Genetic Analysis Technology Research Articles

Genetic Analysis of Cerebrovascular Diseases

With advances in genetic analysis technology, the genetic and molecular backgrounds of cerebrovascular diseases have become clearer. In moyamoya disease and intracranial artery stenosis, RNF213 p.Arg4810Lys has been identified as a disease susceptibility gene variant(germline variant), and various analyses have been conducted. PDGFRB mutations have been identified as characteristic somatic variants in cerebral aneurysms and are attracting attention. In addition, PIK3CA and MAP3K3 mutaions have been identified in cerebral cavernous malformations as somatic variants. Moreover, KRAS and BRAF mutations have been identified in arteriovenous malformations as somatic variants, respectively. Further studies are in progress. We reviewed the results of recent genetic analyses of cerebrovascular diseases, focusing particularly on genetic mutations.

Understanding the intrinsic biology of diffuse large B-cell lymphoma: recent advances and future prospects.

Diffuse large B-cell lymphoma (DLBCL) is the most common type of lymphoid tumor, and accounts for approximately 30-40% of non-Hodgkin lymphomas. Although the prognosis has significantly improved with the advent of rituximab combination chemotherapy in the early 2000s, recurrence still occurs in about 40% of cases. Even though chemotherapy with increased dose-intensity is used in recurrent cases, the prognosis of such patients remains poor. Thus, the development of personalized medicine, including molecular-targeted drugs, is required to improve the prognosis of DLBCL patients, and further understanding of the molecular pathogenesis of DLBCL is essential for this purpose. With recent advances in genetic analysis technology, unknown genetic abnormalities and gene expression patterns have been discovered, and based on these discoveries, progress is being made in elucidating and subdividing molecular pathologies. This article summarizes recent findings regarding molecular pathogenesis in DLBCL using transcriptome and genomics technologies, and outlines the path to personalized medicine.

Genetic Analyses of Primary Liver Cancer Cell Lines: Correspondence With Morphological Features of Original Tumors.

Advancements in genetic analysis technologies have led to establishment of molecular classifications systems for primary liver cancers. The correlation between pathological morphology and genetic mutations in hepatocellular carcinoma (HCC) is becoming increasingly evident. To construct appropriate experimental models, it is crucial to select cell lines based on their morphology and genetic mutations. In this study, we conducted comprehensive genetic analyses of primary liver cancer cell lines and examined their correlations with morphology. Thirteen primary liver cancer cell lines established in our Department were investigated. Eleven cell lines were HCC cell lines, whereas 2 were combined hepatocellular-cholangiocarcinoma (CHC) cell line characteristics. Whole exome sequencing and fusion gene analyses were conducted using a next generation sequencing platform. We also examined correlations between cell mutations and morphological findings and conducted experiments to clarify the association between morphological findings and genetic alterations. Mutations in TP53, HMCN1, PCLO, HYDIN, APOB, and EYS were found in 11, 5, 4, 4, 3, and 3 cell lines, respectively. CTNNB1 mutation was not identified in any cell line. The original tumor of four cell lines (KYN-1, KYN-2, KYN-3, and HAK-6) showed morphologically macrotrabecular massive patterns and these cell lines harbor TP53 mutations. Two cell lines (KYN-2 and KMCH-2) showed an extremely high tumor mutation burden. These two cell lines possess ultra-mutations associated with DNA repair and/or DNA polymerase. The study identified correlations between morphological findings and genetic mutations in several HCC cell lines. Cell lines with unique genetic mutations were found. This information will be a valuable tool for the selection of suitable experimental models in HCC research.

Impact of Genetic Variation on Human Disease: From Genomics to Phenotype

The impact of genetic variation on human disease is a multifaceted phenomenon involving genomics, epigenetics, and phenotype relationships. Advances in human genetic analysis technology have revolutionized our understanding of genetic disorders, enabling large-scale screens to explore associations between sequence variants and disease susceptibility. Environmental stimuli also contribute to epigenetic modifications that shape gene expression and phenotypic traits. Recent research on monozygotic twins highlights the role of non-genetic factors in phenotypic differences, underscoring the complex interplay between genetics and environment. The literature review highlights seminal studies that elucidate genetic and evolutionary mechanisms underlying human disease, providing insights for personalized medicine. Methods such as pharmacogenetics and immunomics provide avenues for precision therapies and immunological research. The Heterogeneity Weighted U test and the Gaussian random effects model facilitate the exploration of phenotype-genotype relationships and help to assess genetic heterogeneity in disease models. Simulation experiments deepen our understanding of the impact of genetic variation on disease manifestation. The interdisciplinary approaches are crucial to unravel the complex genetic landscape of human diseases and to develop tailored therapeutic interventions.

Pathobiology and its clinical relevance in diffuse large B-cell lymphoma

Diffuse large B-cell lymphoma (DLBCL) is a clinically and biologically heterogeneous disease. Remarkable effort has been exerted in the classification of DLBCL and the development of its corresponding treatment. The prognosis of patients with DLBCL receiving rituximab combination chemotherapy significantly improved in the early 2000s. However, approximately 40% of patients still develop recurrence, and the prognosis of these patients is extremely poor. Recently, the use of polatuzumab vedotin and CAR T-cell therapies has improved patient prognosis. However, it is extremely important to identify the patient group who can benefit from the efficacy of these treatments. In relation to this, the molecular pathogenesis of DLBCL should be further evaluated. Recent advancements in the genetic analysis technology have led to the discovery of unknown genetic abnormalities and gene expression patterns. The elucidation and subdivision of the molecular pathology based on these findings will be the foundation of future personalized medicine.

Association of Familial Fanconi Syndrome with a Novel GATM Variant.

Fanconi syndrome is a disorder of the proximal renal tubule. Recently, advanced genetic analysis technology has revealed that several genes cause familial Fanconi syndrome. We identified a family with autosomal dominant Fanconi syndrome and chronic kidney disease with a novel glycine amidinotransferase (GATM) variant. Case 1 was a 57-year-old Japanese woman. Her father and two siblings had Fanconi syndrome or chronic kidney disease. She presented to our hospital at the age of 34 years with recurrent glucosuria. Her height and weight were 151 cm and 46.6 kg, respectively. Laboratory tests showed glucosuria, hypophosphatemia, hypouricemia, and normal renal function. Her serum creatinine level gradually increased over the following next two decades, and she developed end-stage renal disease. Case 2, the daughter of Case 1, was a 26-year-old woman. Her height and weight were 151 cm and 37.5 kg, respectively. Glucosuria was detected at the age of 13 years, which led to a referral to our hospital. Urinalysis showed low-molecular-weight proteinuria. She was diagnosed with Fanconi syndrome. At the age of 26 years, she had glucosuria, low-molecular-weight proteinuria, hypouricemia, and normal renal function. Genetic testing of both cases revealed a novel missense variant in GATM. The heterozygous missense variants in GATM have been reported to cause familial Fanconi syndrome, which manifests early in life and progresses to renal glomerular failure by mid-adulthood. The novel GATM variant detected in our cases was suspected to be associated with the development of Fanconi syndrome. GATM variants should be tested in patients with idiopathic Fanconi syndrome.

Development of a clinical microarray system for genetic analysis screening

ObjectivesResearch on the relationship between diseases and genes and the advancement of genetic analysis technologies have made genetic testing in medical care possible. There are various methods for genetic testing, including PCR-based methods and next-generation sequencing; however, screening tests in clinical laboratories are becoming more diverse; therefore, novel measurement systems and equipment are required to meet the needs of each situation. In this study, we aimed to develop a novel microarray-based genetic analysis system that uses a Peltier element to overcome the issues of conventional microarrays, such as the long measurement time and high cost. MethodsWe constructed a microarray system to detect the UDP-glucuronosyltransferase gene polymorphisms UGT1A1*6 and UGT1A1*28 in patients eligible for irinotecan hydrochloride treatment for use in clinical laboratories. To evaluate the performance of the system, the hybridization temperature and reaction time were determined, and the results were compared with those obtained using a conventional hybridization oven. ResultsThe hybridization temperature reached its target in 1/27th of the time required by the conventional system. We assessed 111 human clinical samples and found that our results agreed with those obtained using existing methods. The total time for the newly developed device was reduced by 85 min compared to that for existing methods, as the automated DNA microarray eliminates the time that existing methods spend on manual operation. ConclusionsThe surface treatment technology used in our system enables high-density and strong DNA fixation, allowing the construction of a measurement system suitable for clinical applications.

DNA Methylation and Epigenetic Events Underlying Renal Cell Carcinomas.

Renal cell carcinoma (RCC) refers to a group of tumors that develop from the epithelium of the kidney tubes, including clear cell RCC, papillary RCC, and chromophobe RCC. Most clear cell renal carcinomas have a large histologic subtype, genetic or epigenetic von Hippel-Lindau (VHL). A comprehensive analysis of the genetic modification genome suggested that chromosome 3p loss and chromosome gains 5q and 7 may be significant copy defects in the development of clear RCC. A more potent RCC may develop if chromosome 1p, 4, 9, 13q, or 14q is also lost. Renal carcinogenesis is not associated with chronic inflammation or histological changes. However,ifregional hypermethylation of DNA in CpG C-type islandshas alreadyaccumulated in cancer-free kidney tissue,it impliesthat the presence of malignant kidney lesions may also be detected by modified DNA methylation. Modification of DNA methylation in cancerous kidney tissue may advance kidney tissue to epigenetic mutations and genes, leading to more serious cancers and even determining a patient's outcome. The genetic and epigenetic profile provides accurate predictors for patients with kidney cancer. New genetic and epigenetic analysis technologies will help to speed up the identification of vital cells for kidney cancer prevention, diagnosis, and treatment.

Genetics and Epigenetics in Personalized Nutrition: Evidence, Expectations, and Experiences.

With the presentation of the blueprint of the first human genome in 2001 and the advent of technologies for high-throughput genetic analysis, personalized nutrition (PN) becomes a new scientific field and the first commercial offerings of genotype-based nutrition advice emerge at the same time. Here, the state of evidence for the effect of genetic and epigenetic factors in the development of obesity, the metabolic syndrome, and resulting illnesses such as non-insulin-dependent diabetes mellitus and cardiovascular diseases is summarized. This study also critically value the concepts of PN that are built around the new genetic avenue from both the academic and a commercial perspective and their effectiveness in causing sustained changes in diet, lifestyle, and for improving health. Despite almost 20 years of research and commercial direct-to-consumer offerings, evidence for the success of gene-based dietary recommendations is still generally lacking. This calls for new concepts of future PN solutions that incorporate more phenotypic measures and provide a panel of instruments (e.g., self- and bio-monitoring tools, feedback systems, algorithms based on artificial intelligence) that increases compliance based on the individual´s physical and social environment and value system.

Application of Patient-Derived Cancer Organoids to Personalized Medicine.

Cell models are indispensable for the research and development of cancer therapies. Cancer medications have evolved with the establishment of various cell models. Patient-derived cell lines are very useful for identifying characteristic phenotypes and susceptibilities to anticancer drugs as well as molecularly targeted therapies for tumors. However, conventional 2-dimensional (2D) cell cultures have several drawbacks in terms of engraftment rate and phenotypic changes during culture. The organoid is a recently developed in vitro model with cultured cells that form a three-dimensional structure in the extracellular matrix. Organoids have the capacity to self-renew and can organize themselves to resemble the original organ or tumor in terms of both structure and function. Patient-derived cancer organoids are more suitable for the investigation of cancer biology and clinical medicine than conventional 2D cell lines or patient-derived xenografts. With recent advances in genetic analysis technology, the genetic information of various tumors has been clarified, and personalized medicine based on genetic information has become clinically available. Here, we have reviewed the recent advances in the development and application of patient-derived cancer organoids in cancer biology studies and personalized medicine. We have focused on the potential of organoids as a platform for the identification and development of novel targeted medicines for pancreatobiliary cancer, which is the most intractable cancer.

“Rapid DNA” ― new horizons of forensic DNA profiling using full cycle genetic analyzers

BACKGROUND: The study is a part of a comprehensive research conducted at the Russian Center of Forensic Medical Expertise of the Ministry of Health of the Russian Federation with the ultimate goal of development and implementation of new effective technologies for molecular genetic identity testing and kinship analysis in the realm of national forensic expertise.
 AIMS: The objective of the study is to evaluate the possibilities of the Rapid DNA technology in application to forensic DNA analysis. One may expect that this technology in the future will ensure the creation of an operational and highly effective procedure for forensic DNA profiling.
 MATERIAL AND METHODS: The methodical basis of the work is the genotyping of polymorphic STR-loci of chromosomal DNA in biological material using high-tech fully automated (full cycle) analytical platform RapidHIT: namely, RapidHIT 200 (Integenx, USA) and RapidHIT-ID (Life Technologies/Thermo Fisher, IntegenX (USA) instruments.
 RESULTS: Buccal epithelium and blood samples, while being the most common in a forensic DNA testing, have been examined as a validation test objects. Then experiments were carried out with real expert material such as cigarette butts, hair, nails, bones, chewing gum; swabs from different surfaces were examined as test samples as well.
 CONCLUSIONS: Based on the data obtained, it can be stated that the Rapid DNA technology as it is implemented in the full-cycle genetic analyzers IntegenX RapidHIT 200 (Integenx, USA) and RapidHIT ID (Thermo Fisher, IntegenX, USA), according to the criteria studied, in general supports the idea of to be introduced into the practice of forensic DNA analysis.

Rice contains a biosynthetic gene cluster associated with production of the casbane-type diterpenoid phytoalexin ent-10-oxodepressin.

Diterpenoids play important roles in rice microbial disease resistance as phytoalexins, as well as acting in allelopathy and abiotic stress responses. Recently, the casbane-type phytoalexin ent-10-oxodepressin was identified in rice, but its biosynthesis has not yet been elucidated. Here ent-10-oxodepressin biosynthesis was investigated via co-expression analysis and biochemical characterisation, with use of the CRISPR/Cas9 technology for genetic analysis. The results identified a biosynthetic gene cluster (BGC) on rice chromosome 7 (c7BGC), containing the relevant ent-casbene synthase (OsECBS), and four cytochrome P450 (CYP) genes from the CYP71Z subfamily. Three of these CYPs were shown to act on ent-casbene, with CYP71Z2 able to produce a keto group at carbon-5 (C5), while the closely related paralogues CYP71Z21 and CYP71Z22 both readily produce a keto group at C10. Together these C5 and C10 oxidases can elaborate ent-casbene to ent-10-oxodepressin (5,10-diketo-ent-casbene). OsECBS knockout lines no longer produce casbane-type diterpenoids and exhibit impaired resistance to the rice fungal blast pathogen Magnaporthe oryzae. Elucidation of ent-10-oxodepressin biosynthesis and the associated c7BGC provides not only a potential target for molecular breeding, but also, gives the intriguing parallels to the independently assembled BGCs for casbene-derived diterpenoids in the Euphorbiaceae, further insight into plant BGC evolution, as discussed here.

Genomic Analysis for Citrus Disease Detection

Citrus is an important group of globally produced fruit crops, holding great economic, cultural, and health value. Belonging to the Rutacaeae family, the genus Citrus includes some of the most iconic and widely appreciated variants of fruits such as the orange, lemon, lime, grapefruit, and tangerine. The spread of various diseases threatens the worldwide production of citrus fruit crops. Diseases such as Asiatic citrus canker, citrus tristeza virus, citrus leprosis, and especially citrus greening disease (also known as Huanglongbing) cause various symptoms harmful to plant growth and fruit production, inflicting tremendous economic damages. Advancements in genetic analysis technologies have offered new tools to investigate the molecular mechanisms underlining these diseases. In this review, we briefly overview the utility of genetic analysis in detection and monitoring of citrus disease-causing pathogens. We then focus our discussion on one of the most damaging citrus diseases, citrus greening disease (Huanglongbing). Genomic and gene expression analysis of citrus plants and their disease-causing microbes, along with tissue and cellular structural imaging, has demonstrated the potential allowing early pathogen detection and disease monitoring. These will eventually help to develop effective treatments to protect and promote citrus crop production.

PRD1, a homologous recombination initiation factor, is involved in spindle assembly in rice meiosis.

The bipolar spindle structure in meiosis is essential for faithful chromosome segregation. PUTATIVE RECOMBINATION INITIATION DEFECT 1 (PRD1) previously has been shown to participate in the formation of DNA double strand breaks (DSBs). However, the role of PRD1 in meiotic spindle assembly has not been elucidated. Here, we reveal by both genetic analysis and immunostaining technology that PRD1 is involved in spindle assembly in rice (Oryza sativa) meiosis. We show that DSB formation and bipolar spindle assembly are disturbed in prd1 meiocytes. PRD1 signals display a dynamic pattern of localization from covering entire chromosomes at leptotene to congregating at the centromere region after leptotene. Centromeric localization of PRD1 signals depends on the organization of leptotene chromosomes, but not on DSB formation and axis establishment. PRD1 exhibits interaction and co-localization with several kinetochore components. We also find that bi-orientation of sister kinetochores within a univalent induced by mutation of REC8 can restore bipolarity in prd1. Furthermore, PRD1 directly interacts with REC8 and SGO1, suggesting that PRD1 may play a role in regulating the orientation of sister kinetochores. Taken together, we speculate that PRD1 promotes bipolar spindle assembly, presumably by modulating the orientation of sister kinetochores in rice meiosis.

High genetic similarity of Salmonella Enteritidis as a predominant serovar by an independent survey in 3 large-scale chicken farms in China

Salmonella Enteritidis (SE) are important zoonotic pathogens, and can be easily transferred to humans by contaminated animal products. Epidemic surveys of SE are necessary in current modern large-scale chicken farms. In this study, Salmonella strains were isolated from possibly infected samples collected at 3 independent farms, and their serotype, drug resistances, virulence genes, and genetic similarity were analyzed by molecular genetic analysis technologies including multilocus sequence typing (MLST), clustered regularly interspaced short palindromic repeats (CRISPR), pulsed-field gel electrophoresis (PFGE), and whole-genome sequencing (WGS). A total of 346 Salmonella strains were isolated from 3,598 samples (9.61%); 329 isolates were identified as SE (95.09%) and 308 isolates were multidrug resistant (93.62%). Virulotyping based on 6 virulence genes showed high similarity in SE isolates of each farm, with the exception of 2 isolates. All SE isolates were found to be the same ST11 type by MLST, and 22 strains of 150 SE isolates selected at random were found to belong to 1 cluster by PFGE and the same SET1 type by CRISPR. WGS results further revealed that these isolates belonged to the same clonal cluster, with high genetic similarity of 99.80 to 100.00%. All these results indicated that these SE isolates were overwhelmingly dominant and demonstrated high genetic similarity, which revealed that the same SE clone might be transmitted in these farms.

Технология генетической идентификации сортов и диких форм винограда на основе мультилокусного микросателлитного анализа

The genotyping technology has been developed based on 9 microsatellite loci (VVS2, VVMD5, VVMD7, VVMD27, VVMD28, VVMD25, VVMD32, VrZAG62, VrZAG79). It can be used for efficient, accurate and fast identification of grape varieties and forms (genus Vitis). The proposed approach includes a multiplex PCR of all loci followed by electrophoretic analysis of DNA fragments in one capillary of a genetic analyzer. The application of an additional length standard, an allelic ladder, consisting of all possible DNA fragments of the analyzed microsatellite loci, is one of the key features of the technology, which ensures the accuracy and reproducibility of the results. The advantage of the proposed technology is the ability to standardize and automate the procedure using 96-well plates, which opens up the possibility of conducting mass analyses. As a result of the study of varieties and forms of Vitis genus species, genetic passports were obtained, according to which a dendrogram was constructed, reflecting the genetic relationship of the studied samples. The developed technology makes it possible to distinguish varieties and wild forms of grapes; it can be used for their identification and determination of genetic distance between them, as well as for assessment of planting material and protection of breeders' rights. Vitis, grapes, microsatellites, SSR, genotyping, multiplex PCR, DNA fragment analysis The work was carried out within the framework of the state assignment on the topic "Development of New Technologies for Genetic Analysis of Forms of Agricultural Plants to Accelerate and Control the Selection Process" (project no. 0574-2019-0003).

Prognostic Impact of Cytogenetic Evolution on the Outcome of Allogeneic Stem Cell Transplantation in Patients with Acute Myeloid Leukemia in Nonremission: A Single-Institute Analysis of 212 Recipients.

Recent progress in genetic analysis technology has helped researchers understand the pathogenesis of acute myeloid leukemia (AML). Considering this progress, AML karyotype is still one of the most significant prognostic factors that provides risk-adapted treatment approaches. Karyotype changes during treatment have been observed at times, but their prognostic impact is sparse, especially on allogeneic stem cell transplantation (allo-SCT). Here, we retrospectively investigated the effect of chromosomal changes between diagnosis and pretransplantation on the prognosis of allo-SCT by analyzing the outcomes of 212 consecutive patients who underwent allo-SCT for the first time at Toranomon Hospital, Tokyo, Japan, between 2008 and 2018. Cytogenetic abnormalities at diagnosis and pretransplantation were categorized based on the 2017 European Leukemia Net risk stratification. Genetic abnormalities such as FLT3-ITD and NPM1 were not considered in this study due to lack of genetic information in most patients. We defined cytogenetic evolution as chromosomal changes classified from lower category to higher category. Seventeen patients (8%) had cytogenetic evolution between diagnosis and pretransplantation, and they showed a significantly worse relapse rate than those who were categorized in the intermediate group based on the karyotype at diagnosis (3-year confidence interval [CI] of relapse, 57.4% versus 24.9%; P < .01). In multivariate analysis, cytogenetic evolution before allo-SCT had a significant impact on the CI of relapse (hazard ratio [HR], 3.89; CI, 1.75 to 8.67; P < .01), as well as the high score of the hematopoietic cell transplantation-specific comorbidity index (HR, 0.54; CI, 0.31 to 0.94; P=.03), but had no significant impact on overall survival or nonrelapse mortality. These results indicate that cytogenetic evolution has a significant impact after allo-SCT and should be considered during AML treatment.

Abstract 774: Highly effective isolation of circulating tumor cells using lateral magnetophoretic technology for precise genetic analysis of prostate cancer

Abstract Introduction Prostate cancer is the second leading cancer among men in the world and known as the most frequently diagnosed cancer in more than half of countries with estimated 1.3 million new emerging cases. Because of low specificity of serum PSA screening for prostate cancer, it raised concerns in keep management of clinical prognosis and therapeutic decision and thereby, there are still needed precise real-time biomarkers. Many recent studies have reported that circulating tumor cells (CTCs) can be used as important biomarkers for prostate cancer at cellular and molecular levels. However, due to their extreme rarity in the circulatory system and the lack of technology to isolate them precisely, it is still difficult to isolate them with high purity and efficiency that can be applied for clinical purposes. In this study, we introduce a microfluidic technology using lateral magnetophoresis for isolating CTCs from blood of prostate cancer patients (n=62) with high purity and efficiency and demonstrate clinical usefulness of isolated CTCs at cellular and molecular levels. Materials and methods By the lateral magnetophoretic technology using immunomagnetic nanobeads, highly pure CTCs were isolated from 5 ml blood, drawn from patients with localized (n=25) and metastatic (n=37) prostate cancer. Then, prostate cancer related genes (AR, AR-V7, EpCAM, KRT-19, PSA and PSMA) were quantified by droplet digital PCR (ddPCR) method using mRNA in CTCs. Results and discussion CTCs were detected from all blood samples (n=62). In localized cases, the average number of isolated CTCs and purity were 6.58 per ml and 2.09%. In metastatic cases, the average number of isolated CTCs and purity were 27.55 per ml and 6.00%, higher than those of localized cases. Then, mRNA was extracted from isolated CTCs and used for detecting prostate cancer specific genes. Their detection rate and expression level were increased from localized to metastatic stages. AR was detected at a similar rate of 76.00% in both localized and mHSPC stages and increased to 100% in mCRPC stage. AR-V7 was detected only in metastatic stages, mainly in mCRPC stages. EpCAM showed the highest detection rate in all stages, such as 96.00% in localized, 88.89% in mHSPC and 100% in mCRPC. This result indicates that CTCs are isolated from all blood of patients. KRT-19 showed slightly lower detection rate than EpCAM, but tend to be similar to EpCAM expression for cancer stages. PSA and PSMA were expressed very high in metastatic stage. PSA expression level in metastatic stage was 198.28 times higher than that in localized stage. Interestingly, the expression of PSA mRNA measured from CTCs was linearly proportional to serum PSA protein level, which is a commonly used diagnostic biomarker for prostate cancer. Conclusions In all stages of prostate cancer patients, the proposed microfluidic technology using the lateral magnetophoresis is a high-performance method for isolating CTCs with high detection rate and purity that can be used for precision genetic analysis. Citation Format: Hyungseok Cho, Ki-Ho Han, Jae-Seung Chung. Highly effective isolation of circulating tumor cells using lateral magnetophoretic technology for precise genetic analysis of prostate cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 774.

Trends in the Development of Diagnostic Tools for Red Blood Cell-Related Diseases and Anemias.

In the recent years, the progress in genetic analysis and next-generation sequencing technologies have opened up exciting landscapes for diagnosis and study of molecular mechanisms, allowing the determination of a particular mutation for individual patients suffering from hereditary red blood cell-related diseases or anemia. However, the huge amount of data obtained makes the interpretation of the results and the identification of the pathogenetic variant responsible for the diseases sometime difficult. Moreover, there is increasing evidence that the same mutation can result in varying cellular properties and different symptoms of the disease. Even for the same patient, the phenotypic expression of the disorder can change over time. Therefore, on top of genetic analysis, there is a further request for functional tests that allow to confirm the pathogenicity of a molecular variant, possibly to predict prognosis and complications (e.g., vaso-occlusive pain crises or other thrombotic events) and, in the best case, to enable personalized theranostics (drug and/or dose) according to the disease state and progression. The mini-review will reflect recent and future directions in the development of diagnostic tools for red blood cell-related diseases and anemias. This includes point of care devices, new incarnations of well-known principles addressing physico-chemical properties, and interactions of red blood cells as well as high-tech screening equipment and mobile laboratories.

Genetic Landscape of Nonobstructive Azoospermia and New Perspectives for the Clinic.

Nonobstructive azoospermia (NOA) represents the most severe expression of male infertility, involving around 1% of the male population and 10% of infertile men. This condition is characterised by the inability of the testis to produce sperm cells, and it is considered to have an important genetic component. During the last two decades, different genetic anomalies, including microdeletions of the Y chromosome, karyotype defects, and missense mutations in genes involved in the reproductive function, have been described as the primary cause of NOA in many infertile men. However, these alterations only explain around 25% of azoospermic cases, with the remaining patients showing an idiopathic origin. Recent studies clearly suggest that the so-called idiopathic NOA has a complex aetiology with a polygenic inheritance, which may alter the spermatogenic process. Although we are far from a complete understanding of the molecular mechanisms underlying NOA, the use of the new technologies for genetic analysis has enabled a considerable increase in knowledge during the last years. In this review, we will provide a comprehensive and updated overview of the genetic basis of NOA, with a special focus on the possible application of the recent insights in clinical practice.