The nasopharynx (NP) is a reservoir for microbes associated with acute respiratory infections (ARIs). Lung inflammation resulting from ARIs during infancy is linked to asthma development. We examined ...the NP microbiome during the critical first year of life in a prospective cohort of 234 children, capturing both the viral and bacterial communities and documenting all incidents of ARIs. Most infants were initially colonized with Staphylococcus or Corynebacterium before stable colonization with Alloiococcus or Moraxella. Transient incursions of Streptococcus, Moraxella, or Haemophilus marked virus-associated ARIs. Our data identify the NP microbiome as a determinant for infection spread to the lower airways, severity of accompanying inflammatory symptoms, and risk for future asthma development. Early asymptomatic colonization with Streptococcus was a strong asthma predictor, and antibiotic usage disrupted asymptomatic colonization patterns. In the absence of effective anti-viral therapies, targeting pathogenic bacteria within the NP microbiome could represent a prophylactic approach to asthma.
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•The nasopharynx microbiome of infants has a simple structure dominated by six genera•Microbiome composition affects infection severity and pathogen spread to lower airways•Early asymptomatic colonization with Streptococcus increases risk of asthma•Antibiotic usage disrupts asymptomatic colonization patterns
Teo et al. characterize bacterial and viral communities within the infant nasopharynx during the first year of life, comparing between asymptomatic colonization and episodes of acute respiratory infections. Microbiome composition affects infection severity and spread to lower airways and risk for future asthma development.
To examine the transmission dynamics of Mycobacterium tuberculosis (Mtb) isolated from tuberculosis patients in Ho Chi Minh City, Vietnam, we sequenced the whole genomes of 1,635 isolates and ...compared these with 3,144 isolates from elsewhere. The data identify an underlying burden of disease caused by the endemic Mtb lineage 1 associated with the activation of long-term latent infection, and a threefold higher burden associated with the more recently introduced Beijing lineage and lineage 4 Mtb strains. We find that Beijing lineage Mtb is frequently transferred between Vietnam and other countries, and detect higher levels of transmission of Beijing lineage strains within this host population than the endemic lineage 1 Mtb. Screening for parallel evolution of Beijing lineage-associated SNPs in other Mtb lineages as a signal of positive selection, we identify an alteration in the ESX-5 type VII-secreted protein EsxW, which could potentially contribute to the enhanced transmission of Beijing lineage Mtb in Vietnamese and other host populations.
Repeated cycles of infection-associated lower airway inflammation drive the pathogenesis of persistent wheezing disease in children. In this study, the occurrence of acute respiratory tract illnesses ...(ARIs) and the nasopharyngeal microbiome (NPM) were characterized in 244 infants through their first five years of life. Through this analysis, we demonstrate that >80% of infectious events involve viral pathogens, but are accompanied by a shift in the NPM toward dominance by a small range of pathogenic bacterial genera. Unexpectedly, this change frequently precedes the detection of viral pathogens and acute symptoms. Colonization of illness-associated bacteria coupled with early allergic sensitization is associated with persistent wheeze in school-aged children, which is the hallmark of the asthma phenotype. In contrast, these bacterial genera are associated with “transient wheeze” that resolves after age 3 years in non-sensitized children. Thus, to complement early allergic sensitization, monitoring NPM composition may enable early detection and intervention in high-risk children.
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•Six genera dominate airway microbiota from birth to 2 years, but diversifies thereafter•Acute respiratory illness associates with pathogenic bacteria in the airway microbiota•Pathogenic airway bacteria may precede viral incursions and acute respiratory illness•Colonization with pathogens predicts chronic wheeze in allergic-sensitized children
Teo et al. characterize nasopharyngeal microbiota (NPM) in 244 children from birth to age 5 years during periods of illness and health. NPM colonization with illness-associated bacteria may promote acute respiratory illness independent of viral infection, and is associated with chronic or transient wheeze in allergic-sensitized or non-sensitized children, respectively.
Background: Epithelioid haemangioendothelioma (EHE) is an ultra-rare malignant vascular tumour with a prevalence of 1 per 1,000,000. It is typically molecularly characterised by a WWTR1::CAMTA1 gene ...fusion in approximately 90% of cases, or a YAP1::TFE3 gene fusion in approximately 10% of cases. EHE cases are typically refractory to therapies, and no anticancer agents are reimbursed for EHE in Australia. Methods: We report a cohort of nine EHE cases with comprehensive histologic and molecular profiling from the Walter and Eliza Hall Institute of Medical Research Stafford Fox Rare Cancer Program (WEHI-SFRCP) collated via nation-wide referral to the Australian Rare Cancer (ARC) Portal. The diagnoses of EHE were confirmed by histopathological and immunohistochemical (IHC) examination. Molecular profiling was performed using the TruSight Oncology 500 assay, the TruSight RNA fusion panel, whole genome sequencing (WGS), or whole exome sequencing (WES). Results: Molecular analysis of RNA, DNA or both was possible in seven of nine cases. The WWTR1::CAMTA1 fusion was identified in five cases. The YAP1::TFE3 fusion was identified in one case, demonstrating unique morphology compared to cases with the more common WWTR1::CAMTA1 fusion. All tumours expressed typical endothelial markers CD31, ERG, and CD34 and were negative for pan-cytokeratin. Cases with a WWTR1::CAMTA1 fusion displayed high expression of CAMTA1 and the single case with a YAP1::TFE3 fusion displayed high expression of TFE3. Survival was highly variable and unrelated to molecular profile. Conclusions: This cohort of EHE cases provides molecular and histopathological characterisation and matching clinical information that emphasises the molecular patterns and variable clinical outcomes and adds to our knowledge of this ultra-rare cancer. Such information from multiple studies will advance our understanding, potentially improving treatment options.
Drug-resistant tuberculosis (TB) infection is a growing and potent concern, and combating it will be necessary to achieve the WHO's goal of a 95% reduction in TB deaths by 2035. While prior studies ...have explored the evolution and spread of drug resistance, we still lack a clear understanding of the fitness costs (if any) imposed by resistance-conferring mutations and the role that
genetic lineage plays in determining the likelihood of resistance evolution. This study offers insight into these questions by assessing the dynamics of resistance evolution in a high-burden Southeast Asian setting with a diverse lineage composition. It demonstrates that there are clear lineage-specific differences in the dynamics of resistance acquisition and transmission and shows that different lineages evolve resistance via characteristic mutational pathways.
ABSTRACT
Conventional means of identifying variants in high‐throughput sequencing align each read against a reference sequence, and then call variants at each position. Here, we demonstrate an ...orthogonal means of identifying sequence variation by grouping the reads as amplicons prior to any alignment. We used AmpliVar to make key‐value hashes of sequence reads and group reads as individual amplicons using a table of flanking sequences. Low‐abundance reads were removed according to a selectable threshold, and reads above this threshold were aligned as groups, rather than as individual reads, permitting the use of sensitive alignment tools. We show that this approach is more sensitive, more specific, and more computationally efficient than comparable methods for the analysis of amplicon‐based high‐throughput sequencing data. The method can be extended to enable alignment‐free confirmation of variants seen in hybridization capture target‐enrichment data.
In this study, AmpliVar was applied to amplicon sequencing data derived from acute myeloid leukemia, breast and ovarian cancer samples. Using a grouped read and quality filtering approach, AmpliVar identified variants with the highest sensitivity and effective false positive reduction. It displayed superior processing speed and was able to be used as an orthogonal means of validation for hybridization capture data.
Abstract Non-Human Leukocyte Antigen (HLA) genes have concomitant, although modest, effects on multiple sclerosis (MS) susceptibility; however findings have varied in different populations. Here we ...present the results of an association study of 16 single nucleotide polymorphisms (SNPs) in 10 non-HLA genes ( IL7R , IL2RA , CLEC-16A , TYK2 , CD58 , IRF5 , STAT3 , CTLA-4 , APOE , ICAM-1 ) in a Western Australian cohort of 350 MS patients and 498 population control subjects. Our results indicate that in this population, SNPs in IL7R , TYK2 , IRF5 and APOE have modifying effects on MS susceptibility. We also found evidence of interactive protective effects between polymorphisms in the IL7R / CD58 , CLEC-16A / CTLA-4 , and TYK2 / IRF5 genes, which in some instances are restricted within HLA- or gender-defined groups.
ObjectiveAlthough counting of circulating tumour cells (CTC) has attracted a broad interest as potential markers of tumour progression and treatment response, the lack of functional characterisation ...of these cells had become a bottleneck in taking these observations to the clinic. Our objective was to culture these cells in order to understand them and exploit their therapeutic potential to the full.DesignHere, hypothesising that some CTC potentially have cancer stem cell (CSC) phenotype, we generated several CTC lines from the blood of patients with advanced metastatic colorectal cancer (CRC) based on their self-renewal abilities. Multiple standard tests were then employed to characterise these cells.ResultsOur CTC lines self-renew, express CSC markers and have multilineage differentiation ability, both in vitro and in vivo. Patient-derived CTC lines are tumorigenic in subcutaneous xenografts and are also able to colonise the liver after intrasplenic injection. RNA sequencing analyses strikingly demonstrate that drug metabolising pathways represent the most upregulated feature among CTC lines in comparison with primary CRC cells grown under similar conditions. This result is corroborated by the high resistance of the CTC lines to conventional cytotoxic compounds.ConclusionsTaken together, our results directly demonstrate the existence of patient-derived colorectal CTCs that bear all the functional attributes of CSCs. The CTC culture model described here is simple and takes <1 month from blood collection to drug testing, therefore, routine clinical application could facilitate access to personalised medicine.Clinical Trial RegistrationClinicalTrial.gov NCT01577511.
Abstract
High-grade serous endometrial carcinoma (HGSEC) accounts for just 10% of endometrial cancer (EC) cases but is responsible for at least 40% of all EC-related deaths. It typically arises in ...post-menopausal women, with 70% of patients presenting with stage III or IV disease, does not respond to hormone therapy unlike the less aggressive forms of EC, and has a lower overall survival rate of just 18-27%, which has not improved over the past two decades. The primary treatment for HGSEC is surgery, followed by a combination of standard chemotherapies (platinum and taxane) with or without localised radiotherapy. However, recurrent HGSEC is less responsive to chemotherapy than are other subtypes of EC and even initial responses to chemotherapy are poor. Therefore, there is a great unmet clinical need to find better treatment options for women with this aggressive cancer. Apart from TP53 (mutated in up to 90% of cases), the other most frequently mutated genes in HGSEC are PPP2R1A (31%), PIK3CA (22%), FBXW7 (28%), CHD4 (17%) and BRCA2 (12%). Focal amplifications of the genes MYC, ERBB2, CCNE1, FGFR3 and SOX17 are also common. The presence of ERBB2 amplification and/or HER2 over-expression in around 30% of HGSEC suggests these patients may respond to HER2-targeting drugs, such as trastuzumab. However, only modest benefit has so far been seen for single-agent HER2-targeted therapies (ie trastuzumab or lapatinib) against HGSEC, suggesting resistance mechanisms are present. Another feature of HGSEC that could be exploited therapeutically is homologous recombination deficiency (HRD), which may be targeted with PARP inhibitors (PARPi). It is not clear what proportion of HGSEC are HRD and neither HER2-targeting drugs or PARPi have been approved for the treatment of HGSEC. Due to its rarity and a lack of pre-clinical models, HGSEC has so far been understudied, resulting in a lack of effective treatment options. We currently have 33 HGSEC patients consented to the WEHI-Stafford Fox Rare Cancer Program and have developed pre-clinical models from fresh patient tumour samples received (4 patient-derived xenograft (PDX) models validated, with 3 pending). Preliminary molecular analysis of whole-genome sequencing (5 samples, one of which gave rise to a PDX model), whole-exome sequencing (4 samples), and cancer panel sequencing (3 samples, 2 of which gave rise to PDX models; one harbouring ERBB2 amplification and one harbouring an AKT mutation) data from our HGSEC cohort has been performed. This has identified potential treatment targets, including ERBB2 amplifications and mutations in HR genes. I am using the PDX models for initial in vivo therapeutic characterization studies and to develop organoid models for use in high-throughput drug assays in vitro. This will guide subsequent novel drug combination testing in our PDX models. By combining specific targeted drugs I hope to overcome de novo resistance mechanisms and prevent acquired resistance. Results from this study will guide future decisions about therapeutic strategies to improve survival of women with HGSEC.
Citation Format: Holly E. Barker, Ratana Lim, Amandine Carmagnac, Cassandra Vandenberg, Gayanie Ratnayake, Genevieve Dall, Briony Milesi, Angela Komiti, Emily O'Grady, Joshua Tram, Kym Pham Stewart, Justin Bedo, Jocelyn Penington, Joep Vissers, Sean Grimmond, Matthew Wakefield, Tony Papenfuss, Clare Scott. Identifying effective combinations of targeted therapies, using novel pre-clinical models, to improve treatment options for high-grade serous endometrial cancer abstract. In: Proceedings of the AACR Virtual Special Conference: Endometrial Cancer: New Biology Driving Research and Treatment; 2020 Nov 9-10. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(3_Suppl):Abstract nr PO037.
Abstract
Uterine sarcomas make up 1-4% of uterine malignancies. Of these 60% are classified as leiomyosarcoma (uLMS). The 5-year survival rate of uLMS is 35-65.2% for tumours that have not spread ...beyond the uterus. However, women are often diagnosed at a late stage due to a lack of screening options by which time the tumour has often spread to adjacent and distant tissues. Current standard of care for uLMS patients is surgical de-bulking followed by adjuvant chemotherapy, but significant improvement in progression free survival and overall survival is not consistently observed. The lack of advances for the treatment and screening of uLMS is due in part to the scarcity of appropriate research resources for this rare disease. Genetic analyses have been performed but on relatively small samples and there are just 14 reported patient-derived xenograft (PDX) models of uLMS in the literature to date. Through the WEHI Stafford Fox Rare Cancer Program, as well as collaborations (facilitated by ANZGOG) throughout the country and internationally, we have access to a large biobank of uLMS tissue. We have received 8 fresh uLMS samples in the laboratory, 2 of which have established PDX lines that were validated as uLMS by our anatomical pathologist. All fresh samples received into the laboratory are snap frozen for whole-genome sequencing as well as viably frozen to enable regeneration of the tissue for future applications. One application is organoid culturing, which allows the tissue to retain its 3D growth properties and is significantly cheaper than growing tissue as a PDX. Organoid culturing also allows for higher throughput of samples in drug screening assays, enabling us to fast-track the selection of drugs for validation in our PDX models. In addition to these fresh samples we also have 23 archival uLMS samples (formalin fixed, paraffin embedded) that can be used for lower coverage genetic analysis, and protein expression by immunohistochemistry. This unique biobank of uLMS tissue is the first of its kind in Australia and with it we will endeavour to gain a comprehensive understanding of this disease. Through our PDX modelling we also have the opportunity to predict resistance to therapy and test emerging therapies in a clinically relevant context. We believe this biobank will provide a critical resource which will ultimately lead to better outcomes for uLMS patients.
Citation Format: Genevieve Dall, Cassandra Vandenberg, Amandine Carmagnac, Ratana Lim, Briony Milesi, Angela Komiti, Emily O'Grady, Joshua Tram, Gayanie Ratnayake, Kym Pham Stewart, Justin Bedo, Jocelyn Penington, Joep Vissers, Inger Olesen, Sean Grimmond, Holly Barker, Tony Papenfuss, Clare Scott. Developing pre-clinical models of uterine leiomyosarcoma abstract. In: Proceedings of the AACR Virtual Special Conference: Endometrial Cancer: New Biology Driving Research and Treatment; 2020 Nov 9-10. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(3_Suppl):Abstract nr PO021.
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