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Abstract Background Despite significant advancements in multiple myeloma (MM) therapy, the highly heterogenous treatment response hinders reliable prognosis and tailored therapeutics. Herein, we have studied the clinical utility of miRNAs in ameliorating patients’ management. Methods miRNA-seq was performed in bone marrow CD138+ plasma cells (PCs) of 24 MM and smoldering MM (sMM) patients to analyze miRNAs profile. CD138+ and circulating miR-25 levels were quantified using in house RT-qPCR assays in our screening MM/sMM cohort (CD138+ plasma cells n = 167; subcohort of MM peripheral plasma samples n = 69). Two external datasets (Kryukov et al. cohort n = 149; MMRF CoMMpass study n = 760) served as institutional-independent validation cohorts. Patients’ mortality and disease progression were assessed as clinical endpoints. Internal validation was performed by bootstrap analysis. Clinical benefit was estimated by decision curve analysis. Results miRNA-seq highlighted miR-25 of CD138+ plasma cells to be upregulated in MM vs. sMM, R-ISS II/III vs. R-ISS I, and in progressed compared to progression-free patients. The analysis of our screening cohort highlighted that CD138+ miR-25 levels were correlated with short-term progression (HR = 2.729; p = 0.009) and poor survival (HR = 4.581; p = 0.004) of the patients; which was confirmed by Kryukov et al. cohort (HR = 1.878; p = 0.005) and MMRF CoMMpass study (HR = 1.414; p = 0.039) validation cohorts. Moreover, multivariate miR-25-fitted models contributed to superior risk-stratification and clinical benefit in MM prognostication. Finally, elevated miR-25 circulating levels were correlated with poor survival of MM patients (HR = 5.435; p = 0.021), serving as a potent non-invasive molecular prognostic tool. Conclusions Our study identified miR-25 overexpression as a powerful independent predictor of poor treatment outcome and post-treatment progression, aiding towards modern non-invasive disease prognosis and personalized treatment decisions.

Abstract Background Multiple Myeloma (MM) is a progressive plasma cell neoplasm characterized by heterogeneous clonal expansion. Despite promising response rates achieved with anti-BCMA CAR-T cell therapy, patients may still relapse and there are currently no clear therapeutic options in post-CAR-T settings. In this report, we present a case of a post-BCMA CAR-T relapsed/refractory (RR) MM patient with skin extramedullary disease (EMD) in which a novel MAPK inhibition combinatorial strategy was implemented based on next-generation sequencing and in vitro experiments. Case presentation A 61-year-old male with penta-refractory MM penta- (IgA lambda), ISS stage 3 with hyperdiploidy, gain of 1q21 and del13 was treated with anti-BCMA CAR-T cell therapy, achieving a best response of VGPR. He progressed after 6 months and was salvaged for a short period with autologous stem cell transplantation. Eventually, he progressed with extramedullary disease manifested as subcutaneous nodules. Based on whole-exome sequencing, we identified a BRAF (V600E) dominant subclone in both bone marrow and cutaneous plasmacytoma. Following in vitro experiments, and according to our previous studies, we implemented a triple MAPK inhibition strategy under which the patient achieved a very good partial response for 110 days, which allowed to bridge him to subsequent clinical trials and eventually achieve a stringent complete response (sCR). Conclusion Here, we show the applicability, effectiveness, and tolerability the triple MAPK inhibition strategy in the context of post-BCMA CAR-T failure in specific subset of patients. The triple therapy could bridge our hospice bound RRMM patient with BRAF (V600E) to further therapeutic options where sCR was achieved. We will further evaluate triple MAPK inhibition in patients with BRAF V600E in a precision medicine clinical trial launching soon.

Abstract Background Technological advancements in the era of massive parallel sequencing have enabled the functional dissection of the human transcriptome. However, 5′ ends of mRNAs are significantly underrepresented in these datasets, hindering the efficient analysis of the complex human transcriptome. The implementation of the template-switching mechanism at the reverse transcription stage along with 5′ rapid amplification of cDNA ends (RACE) constitutes the most prominent and efficient strategy to specify the actual 5′ ends of cDNAs. In the current study, we developed a 5′ RACE-seq method by coupling a custom template-switching and 5′ RACE assay with targeted nanopore sequencing, to accurately unveil 5′ termini of mRNA targets. Results The optimization of the described 5′ RACE-seq method was accomplished using the human BCL2L12 as control gene. We unveiled that the selection of hybrid DNA/RNA template-switching oligonucleotides as well as the complete separation of the cDNA extension incubation from the template-switching process, significantly increase the overall efficiency of the downstream 5′ RACE. Collectively, our results support the existence of two distinct 5′ termini for BCL2L12, being in complete accordance with the results derived from both direct RNA and PCR-cDNA sequencing approaches from Oxford Nanopore Technologies. As proof of concept, we implemented the described 5′ RACE-seq methodology to investigate the 5′ UTRs of several kallikrein-related peptidases (KLKs) gene family members. Our results confirmed the existence of multiple annotated 5′ UTRs of the human KLK gene family members, but also identified novel, previously uncharacterized ones. Conclusions In this work we present an in-house developed 5′ RACE-seq method, based on the template-switching mechanism and targeted nanopore sequencing. This approach enables the broad and in-depth study of 5′ UTRs of any mRNA of interest, by offering a tremendous sequencing depth, while significantly reducing the cost-per reaction compared to commercially available kits.

Abstract Background Myocardial iron overload in patients with thalassemia major (TM) is one of the most important complications. The purpose of the study was to identify advanced echocardiography parameters for early identification of myocardial dysfunction during follow-up of patients with TM. Methods Forty TM patients who were 41 ± 5 years old were included in the study and divided into two groups according to cardiac magnetic resonance T2* results (Group 1: Τ2* > 25 ms, Group 2: Τ2* ≤ 25 ms). Liver T2* parameters were also measured. Conventional and deformational echocardiographic parameters were measured at baseline and approximately 2 years later. Results Thirty-two patients had Τ2* = 34 ± 4 ms (Group 1), and 8 had Τ2* = 17 ± 9 ms (Group 2). Blood consumption was 185 ± 60 and 199 ± 37 ml/kg/yr (p = 0.64), and liver T2* was 4 ± 5 and 17 ± 21 ms (p = 0.01) in Groups 1 and 2, respectively. At baseline, Group 1 had better left ventricular global longitudinal strain (GLS) (− 22 ± 3 vs. − 18 ± 5, p = 0.01) and similar left ventricular ejection fraction (LVEF) (62 ± 5% vs. 58 ± 10%, p = 0.086) than Group 2. At the 28 ± 11-month follow-up, LVEF, GLS, and T2* values in Group 1 (63 ± 3%, − 21 ± 3%, 34 ± 4 ms) and Group 2 (56 ± 11%, − 17 ± 4%, 17 ± 9 ms) did not change significantly compared to their corresponding baseline values. In 8 patients from Group 1, a worsening (> 15%) in LS (p = 0.001) was detected during follow-up, with a marginal reduction in LVEF. Conclusions GLS seems to be an efficient echocardiographic parameter for detecting hemochromatosis-related cardiac dysfunction earlier than LVEF. It also seems to be affected by other factors (free radical oxygen, immunogenetic mechanisms or viral infections) in a minority of patients, underscoring the multifactorial etiology of cardiomyopathy.

Abstract Background Restrictive cardiomyopathy is a rare cardiac disease, for which several genes including TNNT2, MYPN, FLNC and TNNI3 have been associated with its familial form. Case presentation Here we describe a female proband with a severely manifested restrictive phenotype leading to heart transplantation at the age of 41, who was found homozygous for the novel TNNI3 mutation: NM_000363.4:c.586G > C, p.(Asp196His). Her parents were third-degree cousins originating from a small village and although they were found heterozygous for the same variant they displayed no symptoms of the disease. Her older sister who was also found heterozygous was asymptomatic. Her twin sister and her brother who were homozygous for the same variant displayed a restrictive and a hypertrophic phenotype, respectively. Their children are all carriers of the mutation and remain asymptomatic until the age of 21. Conclusion These observations point to a recessive mode of inheritance reported for the first time for this combination of gene/disease.

Abstract Background Primary cardiac lymphomas (PCL) represent extremely rare cardiac tumors which are accompanied by poor prognosis, unless they are timely diagnosed and treated. Case presentation Herein we present a 28-year-old, immunocompetent man who presented to our hospital due to progressively worsening symptoms and signs of superior vena cava syndrome. Multi-modality imaging demonstrated a large intracardiac tumor, which was proven, by biopsy, to be a PCL. The patient received targeted chemotherapy which led to total remission of his disease, with no relapse over a 15-month follow-up period. Conclusions Although PCLs are rare, they should always be kept in mind in the differential diagnosis of cardiac tumors. Timely diagnosis of PCLs and appropriate chemotherapy, alone or in combination with radiotherapy, seems to provide the best results.

Background: Augmentation index (AIx) serves as an independent predictor of cardiovascular events and mortality. It is often clinically used as a measure of arterial stiffness and reflections, a practice that was recently challenged. In this work, we sought to investigate the association between AIx and wave reflections in patients with severe aortic stenosis. Methods: The final cohort comprised of 18 subjects (86 ± 4 years, 28% males) with severe aortic stenosis who underwent a Transcatheter Aortic Valve Implantation (TAVI). Invasive aortic and left ventricular pressure waveforms were obtained simultaneously before and after the TAVI. Key pressure waveform features were extracted and wave separation analysis was performed by combining aortic pressure and flow (Doppler echocardiography) data before and after the valve replacement. Results: After the TAVI, aortic SBP and maximal blood flow velocity increased (from 134 ± 22 to 151 ± 30 mmHg, p = 0.04 and from 298 ± 65 to 376 ± 81 mL/sec, p < 0.01 respectively), while the ejection duration decreased (from 0.42 ± 0.07 to 0.35 ± 0.06 sec, p < 0.01). A significant decrease in AIx and augmented pressure was noted (from 44 ± 16 to 17 ± 12% and from 34 ± 13 to 17 ± 14 mmHg respectively, p < 0.01 for all). Wave separation analysis however showed a comparable increase in both forward (from 61 ± 14 to 79 ± 22 mmHg, p < 0.01) and backward (from 31 ± 11 to 39 ± 12 mmHg, p < 0.05) wave amplitudes. Reflection magnitude and backward wave transit time remained unchanged (p = NS). Conclusion: Decrease in AIx after TAVI is not associated with changes in wave reflection amplitude or timing. For patients with severe aortic stenosis, interpretation of AIx should be done with caution.

Abstract Background Cornu aspersum is a quite intriguing species from the point of view of ecology and evolution and its potential use in medical and environmental applications. It is a species of economic importance since it is farmed and used for culinary purposes. However, the genomic tools that would allow a thorough insight into the ecology, evolution, nutritional and medical properties of this highly adaptable organism, are missing. In this work, using next-generation sequencing (NGS) techniques we assessed a significant portion of the transcriptome of this non-model organism. Results Out of the 9445 de novo assembled contigs, 2886 (30.6%) returned significant hits and for 2261 (24%) of them Gene Ontology (GO) terms associated to the hits were retrieved. A high percentage of the contigs (69.4%) produced no BLASTx hits. The GO terms were grouped to reflect biological processes, molecular functions and cellular components. Certain GO terms were dominant in all groups. After scanning the assembled transcriptome for microsatellites (simple sequence repeats, SSRs), a total of 563 SSRs were recovered. Among the identified SSRs, trinucleotide repeats were the predominant followed by tetranucleotide and dinucleotide repeats. Conclusion The annotation success of the transcriptome of C. aspersum was relatively low. This is probably due to the very limited number of annotated reference genomes existing for mollusc species, especially terrestrial ones. Several biological processes being active in the aestivating species were revealed through the association of the transcripts to enzymes relating to the pathways. The genomic tools provided herein will eventually aid in the study of the global genomic diversity of the species and the investigation of aspects of the ecology, evolution, behavior, nutritional and medical properties of this highly adaptable organism.

Background and Aims: Arterial volume compliance is a major determinant of cardiac afterload. More than 50% of the arterial volume compliance resides in the proximal aorta. Researchers often use area compliance for the estimation of volume compliance, assuming an invariant vessel length over pressure changes. Recent studies have provided evidence to question this simplification, showing that the extension of the proximal aorta along its long axis during systole produces significant longitudinal strain, which could lead to erroneous estimation of arterial stiffness. The aim of the present study was to test this hypothesis in a computational environment. Methods: The 3-D proximal aortic geometry of a healthy young male was reconstructed and meshed and the original zero-pressure geometry was restored. Material behavior was approximated based on the model of (1). Viscoelastic support conditions were introduced along the aortic wall and aortic root motion, estimated from the cardiacgated CT data of a healthy subject, was enforced at the proximal boundary. The simulation was run for an input pressure ranging from 80-110mmHg. Volume compliance of the vessel as obtained by integrating the area compliance over the centerline length (both variable and invariable) was subsequently compared to the ground truth (which was imposed by the material stiffness). Results: Integration of the area compliance over an invariable centerline length led to an underestimated average distensibility by −68%. After taking into account the elongation, the error was improved to −25% (Figure 1). Conclusion: The elongation of the aorta during cardiac cycle was found to affect significantly the estimation of arterial compliance.