Metastatic and non-metastatic melanoma imaging using Sgc8-c aptamer PTK7-recognizer

Melanoma is one of the most aggressive and deadly skin cancers, and although histopathological criteria are used for its prognosis, biomarkers are necessary to identify the different evolution stages. The applications of molecular imaging include the in vivo diagnosis of cancer with probes that recognize the tumor-biomarkers specific expression allowing external image acquisitions and evaluation of the biological process in quali-quantitative ways. Aptamers are oligonucleotides that recognize targets with high affinity and specificity presenting advantages that make them interesting molecular imaging probes. Sgc8-c (DNA-aptamer) selectively recognizes PTK7-receptor overexpressed in various types of tumors. Herein, Sgc8-c was evaluated, for the first time, in a metastatic melanoma model as molecular imaging probe for in vivo diagnostic, as well as in a non-metastatic melanoma model. Firstly, two probes, radio- and fluorescent-probe, were in vitro evaluated verifying the high specific PTK7 recognition and its internalization in tumor cells by the endosomal route. Secondly, in vivo proof of concept was performed in animal tumor models. In addition, they have rapid clearance from blood exhibiting excellent target (tumor)/non-target organ ratios. Furthermore, optimal biodistribution was observed 24 h after probes injections accumulating almost exclusively in the tumor tissue. Sgc8-c is a potential tool for their specific use in the early detection of melanoma.

Aptamer Applications in Emerging Viral Diseases

Aptamers are single-stranded DNA or RNA molecules which are submitted to a process denominated SELEX. SELEX uses reiterative screening of a random oligonucleotide library to identify high-affinity binders to a chosen target, which may be a peptide, protein, or entire cells or viral particles. Aptamers can rival antibodies in target recognition, and benefit from their non-proteic nature, ease of modification, increased stability, and pharmacokinetic properties. This turns them into ideal candidates for diagnostic as well as therapeutic applications. Here, we review the recent accomplishments in the development of aptamers targeting emerging viral diseases, with emphasis on recent findings of aptamers binding to coronaviruses. We focus on aptamer development for diagnosis, including biosensors, in addition to aptamer modifications for stabilization in body fluids and tissue penetration. Such aptamers are aimed at in vivo diagnosis and treatment, such as quantification of viral load and blocking host cell invasion, virus assembly, or replication, respectively. Although there are currently no in vivo applications of aptamers in combating viral diseases, such strategies are promising for therapy development in the future.

Cerebrospinal Fluid and Plasma Small Extracellular Vesicles and miRNAs as Biomarkers for Prion Diseases

Diagnosis of transmissible spongiform encephalopathies (TSEs), or prion diseases, is based on the detection of proteinase K (PK)-resistant PrPSc in post-mortem tissues as indication of infection and disease. Since PrPSc detection is not considered a reliable method for in vivo diagnosis in most TSEs, it is of crucial importance to identify an alternative source of biomarkers to provide useful alternatives for current diagnostic methodology. Ovine scrapie is the prototype of TSEs and has been known for a long time. Using this natural model of TSE, we investigated the presence of PrPSc in exosomes derived from plasma and cerebrospinal fluid (CSF) by protein misfolding cyclic amplification (PMCA) and the levels of candidate microRNAs (miRNAs) by quantitative PCR (qPCR). Significant scrapie-associated increase was found for miR-21-5p in plasma-derived but not in CSF-derived exosomes. However, miR-342-3p, miR-146a-5p, miR-128-3p and miR-21-5p displayed higher levels in total CSF from scrapie-infected sheep. The analysis of overexpressed miRNAs in this biofluid, together with plasma exosomal miR-21-5p, could help in scrapie diagnosis once the presence of the disease is suspected. In addition, we found the presence of PrPSc in most CSF-derived exosomes from clinically affected sheep, which may facilitate in vivo diagnosis of prion diseases, at least during the clinical stage.

In Vivo Diagnosis of Synucleinopathies: A Comparative Study of Skin Biopsy and RT-QuIC

Objective.To determine whether: 1) the immunofluorescence (IF) is a reproducible technique in detecting misfolded α-synuclein (α-syn) in skin nerves; and subsequently 2) IF and RT-QuIC (both in skin and CSF) show a comparable in vivo diagnostic accuracy in distinguish synucleinopathies (SOPs) from non-synucleinopathies (non-SOPs) in a large cohort of patients.Methods.We prospectively recruited 90 patients fulfilling clinical and instrumental diagnostic criteria for all SOPs variants and non-SOPs (mainly including Alzheimer disease, tauopathies, and vascular parkinsonism or dementia). Twenty-four patients with mainly peripheral neuropathies were used as controls. Patients underwent skin biopsy for IF and RT-QuIC whereas CSF was performed in patients who underwent lumbar puncture for diagnostic purposes. IF and RT-QuIC analysis were made blinded to the clinical diagnosis.Results.IF showed reproducible results between two pairs of neighbouring skin samples. Furthermore, both IF and RT-QuIC showed high sensitivity and specificity in discriminating SOPs from non-SOPs and controls but IF presented the higher diagnostic accuracy. IF presented a good level of agreement with RT-QuIC both skin and CSF in SOPs.Conclusions:1) Both IF and RT-QuIC showed a high diagnostic accuracy although IF displayed the better value as well as an optimal reproducibility; 2) they presented a good level of agreement in SOPs supporting the use of a less invasive tests such as skin IF or RT-QuIC instead of CSF RT-QuIC as diagnostic tool for synucleinopathies.Classification of evidence:This study provides Class III evidence that IF or RT-QuIC accurately distinguish SOPs from non-SOPs patients.

NEUROIMAGING FOR THE DIAGNOSIS OF DEMENTIA

В результате совершенствования методов лабораторной диагностики и нейровизуализации стало возможным определение ряда биомаркеров, которые позволили улучшить прижизненную диагностику наиболее распространенных вариантов деменции (болезни Альцгеймера, деменции с тельцами Леви, сосудистой деменции). Структурные изменения, которые выявляются при МРТ головного мозга, являются значимыми для прогноза изменений в когнитивной сфере. Дополнительным доступным маркером, который повышает диагностическую значимость лейкоареоза и свидетельствует в пользу гипертонической микроангиопатии или церебральной амилоидной ангиопатии, является церебральное микрокровоизлияние, особенно в случаях тяжелого когнитивного дефицита и смешанной патологии. Приводится собственное наблюдение и данные цифровой гиппокампометрии как пример эффективного использования нейровизуализационных методов в дифференциальной диагностике деменции. Development of laboratory diagnosis and neuroimaging revealed some biomarkers for in vivo diagnosis of the most common forms of dementia (Alzheimer’s disease, Lewy body dementia and vascular dementia) for their differential diagnosis. Structural changes found using magnetic resonance imaging (MRI) are significant for the prognosis of the cognitive decline. Cerebral microbleeds are an available adjuvant diagnostic marker, which increases the diagnostic value of leukoaraiosis that suggests the development of cerebral amyloid angiopathy or hypertensive microangiopathy, especially in cases of mixed causes of dementia and severe cognitive deficits. We describe our own clinical case and the results of digital voxel hippocampometry as an example of effective usage of neuroimaging for the differential diagnosis of dementia.