The next horizon in precision oncology: Proteogenomics to inform cancer diagnosis and treatment.

When it comes to precision oncology, proteogenomics may provide better prospects to the clinical characterization of tumors, help make a more accurate diagnosis of cancer, and improve treatment for patients with cancer. This perspective describes the significant contributions of The Cancer Genome Atlas and the Clinical Proteomic Tumor Analysis Consortium to precision oncology and makes the case that proteogenomics needs to be fully integrated into clinical trials and patient care in order for precision oncology to deliver the right cancer treatment to the right patient at the right dose and at the right time.

Origins and molecular effects of hypoxia in cancer.

Hypoxia (insufficient O2) is a pivotal factor in cancer progression, triggering genetic, transcriptional, translational and epigenetic adaptations associated to therapy resistance, metastasis and patient mortality. In this review, we outline the microenvironmental origins and molecular mechanisms responsible for hypoxic cancer cell adaptations in situ and in vitro, whilst outlining current approaches to stratify, quantify and therapeutically target hypoxia in the context of precision oncology.

Spectrum of LYST mutations in Chediak-Higashi syndrome: a report of novel variants and a comprehensive review of the literature.

INTRODUCTION: Chediak-Higashi syndrome (CHS) is a rare autosomal recessive disorder characterised by partial oculocutaneous albinism, a bleeding diathesis, immunological dysfunction and neurological impairment. Bi-allelic loss-of-function variants in LYST cause CHS. LYST encodes the lysosomal trafficking regulator, a highly conserved 429 kDa cytoplasmic protein with an unknown function. METHODS: To further our understanding of the pathogenesis of CHS, we conducted clinical evaluations on individuals with CHS enrolled in our natural history study. Using genomic DNA Sanger sequencing, we identified novel pathogenic LYST variants. Additionally, we performed an extensive literature review to curate reported LYST variants and classified these novel and reported variants according to the American College of Medical Genetics/Association for Molecular Pathology variant interpretation guidelines. RESULTS: Our investigation unveiled 11 novel pathogenic LYST variants in eight patients with a clinical diagnosis of CHS, substantiated by the presence of pathognomonic giant intracellular granules. From these novel variants, together with a comprehensive review of the literature, we compiled a total of 147 variants in LYST, including 61 frameshift variants (41%), 44 nonsense variants (30%), 23 missense variants (16%), 13 splice site variants or small genomic deletions for which the coding effect is unknown (9%), 5 in-frame variants (3%) and 1 start-loss variant (1%). Notably, a genotype-phenotype correlation emerged, whereby individuals harbouring at least one missense or in-frame variant generally resulted in milder disease, while those with two nonsense or frameshift variants generally had more severe disease. CONCLUSION: The identification of novel pathogenic LYST variants and improvements in variant classification will provide earlier diagnoses and improved care to individuals with CHS.

Variant characterisation and clinical profile in a large cohort of patients with Ellis-van Creveld syndrome and a family with Weyers acrofacial dysostosis.

BACKGROUND: Ellis-van Creveld syndrome (EvC) is a recessive disorder characterised by acromesomelic limb shortening, postaxial polydactyly, nail-teeth dysplasia and congenital cardiac defects, primarily caused by pathogenic variants in EVC or EVC2. Weyers acrofacial dysostosis (WAD) is an ultra-rare dominant condition allelic to EvC. The present work aimed to enhance current knowledge on the clinical manifestations of EvC and WAD and broaden their mutational spectrum. METHODS: We conducted molecular studies in 46 individuals from 43 unrelated families with a preliminary clinical diagnosis of EvC and 3 affected individuals from a family with WAD and retrospectively analysed clinical data. The deleterious effect of selected variants of uncertain significance was evaluated by cellular assays. MAIN RESULTS: We identified pathogenic variants in EVC/EVC2 in affected individuals from 41 of the 43 families with EvC. Patients from each of the two remaining families were found with a homozygous splicing variant in WDR35 and a de novo heterozygous frameshift variant in GLI3, respectively. The phenotype of these patients showed a remarkable overlap with EvC. A novel EVC2 C-terminal truncating variant was identified in the family with WAD. Deep phenotyping of the cohort recapitulated 'classical EvC findings' in the literature and highlighted findings previously undescribed or rarely described as part of EvC. CONCLUSIONS: This study presents the largest cohort of living patients with EvC to date, contributing to better understanding of the full clinical spectrum of EvC. We also provide comprehensive information on the EVC/EVC2 mutational landscape and add GLI3 to the list of genes associated with EvC-like phenotypes.

Exploring the current landscape of single-cell RNA sequencing applications in gastric cancer research.

Gastric cancer (GC) represents a major global health burden and is responsible for a significant number of cancer-related fatalities. Its complex nature, characterized by heterogeneity and aggressive behaviour, poses considerable challenges for effective diagnosis and treatment. Single-cell RNA sequencing (scRNA-seq) has emerged as an important technique, offering unprecedented precision and depth in gene expression profiling at the cellular level. By facilitating the identification of distinct cell populations, rare cells and dynamic transcriptional changes within GC, scRNA-seq has yielded valuable insights into tumour progression and potential therapeutic targets. Moreover, this technology has significantly improved our comprehension of the tumour microenvironment (TME) and its intricate interplay with immune cells, thereby opening avenues for targeted therapeutic strategies. Nonetheless, certain obstacles, including tumour heterogeneity and technical limitations, persist in the field. Current endeavours are dedicated to refining protocols and computational tools to surmount these challenges. In this narrative review, we explore the significance of scRNA-seq in GC, emphasizing its advantages, challenges and potential applications in unravelling tumour heterogeneity and identifying promising therapeutic targets. Additionally, we discuss recent developments, ongoing efforts to overcome these challenges, and future prospects. Although further enhancements are required, scRNA-seq has already provided valuable insights into GC and holds promise for advancing biomedical research and clinical practice.

Sudden Cardiac Death in the Young: State-of-the-Art Review in Molecular Autopsy.

Sudden cardiac death (SCD) is defined as unexpected death due to a cardiac cause that occurs rapidly. Despite the identification of prevention strategies, SCD remains a serious public health problem worldwide, accounting for 15-20% of all deaths, and is therefore a challenge for modern medicine, especially when it affects young people. Sudden cardiac death in young people affects the population aged ≤ 35 years, including athletes and non-athletes, and it is due to various hereditary and non-hereditary causes. After an autopsy, if the cause remains unknown, it is called sudden unexplained death, often attributable to genetic causes. In these cases, molecular autopsy-post-mortem genetic testing-is essential to facilitate diagnostic and therapeutic pathways and/or the monitoring of family members of the cases. This review aims to elaborate on cardiac disorders marked by genetic mutations, necessitating the post-mortem genetic investigation of the deceased for an accurate diagnosis in order to facilitate informed genetic counseling and to implement preventive strategies for family members of the cases.

Unraveling the role of HIF-1α in sepsis: from pathophysiology to potential therapeutics-a narrative review.

Sepsis is characterized by organ dysfunction resulting from a dysregulated inflammatory response triggered by infection, involving multifactorial and intricate molecular mechanisms. Hypoxia-inducible factor-1α (HIF-1α), a notable transcription factor, assumes a pivotal role in the onset and progression of sepsis. This review aims to furnish a comprehensive overview of HIF-1α's mechanism of action in sepsis, scrutinizing its involvement in inflammatory regulation, hypoxia adaptation, immune response, and organ dysfunction. The review encompasses an analysis of the structural features, regulatory activation, and downstream signaling pathways of HIF-1α, alongside its mechanism of action in the pathophysiological processes of sepsis. Furthermore, it will delve into the roles of HIF-1α in modulating the inflammatory response, including its association with inflammatory mediators, immune cell activation, and vasodilation. Additionally, attention will be directed toward the regulatory function of HIF-1α in hypoxic environments and its linkage with intracellular signaling, oxidative stress, and mitochondrial damage. Finally, the potential therapeutic value of HIF-1α as a targeted therapy and its significance in the clinical management of sepsis will be discussed, aiming to serve as a significant reference for an in-depth understanding of sepsis pathogenesis and potential therapeutic targets, as well as to establish a theoretical foundation for clinical applications.

Genotypes and phenotypes heterogeneity in PIK3CA-related overgrowth spectrum and overlapping conditions: 150 novel patients and systematic review of 1007 patients with PIK3CA pathogenetic variants.

BACKGROUND: Postzygotic activating PIK3CA variants cause several phenotypes within the PIK3CA-related overgrowth spectrum (PROS). Variant strength, mosaicism level, specific tissue involvement and overlapping disorders are responsible for disease heterogeneity. We explored these factors in 150 novel patients and in an expanded cohort of 1007 PIK3CA-mutated patients, analysing our new data with previous literature to give a comprehensive picture. METHODS: We performed ultradeep targeted next-generation sequencing (NGS) on DNA from skin biopsy, buccal swab or blood using a panel including phosphatidylinositol 3-kinase/AKT/mammalian target of rapamycin pathway genes and GNAQ, GNA11, RASA1 and TEK. Additionally, 914 patients previously reported were systematically reviewed. RESULTS: 93 of our 150 patients had PIK3CA pathogenetic variants. The merged PROS cohort showed that PIK3CA variants span thorough all gene domains, some were exclusively associated with specific PROS phenotypes: weakly activating variants were associated with central nervous system (CNS) involvement, and strongly activating variants with extra-CNS phenotypes. Among the 57 with a wild-type PIK3CA allele, 11 patients with overgrowth and vascular malformations overlapping PROS had variants in GNAQ, GNA11, RASA1 or TEK. CONCLUSION: We confirm that (1) molecular diagnostic yield increases when multiple tissues are tested and by enriching NGS panels with genes of overlapping 'vascular' phenotypes; (2) strongly activating PIK3CA variants are found in affected tissue, rarely in blood: conversely, weakly activating mutations more common in blood; (3) weakly activating variants correlate with CNS involvement, strong variants are more common in cases without; (4) patients with vascular malformations overlapping those of PROS can harbour variants in genes other than PIK3CA.

Different Strategies to Overcome Resistance to Proteasome Inhibitors-A Summary 20 Years after Their Introduction.

Proteasome inhibitors (PIs), bortezomib, carfilzomib, and ixazomib, are the first-line treatment for multiple myeloma (MM). They inhibit cytosolic protein degradation in cells, which leads to the accumulation of misfolded and malfunctioned proteins in the cytosol and endoplasmic reticulum, resulting in cell death. Despite being a breakthrough in MM therapy, malignant cells develop resistance to PIs via different mechanisms. Understanding these mechanisms drives research toward new anticancer agents to overcome PI resistance. In this review, we summarize the mechanism of action of PIs and how MM cells adapt to these drugs to develop resistance. Finally, we explore these mechanisms to present strategies to interfere with PI resistance. The strategies include new inhibitors of the ubiquitin-proteasome system, drug efflux inhibitors, autophagy disruption, targeting stress response mechanisms, affecting survival and cell cycle regulators, bone marrow microenvironment modulation, and immunotherapy. We list potential pharmacological targets examined in in vitro, in vivo, and clinical studies. Some of these strategies have already provided clinicians with new anti-MM medications, such as panobinostat and selinexor. We hope that further exploration of the subject will broaden the range of therapeutic options and improve patient outcomes.

Next generation sequencing (NGS)-based approach to diagnosing Algerian patients with suspected inborn errors of immunity (IEIs).

The advent of next-generation sequencing (NGS) technologies has greatly expanded our understanding of both the clinical spectra and genetic landscape of inborn errors of immunity (IEIs). Endogamous populations may be enriched for unique, ancestry-specific disease-causing variants, a consideration that significantly impacts molecular testing and analysis strategies. Herein, we report on the application of a 2-step NGS-based testing approach beginning with targeted gene panels (TGPs) tailored to specific IEI subtypes and reflexing to whole exome sequencing (WES) if negative for Northwest Algerian patients with suspected IEIs. Our overall diagnostic yield of 57% is comparable to others broadly applying short-read NGS to IEI detection, but data from our localized cohort show some similarities and differences from NGS studies performed on larger regional IEI cohorts. This suggests the importance of tailoring diagnostic strategies to local demographics and needs, but also highlights ongoing concerns inherent to the application of genomics for clinical IEI diagnostics.

Heterozygous gain of function variants in a critical region of RNF13 cause congenital microcephaly, epileptic encephalopathy, blindness, and failure to thrive.

Missense variants in the RNF13 gene have been previously known to cause congenital microcephaly, epileptic encephalopathy, blindness, and failure to thrive through a gain-of-function disease mechanism. Here, we identify a nonsense variant, expected to result in protein truncation, in a similarly affected patient. We show that this nonsense variant, residing in the terminal exon, is likely to escape nonsense-mediated decay while removing a critical region for protein function, thus resulting in a gain-of-function effect. We review the literature and disease databases and identify several other affected individuals with overlapping phenotypes carrying distinct truncating variants in the terminal exon upstream of the putative critical region. Furthermore, we analyze truncating variants from the general population, namely, the Genome Aggregation Database (gnomAD), and provide additional evidence supporting our hypothesis, and ruling out haploinsufficiency as an alternative disease mechanism. In summary, our case report, literature review, and analysis of disease and population databases strongly support the hypothesis that heterozygous gain-of-function variants in a critical region of RNF13 cause congenital microcephaly, epileptic encephalopathy, blindness, and failure to thrive.

MELAS-associated m.5541C>T mutation caused instability of mitochondrial tRNATrp and remarkable mitochondrial dysfunction.

BACKGROUND: Mitochondrial encephalomyopathy with lactic acidosis and stroke-like episode (MELAS) is a group of genetic diseases caused by mutations in mitochondrial DNA and nuclear DNA. The causative mutations of MELAS have drawn much attention, among them, mutations in mitochondrial tRNA genes possessing prominent status. However, the detailed molecular pathogenesis of these tRNA gene mutations remains unclear and there are very few effective therapies available to date. METHODS: We performed muscle histochemistry, genetic analysis, molecular dynamic stimulation and measurement of oxygen consumption rate and respiratory chain complex activities to demonstrate the molecular pathomechanisms of m.5541C>T mutation. Moreover, we use cybrid cells to investigate the potential of taurine to rescue mitochondrial dysfunction caused by this mutation. RESULTS: We found a pathogenic m.5541C>T mutation in the tRNATrp gene in a large MELAS family. This mutation first affected the maturation and stability of tRNATrp and impaired mitochondrial respiratory chain complex activities, followed by remarkable mitochondrial dysfunction. Surprisingly, we identified that the supplementation of taurine almost completely restored mitochondrial tRNATrp levels and mitochondrial respiration deficiency at the in vitro cell level. CONCLUSION: The m.5541C>T mutation disturbed the translation machinery of mitochondrial tRNATrp and taurine supplementation may be a potential treatment for patients with m.5541C>T mutation. Further studies are needed to explore the full potential of taurine supplementation as therapy for patients with this mutation.

Biallelic cGMP-dependent type II protein kinase gene (PRKG2) variants cause a novel acromesomelic dysplasia.

BACKGROUND: C-type natriuretic peptide (CNP), its endogenous receptor, natriuretic peptide receptor-B (NPR-B), as well as its downstream mediator, cyclic guanosine monophosphate (cGMP) dependent protein kinase II (cGKII), have been shown to play a pivotal role in chondrogenic differentiation and endochondral bone growth. In humans, biallelic variants in NPR2, encoding NPR-B, cause acromesomelic dysplasia, type Maroteaux, while heterozygous variants in NPR2 (natriuretic peptide receptor 2) and NPPC (natriuretic peptide precursor C), encoding CNP, cause milder phenotypes. In contrast, no variants in cGKII, encoded by the protein kinase cGMP-dependent type II gene (PRKG2), have been reported in humans to date, although its role in longitudinal growth has been clearly demonstrated in several animal models. METHODS: Exome sequencing was performed in two girls with severe short stature due to acromesomelic limb shortening, brachydactyly, mild to moderate platyspondyly and progressively increasing metaphyseal alterations of the long bones. Functional characterisation was undertaken for the identified variants. RESULTS: Two homozygous PRKG2 variants, a nonsense and a frameshift, were identified. The mutant transcripts are exposed to nonsense-mediated decay and the truncated mutant cGKII proteins, partially or completely lacking the kinase domain, alter the downstream mitogen activation protein kinase signalling pathway by failing to phosphorylate c-Raf 1 at Ser43 and subsequently reduce ERK1/2 activation in response to fibroblast growth factor 2. They also downregulate COL10A1 and upregulate COL2A1 expression through SOX9. CONCLUSION: In conclusion, we have clinically and molecularly characterised a new acromesomelic dysplasia, acromesomelic dysplasia, PRKG2 type (AMDP).

An Exploratory Study Using Next-Generation Sequencing to Identify Prothrombotic Variants in Patients with Cerebral Vein Thrombosis.

Prothrombotic hereditary risk factors for cerebral vein thrombosis (CVT) are of clinical interest to better understand the underlying pathophysiology and stratify patients for the risk of recurrence. This study explores prothrombotic risk factors in CVT patients. An initial screening in patients of the outpatient clinic of the Department of Transfusion Medicine and Hemostaseology of the University Hospital Erlangen, Germany, revealed 183 patients with a history of CVT. An initial screening identified a number of common prothrombic risk factors, including Factor V Leiden (rs6025) and Prothrombin G20210A (rs1799963). All patients without relevant findings (58 individuals) were invited to participate in a subsequent genetic analysis of 55 relevant genes using next-generation sequencing (NGS). Three intron variants (ADAMTS13: rs28446901, FN1: rs56380797, rs35343655) were identified to occur with a significantly higher frequency in the CVT patient cohort compared to the general European population. Furthermore, the combined prevalence of at least two of four potentially prothrombic variants (FGA (rs6050), F13A1 (rs5985), ITGB3 (rs5918), and PROCR (rs867186)) was significantly higher in the CVT subjects. The possible impact of the identified variants on CVT is discussed.

The liver in sepsis: molecular mechanism of liver failure and their potential for clinical translation.

Liver failure is a life-threatening complication of infections restricting the host's response to infection. The pivotal role of the liver in metabolic, synthetic, and immunological pathways enforces limits the host's ability to control the immune response appropriately, making it vulnerable to ineffective pathogen resistance and tissue damage. Deregulated networks of liver diseases are gradually uncovered by high-throughput, single-cell resolved OMICS technologies visualizing an astonishing diversity of cell types and regulatory interaction driving tolerogenic signaling in health and inflammation in disease. Therefore, this review elucidates the effects of the dysregulated host response on the liver, consequences for the immune response, and possible avenues for personalized therapeutics.

Self-assembling Molecular Medicine for the Subacute Phase of Ischemic Stroke.

Ischemic stroke leads to acute neuron death and forms an injured core, triggering delayed cell death at the penumbra. The impaired brain functions after ischemic stroke are hardly recovered because of the limited regenerative properties. However, recent rodent intervention studies manipulating the extracellular environments at the subacute phase shed new light on the regenerative potency of the injured brain. This review introduces the rational design of artificial extracellular matrix (ECM) mimics using supramolecular peptidic scaffolds, which self-assemble via non-covalent bonds and form hydrogels. The facile customizability of the peptide structures allows tuning the hydrogels' physical and biochemical properties, such as charge states, hydrophobicity, cell adhesiveness, stiffness, and stimuli responses. Supramolecular peptidic materials can create safer and more economical drugs than polymer materials and cell transplantation. We also discuss the importance of activating developmental programs for the recovery at the subacute phase of ischemic stroke. Self-assembling molecular medicine mimicking the ECMs and activating developmental programs may stand as a new drug modality of regenerative medicine in various tissues.

The clinical and genetic features of hereditary pancreatitis in South Australia.

OBJECTIVE: To characterise the clinical phenotypes and genetic variants of hereditary pancreatitis in people diagnosed in South Australia. DESIGN, SETTING, PARTICIPANTS: Cross-sectional study of people who received molecular diagnoses of hereditary pancreatitis from one of four major diagnostic services in South Australia, 1 January 2006 - 30 June 2021. MAIN OUTCOME MEASURES: Genotypic and clinical features of people with hereditary pancreatitis, including age at onset, attack frequency, pain indices, use of opioid medications, and physical and mental health impact of hereditary pancreatitis. RESULTS: We identified 44 people from ten families who received molecular diagnoses of hereditary pancreatitis during 2006-21 (including 25 Indigenous people [57%] and 27 women [61%]): 36 with PRSS1, five with SPINK1, and three with PRSS1 and SPINK1 mutations (determined by whole exome sequencing). Symptom onset before the age of ten years was reported by 37 people (84%). Pancreatitis-related pain during the preceding four weeks was described as moderate or high by 35 people (79%); 38 people regularly used opioids (86%). Fifteen patients had diabetes mellitus (34%), and eight had undergone pancreatic surgery (18%). The estimated prevalence of hereditary pancreatitis was 1.1 (95% CI, 0.72-1.4) cases per 100 000 population for non-Indigenous and 71 (95% CI, 66-77) cases per 100 000 population for Indigenous South Australians. Among people with adult-onset chronic pancreatitis admitted to South Australian public hospitals during 2001-2019, the proportions of Indigenous people (12%) and women (38%) were smaller than we report for hereditary pancreatitis. CONCLUSION: The estimated prevalence of hereditary pancreatitis in South Australia is higher than in Europe. PRSS1 gene mutations are important causes, particularly among Indigenous young people.

Emerging roles of rare and low-frequency genetic variants in type 1 diabetes mellitus.

Type 1 diabetes mellitus (T1DM) is defined as an autoimmune disorder and has enormous complexity and heterogeneity. Although its precise pathogenic mechanisms are obscure, this disease is widely acknowledged to be precipitated by environmental factors in individuals with genetic susceptibility. To date, the known susceptibility loci, which have mostly been identified by genome-wide association studies, can explain 80%-85% of the heritability of T1DM. Researchers believe that at least a part of its missing genetic component is caused by undetected rare and low-frequency variants. Most common variants have only small to modest effect sizes, which increases the difficulty of dissecting their functions and restricts their potential clinical application. Intriguingly, many studies have indicated that rare and low-frequency variants have larger effect sizes and play more significant roles in susceptibility to common diseases, including T1DM, than common variants do. Therefore, better recognition of rare and low-frequency variants is beneficial for revealing the genetic architecture of T1DM and for providing new and potent therapeutic targets for this disease. Here, we will discuss existing challenges as well as the great significance of this field and review current knowledge of the contributions of rare and low-frequency variants to T1DM.

MicroRNA Modulation during Orthodontic Tooth Movement: A Promising Strategy for Novel Diagnostic and Personalized Therapeutic Interventions.

The Orthodontic Tooth Movement (OTM) is allowed through a mediated cell/tissue mechanism performed by applying a force or a pair of forces on the dental elements, and the tooth movement is a fundamental requirement during any orthodontic treatment. In this regard, it has been widely shown that each orthodontic treatment has a minimum duration required concerning numerous factors (age, patient compliance, type of technique used, etc.). In this regard, the aim of the following revision of the literature is to give readers a global vision of principal microRNAs (miRNAs) that are most frequently associated with OTM and their possible roles. Previously published studies of the last 15 years have been considered in the PubMed search using "OTM" and "miRNA" keywords for the present review article. In vitro and in vivo studies and clinical trials were mainly explored. Correlation between OTM and modulation of several miRNAs acting through post-transcriptional regulation on target genes was observed in the majority of previous studied. The expression analysis of miRNAs in biological samples, such as gingival crevicular fluid (GCF), can be considered a useful tool for novel diagnostic and/or prognostic approaches and for new personalized orthodontic treatments able to achieve a better clinical response rate. Although only a few studies have been published, the data obtained until now encourage further investigation of the role of miRNA modulation during orthodontic treatment. The aim of this study is to update the insights into the role and impact of principal micro-RNAs (miRNAs) that are most frequently associated during OTM.

Mechanisms of the FMR1 Repeat Instability: How Does the CGG Sequence Expand?

A dynamic mutation in exon 1 of the FMR1 gene causes Fragile X-related Disorders (FXDs), due to the expansion of an unstable CGG repeat sequence. Based on the CGG sequence size, two types of FMR1 alleles are possible: "premutation" (PM, with 56-200 CGGs) and "full mutation" (FM, with >200 triplets). Premutated females are at risk of transmitting a FM allele that, when methylated, epigenetically silences FMR1 and causes Fragile X syndrome (FXS), a very common form of inherited intellectual disability (ID). Expansions events of the CGG sequence are predominant over contractions and are responsible for meiotic and mitotic instability. The CGG repeat usually includes one or more AGG interspersed triplets that influence allele stability and the risk of transmitting FM to children through maternal meiosis. A unique mechanism responsible for repeat instability has not been identified, but several processes are under investigations using cellular and animal models. The formation of unusual secondary DNA structures at the expanded repeats are likely to occur and contribute to the CGG expansion. This review will focus on the current knowledge about CGG repeat instability addressing the CGG sequence expands.