Chromosome 9p21 Research Articles

EPCO-54. MULTIOMIC ANALYSIS OF SPINAL EPENDYMOMAS REVEALS MECHANISMS OF TUMOR AGGRESSIVENESS WITH MYCN AMPLIFICATION

Abstract Grade-3 spinal ependymomas, particularly those harboring MYCN amplification, have dismal prognoses despite optimal treatment regimens. MYCN functions as a transcription factor (TF) to drive the expression of key tumor driver genes. Our aim is to identify mechanisms underlying tumor aggressiveness and treatment resistance in MYCN-amplified ependymomas. MYCN amplification in spinal ependymomas was identified with custom next generation sequencing panels and bulk methylation analysis. We then conducted paired single-nucleus RNA (snRNA-seq) and chromatin accessibility (snATAC-seq) sequencing of grade-2 (n=2, 1 patient) and grade-3 spinal ependymomas (n=6, 6 patients, 4 MYCN-amplified). Comparator snRNA-seq datasets (grade-2/grade-3; 8/2) were included from a publicly available dataset (GSE163686; all non-MYCN-amplified). Canonical cell classes were identified with a combination of cluster- and cell-based strategies. Downstream analysis was conducted using R 4.3 (Seurat, Signac, clusterProfiler, CellChat), and Python 3.11 (scanpy) packages. We identified ependymoma tumor cells distinct from canonical immune cell classes with gene expression analysis and confirmed a distinct copy-number-variation pattern in these cells with chromosome 1 and 2p gain, and chromosome 6p and 22q loss. MYCN-amplified tumor cells (compared to non-MYCN-amplified grade2/3 tumor cells) showed increased differential accessibility at genes including VEGFA. Increased MYCN TF activity was confirmed with gene overexpression of downstream targets including NOTCH3, CREB5, PLK1, and VEGFA. Other genes that were highly accessible and overexpressed in MYCN-amplified tumor cells were related to oxidative stress, differentiation, and anti-apoptotic pathways. MYCN-amplified tumor cells modified the tumor microenvironment with increased VISTA signaling between MYCN-amplified tumor cells and tumor-associated macrophages, creating an immunosuppressive environment. We confirmed these effects by observing a higher proportion of M2-like macrophages in MYCN-amplified tumors (29.6% vs 18.2%). In conclusion, MYCN amplification in spinal ependymomas upregulates tumor cell proliferation, angiogenesis, and M2-like macrophage recruitment/programming.

A familial chromosome 4p16.3 terminal microdeletion that does not cause Wolf-Hirschhorn (4p-) syndrome

A familial chromosome 4p16.3 terminal microdeletion that does not cause Wolf-Hirschhorn (4p-) syndrome

Heterozygous Beta-Thalassaemia in Pregnancy: Two Rare Causes of Severe Fetal Anemia Requiring Intrauterine Blood Transfusions.

In this article, we present two cases of severe fetal hemolytic anemia based on a beta-thalassaemia trait inherited from a single parent. These cases, presented at 20 and 28weeks' gestation, necessitated intra-uterine blood transfusions. This occurrence is remarkable because it challenges the common assumption that beta-thalassaemia typically has no prenatal implications regarding fetal anemia. Both fetuses inherited a rare heterozygous mutation from their mother, resulting in gamma-thalassaemia-related anemia. In the first case, the anemia was related to a deletion in the beta locus control region (βLCR) and in the second case, a deletion on chromosome 11p15.4 was the cause. These mutations not only affect the beta chain production, but also the gamma chain production, leading to a reduction in the synthesis of HbF, ineffective erythropoiesis and consequently, perinatal hemolytic anemia. Clinicians should be vigilant regarding these rare mutations in families with a history of beta-thalassaemia as the fetal clinical consequences can be severe and intra-uterine blood transfusions may prove life-saving for these fetuses.

Update on surveillance guidelines in emerging Wilms tumor predisposition syndromes.

Wilms tumors are commonly associated with predisposition syndromes. Many of these syndromes are associated with specific phenotypic features and are discussed in the related paper from the AACR Pediatric Cancer Working Group. Guidelines for surveillance in this population were published in 2017 but since then several studies have identified new genes with recurrent pathogenic variants associated with increased risk for Wilms tumor development. In general, variants in these genes are less likely to be associated with other phenotypic features. Recently, members of the AACR Pediatric Cancer Working Group met to update surveillance guidelines for patients with a predisposition to Wilms tumors with a review of recently published evidence and risk estimates. Risk estimates for Wilms tumor for the more recently described genes are discussed here along with suggested surveillance guidelines for these populations. Several other emerging clinical scenarios associated with Wilms tumor predisposition are also discussed including patients with family histories of Wilms tumor and no identified causative gene, patients with bilateral tumors, and patients with somatic mosaicism for chromosome 11p15.5 alterations. This perspective serves to update pediatric oncologists, geneticists, radiologists, counselors and other healthcare professionals on emerging evidence and harmonize updated surveillance recommendations in the North American and Australian context for patients with emerging forms of Wilms tumor predisposition.

The NIPBL-gene mutation of a Cornelia de Lange Syndrome patient causes deficits in the hepatocyte differentiation of induced Pluripotent Stem Cells via altered chromatin-accessibility

The Cornelia de Lange syndrome (CdLS) is a rare genetic disease, which is characterized by a cohesinopathy. Mutations of the NIPBL gene are observed in 65% of CdLS patients. A novel iPSC (induced Pluripotent Stem Cell) line was reprogrammed from the leukocytes of a CdLS patient carrying a missense mutation of the NIPBL gene. A mutation-corrected isogenic iPSC-line and two iPSC-lines generated from the healthy parents were used as controls. The iPSC lines were differentiated along the hepatocyte-lineage. Comparative immunofluorescence, RNA-seq and ATAC-seq analyses were performed on undifferentiated and differentiated iPSCs. In addition, chromatin organization was studied by ChIP-Seq analysis on the patient derived iPSCs as well as the respective controls. Relative to the mutation-corrected and the healthy-parents iPSCs, the patient-derived counterparts are defective in terms of differentiation along the hepatocyte-lineage. One-third of the genes selectively up-regulated in CdLS-derived iPSCs and hepatic cells are non-protein-coding genes. By converse, most of the selectively down-regulated genes code for transcription factors and proteins regulating neural differentiation. Some of the transcriptionally silenced loci, such as the DPP6 gene on chromosome 7q36.2 and the ZNF gene cluster on chromosome 19p12, are located in closed-chromatin regions. Relative to the corresponding controls, the global transcriptomic differences observed in CdLS undifferentiated iPSCs are associated with altered chromatin accessibility, which was confirmed by ChIP-Seq analysis. Thus, the deficits in the differentiation along the hepatocyte lineage observed in our CdLS patient is likely to be due to a transcriptional dysregulation resulting from a cohesin-dependent alteration of chromatin accessibility.

Morphometric measurements of intraoral anatomy in children with Beckwith-Wiedemann syndrome: a novel approach

BackgroundAn easy-to-use tool to objectively measure intraoral anatomy with meaningful clinical correlations may improve care for patients with Beckwith-Wiedemann syndrome (BWS), who commonly have symptomatic macroglossia.MethodsChildren aged 2–17 years with BWS were enrolled between 12/2021 and 01/2024. Digital intraoral photographs with a laser ruler were taken, and morphometric measurements were made using ImageJ software. Relationships between morphometrics and outcomes including BWS clinical score, percentage mosaicism, and incidence of tongue reduction surgery were examined using t-tests and multivariate linear models.ResultsPharyngeal morphometric measurements were obtained in 49 patients with BWS. Mouth area, width, and height differed significantly across BWS molecular subtypes. Right-to-left tongue width and mouth width were larger in those with loss of methylation at imprinting control region 2 (IC2 LOM) than other BWS variants. Patients with paternal uniparental isodisomy of chromosome 11p15 (pUPD11) had narrower mouths than others. Those with tongue reduction surgery had more tongue ridging than those without surgery. There were correlations between mouth area and BWS clinical score, tongue width and BWS clinical score, and tongue length and percentage mosaicism.ConclusionIntraoral morphometric measurements are associated with phenotypic burden in BWS. Tongue morphology varies across the BWS spectrum, with IC2 LOM having wider tongues and mouths, and pUPD11 having narrower mouths. Tongue ridging is more common in those selected for surgery. Intraoral morphometric measurements may be safely obtained at low costs across centers caring for children with BWS or others at risk of upper airway obstruction.

A genome-wide association study of adults with community-acquired pneumonia

BackgroundCommunity-acquired pneumonia (CAP) is associated with high morbidity and hospitalization rate. In infectious diseases, host genetics plays a critical role in susceptibility and immune response, and the immune pathways involved are highly dependent on the microorganism and its route of infection. Here we aimed to identify genetic risk loci for CAP using a case-control genome-wide association study (GWAS).MethodsWe performed a GWAS on 3,765 Spanish individuals, including 257 adult patients hospitalized with CAP and 3,508 population controls. Pneumococcal CAP was documented in 30% of patients; the remaining 70% were selected among patients with unidentified microbiological etiology. We tested 7,6 million imputed genotypes using logistic regressions. UK Biobank GWAS of bacterial pneumonia were used for results validation. Subsequently, we prioritized genes and likely causal variants based on Bayesian fine mapping and functional evidence. Imputation and association of classical HLA alleles and amino acids were also conducted.ResultsSix independent sentinel variants reached the genome-wide significance (p < 5 × 10-8), three on chromosome 6p21.32, and one for each of the chromosomes 4q28.2, 11p12, and 20q11.22. Only one variant at 6p21.32 was validated in independent GWAS of bacterial and pneumococcal pneumonia. Our analyses prioritized C4orf33 on 4q28.2, TAPBP on 6p21.32, and ZNF341 on 20q11.22. Interestingly, genetic defects of TAPBP and ZNF341 are previously known inborn errors of immunity predisposing to bacterial pneumonia, including pneumococcus and Haemophilus influenzae. Associations were all non-significant for the classical HLA alleles.ConclusionsWe completed a GWAS of CAP and identified four novel risk loci involved in CAP susceptibility.

CD5+ Large B cell lymphoma with IRF4 rearrangement: case report and review of the literature

Abstract Introduction/Objective Large B cell lymphoma with IRF4 rearrangement was introduced as a diagnosis by WHO in 2017. It is a rare entity that affects Waldeyer’s ring in mostly males, children and young adults. The lesional immunophenotype may be positive for CD10. FISH is positive for IRF4 rearrangement on chromosome 6p25.Among reported cases in the English literature only 6 cases have reported specifically CD5 being negative. We present review of the literature during years 2020-2024, and a case of large B cell lymphoma with IRF4 rearrangement, specifically positive for CD5. Methods/Case Report The patient is a 5 years old male, since 2017 suffering from recurring ear infections with approximately 4-5 episodes of otitis media during every 6 months. In 2019 the patient’s mother reported snoring, constant sore throat, bad breath for 2 years. On physical examination the left tonsil appeared enlarged; exudative tonsillitis was present. Later on, an asymmetric, firm mass in the location of the left tonsil with induration of the left palate was identified. CT scan showed mild asymmetric soft tissue thickening of the left aspect of the soft palate and left palatine tonsil. Mildly prominent left-sided levels IIa and IIb lymph nodes, measuring up to 1 cm were identified. H&amp;E sections revealed an expansile proliferation of neoplastic cells, large with void to indented nuclei, vesicular chromatin, small nucleoli, visible cytoplasms. Frequent mitoses, apoptosis and individual cell necrosis were seen. Immunohistochemistry revealed CD20+ B cell neoplasm with high expression for MUM1 and high proliferation rate along with strong CD5+ and CD10-. By multicolor flow cytometric studies, an abnormal B cell population was found (40%) with increased forward scatter along with expression for CD5, CD19, CD20, CD22, CD45, CD79a (cytoplasmic), FMC7, (dim) lambda-restriction. Fluorescence In Situ Hybridization Study revealed IRF4 (DUSP22) rearrangement on chromosome 6p25. Results (if a Case Study enter NA) NA Conclusion We analyzed 24 cases reported in the English literature. The mean age of the patients is 7 years old. The majority are males (14/24 = 58%).The most common location is tonsil/ neck mass (15/24 = 62%).CD5 was negative in 6/24 cases.

The discovery of the RCC1::IRF4 Fusion in CLL patients with t(1;6)(p35.3;p25.2) chromosomal translocation.

Jayne etal. provide a molecular characterization of the t(1;6)(p35.3;p25.2) chromosomal translocation in patients with chronic lymphocytic leukaemia. They indicate that this translocation involves the gene encoding interferon regulatory factor 4 (IRF4) on chromosome 6p25.2 with the regulator of chromosome condensation 1 (RCC1) gene on chromosome 1p35.3. This translocations may have important prognostic value. Commentary on: Jayne etal. The chromosomal translocation t(1;6)(p35.3;p25.2), recurrent in chronic lymphocytic leukaemia leads to RCC1::IRF4 fusion. Br J Haematol 2024 (Online ahead of print). doi: 10.1111/bjh.19790.

Event-free survival in neuroblastoma with MYCN amplification and deletion of 1p or 11q may be associated with altered immune status

BackgroundNeuroblastoma exhibits substantial heterogeneity, which is intricately linked to various genetic alterations. We aimed to explore immune status in the peripheral blood and prognosis of patients with neuroblastoma with different genetic characteristics.MethodsWe enrolled 31 patients with neuroblastoma and collected samples to detect three genetic characteristics. Peripheral blood samples were tested for immune cells and cytokines by fluorescent microspheres conjugated with antibodies and flow cytometry. Event-free survival (EFS) was analyzed using the Kaplan‒Meier method.ResultsTwenty-two patients had genetic aberrations, including MYCN amplification in 6 patients, chromosome 1p deletion in 9 patients, and chromosome 11q deletion in 14 patients. Two genetic alterations were present in seven patients. The EFS was worse in patients with MYCN amplification or 1p deletion than in the corresponding group, whereas 11q deletion was a prognostic factor only in patients with unamplified MYCN. Changes in immune status revealed a decrease in the proportion of T cells in blood, and an increase in regulatory T cells and immunosuppression-related cytokines such as interleukin (IL)-10. The EFS of the IL-10 high-level group was lower than that of the low-level group. Patients with concomitant genetic alterations and a high level of IL-10 had worse EFS than other patients.ConclusionsPatients with neuroblastoma characterized by these genetic characteristics often have suppressed T cell response and an overabundance of immunosuppressive cells and cytokines in the peripheral blood. This imbalance is significantly associated with poor EFS. Moreover, if these patients show an elevated levels of immunosuppressive cytokines such as IL-10, the prognosis will be worse.

The NOTCH1 and miR-34a signaling network is affected by TP53 alterations in CLL

In chronic lymphocytic leukemia (CLL), TP53 mutations or deletions on chromosome 17p lead to adverse prognosis and reduced levels of miR-34a, which targets NOTCH1. Also, hyperactivated NOTCH1 signaling is crucial for CLL progression. Here we explored the interaction between p53, miR-34a, and NOTCH1 in CLL. We investigated the effect of p53 and miR-34a on NOTCH1 signaling and expression in CLL cells with altered TP53. Our results indicate that miR-34a reduces NOTCH1 3’ UTR activity but might not be a mediator between p53 signaling and NOTCH1. p53 activation increases miR-34a expression and NOTCH1 protein levels, correlating with decreased NOTCH1 and miR-34a levels in primary CLL cells with TP53 alterations. Some samples with high NOTCH1 levels presented increased BCL-2, suggesting an anti-apoptotic mechanism of a potentially direct p53-NOTCH1 relation in CLL. This study deepens the understanding of the p53-miR-34a-NOTCH1 signaling network, providing insights that could guide future therapeutic strategies for CLL.

Chromosome 8p Syndromes Clinical Presentation and Management Guidelines.

Rearrangements of the p-arm of Chromosome 8 can result in a spectrum of neurodevelopmental challenges, along with increased risk of epilepsy, structural brain and cardiac malformations, persisting developmental delays, and other health challenges. The majority of patients reported on in this sample are characterized by an inverted-duplication deletion rearrangement, but deletions, duplications, and mosaic ring changes in 8p result in similar phenotype. In this report, we add to the phenotypic and functional description of these patients according to their specific chromosomal rearrangement, share neuro-psychometric values, and propose surveillance care guidelines for caregivers and medical providers of patients with Chromosome 8p Syndromes. Observations from clinical experience with 24 patients seen at our 8p-dedicated Multi-Disciplinary Neurogenetics program are shared.

Low-level mosaic GCK mutations in children with diazoxide-unresponsive congenital hyperinsulinism.

Some children with diazoxide-unresponsive congenital hyperinsulinism (HI) lack any detectable disease-causing mutation in peripheral blood DNA. To examine whether somatic post-zygotic mutations of known HI genes are responsible for disease in children with diazoxide-unresponsive HI requiring surgery with histology not classified as focal or Localized Islet Nuclear Enlargement (LINE), and without detectable mutations by standard genetic testing of peripheral blood DNA. Next-generation sequencing (NGS) was performed on specimens of pancreas from 10 children with diazoxide-unresponsive HI. Four unique GCK mutations were identified at low levels of mosaicism ranging from 4.4-10.1% in pancreatic DNA from five of these 10 children. The GCK mutations were not detectable in peripheral blood DNA by NGS in three cases from which peripheral blood DNA was available for testing. All four GCK mutations have been previously published as activating HI mutations. The histology was consistent with diffuse-HI in four of the five cases with mosaic GCK mutations. In one of these, hypomethylation of IC2 on chromosome 11p was identified in pancreatic and peripheral blood DNA. Histology of the fifth case revealed minor islet abnormalities suggestive of Beckwith Wiedemann Spectrum (BWSp) although molecular analysis for 11pUPD was negative in pancreas. These results indicate that post-zygotic somatic GCK mutations are responsible for some cases of non-focal diazoxide-unresponsive hyperinsulinism.

Sarcomatoid Urothelial Carcinoma Arising in Autosomal Dominant Polycystic Kidney Disease: A Case Report and Literature Review.

Autosomal dominant polycystic kidney disease (ADPKD) may be associated with various epithelial malignancies. The most reported ones are papillary renal cell carcinoma (RCC) and clear cell RCC. Only one noninvasive urothelial carcinoma arising in the renal pelvis has been previously reported in the setting of ADPKD in the English literature. A 52-year-old patient with ADPKD and a history of renal transplant presented with a poorly differentiated sarcomatoid neoplasm in his native left polycystic kidney. A recognizable urothelial or renal cell carcinoma differentiation was not identified in the resected neoplasm microscopically. The initial diagnosis for this specimen was challenging on morphology and immunohistochemistry, but targeted next-generation sequencing provided molecular evidence in support of urothelial origin, indicating a hotspot mutation -124 C > T in the TERT promoter (C228 T) and loss of heterozygosity on chromosomes 9p and 8p. This tumor is unique because, to our knowledge, this is the first report of upper tract sarcomatoid urothelial carcinoma in a patient with ADPKD.

7238 A rare cause of primary ovarian failure in an adolescent due to a genetic mutation in mitochondrial poly-A-polymerase (mTPAP) mRNA

Abstract Disclosure: J. Epstein: None. S. Veera: None. J. Pappas: None. B.C. Shah: None. Primary ovarian insufficiency (POI) is a dysfunction or loss of ovarian follicles with associated amenorrhea. While most causes are idiopathic, approximately 15% of POI cases are due to single gene or chromosomal abnormalities, such as X chromosome abnormalities. Many of these nuclear genes encode for proteins that function within the mitochondria. Mitochondrial disorders affect tissues with high energy demand, such as the neurological, skeletal, cardiac, and endocrine systems. Furthermore, mature ovaries have a relatively higher concentration of mitochondria, which play a role in cellular respiration along with steroidogenesis. Mutations that disrupt these pathways can therefore lead to ovarian dysfunction. We present a case of a 17-year-old female with concern for primary amenorrhea. She underwent pubarche at age 12 years and thelarche at 14 years, with slowed progression. Height Z score was -1.61 and weight Z score was -0.41. Lab work at presentation was significant for LH 18.6 IU/L (N 2.4-12.6 IU/L), FSH 43.9 IU/L (N 1.1-9.6 IU/L), low estradiol &amp;lt;2.5 pg/mL (N 30-100 pg/mL), low total testosterone 8.3 ng/dL (N 15-31 ng/dL), unremarkable HbA1C, cortisol, thyroid levels, and a 46 XX karyotype. Pelvic ultrasound showed small ovarian volumes with a 1.9 cc right ovary, 1.5 cc left ovary consistent with lack of hormonal stimulation, and uterus length of 4.9 cm (N &amp;gt; 4 cm) consistent with early hormonal stimulation. She was started on transdermal estrogen replacement. Past medical history included stable pericardial effusion, episodes of supraventricular tachycardia, pre-cataracts, speech delays, mild learning disabilities, spastic diplegia, progressive gait abnormalities, ligamentous laxity, and recurrent patellar dislocations requiring multiple orthopedic limb surgeries. Family history was significant for parental consanguinity as first cousins. As the etiology of the POI remained unknown, genetic testing was completed. Whole exome sequencing (WES) revealed a homozygous pathogenic variant in the MTPAP gene: p.(Leu495Arg) (CTG&amp;gt;CGG): c.1484 T&amp;gt;G in exon 9 on chromosome 10p11.23. MTPAP, also known as SPAX4, TENT6, or PAPD1, is a nuclear-encoded polymerase involved in synthesizing the 3’ poly(A) tail of mitochondrial transcripts as well as in mRNA degradation. She was diagnosed with autosomal recessive MTPAP-related neurodevelopmental and movement disorder. Monogenic disorders affecting mitochondrial function are rare causes of POI. A high index of suspicion should be raised when a constellation of neurological, musculoskeletal, and cardiac problems co-exist. WES can be a very useful tool for uncovering rare genetic disorders. Further studies are needed to clarify details of organ specific mitochondrial dysfunction and possible therapeutics in individuals affected with MTPAP-related neurodevelopmental and movement disorders. Presentation: 6/1/2024

Ancient viral DNA in the human genome linked to neurodegenerative diseases

BackgroundHuman endogenous retroviruses (HERVs) are sequences in the human genome that originated from infections with ancient retroviruses during our evolution. Previous studies have linked HERVs to neurodegenerative diseases, but defining their role in aetiology has been challenging. Here, we used a retrotranscriptome-wide association study (rTWAS) approach to assess the relationships between genetic risk for neurodegenerative diseases and HERV expression in the brain, calculated with genomic precision. MethodsWe analysed genetic association statistics pertaining to Alzheimer’s disease, amyotrophic lateral sclerosis, multiple sclerosis, and Parkinson’s disease, using HERV expression models calculated from 792 cortical samples. Robust risk factors were considered those that survived multiple testing correction in the primary analysis, which were also significant in conditional and joint analyses, and that had a posterior inclusion probability above 0.5 in fine-mapping analyses. ResultsThe primary analysis identified 12 HERV expression signatures associated with neurodegenerative disease susceptibility. We found one HERV expression signature robustly associated with amyotrophic lateral sclerosis on chromosome 12q14 (MER61_12q14.2) and one robustly associated with multiple sclerosis on chromosome 1p36 (ERVLE_1p36.32a). A co-expression analysis suggested that these HERVs are involved in homophilic cell adhesion via plasma membrane adhesion molecules. ConclusionsWe found HERV expression profiles robustly associated with amyotrophic lateral sclerosis and multiple sclerosis susceptibility, highlighting novel risk mechanisms underlying neurodegenerative disease, and offering potential new targets for therapeutic intervention.

Microsatellite Instability and Loss of Heterozygosity as Prognostic Markers in Oral Squamous Cell Carcinoma: Molecular Mechanisms, Detection Techniques, and Therapeutic Strategies.

The aim of this study was to conduct a systematic review of research investigating the potential role of microsatellite instability (MSI) and loss of heterozygosity (LOH) in oral squamous cell carcinoma (OSCC), with a focus on molecular mechanisms, detection methods, and therapeutic approaches. Search for articles involved the PubMed and Scopus. Previous retrospective and prospective studies identified variations between oral cancers that exhibit microsatellite stability and LOH. In this search, 294 articles were initially retrieved. Of these, 70 were excluded due to duplication, 106 were identified as ineligible by automated tools, and 24 were excluded as they were published in languages other than English. An additional 94 articles were excluded, 32 of which focused on head and neck cancers broadly, and 8 could not be accessed due to withdrawal. Ultimately, a systematic review was conducted based on 54 selected articles. Among the chromosomes analyzed for MSI and LOH, the highest frequency of LOH was observed on chromosome 9p. The MSI subtype is particularly susceptible to immunotherapeutic methods, such as the use of anti-PD-L1 and anti-CTLA4 antibodies, owing to its strong immunogenicity and ubiquitous expression of immune checkpoint ligands. Given the distinct characteristics and clinical behavior of oral cancer with MSI compared to microsatellite stable disease, it is advisable to incorporate MSI testing into the diagnostic process for all stages of tumor development. This ensured that each patient had received precise and effective treatment.

GENETIC MAPPING OF THREE LARGE CONSANGUINEOUS KASHMIRI FAMILIES WITH AUTOSOMAL RECESSIVE POLYDACTYLY SCREENED FOR FIVE PREVALENT GENES

Background: Polydactyly, a congenital hand defect characterized by extra digits, is more complicated than simple duplication, delicately weaving aberrant anatomical components with hypoplasia, uneven joint shapes, and unusual tendon and ligament placements. The dominant theory attributes its genesis to a group of five genes: GLI1 (chromosome 12q13.3), ZNF141 (chromosome 4p16.3), IQCE (chromosome 7p22.2), KIAA0825 (chromosome 5q15), and FAM92A (chromosome 8q22.1). The objective of this study is to identify the most prevalent genes responsible for polydactyly in the population of Azad Jammu and Kashmir. Method: The microsatellite markers used for PCR amplification and subsequently testing linkage to known genes are presented below. Represent electropherograms of ethidium bromide-stained 8% non-denaturing polyacrylamide gels (PAGEs) obtained by genotyping microsatellite markers linked on chromosome 8q22.1, 5q15, 12q13.3, 4p16.3, and 7p22.3 in family A, B, C. Genetic positions (in centiMorgan) for these marker loci were obtained from Rutgers combined linkage-physical map of the human genome. Results: Screening of most prevalent genes that include GLI1 (chromosome 12q13.3), ZNF141 (chromosome 4p16.3), IQCE (Chromosome 7p22.2), KIAA0825 (chromosome 5q15) and FAM92A (8q22.1) showed heterozygosity on every locus. Already known disease loci were further narrowed down with highly polymorphic markers which failed to find any linkage. Conclusion: The previously reported genotypic-phenotypic relation was not revealed in these 3 families signifying the probable involvement of unexplored genetic segments in intricate pathogenesis of this condition, emphasizing the need for further exploration beyond the established genetic association. Pak J Physiol 2024;20(3):44–8, DOI: https://doi.org/10.69656/pjp.v20i3.1611

Upregulation of Apoptosis Related Genes in Clinically Normal Tongue Contralateral to Squamous Cell Carcinoma of the Oral Tongue, an Effort to Maintain Tissue Homeostasis

PurposeThe field cancerization concept indicates the presence of pre-cancerous changes in clinically normal tissue surrounding the tumor. In squamous cell carcinoma of the oral tongue (SCCOT) which is infrequently linked to human papillomavirus infection, we have previously reported that clinically normal tongue contralateral to tumor (NTCT) is molecularly abnormal. Here, combining our transcriptomic and genomic data, we aimed to investigate the contribution of molecular changes in NTCT to cancer development.MethodsMicroarray gene expression data of 14 healthy controls, 23 NTCT and 29 SCCOT samples were investigated to characterize transcriptional profiles in NTCT. Whole exome sequencing and RNA-sequencing data of paired NTCT and tumor samples from 15 SCCOT patients were used to study correlation between copy number variation and differential gene expression.ResultsUsing supervised multivariate partial least squares discriminant analysis, a total of 61 mRNAs that distinguish NTCT from healthy tongue were selected. Functional enrichment analysis of the 22 upregulated genes showed increased “positive regulation of nitrogen compound metabolic process” in NTCT. All 12 genes involved in this process have roles in apoptosis (anti- and/or pro-apoptotic). Compared to healthy controls, Zinc Finger Protein 395 (ZNF395), a pro-apoptotic tumor suppressor located on chromosome 8p, was the only gene showing increased mRNA level in NTCT whereas decreased in SCCOT. Given the frequent loss of chromosome 8p in SCCOT, the impact of ZNF395 copy number variation on gene expression was further examined, revealing a positive correlation between copy number and mRNA level (correlation coefficient = 0.572, p < 0.001).ConclusionNTCT is susceptible to malignant transformation, where tissue homeostasis is maintained at least partly through regulation of apoptosis. Loss of the pro-apoptotic gene ZNF395 could thus initiate cancer development.

MTAP protein status is highly concordant with CDKN2A fluorescent in situ hybridization and allows stratification of the luminal subtype in muscle-invasive bladder cancer.

Loss of heterozygosity in chromosome 9p21, common in urothelial carcinoma (UC), typically involves deletion of CDKN2A and MTAP genes. MTAP loss is emerging as a promising therapeutic target and predictive biomarker in UC. This single-centrre retrospective study examined the incidence of CDKN2A deletions and MTAP loss in muscle-invasive bladder cancer (MIBC) and metastatic urothelial carcinoma (mUC), investigating their correlations with clinical, pathological, and genomic features, as well as patient outcomes. Fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC) were performed on 302 MIBC specimens and 63 biopsy-proven metachronous urothelial metastases to assess CDKN2A deletions and MTAP protein expression. CDKN2A homozygous deletion (HD), identified in 30.3% of MIBCs, and MTAP loss, found in 28.8% of MIBCs, were both significantly associated with the luminal-URO subtype, FGFR3 mutations, and normal/wildtype p53 IHC (P < 0.05). Loss of MTAP expression was significantly correlated with CDKN2A HD, with 84.0% sensitivity, 92.3% negative predictive value (NPV), 96.3% specificity, and 91.9% positive predictive value (PPV). MTAP expression was 100% concordant between primary tumours and nodal metastases. Patients with MTAP loss had a higher incidence of visceral metastases (50%) compared to bone/soft tissue (35.7%) and nodes (14.3%). Mean progression-free survival and overall survival were shorter for patients with MTAP loss, although not statistically significant. Our findings highlight CDKN2A HD and MTAP loss as prevalent genetic alterations in MIBC and mUC, particularly within the luminal-URO subtype and FGFR3-mutated, p53-normal/wildtype tumours. MTAP IHC can serve as a surrogate marker for 9p21.3 HD, highlighting its clinical relevance and potential as a therapeutic target and predictive biomarker in MIBC.