Studies investigating microRNAs as potential biomarkers for cancer, immune-related diseases, or cardiac pathogenic diseases, among others, have exponentially increased in the last years. In particular, altered expression of specific miRNAs correlates with the occurrence of several diseases, making these molecules potential molecular tools for non-invasive diagnosis, prognosis, and response to therapy. Nonetheless, microRNAs are not in clinical use yet, due to inconsistencies in the literature regarding the specific miRNAs identified as biomarkers for a specific disease, which in turn can be attributed to several reasons, including lack of assay standardization and reproducibility. Technological limitations in circulating microRNAs measurement have been, to date, the biggest challenge for using these molecules in clinical settings. In this review we will discuss pre-analytical, analytical, and post-analytical challenges to address the potential technical biases and patient-related parameters that can have an influence and should be improved to translate miRNA biomarkers to the clinical stage. Moreover, we will describe the currently available methods for circulating miRNA expression profiling and measurement, underlining their advantages and potential pitfalls.

Over the past 15 years, zebrafish have emerged as a powerful tool for studying human cancers. Transgenic techniques have been employed to model different types of tumors, including leukemia, melanoma, glioblastoma and endocrine tumors. These models present histopathological and molecular conservation with their human cancer counterparts and have been fundamental for understanding mechanisms of tumor initiation and progression. Moreover, xenotransplantation of human cancer cells in embryos or adult zebrafish offers the advantage of studying the behavior of human cancer cells in a live organism. Chemical-genetic screens using zebrafish embryos have uncovered novel druggable pathways and new therapeutic strategies, some of which are now tested in clinical trials. In this review, we will report on recent advances in using zebrafish as a model in cancer studies—with specific focus on four cancer types—where zebrafish has contributed to novel discoveries or approaches to novel therapies.

Cutaneous melanoma is the most aggressive form of skin cancer. Despite the significant advances in the management of melanoma in recent decades, it still represents a challenge for clinicians. The TP53 gene, the guardian of the genome, which is altered in more than 50% of human cancers, is rarely mutated in melanoma. More recently, researchers started to appreciate the importance of shorter p53 isoforms as potential modifiers of the p53-dependent responses. We analyzed the expression of p53 and p73 isoforms both at the RNA and protein level in a panel of melanoma-derived cell lines with different TP53 and BRAF status, in normal conditions or upon treatment with common anti-cancer DNA damaging agents or targeted therapy. Using lentiviral vectors, we also generated stable clones of H1299 p53 null cells over-expressing the less characterized isoforms Δ160p53α, Δ160p53β, and Δ160p53γ. Further, we obtained two melanoma-derived cell lines resistant to BRAF inhibitor vemurafenib. We observed that melanoma cell lines expressed a wide array of p53 and p73 isoforms, with Δ160p53α as the most variable one. We demonstrated for the first time that Δ160p53α, and to a lesser extent Δ160p53β, can be recruited on chromatin, and that Δ160p53γ can localize in perinuclear foci; moreover, all Δ160p53 isoforms can stimulate proliferation and in vitro migration. Lastly, vemurafenib-resistant melanoma cells showed an altered expression of p53 and p73 isoforms, namely an increased expression of potentially pro-oncogenic Δ40p53β and a decrease in tumor-suppressive TAp73β. We therefore propose that p53 family isoforms can play a role in melanoma cells’ aggressiveness.

Abiraterone acetate (AA) serves as a medication for managing persistent testosterone production in patients with metastatic castration-resistant prostate cancer (mCRPC). However, its efficacy varies among individuals; thus, the identification of biomarkers to predict and follow treatment response is required. In this pilot study, we explored the potential of circulating microRNAs (c-miRNAs) to stratify patients based on their responsiveness to AA. We conducted an analysis of plasma samples obtained from a cohort of 33 mCRPC patients before and after three, six, and nine months of AA treatment. Using miRNA RT-qPCR panels for candidate discovery and TaqMan RT-qPCR for validation, we identified promising miRNA signatures. Our investigation indicated that a signature based on miR-103a-3p and miR-378a-5p effectively discriminates between non-responder and responder patients, while also following the drug’s efficacy over time. Additionally, through in silico analysis, we identified target genes and transcription factors of the two miRNAs, including PTEN and HOXB13, which are known to play roles in AA resistance in mCRPC. In summary, our study highlights two c-miRNAs as potential companion diagnostics of AA in mCRPC patients, offering novel insights for informed decision-making in the treatment of mCRPC.

Bovine alphaherpesviruses, BoAHV, can cause respiratory, genital and neurological disorders. In particular, bovine alphaherpesvirus type 1 (BoAHV1) is one of the most significant ruminant pathogens worldwide and it can heavily damage the livestock industry. BoAHV1 can cause infectious bovine rhinotracheitis (IBR) along with fertility disorders. Bovine alphaherpesvirus type 2 (BoAHV2) can cause two different conditions as well: pseudo-lumpy skin disease (PSLD) and bovine herpetic mammillitis (BHM). The autonomous province of Bolzano (Italy) has adopted several strategies to control and eradicate IBR, and it was declared in 2000 to be IBR-free by the European Commission. Since 2001, a post-eradication monitoring program has overseen the serological analysis of bulk milk and, in the presence of a positive result, a follow-up examination is performed on the individual blood serum of all bovines older than 24 months that belong to bulk milk-positive herds. Despite the detection of positives in both bulk milk and serum samples, South Tyrol has been declared IBR-free, as these positives have never been confirmed through seroneutralization. Between 2014 and 2022, approximately 41,000 bulk milk (averaging 4300 samples/year) and 3229 serum samples were tested for BoAHV1. The aim of this study was to evaluate the post-eradication program for IBR with a particular focus on the potential cross-reactivity with BoAHV2; for this reason, serum samples were also tested for BoAHV2 antibodies. This study could be of great importance for those countries that submit herds to an IBR monitoring and eradication program; performing further analyses to confirm and explain false positive outcomes would increase the reliability of the obtained results.

Tetracapsuloides bryosalmonae is a myxozoan endoparasite responsible for Proliferative Kidney Disease (PKD) among salmonids. The myxozoan life cycle and the manifestation of clinical signs of infection in fish are water temperature related. PKD is considered an emerging disease, as climate changes could increase the possibility of outbreaks. In South Tyrol (Italy), the protection of the environment and ecosystems is considered of primary importance. The population of fish is therefore routinely monitored to verify the health status of the local waterways. Based on this knowledge, a two-year study (2022-2023) was conducted by the Institute for Animal Disease Control of Bolzano, with the contribution of the Wildlife Management Office of Bolzano. The aim of the study was to investigate the prevalence and the geographic distribution of T. bryosalmonae in trout from four of the main watercourses in South Tyrol: Isarco, Rienza and Adige rivers and the Passirio stream, with a particular focus on identifying clinical and subclinical forms of infection. A total of 143 trout was collected in 2022 (102 juveniles and 41 adults) and a total of 137 in 2023 (76 juveniles and 61 adults) in 30 different sites from the abovementioned watercourses and their adjacent irrigation ditches. All fish were subjected to anatomical inspections and kidney samples were processed to perform cytological, histological and molecular examinations. Results showed a positivity rate of T. bryosalmonae of 11 % within the sampled specimens, specifically affecting adult trout, and mainly distributed in a limited geographic area.