Amyotrophic Lateral Sclerosis (ALS), a progressive neurodegenerative disease, primarily impairs upper and lower motor neurons, leading to debilitating motor dysfunction and eventually respiratory failure, widely known as Lou Gehrig's disease. ALS presents with diverse symptomatology, including dysarthria, dysphagia, muscle atrophy, and hyperreflexia. The prevalence of ALS varies globally, with incidence rates ranging from 1.5 to 3.8 per 100,000 individuals, significantly affecting populations aged 45-80. A complex interplay of genetic and environmental factors underpins ALS pathogenesis. Key genetic contributors include mutations in chromosome 9 open reading frame 72 (C9ORF72), superoxide dismutase type 1 (SOD1), Fusedin sarcoma (FUS), and TAR DNA-binding protein (TARDBP) genes, accounting for a considerable fraction of both familial (fALS) and sporadic (sALS) cases. The disease mechanism encompasses aberrant protein folding, mitochondrial dysfunction, oxidative stress, excitotoxicity, and neuroinflammation, contributing to neuronal death. This review consolidates current insights into ALS's multifaceted etiology, highlighting the roles of environmental exposures (e.g., toxins, heavy metals) and their interaction with genetic predispositions. We emphasize the polygenic nature of ALS, where multiple genetic variations cumulatively influence disease susceptibility and progression. This aspect underscores the challenges in ALS diagnosis, which currently lacks specific biomarkers and relies on symptomatology and familial history. Therapeutic strategies for ALS, still in nascent stages, involve symptomatic management and experimental approaches targeting molecular pathways implicated in ALS pathology. Gene therapy, focusing on specific ALS mutations, and stem cell therapy emerge as promising avenues. However, effective treatments remain elusive, necessitating a deeper understanding of ALS's genetic architecture and the development of targeted therapies based on personalized medicine principles. This review aims to provide a comprehensive understanding of ALS, encouraging further research into its complex genetic underpinnings and the development of innovative, effective treatment modalities.

The current radiation protection reference standards on stochastic cancer risk, drafted by the International Committee on Radiation Protection, are mostly based on the Life Span Study (LSS), though sufficient epidemiological and basic research evidence is lacking. The relationship between low-dose ionizing radiation (LDIR) and cancer risk is currently modeled with linear non-threshold (LNT) models. However, with the widespread use of medical examinations, the demand for substantial evidence of cancer risk under LDIR and the establishment of a threshold has become more significant. In the first part of the review, we summarize pivotal research in epidemiology, which includes the LSS, medical radiation studies, and occupational and environmental exposure studies. We describe and discuss solid cancers and hematopoietic malignancies induced by LDIR separately, attempting to identify the consistency and differences in the research results, and offering suggestions for future research directions. In the second part, we review recent progress in the underlying biology of cancer associated with LDIR. Besides the obvious harmful effect of DNA damage, chromosome aberrations caused by LDIR, epigenetic regulation also requires attention due to their relationship with carcinogenic and genetic risk. The multistage carcinogenesis model of stem cells, along with the varying effects of radiation on different tumors, may challenge the LNT model. Related research of stem cells, mitochondria and omic biology also offers promising directions for future research in this field.

Advanced sequencing technologies (ASTs) have revolutionized the quantitation of cancer driver mutations (CDMs) as rare events, which has utility in clinical oncology, cancer research, and cancer risk assessment. This review focuses on studies that have used ASTs to characterize clonal expansion (CE) of cells carrying CDMs and to explicate the selective pressures that shape CE. Importantly, high-sensitivity ASTs have made possible the characterization of mutant clones and CE in histologically normal tissue samples, providing the means to investigate nascent tumor development. Some ASTs can identify mutant clones in a spatially defined context; others enable integration of mutant data with analyses of gene expression, thereby elaborating immune, inflammatory, metabolic, and/or stromal microenvironmental impacts on CE. As a whole, these studies make it clear that a startlingly large fraction of cells in histologically normal tissues carry CDMs, CDMs may confer a context-specific selective advantage leading to CE, and only a small fraction of cells carrying CDMs eventually result in neoplasia. These observations were integrated with available literature regarding the mechanisms underlying clonal selection to interpret how measurements of CDMs and CE can be interpreted as biomarkers of cancer risk. Given the stochastic nature of carcinogenesis, the potential functional latency of driver mutations, the complexity of potential mutational and microenvironmental interactions, and involvement of other types of genetic and epigenetic changes, it is concluded that CDM-based measurements should be viewed as probabilistic rather than deterministic biomarkers. Increasing inter-sample variability in CDM levels (as a consequence of CE) may be interpretable as a shift away from normal tissue homeostasis and an indication of increased future cancer risk, a process that may reflect normal aging or carcinogen exposure. Consequently, analyses of variability in levels of CDMs have the potential to bolster existing approaches for carcinogenicity testing.

Single nucleotide variants (SNVs) constitute the most frequent variants that cause human genetic diseases. Base editors (BEs) comprise a new generation of CRISPR-based technologies, which are considered to have a promising future for curing genetic diseases caused by SNVs as they enable the direct and irreversible correction of base mutations. Two of the early types of BEs, cytosine base editor (CBE) and adenine base editor (ABE), mediate C-to-T, T-to-C, A-to-G, and G-to-A base transition mutations. Together, these represent half of all the known disease-associated SNVs. However, the remaining transversion (i.e., purine-pyrimidine) mutations cannot be restored by direct deamination and so these require the replacement of the entire base. Recently, a variety of base transversion editors were developed and so these add to the currently available BEs enabling the correction of all types of point mutation. However, compared to the base transition editors (including CBEs and ABEs), base transversion editors are still in the early development stage. In this review, we describe the basics and advances of the various base transversion editors, highlight their limitations, and discuss their potential for treating human diseases.

The human genome is constantly threatened by endogenous and environmental DNA damaging agents that can induce a variety of chemically modified DNA lesions including 8-oxo-7,8-dihydroguanine (OG). Increasing evidence has indicated that OG is not only a biomarker for oxidative DNA damage but also a novel epigenetic-like modification involved in regulation of gene expression in mammalian cells. Here we summarize the recent progress in OG research focusing on the following points: (i) the mechanism of OG production in organisms and its biological consequences in cells, (ii) the accurate identification of OG in low-abundance genomes and complex biological backgrounds, (iii) the development of OG sequencing methods. These studies will be helpful for further understanding of the molecular mechanisms of OG-induced mutagenesis and its potential roles in human development and diseases such as cancer.

Micronuclei (MN) are a nuclear abnormality that occurs when chromosome fragments or whole chromosomes are not properly segregated during mitosis and consequently are excluded from the main nuclei and wrapped within nuclear membrane to form small nuclei. This maldistribution of genetic material leads to abnormal cellular genomes which may increase risk of developmental defects, cancers, and accelerated aging. Despite the potential importance of MN as biomarkers of genotoxicity, very little was known about the optimal way to measure MN in humans, the normal ranges of values of MN in healthy humans and the prospective association of MN with developmental and degenerative diseases prior to the 1980's. In the early 1980's two important methods to measure MN in humans were developed namely, the cytokinesis-block MN (CBMN) assay using peripheral blood lymphocytes and the Buccal MN assay that measures MN in epithelial cells from the oral mucosa. These discoveries greatly increased interest to use MN assays in human studies. In 1997 the Human Micronucleus (HUMN) project was founded to initiate an international collaboration to (i) harmonise and standardise the techniques used to perform the lymphocyte CBMN assay and the Buccal MN assay; (ii) establish and collate databases of MN frequency in human populations world-wide which also captured demographic, lifestyle and environmental genotoxin exposure data and (iii) use these data to identify the most important variables affecting MN frequency and to also determine whether MN predict disease risk. In this paper we briefly describe the achievements of the HUMN project during the period from the date of its foundation on 9th September 1997 until its 26th Anniversary in 2023, which included more than 200 publications and 23 workshops world-wide.

Breast cancer continues to be a major global health challenge, driving the need for effective therapeutic strategies. Cisplatin, a powerful chemotherapeutic agent, is widely used in breast cancer treatment. However, its effectiveness is often limited by systemic toxicity and the development of drug resistance. This review examines the molecular factors that influence cisplatin response and resistance, offering crucial insights for the scientific community. It highlights the significance of understanding cisplatin resistance's genetic and epigenetic contributors, which could lead to more personalized treatment approaches. Additionally, the review explores innovative strategies to counteract cisplatin resistance, including combination therapies, nanoparticle-based drug delivery systems, and targeted therapies. These approaches are under intensive investigation and promise to enhance breast cancer treatment outcomes. This comprehensive discussion is a valuable resource to advance breast cancer therapeutics and address the challenge of cisplatin resistance.

Mutation spectra and mutational signatures in cancerous and non-cancerous tissues can be identified by various established techniques of massively parallel sequencing (or next-generation sequencing) including whole-exome or whole-genome sequencing, and more recently by error-corrected/duplex sequencing. One rather underexplored area has been the genome-scale analysis of mutational signatures as markers of mutagenic exposures, and their impact on cancer driver events applied to formalin-fixed or alcohol-fixed paraffin embedded archived biospecimens. This review showcases successful applications of the next-generation sequencing methodologies in archived fixed tissues, including the delineation of the specific tissue fixation-related DNA damage manifesting as artifactual signatures, distinguishable from the true signatures that arise from biological mutagenic processes. Overall, we discuss and demonstrate how next-generation sequencing techniques applied to archived fixed biospecimens can enhance our understanding of cancer causes including mutagenic effects of extrinsic cancer risk agents, and the implications for prevention efforts aimed at reducing avoidable cancer-causing exposures.

Autism spectrum disorder (ASD) is a complex neurodevelopmental disorder (NDD) influenced by genetic, epigenetic, and environmental factors. Recent advancements in genomic analysis have shed light on numerous genes associated with ASD, highlighting the significant role of both common and rare genetic mutations, as well as copy number variations (CNVs), single nucleotide polymorphisms (SNPs) and unique de novo variants. These genetic variations disrupt neurodevelopmental pathways, contributing to the disorder's complexity. Notably, CNVs are present in 10 %-20 % of individuals with autism, with 3 %-7 % detectable through cytogenetic methods. While the role of submicroscopic CNVs in ASD has been recently studied, their association with genomic loci and genes has not been thoroughly explored. In this review, we focus on 47 CNV regions linked to ASD, encompassing 1632 genes, including protein-coding genes and long non-coding RNAs (lncRNAs), of which 659 show significant brain expression. Using a list of ASD-associated genes from SFARI, we detect 17 regions harboring at least one known ASD-related protein-coding gene. Of the remaining 30 regions, we identify 24 regions containing at least one protein-coding gene with brain-enriched expression and a nervous system phenotype in mouse mutants, and one lncRNA with both brain-enriched expression and upregulation in iPSC to neuron differentiation. This review not only expands our understanding of the genetic diversity associated with ASD but also underscores the potential of lncRNAs in contributing to its etiology. Additionally, the discovered CNVs will be a valuable resource for future diagnostic, therapeutic, and research endeavors aimed at prioritizing genetic variations in ASD.

Micronucleus (MN) assays with buccal cells are at present widely used to investigate occupational exposures to genotoxic carcinogens. This article describes their use for the monitoring of metal exposed workers. We found in total 73 relevant articles, in the majority (97 %) increased MN and/or other nuclear anomalies were reported. Most studies were realized in South East Asia and South America. A variety of different occupations was studied including welders, electroplaters, painters, workers in battery recycling and production, tannery workers, dental technicians, miners, workers in foundries and smelters, and also subjects working in waste recycling, glass, aluminum and steel production. In many investigations the effects increased with the duration of the working period. The quality of individual studies was evaluated with a quality score tool. The number of cells was in most studies sufficient and DNA-specific stains were used. However, many studies have shortcomings, e.g. they focused solely on MN formation and did not evaluate anomalies, which provide additional information about the stability of the genetic material and acute cytotoxic effects. Only 35 % of the investigations contain quantitative information about exposures to metals and other toxicants. In 6 of these studies, correlations were observed between the concentrations of specific metals (As, Pb, Cr, Cd) in body fluids and MN frequencies. Taken together, the available data indicate that the MN assay can be used to detect chromosomal damage in metal exposed groups; furthermore, it enables also comparisons between subgroups differing in regard to their exposure and allows an estimation of the efficiency of protective measures. The exposure of workers to metals is currently controlled with chemical analytical measurements only, MN assays with buccal cells could contribute to further improve the safety at workplaces as they reflect the biological consequences including synergistic and antagonistic interactions between toxicants.

Oral squamous cell carcinoma (OSCC) is the most common oral malignancy, often preceded by oral potentially malignant disorders (OPMDs). Currently, no clinical biomarker exists to predict malignancy, necessitating OPMD follow-up. Habits and environmental factors, such as smoking, and alcohol consumption, influence OSCC onset. Increased micronuclei (MNs) formation has been observed in the development of OSCC. Non-invasive diagnostic tests like exfoliative cytology offer painless and regular monitoring options. This study evaluates the impact of tobacco, alcohol, and pesticide exposure on MNs occurrence in exfoliative cytology-collected oral mucosal cells, assessing their potential as non-invasive biomarker for OSCC development prediction and monitoring in high-risk patients. Despite results from this meta-analysis supporting the existence of a stepwise increase from controls to patients with OPMD to OSCC, the translation of these findings into clinical practice is limited due to intra- and inter-individual heterogeneity, as well as methodological variability in MNs quantification. Various factors contribute to this heterogeneity, including demographic variables, methodological variability of different laboratories, staining techniques, sample collection location, and patient characteristics. All these points were discussed to provide further insights and improve standardization for future studies.

Given that telomeres play a fundamental role in maintaining genomic stability, the study of the chromosomal aberrations involving telomeric sequences is a topic of considerable research interest. In recent years, the scoring of these types of aberrations has been used in vertebrate cells, particularly human cells, to evaluate the effects of genotoxic agents on telomeres and the involvement of telomeric sequences on chromosomal aberrations. Currently, chromosomal aberrations involving telomeric sequences are evaluated in peripheral blood lymphocytes or immortalized cell lines, using telomere or telomere plus centromere fluorescence in situ hybridization (FISH) with Peptide Nucleic Acid (PNA) probes (PNA-FISH). The telomere PNA probe is more efficient in the detection of telomeric sequences than conventional FISH with a telomere DNA probe. In addition, the intensity of the telomeric PNA-FISH probe signal is directly correlated with the number of telomeric repeats. Therefore, use of this type of probe can identify chromosomal aberrations involving telomeres as well as determine the telomere length of the sample. There are several mistakes and inconsistencies in the literature regarding the identification of telomere aberrations, which prevent accurate scoring and data comparison between different publications concerning these types of aberrations. The aim of this review is to clarify these issues, and provide proper terminology and criteria for the identification, scoring, and analysis of telomere aberrations.

Chromosomal structural variation (SV) encompasses a heterogenous class of genetic variants that exerts strong influences on human health and disease. Despite their importance, many structural variants (SVs) have remained poorly characterized at even a basic level, a discrepancy predicated upon the technical limitations of prior genomic assays. However, recent advances in genomic technology can identify and localize SVs accurately, opening new questions regarding SV risk factors and their impacts in humans. Here, we first define and classify human SVs and their generative mechanisms, highlighting characteristics leveraged by various SV assays. We next examine the first-ever gapless assembly of the human genome and the technical process of assembling it, which required third-generation sequencing technologies to resolve structurally complex loci. The new portions of that "telomere-to-telomere" and subsequent pangenome assemblies highlight aspects of SV biology likely to develop in the near-term. We consider the strengths and limitations of the most promising new SV technologies and when they or longstanding approaches are best suited to meeting salient goals in the study of human SV in population-scale genomics research, clinical, and public health contexts. It is a watershed time in our understanding of human SV when new approaches are expected to fundamentally change genomic applications.

Head and neck cancer is the seventh most common malignancy globally. Head and neck squamous cell carcinoma (HNSCC) originates from squamous cells and 90% of HNC are HNSCC. The gold standard for diagnosing HNSCC is tissue biopsy. However, given tumour heterogeneity, biopsies may miss important cancer-associated molecular signatures, and more importantly, after the tumour is excised, there is no means of tracking response to treatment in patients. Captured under liquid biopsy, circulating tumour DNA (ctDNA), may identify in vivo molecular genotypes and complements tumour tissue analysis in cancer management. A systematic search was conducted in PubMed, Embase, Scopus and the Cochran Library between 2012 to early 2023 on ctDNA in HNSCC using publications written in English. We summarise 20 studies that compared mutational profiles between tumour tissue DNA (tDNA) and ctDNA, using a cohort of 631 HNSCC patients and 139 controls. Among these studies, the concordance rates varied greatly and the most mutated and the most concordant gene was TP53, followed by PIK3CA, CDKN2A, NOTCH1 and FAT1. Concordant variants were mainly found in Stage IV tumours, and the mutation type is mostly single nucleotide variants (SNV). We conclude that, as a biomarker for HNSCC, ctDNA demonstrates great promise as it recapitulates tumour genotypes, however additional multi-central trials are needed.

Neutropenia is a hematological condition characterized by a decrease in absolute neutrophil count (ANC) in peripheral blood, typically classified in adults as mild (1-1.5 × 10/L), moderate (0.5-1 × 10/L), or severe (< 0.5 × 10/L). It can be categorized into two types: congenital and acquired. Congenital severe chronic neutropenia (SCN) arises from mutations in various genes, with different inheritance patterns, including autosomal recessive, autosomal dominant, and X-linked forms, often linked to mitochondrial diseases. The most common genetic cause is alterations in the ELANE gene. Some cases exist as non-syndromic neutropenia within the SCN spectrum, where genetic origins remain unidentified. The clinical consequences of congenital neutropenia depend on granulocyte levels and dysfunction. Infants with this condition often experience recurrent bacterial infections, with approximately half facing severe infections within their first six months of life. These infections commonly affect the respiratory system, digestive tract, and skin, resulting in symptoms like fever, abscesses, and even sepsis. The severity of these symptoms varies, and the specific organs and systems affected depend on the genetic defect. Congenital neutropenia elevates the risk of developing acute myeloid leukemia (AML) or myelodysplastic syndromes (MDS), particularly with certain genetic variants. SCN patients may acquire CSF3R and RUNX1 mutations, which can predict the development of leukemia. It is important to note that high-dose granulocyte colony-stimulating factor (G-CSF) treatment may have the potential to promote leukemogenesis. Treatment for neutropenia involves antibiotics, drugs that boost neutrophil production, or bone marrow transplants. Immediate treatment is essential due to the heightened risk of severe infections. In severe congenital or cyclic neutropenia (CyN), the primary therapy is G-CSF, often combined with antibiotics. The G-CSF dosage is gradually increased to normalize neutrophil counts. Hematopoietic stem cell transplants are considered for non-responders or those at risk of AML/MDS. In cases of WHIM syndrome, CXCR4 inhibitors can be effective. Future treatments may involve gene editing and the use of the diabetes drug empagliflozin to alleviate neutropenia symptoms.

GNAS (guanine nucleotide-binding protein, alpha stimulating) is an imprinted gene that encodes Gα, the α subunit of the heterotrimeric stimulatory G protein. This subunit mediates the signalling of a diverse array of G protein-coupled receptors (GPCRs), including the melanocortin 4 receptor (MC4R) that serves a pivotal role in regulating food intake, energy homoeostasis, and body weight. Genetic or epigenetic alterations in GNAS are known to cause pseudohypoparathyroidism in its different subtypes and have been recently associated with isolated, early-onset, severe obesity. Given the diverse biological functions that Gα serves, multiple molecular mechanisms involving various GPCRs, such as MC4R, β- and β-adrenoceptors, and corticotropin-releasing hormone receptor, have been implicated in the pathophysiology of severe, early-onset obesity that results from genetic or epigenetic GNAS changes.

The DNA damage response (DDR) is a complex and highly regulated cellular process that detects and repairs DNA damage. The integrity of the DNA molecule is crucial for the proper functioning and survival of cells, as DNA damage can lead to mutations, genomic instability, and various diseases, including cancer. The DDR safeguards the genome by coordinating a series of signaling events and repair mechanisms to maintain genomic stability and prevent the propagation of damaged DNA to daughter cells. The study of an ion channels in the context of DDR is a promising avenue in biomedical research. Lately, it has been reported that the movement of ions through channels plays a crucial role in various physiological processes, including nerve signaling, muscle contraction, cell signaling, and maintaining cell membrane potential. Knowledge regarding the involvement of ion channels in the DDR could support refinement of our approach to several pathologies, mainly cancer, and perhaps lead to innovative therapies. In this review, we focused on the ion channel's possible role in the DDR. We present an analysis of the involvement of ion channels in DDR, their role in DNA repair mechanisms, and cellular outcomes. By addressing these areas, we aim to provide a comprehensive perspective on ion channels in the DDR and potentially guide future research in this field. It is worth noting that the interplay between ion channels and the cellular DDR is complex and multifaceted. More research is needed to fully understand the underlying mechanisms and potential therapeutic implications of these interactions.

Gastric cancer (GC) is the fifth most prevalent malignancy and the third leading cause of cancer-related mortality globally. Despite intensive efforts to enhance the efficiencies of various therapeutics (chemotherapy, surgical interventions, molecular-targeted therapies, immunotherapies), the prognosis for patients with GC remains poor. This might be predominantly due to the limited understanding of the complicated etiology of GC. Importantly, epigenetic modifications and alterations are crucial during GC development. Super-enhancers (SEs) are a large cluster of adjacent enhancers that greatly activate transcription. SEs sustain cell-specific identity by enhancing the transcription of specific oncogenes. In this review, we systematically summarize how SEs are involved in GC development, including the SE landscape in GC, the SE target genes in GC, and the interventions related to SE functions for treating GC.

The diversified impacts of mitochondrial function vs. dysfunction have been observed in almost all disease conditions including cancers. Mitochondria play crucial roles in cellular homeostasis and integrity, however, mitochondrial dysfunctions influenced by alterations in the mtDNA can disrupt cellular balance. Many external stimuli or cellular defects that cause cellular integrity abnormalities, also impact mitochondrial functions. Imbalances in mitochondrial activity can initiate and lead to accumulations of genetic mutations and can promote the processes of tumorigenesis, progression, and survival. This comprehensive review summarizes epigenetic and genetic alterations that affect the functionality of the mitochondria, with considerations of cellular metabolism, and as influenced by ethnicity. We have also reviewed recent insights regarding mitochondrial dynamics, miRNAs, exosomes that play pivotal roles in cancer promotion, and the impact of mitochondrial dynamics on immune cell mechanisms. The review also summarizes recent therapeutic approaches targeting mitochondria in anti-cancer treatment strategies.

Humans ingest particles and fibers on daily basis. Non-digestible carbohydrates are beneficial to health and food additives are considered safe. However, titanium dioxide (E171) has been banned in the European Union because the European Food Safety Authority no longer considers it non-genotoxic. Ingestion of microplastics and nanoplastics are novel exposures; their potential hazardous effects to humans have been under the radar for many years. In this review, we have assessed the association between oral exposure to man-made particles/fibers and genotoxicity in gastrointestinal tract cells and secondary tissues. We identified a total of 137 studies on oral exposure to particles and fibers. This was reduced to 49 papers with sufficient quality and relevance, including exposures to asbestos, diesel exhaust particles, titanium dioxide, silver nanoparticles, zinc oxide, synthetic amorphous silica and certain other nanomaterials. Nineteen studies show positive results, 25 studies show null results, and 5 papers show equivocal results on genotoxicity. Recent studies seem to show null effects, whereas there is a higher proportion of positive genotoxicity results in early studies. Genotoxic effects seem to cluster in studies on diesel exhaust particles and titanium dioxide, whereas studies on silver nanoparticles, zinc oxide and synthetic amorphous silica seem to show mainly null effects. The most widely used genotoxic tests are the alkaline comet assay and micronucleus assay. There are relatively few results on genotoxicity using reliable measurements of oxidatively damaged DNA, DNA double strand breaks (γH2AX assay) and mutations. In general, evidence suggest that oral exposure to particles and fibers is associated with genotoxicity in animals.

A pathology-supported genetic testing (PSGT) framework was established in South Africa to improve access to precision medicine for patients with breast carcinomas. Nevertheless, the frequent identification of variants of uncertain significance (VUSs) with the use of genome-scale next-generation sequencing has created a bottleneck in the return of results to patients. This review highlights the importance of incorporating functional genomics into the PSGT framework as a proposed initiative. Here, we explore various model systems and experimental methods available for conducting functional studies in South Africa to enhance both variant classification and clinical interpretation. We emphasize the distinct advantages of using in vitro, in vivo, and translational ex vivo models to improve the effectiveness of precision oncology. Moreover, we highlight the relevance of methodologies such as protein modelling and structural bioinformatics, multi-omics, metabolic activity assays, flow cytometry, cell migration and invasion assays, tube-formation assays, multiplex assays of variant effect, and database mining and machine learning models. The selection of the appropriate experimental approach largely depends on the molecular mechanism of the gene under investigation and the predicted functional effect of the VUS. However, before making final decisions regarding the pathogenicity of VUSs, it is essential to assess the functional evidence and clinical outcomes under current variant interpretation guidelines. The inclusion of a functional genomics infrastructure within the PSGT framework will significantly advance the reclassification of VUSs and enhance the precision medicine pipeline for patients with breast carcinomas in South Africa.