Integrating extracellular vesicle and circulating cell-free DNA analysis using a single plasma aliquot improves the detection of HER2 positivity in breast cancer patients.

Multi-analyte liquid biopsies represent an emerging opportunity for non-invasive cancer assessment. We developed ONCE (ONe Aliquot for Circulating Elements), an approach for the isolation of extracellular vesicles (EV) and cell-free DNA (cfDNA) from a single aliquot of blood. We assessed ONCE performance to classify HER2-positive early-stage breast cancer (BrCa) patients by combining EV-associated RNA (EV-RNA) and cfDNA signals on n=64 healthy donors (HD) and non-metastatic BrCa patients. Specifically, we isolated EV-enriched samples by a charge-based (CB) method and investigated EV-RNA and cfDNA by next-generation sequencing (NGS) and by digital droplet PCR (ddPCR). Sequencing of cfDNA and EV-RNA from HER2- and HER2+ patients demonstrated concordance with in situ molecular analyses of matched tissues. Combined analysis of the two circulating analytes by ddPCR showed increased sensitivity in ERBB2/HER2 detection compared to single nucleic acid components. Multi-analyte liquid biopsy prediction performance was comparable to tissue-based sequencing results from TCGA. Also, imaging flow cytometry analysis revealed HER2 protein on the surface of EV isolated from the HER2+ BrCa plasma, thus corroborating the potential relevance of studying EV as companion analyte to cfDNA. This data confirms the relevance of combining cfDNA and EV-RNA for HER2 cancer assessment and supports the ONCE as a valuable tool for multi-analytes liquid biopsies' clinical implementation.

Prevalence of Otolaryngology Diseases in an Urban Homeless Population.

OBJECTIVE: Otolaryngology diseases are common among people experiencing homelessness; however, they are seldom evaluated in a specialist setting, and investigations on their prevalence have rarely been conducted. The aim of this retrospective study was to evaluate the prevalence of otolaryngology conditions in an urban homeless population. STUDY DESIGN: Retrospective study. SETTING: Primary health care facility. METHODS: The clinical records of patients referred to the medical facilities of the Primary Care Services of the Eleemosynaria Apostolica, Vatican City, between October 1, 2019, and July 31, 2021, were retrospectively reviewed; those reporting at least 1 otolaryngology disease were included in the study. RESULTS: A total of 2516 records were retrospectively reviewed, and 484 (19.24%) were included in the study. The most common otolaryngology disease was pharyngotonsillitis (n = 118, 24.13%), followed by rhinitis with nasal obstruction (n = 107, 21.88%), hearing loss (n = 93, 19.01%), otitis (n = 81, 16.56%), abscess (n = 46, 9.40%), and sinusitis (n = 33, 6.74%). Head and neck cancer or precancerous lesions were reported in 34 subjects (7.02%). More than 1 simultaneous otolaryngology disorder was found in nearly 50% of our sample. A wide range of comorbidities was also reported. CONCLUSIONS: Our results confirm an elevated otolaryngology demand in the homeless population and encourage the development of more efficient and effective strategies for a population-tailored diagnosis and treatment of these conditions.

Aural carcinoma with chondroid metaplasia at metastatic sites in a dog.

A case of aural carcinoma with chondroid metaplasia at metastatic foci in an 8-year-old male pug is described. Multiple metastases in both lungs and the right submandibular, parotid, retropharyngeal, cervical and prescapular lymph nodes were detected. Histologically, the skin of the right ear canal appeared to be diffusely infiltrated by cords and nests of neoplastic epithelial cells, showing multifocal contiguity with the overlying hyperplastic squamous epithelium. Most of the carcinomatous cells were arranged in a glandular-like pattern, with formation of lumens containing epithelial cells attached to the peripheral cell layer by elongated intercellular bridges. Scattered foci of keratinization with central accumulations of compact, laminated keratin were also observed, and histochemical stains failed to detect mucinous secretory material. Even though histological and histochemical findings were compatible with a diagnosis of acantholytic squamous cell carcinoma, CAM5.2 immunostaining was detectable in the majority, although not all, neoplastic cells, confirming a diagnosis of poorly differentiated ceruminous gland carcinoma. Pulmonary metastatic nodules revealed multifocal areas of cartilaginous metaplasia with apparent transition of carcinomatous cells to chondroid cells, showing nuclear atypia and focal cytokeratin immunostaining. Carcinomatous cells surrounding chondroid areas also revealed focal vimentin and S100 immunoreactivity. Histological evidence of transition between the two components, as well as the presence of intermediate cells displaying both epithelial and mesenchymal immunohistochemical features, strongly indicated a final diagnosis of carcinosarcoma, in which chondrosarcomatous elements were derived from carcinoma cells.

Heat shock protein expression in canine malignant mammary tumours.

BACKGROUND: Abnormal levels of Heat Shock Proteins (HSPs) have been observed in many human neoplasms including breast cancer and it has been demonstrated that they have both prognostic and therapeutic implications. In this study, we evaluated immunohistochemical expression of HSPs in normal and neoplastic canine mammary glands and confronted these results with overall survival (OS), in order to understand the role of HSPs in carcinogenesis and to establish their potential prognostic and/or therapeutic value. METHODS: Immunohistochemical expression of Hsp27, Hsp72, Hsp73 and Hsp90 was evaluated in 3 normal canine mammary glands and 30 malignant mammary tumours (10 in situ carcinomas, 10 invasive carcinomas limited to local structures without identifiable invasion of blood or lymphatic vessels, 10 carcinomas with invasion of blood or lymphatic vessels and/or metastases to regional lymph nodes). A semi-quantitative method was used for the analysis of the results. RESULTS: Widespread constitutive expression of Hsp73 and Hsp90 was detected in normal tissue, Hsp72 appeared to be focally distributed and Hsp27 showed a negative to rare weak immunostaining. In mammary tumours, a significant increase in Hsp27 (P < 0.01), Hsp72 (P < 0.05) and Hsp90 (P < 0.01) expression was observed as well as a significant reduction in Hsp73 (P < 0.01) immunoreactivity compared to normal mammary gland tissue. Hsp27 demonstrated a strong positivity in infiltrating tumour cells and metaplastic squamous elements of invasive groups. High Hsp27 expression also appeared to be significantly correlated to a shorter OS (P = 0.00087). Intense immunolabelling of Hsp72 and Hsp73 was frequently detected in infiltrative or inflammatory tumour areas. Hsp90 expression was high in all tumours and, like Hsp73, it also showed an intense positivity in lymphatic emboli. CONCLUSION: These results suggest that Hsp27, Hsp72 and Hsp90 are involved in canine mammary gland carcinogenesis. In addition, Hsp27 appears to be implicated in tumour invasiveness and its high immunodetection in invasive tumours is indicative of a poorer clinical outcome.