains are classified into the African and Asian genotypes. The higher virulence of the African MR766 strain, which has been used extensively in ZIKV research, in adult IFNα/β receptor knockout (IFNAR-/-) mice is widely viewed as an artifact associated with mouse adaptation due to at least 146 passages in wild-type suckling mouse brains. To gain insights into the molecular determinants of MR766's virulence, a series of genes from MR766 were swapped with those from the Asian genotype PRVABC59 isolate, which is less virulent in IFNAR-/- mice. MR766 causes 100% lethal infection in IFNAR-/- mice, but when the prM gene of MR766 was replaced with that of PRVABC59, the chimera MR/PR(prM) showed 0% lethal infection. The reduced virulence was associated with reduced neuroinvasiveness, with MR766 brain titers ≈3 logs higher than those of MR/PR(prM) after subcutaneous infection, but was not significantly different in brain titers of MR766 and MR/PR(prM) after intracranial inoculation. MR/PR(prM) also showed reduced transcytosis when compared with MR766 in vitro. The high neuroinvasiveness of MR766 in IFNAR-/- mice could be linked to the 10 amino acids that differ between the prM proteins of MR766 and PRVABC59, with 5 of these changes affecting positive charge and hydrophobicity on the exposed surface of the prM protein. These 10 amino acids are highly conserved amongst African ZIKV isolates, irrespective of suckling mouse passage, arguing that the high virulence of MR766 in adult IFNAR-/- mice is not the result of mouse adaptation. Author summary: Contemporary Asian genotype Zika virus (ZIKV) caused a large epidemic in the Americas and was associated with congenital abnormalities and neurological disease in adults, whereas African ZIKV has not been reported to cause neurological malformations in humans. Whether the lack of reported cases caused by African ZIKV simply reflects under-reporting remains unclear. Numerous studies have attempted to explain how and why these severe manifestations are restricted to ZIKV of Asian origin. Although several studies have reported that African ZIKV strains are more virulent in adult IFNα/β receptor knockout (IFNAR-/-) mice, the representative African ZIKV strain MR766 is believed to have artefactually high virulence in adult IFNAR-/- mice due to its extensive passage history in suckling wild-type mouse brains. However, we provide evidence herein that the lethality of MR766 in the adult IFNAR-/- mice is associated with 10 amino acid residues in the prM protein that are associated with an increased ability to cross the blood-brain barrier. These amino acids are highly conserved in African genotype viruses, irrespective of multiple passage history in suckling mouse brains, suggesting that high virulence is a legitimate characteristic of African ZIKVs. [ABSTRACT FROM AUTHOR]

JC polyomavirus (JCPyV, JCV) causes progressive multifocal leukoencephalopathy (PML) in immunocompromised hosts. JCPyV replicates in oligodendrocytes within the brain tissue of patients with PML. The JCPyV genome encodes a microRNA (miRNA) in the region encoding the large T antigen. JCPyV-encoded miRNA (miR-J1) has been detected in the tissue and cerebrospinal fluid samples of patients with PML; however, there are no reports describing the localization of polyomavirus-encoded miRNA in histological samples of patients with virus-associated diseases. In the present study, we detected high miR-J1 expression in the nuclei of JCPyV-infected cells in PML tissue samples via in situ hybridization. Additionally, in situ hybridization also revealed the expression of BK polyomavirus (BKPyV)-encoded miRNA in lesions of BKPyV -associated nephropathy. In situ hybridization for miR-J1-5p and -3p showed positive signals in 24/25 (96%) of PML tissues that were positive for JCPyV by immunohistochemistry. Higher copy numbers of miR-J1 were detected in PML tissues than in non-PML tissues by real-time reverse transcription PCR. Next generation sequencing showed that miR-J1-5p, a mature miRNA of primary miRNA, was predominant in the lesions compared with miR-J1-3p, another mature miRNA. Deletion or mutation of miR-J1 in recombinant JCPyV promoted the production of JCPyV-encoded proteins in cells transfected with JCPyV DNA, suggesting that polyomavirus-encoded miRNA may have a repressive role in viral replication in PML tissues. In situ hybridization for viral miRNA may be a useful diagnostic tool for PML. Author summary: JCPyV causes PML in immunocompromised hosts. In patients with PML, JCPyV replicates in the oligodendrocytes of brain tissue. A JCPyV-encoded miRNA, miR-J1, has been detected by real-time PCR in the brain tissue and cerebrospinal fluid of patients with PML; however, there are no reports describing the localization of polyomavirus-encoded miRNA in histological samples of patients with virus-associated diseases. In this study, in situ hybridization clearly showed that miR-J1 was highly expressed in the nuclei of JCPyV-infected cells within PML tissue samples. A high level of BKPyV-encoded miRNA expression was also found in BKPyV-associated nephropathy. The results of experiments with recombinant JCPyV demonstrate that a defect of miR-J1 promotes JCPyV replication in cells transfected with JCPyV DNA. The observed high expression of viral miRNA, which has a repressive role in virus replication, suggests an autoregulation mechanism in virus replication as well as the possibility that viral miRNA may be a new therapeutic tool for PML. [ABSTRACT FROM AUTHOR]

We investigated the growth properties and virulence in mice of three Zika virus (ZIKV) strains of Asian/American lineage, PRVABC59, ZIKV/Hu/Chiba/S36/2016 (ChibaS36), and ZIKV/Hu/NIID123/2016 (NIID123), belonging to the three distinct subtypes of this lineage. The American-subtype strain, PRVABC59, showed the highest growth potential in vitro, whereas the Southeast Asian-subtype strain, NIID123, showed the lowest proliferative capacity. Moreover, PRVABC59- and NIID123-infected mice showed the highest and lowest viremia levels and infectious virus levels in the testis, respectively, and the rate of damaged testis in PRVABC59-infected mice was higher than in mice infected with the other two strains. Lastly, ZIKV NS1 antigen was detected in the damaged testes of mice infected with PRVABC59 and the Pacific-subtype strain, ChibaS36, at 2 weeks post-inoculation and in the epididymides of PRVABC59-infected mice at 6 weeks post-inoculation. Our results indicate that PRVABC59 and ChibaS36 exhibit increased abilities to grow in vitro and in vivo and to induce testis damage in mice. [ABSTRACT FROM AUTHOR]

Influenza A virus is the respiratory pathogen responsible for influenza. Infection by the 2009 pandemic influenza A (H1N1) virus caused severe lower airway inflammation and pneumonia. Asthma is a chronic inflammatory disorder of the airways that affects the entire brachial tree, and was one of the commonest underlying medical conditions among patients hospitalized with the 2009 pandemic influenza virus infection. Although respiratory virus infections are the major causes of asthma exacerbation, the mechanism by which influenza exacerbates asthma is poorly understood. Animal models of disease comorbidity are crucial to understanding host-pathogen interactions and elucidating complex pathologies. Existing murine models of influenza virus infection in asthmatics show that asthmatic mice are highly resistant to influenza virus infection, which contradicts clinical observations in humans. Here, we developed a murine model of influenza virus/asthma comorbidity using NC/Nga mice, which are highly sensitive to allergic reactions such as atopic dermatitis and allergic airway inflammation. This model was then used to examine the impact of allergic airway inflammation on lung pathology in the 2009 pandemic influenza virus infected mice. The results showed that induction of acute allergic airway inflammation in pre-existing influenza virus infection had additive effects on exacerbation of lung pathology, which mirrors findings in human epidemiological studies. In contrast, pre-existing allergic airway inflammation protected from subsequent influenza virus infection, which was compatible with those of previous murine models of influenza virus infection in asthmatic mice. These variable outcomes of this murine model indicate that the temporal relation between allergic airway inflammation and influenza virus infection might play a critical role in asthma and influenza comorbidity. Thus, this murine model will further our understanding of how influenza virus infection affects an asthmatic host and may aid the development of strategies to improve treatments and outcomes for asthmatics harboring influenza virus infection. [ABSTRACT FROM AUTHOR]

Zika virus (ZIKV) strains are classified into the African and Asian genotypes. The higher virulence of the African MR766 strain, which has been used extensively in ZIKV research, in adult IFNα/β receptor knockout (IFNAR-/-) mice is widely viewed as an artifact associated with mouse adaptation due to at least 146 passages in wild-type suckling mouse brains. To gain insights into the molecular determinants of MR766's virulence, a series of genes from MR766 were swapped with those from the Asian genotype PRVABC59 isolate, which is less virulent in IFNAR-/- mice. MR766 causes 100% lethal infection in IFNAR-/- mice, but when the prM gene of MR766 was replaced with that of PRVABC59, the chimera MR/PR(prM) showed 0% lethal infection. The reduced virulence was associated with reduced neuroinvasiveness, with MR766 brain titers ≈3 logs higher than those of MR/PR(prM) after subcutaneous infection, but was not significantly different in brain titers of MR766 and MR/PR(prM) after intracranial inoculation. MR/PR(prM) also showed reduced transcytosis when compared with MR766 in vitro. The high neuroinvasiveness of MR766 in IFNAR-/- mice could be linked to the 10 amino acids that differ between the prM proteins of MR766 and PRVABC59, with 5 of these changes affecting positive charge and hydrophobicity on the exposed surface of the prM protein. These 10 amino acids are highly conserved amongst African ZIKV isolates, irrespective of suckling mouse passage, arguing that the high virulence of MR766 in adult IFNAR-/- mice is not the result of mouse adaptation., Competing Interests: The authors have declared that no competing interests exist.