Altered Gene Research Articles

Changes in Inflammatory Cytokines, Vascular Markers, Cell Cycle Regulators, and Gonadotropin Receptors in Granulosa Cells of COVID-19 Infected Women

Background: COVID-19 infection can negatively affect multiple organ systems, including the reproductive system. Previous research has indicated altered levels of inflammatory markers in the reproductive tissues of women with chronic diseases. This study aimed to assess the expression of inflammatory, vascular, cell cycle, and gonadotropin receptor genes in the granulosa cells and oocytes of women with recent COVID-19 infection undergoing Assisted Reproductive Technology (ART), compared to healthy controls. Materials and Methods: The study involved 15 women who had tested positive for COVID-19 within three months of ART treatment and 15 age-matched healthy women as controls. Granulosa cells were collected during oocyte retrieval, and RNA was isolated to analyze gene expression using quantitative real-time PCR. The evaluated genes included inflammatory cytokines (IL-1B, TNF-α, IL-6, IL-8), vascular genes (VEGF, ANGPT1), cell cycle regulators (FOXL2, Cyclin D1, Cyclin D2, KLF4), and gonadotropin receptors (LHCGR, FSHR). Results: Results showed significantly higher expression of inflammatory cytokines in the granulosa cells of COVID-19 positive women, including IL-1B (4.2-fold), TNF-α (3.8-fold), IL-8 (2.5-fold), and IL-6 (3.2-fold). Vascular genes VEGF and ANGPT1 were also overexpressed, while FOXL2 was downregulated and Cyclin D1/D2 were upregulated in the study group. However, LH and FSH receptor expression remained similar between both groups. Conclusion: The present study demonstrates altered gene expression of inflammatory cytokines, vascular factors and cell cycle regulators in granulosa cells and oocytes of COVID-19 positive women undergoing ART. The dysregulated molecular pathways could potentially impair folliculogenesis and oocyte development in SARS-CoV-2 infected individuals.

Perivascular Adipose Tissue Becomes Pro-Contractile and Remodels in an IL10-/- Colitis Model of Inflammatory Bowel Disease.

Inflammatory Bowel Diseases (IBDs) are associated with aberrant immune function, widespread inflammation, and altered intestinal blood flow. Perivascular adipose tissue (PVAT) surrounding the mesenteric vasculature can modulate vascular function and control the local immune cell population, but its structure and function have never been investigated in IBD. We used an IL10-/- mouse model of colitis that shares features with human IBD to test the hypothesis that IBD is associated with (1) impaired ability of PVAT to dilate mesenteric arteries and (2) changes in PVAT resident adipocyte and immune cell populations. Pressure myography and electrical field stimulation of isolated mesenteric arteries show that PVAT not only loses its anti-contractile effect but becomes pro-contractile in IBD. Quantitative immunohistochemistry and confocal imaging studies found significant adipocyte hyperplasia and increased PVAT leukocytes, particularly macrophages, in IBD. PCR arrays suggest that these changes occur alongside the altered cytokine and chemokine gene expression associated with altered NF-κB signaling. Collectively, these results show that the accumulation of macrophages in PVAT during IBD pathogenesis may lead to local inflammation, which ultimately contributes to increased arterial constriction and decreased intestinal blood flow with IBD.

6847 Poorly Differentiated Thyroid Cancer: A Rare Entity

Abstract Disclosure: S. Fatima: None. S. Ward: None. A. Champion: None. Introduction: Poorly differentiated thyroid carcinoma (PDTC) is a rare thyroid carcinoma originating from follicular epithelial cells. Regarding the degree of differentiation, PDTC ranks between differentiated thyroid cancer and anaplastic thyroid cancer. We hereby present a rare case of PDTC. Case: 29-year-old female with a history of metastatic papillary thyroid cancer (PTC) presented to the emergency department with shortness of breath. She initially noticed a left thyroid mass in 2021. CT neck in 1/2022 showed a 7 cm left thyroid mass with substernal extension, small right thyroid mass with tracheal compression, left bulky cystic nodal masses extending anterior and posterior to carotid and internal jugular vein. PET scan in 3/2022 showed left 7.6 cm thyroid mass SUV 12.1 with tracheal compression, left level II-IV adenopathy with level IV nodal masses 5.1 cm SUV 4.5. Ultrasound-guided FNA in 4/2022 showed PTC in left 8.7cm thyroid nodule; right 2.8 cm nodule showed Atypia of Undetermined Significance; Left 5.5 cm neck level III lymph node (LN) showed PTC metastasis; right 5.1 cm neck level III LN showed PTC metastasis (+ thyroglobulin and TTF-1). CT chest in 11/2022 showed additional involvement of manubrium. Repeat core needle biopsy of left thyroid mass showed left invasive thyroid carcinoma. She was started on neoadjuvant Lenvatinib in 1/2023. Follow-up CT neck in 5/2023 showed a reduction in the size of the left thyroid mass to 5.5cm x 5.8cm x 6.5cm. At this time, she had symptoms of dysphagia, orthopnea, weight loss of about 60 lbs and neck pain. In 11/2023 at time of current presentation she underwent total thyroidectomy, bilateral modified radical neck dissection and manubriectomy. Surgical pathology showed PDTC in left thyroid mass, PTC in right thyroid mass with metastatic PDTC in left proximal sternocleidomastoid, manubrium and LN involvement in neck and mediastinum. OmniSeq Insight testing showed APC and BRCA1 gene mutations, negative BRAF V600E, RET fusion, RET mutation, NTRK1/2/3 fusion and PD L1 immunohistochemistry score is <1%. The patient is planned to receive [1]3[1]I ablation therapy. Conclusion: PDTC accounts for 3–5% of all thyroid carcinomas. The 5, 10-, and 15-year survival rates of patients are 50–85%, 34–50%, and 0%, respectively. RAS gene alterations are found in 25–35%, BRAF mutation in 15–27%, TERT promoter mutation in 40% and mutant TP53 in 16–28% of PDTCs. TERT promoter gene and TP53 mutation usually indicate an aggressive phenotype and a dismal prognosis. Surgery, [1]3[1]I therapy, and external beam radiotherapy constitute the mainstay of traditional treatment for patients with advanced disease; however, treatment is challenging. Tyrosine kinase inhibitors i.e, sorafenib and lenvatinib, have been used in cases of progressive, recurrent, or metastatic disease not responsive to [1]3[1]I therapy. Immunotherapy may be an option in PD-L1-positive tumors. Presentation: 6/1/2024

Deciphering resistance mechanisms in cancer: final report of MATCH-R study with a focus on molecular drivers and PDX development

BackgroundUnderstanding the resistance mechanisms of tumor is crucial for advancing cancer therapies. The prospective MATCH-R trial (NCT02517892), led by Gustave Roussy, aimed to characterize resistance mechanisms to cancer treatments through molecular analysis of fresh tumor biopsies. This report presents the genomic data analysis of the MATCH-R study conducted from 2015 to 2022 and focuses on targeted therapies.MethodsThe study included resistant metastatic patients (pts) who accepted an image-guided tumor biopsy. After evaluation of tumor content (TC) in frozen tissue biopsies, targeted NGS (10 < TC < 30%) or Whole Exome Sequencing and RNA sequencing (TC > 30%) were performed before and/or after the anticancer therapy.Patient-derived xenografts (PDX) were established by implanting tumor fragments into NOD scid gamma mice and amplified up to five passages.ResultsA total of 1,120 biopsies were collected from 857 pts with the most frequent tumor types being lung (38.8%), digestive (16.3%) and prostate (14.1%) cancer. Molecular targetable driver were identified in 30.9% (n = 265/857) of the patients, with EGFR (41.5%), FGFR2/3 (15.5%), ALK (11.7%), BRAF (6.8%), and KRAS (5.7%) being the most common altered genes. Furthermore, 66.0% (n = 175/265) had a biopsy at progression on targeted therapy. Among resistant cases, 41.1% (n = 72/175) had no identified molecular mechanism, 32.0% (n = 56/175) showed on-target resistance, and 25.1% (n = 44/175) exhibited a by-pass resistance mechanism. Molecular profiling of the 44 patients with by-pass resistance identified 51 variants, with KRAS (13.7%), PIK3CA (11.8%), PTEN (11.8%), NF2 (7.8%), AKT1 (5.9%), and NF1 (5.9%) being the most altered genes. Treatment was tailored for 45% of the patients with a resistance mechanism identified leading to an 11 months median extension of clinical benefit.A total of 341 biopsies were implanted in mice, successfully establishing 136 PDX models achieving a 39.9% success rate. PDX models are available for EGFR (n = 31), FGFR2/3 (n = 26), KRAS (n = 18), ALK (n = 16), BRAF (n = 6) and NTRK (n = 2) driven cancers. These models closely recapitulate the biology of the original tumors in term of molecular alterations and pharmacological status, and served as valuable models to validate overcoming treatment strategies.ConclusionThe MATCH-R study highlights the feasibility of on purpose image guided tumor biopsies and PDX establishment to characterize resistance mechanisms and guide personalized therapies to improve outcomes in pre-treated metastatic patients.Graphical

A human progeria-associated BAF-1 mutation modulates gene expression and accelerates aging in C. elegans.

Alterations in the nuclear envelope are linked to a variety of rare diseases termed laminopathies. A single amino acid substitution at position 12 (A12T) of the human nuclear envelope protein BAF (Barrier to Autointegration Factor) causes Néstor-Guillermo Progeria Syndrome (NGPS). This premature ageing condition leads to growth retardation and severe skeletal defects, but the underlying mechanisms are unknown. Here, we have generated a novel in vivo model for NGPS by modifying the baf-1 locus in C. elegans to mimic the human NGPS mutation. These baf-1(G12T) mutant worms displayed multiple phenotypes related to fertility, lifespan, and stress resistance. Importantly, nuclear morphology deteriorated faster during aging in baf-1(G12T) compared to wild-type animals, recapitulating an important hallmark of cells from progeria patients. Although localization of BAF-1(G12T) was similar to wild-type BAF-1, lamin accumulation at the nuclear envelope was reduced in mutant worms. Tissue-specific chromatin binding andtranscriptome analyses showed reduced BAF-1 association in most genes deregulated by the baf-1(G12T) mutation, suggesting that altered BAF chromatin association induces NGPS phenotypes via altered gene expression.

The three YTHDF paralogs and VIRMA are strong cross-histotype tumor driver candidates among m6A core genes.

N6-Methyladenosine (m6A) is the most abundant internal modification in mRNAs. Despite accumulating evidence for the profound impact of m6A on cancer biology, there are conflicting reports that alterations in genes encoding the m6A machinery proteins can either promote or suppress cancer, even in the same tumor type. Using data from The Cancer Genome Atlas, we performed a pan-cancer investigation of 15 m6A core factors in nearly 10000 samples from 31 tumor types to reveal underlying cross-tumor patterns. Altered expression, largely driven by copy number variations at the chromosome arm level, results in the most common mode of dysregulation of these factors. YTHDF1, YTHDF2, YTHDF3and VIRMA are the most frequently altered factors and the only ones to be uniquely altered when tumors are grouped according to the expression pattern of the m6A factors. These genes are also the only ones with coherent, pan-cancer predictive power for progression-free survival. On the contrary, METTL3, the most intensively studied m6A factor as a cancer target, shows much lower levels of alteration and no predictive power for patient survival. Therefore, we propose the non-enzymatic YTHDF and VIRMA genes as preferred subjects to dissect the role of m6A in cancer and as priority cancer targets.

An integrative single-cell atlas for exploring the cellular and temporal specificity of genes related to neurological disorders during human brain development.

Single-cell technologies have enhanced comprehensive knowledge regarding the human brain by facilitating an extensive transcriptomic census across diverse brain regions. Nevertheless, understanding the cellular and temporal specificity of neurological disorders remains ambiguous due to developmental variations. To address this gap, we illustrated the dynamics of disorder risk gene expression under development by integrating multiple single-cell RNA sequencing datasets. We constructed a comprehensive single-cell atlas of the developing human brain, encompassing 393,060 single cells across diverse developmental stages. Temporal analysis revealed the distinct expression patterns of disorder risk genes, including those associated with autism, highlighting their temporal regulation in different neuronal and glial lineages. We identified distinct neuronal lineages that diverged across developmental stages, each exhibiting temporal-specific expression patterns of disorder-related genes. Lineages of nonneuronal cells determined by molecular profiles also showed temporal-specific expression, indicating a link between cellular maturation and the risk of disorder. Furthermore, we explored the regulatory mechanisms involved in early brain development, revealing enriched patterns of fetal cell types associated with neuronal disorders indicative of the prenatal stage's influence on disease determination. Our findings facilitate unbiased comparisons of cell type‒disorder associations and provide insight into dynamic alterations in risk genes during development, paving the way for a deeper understanding of neurological disorders.

The genomic landscape of transformed splenic diffuse red pulp small B‐cell lymphoma

AbstractThe genetic landscape underlying the transformation of splenic diffuse red pulp small B‐cell lymphoma (SDRPL) is not well understood. The present study aimed to unravel the genomic alterations involved in the progression and transformation of SDRPL. We performed genetic studies on both SDRPL and subsequent or synchronous diffuse large B cell lymphoma (DLBCL) samples in three SDRPL patients who eventually developed DLBCL. Our findings revealed that SDRPL cases progressing to DLBCL acquired genomic alterations in genes related to the cell cycle (CDKN2A/B, TP53, MYC and CCND3) and B cell development (BCL6).

Dissecting the Significance of Acid Phosphatase 1 Gene Alterations in Prostate Cancer.

The acid phosphatase 1 (ACP1) gene encodes low-molecular-weight protein tyrosine phosphatase, which is overexpressed in prostate cancer (PC) and a potential therapeutic target. We analyzed ACP1 expression in primary/metastatic PC and its association with molecular profiles and clinical outcomes. NextGen sequencing of DNA (592-gene/whole-exome sequencing)/RNA(whole-transcriptome sequencing) was performed for 5,028 specimens. ACP1-High/ACP1-Low expression was defined as quartile (Q4/1) of RNA transcripts per million (TPM). DNA mutational profiles were analyzed for ACP1-quartile-stratified samples. Gene set enrichment analysis was used for Hallmark collection of pathways. PD-L1+(≥2+, ≥5%; SP142) was tested by immunohistochemistry. Tumor microenvironment's (TME) immune cell fractions were estimated by RNA deconvolution/quanTIseq. Overall survival (OS) was assessed from initial diagnosis/treatment initiation to death/last follow-up. We included 3,058 (60.8%) samples from the prostate, 634 (12.6%) from lymph node metastases (LNMs), and 1,307 (26.0%) from distant metastases (DMs). ACP1 expression was higher in LNM/DM than prostate (49.8/47.9 v 44.1 TPM; P < .0001). TP53 mutations were enriched in ACP1-Q4 (37.9%[Q4] v 27.0%[Q1]; P < .001) among prostate samples. Pathways associated with cell cycle regulation and oxidative phosphorylation were enriched in ACP1-Q4, whereas epithelial-mesenchymal transition and tumor necrosis factor-alpha signaling via nuclear factor kappa-light-chain-enhancer of activated B-cell pathways were enriched in ACP1-Q1. Neuroendocrine and androgen receptor signaling was increased in ACP1-Q4. M2 macrophages and natural killer cell fractions were increased, whereas T cells and M1 macrophages were decreased in ACP1-Q4. While OS differences between ACP1-Q1/Q4 were not statistically significant, there was a trend for worse OS among ACP1-Q4 prostate samples (Q4 v Q1: hazard ratio [HR], 1.19 [95% CI, 0.99 to 1.42]; P = .06) and DM (HR, 1.12 [95% CI, 0.93 to 1.36]; P = .22) but not LNM (HR, 0.98 [95% CI, 0.74 to 1.29]; P = .87). ACP1-High tumors exhibit a distinct molecular profile and cold TME, highlighting ACP1's potential role in PC pathogenesis and novel therapeutic targeting.

A genome-wide CRISPR/Cas9 knockout screen identifies SEMA3F gene for resistanceto cyclin-dependent kinase 4 and 6 inhibitors in breast cancer.

Palbociclib is a cell-cycle targeted small molecule agent used as one of the standards of care in combination with endocrine therapy for patients with hormone receptor (HR)-positive, human epidermal growth factor receptor 2 (HER2)-negative advanced breast cancer. Although several gene alterations such as loss of Rb gene and amplification of p16 gene are known to be conventional resistance mechanisms to cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitors, the comprehensive landscape of resistance is not yet fully elucidated. The purpose of this study is to identify the novel resistant genes to the CDK4/6 inhibitors in HR-positive HER2-negative breast cancer. The wholegenome knockout screen using CRISPR/Cas9 genome editing was conducted in MCF7 to identify resistant genes to palbociclib. The candidate genes for resistance were selected by NGS analysis and GSEA analysis and validated by cell viability assay and mouse xenograft models. We identified eight genes including RET, TIRAP, GNRH1, SEMA3F, SEMA5A, GATA4, NOD1, SSTR1 as candidate genes from the wholegenome knockout screen. Among those, knockdown of SEMA3F by siRNA significantly and consistently increased the cell viability in the presence of CDK4/6 inhibitors in vitro and in vivo. Furthermore, the level of p-Rb was maintained in the palbociclibtreated SEMA3F-downregulated cells, indicating that the resistance is driven by increased activity of cyclin kinases. Our observation provided the first evidence of SEMA3F as a regulator of sensitivity to CDK4/6 inhibitors in breast cancer. The detailed mechanisms of resistance deserve further functional studies to develop the better strategy to overcome resistance in CDK4/6 inhibitors.

Gene expression profiles in placenta and their association with anesthesia, delivery mode and maternal diabetes

IntroductionFetal development is dependent on placenta and affected by multiple factors including maternal diabetes. Here we aimed to identify maternal diabetes-associated changes in placentas and analyzed placental gene expression to understand its modulation by maternal diabetes and birth mode. MethodsPlacental RNAseq transcriptome analyses were performed on maternally-derived decidua and fetal-derived villous tissue from pregnancies of mothers with type 1 diabetes (n=14), gestational diabetes (n=6) and without diabetes (n=14). Information on delivery mode and anesthesia were included as covariables. Analyses were performed separately for decidua and fetal tissues and adjusted for sex. ResultsSubstantial placenta gene expression variation was associated with factors other than maternal diabetes, including site, sex, anesthesia type and delivery mode. Two dominant gene expression clusters aligned to anesthesia and delivery mode were observed for decidua and villous tissue. Upregulation of genes within pathways related to organ morphogenesis and downregulation of immune response to steroid- and hypoxia pathway genes was characteristic of placentas from primary cesarean section deliveries with spinal anesthesia. Opposite profiles were observed for placentas from secondary cesarean and epidural anesthesia deliveries. Placentas from vaginal delivery had intermediate gene expression profiles. More subtle changes were associated with maternal diabetes: upregulation of ribosome activity, down-regulation of maternally-derived decidua chemokine signaling pathways and for gestational diabetes, alteration in hypoxia response genes. DiscussionThe findings reveal suppression of immune pathways and upregulation of ribosome activity in the placenta by maternal diabetes highlighting the importance of confounding factors when examining cell and tissue expression profiles. Further studies should determine whether the observed gene expression differences are related to underlying causes for cesarean section deliveries.

Targeting the Epidermal Growth Factor Receptor Pathway in Chemotherapy-Resistant Triple-Negative Breast Cancer: A Phase II Study.

Epidermal growth factor receptor (EGFR) pathway activation causes chemotherapy resistance, and inhibition of the EGFR pathway sensitizes triple-negative breast cancer (TNBC) cells to chemotherapy in preclinical models. Given the high prevalence of EGFR overexpression in TNBC, we conducted a single-arm phase II study of panitumumab (anti-EGFR monoclonal antibody), carboplatin, and paclitaxel as the second phase of neoadjuvant therapy (NAT) in patients with doxorubicin and cyclophosphamide (AC)-resistant TNBC (NCT02593175). Patients with early-stage, AC-resistant TNBC, defined as disease progression or ≤80% reduction in tumor volume after four cycles of AC, were eligible for this study and received panitumumab (2.5 mg/kg i.v., every week × 13), paclitaxel (80 mg/m2 i.v. every week × 12), and carboplatin (AUC = 4 i.v., every 3 weeks × 4) as the second phase of NAT. A two-stage Gehan-type design was used to detect an improvement in the pathological complete response (pCR)/residual cancer burden class I (RCB-I) rate from 5% to 20%. Whole-exome sequencing was performed on diagnostic tumor biospecimens, where available. From November 3, 2016, through August 23, 2021, 43 patients with AC-resistant TNBC were enrolled. The combined pCR/RCB-I rate was 30.2%. The most common treatment-related adverse events were neutropenia (72%) and anemia (61%), with 7 (16%), 16 (37%), and 8 (19%) patients experiencing grade 4 neutropenia, grade 3 neutropenia, and grade 3 anemia, respectively. No new safety signals were observed. This study met its primary endpoint (pCR/RCB-I = 30.2% vs. 5% in historical controls), suggesting that panitumumab should be evaluated as a component of NAT in patients with chemotherapy-resistant TNBC in a larger, randomized clinical trial. The epidermal growth factor receptor (EGFR) pathway has been implicated as a driver of chemotherapy resistance in triple-negative breast cancer (TNBC). Here, we evaluate the combination of panitumumab, carboplatin, and paclitaxel as the second phase of neoadjuvant therapy (NAT) in patients with AC-resistant TNBC. This study met its primary efficacy endpoint, and molecular alterations in EGFR pathway genes did not seem to influence response to the study regimen.

Downregulation of type I interferon signalling pathway by urate in primary human PBMCs.

Type I interferons (IFN1s) mediate innate responses to microbial stimuli and regulate interleukin (IL)-1 and IL-1 receptor antagonist (Ra) production in human cells. This study explores interferon-stimulated gene (ISG) alterations in the transcriptome of patients with gout and stimulated human primary cells in vitro in relation to serum urate concentrations. Peripheral blood mononuclear cells (PBMCs) and monocytes of patients with gout were primed in vitro with soluble urate, followed by lipopolysaccharide (LPS) stimulation. Separately, PBMCs were stimulated with various toll-like receptor (TLR) ligands. RNA sequencing and IL-1Ra cytokine measurement were performed. STAT1 phosphorylation was assessed in urate-treated monocytes. Cytokine responses to IFN-β were evaluated in PBMCs cultured with or without urate and restimulated with LPS and monosodium urate (MSU) crystals. Transcriptomics revealed suppressed IFN-related signalling pathways in urate-exposed PBMCs or monocytes which was supported by diminishment of phosphorylated STAT1. The stimulation of PBMCs with IFN-β did not modify the urate-induced inflammation. Interestingly, in vivo, serum urate concentrations were inversely correlated to in vitro ISG expression upon stimulations with TLR ligands. These findings support a deficient IFN1 signalling in the presence of elevated serum urate concentrations, which could translate to increased susceptibility to infections.

P100-1 Clinicopathological and molecular analysis of cancer of unknown primary with chromatin remodeling gene alterations

P100-1 Clinicopathological and molecular analysis of cancer of unknown primary with chromatin remodeling gene alterations

Mechanism of denervation muscle atrophy mediated by Ach/p38/MAPK pathway in rats with erectile dysfunction caused by nerve injury

BackgroundPeripheral nerve injury can result in penile cavernosal denervation muscle atrophy, a primary factor in nerve injury erectile dysfunction (NED). While acetylcholine (Ach) is integral to erectile function, its role and mechanisms in NED need further exploration. ObjectiveTo investigate the inhibition of CCMSCs Apoptosis and Protein Degradation Pathway by Ach in NED rat model. MethodsWe investigated changes in Ach secretion and receptor expression in an NED rat model, followed by the evaluation of apoptosis and ubiquitin proteasome activation in hypoxic Cavernous smooth muscle cells (CCMSCs) and their co-cultures with Schwann cells (SWCs), under Ach influence. Further, key pathways in NED were identified via high-throughput sequencing, focusing on the p38/MAPK signaling pathway. We examined gene alterations related to this pathway using hypoxic cell models and employed p38 inhibitors to verify protein changes. Our findings in vitro were then confirmed in the NED rat model. ResultsNerve injury led to reduced Ach receptors and associated gene expression. Experimentally, Ach was shown to counteract CCMSC apoptosis and muscle protein degradation via the p38/MAPK pathway. Inhibition of the Ach degradation pathway demonstrated a capacity to slow NED progression in vivo. Discussion and conclusionActivation of Ach receptors may decelerate denervation-induced cavernosal muscle atrophy, suggesting a potential therapeutic approach for NED. This study highlights the crucial role of the Ach/p38/MAPK axis in the pathophysiology of penis smooth muscle atrophy and its broader implications in managing NED and male erectile dysfunction.

EP.12A.33 A Comparative Analysis of Outcomes in Patients with Single EGFR Muations for Concurrent Gene Alterations

EP.12A.33 A Comparative Analysis of Outcomes in Patients with Single EGFR Muations for Concurrent Gene Alterations

Multi-omic analysis of a mucolipidosis II neuronal cell model uncovers involvement of pathways related to neurodegeneration and drug metabolism

Defining the molecular consequences of lysosomal dysfunction in neuronal cell types remains an area of investigation that is needed to understand many underappreciated phenotypes associated with lysosomal disorders. Here we characterize GNPTAB-knockout DAOY medulloblastoma cells using different genetic and proteomic approaches, with a focus on how altered gene expression and cell surface abundance of glycoproteins may explain emerging neurological issues in individuals with GNPTAB-related disorders, including mucolipidosis II (ML II) and mucolipidosis IIIα/β (ML IIIα/β). The two knockout clones characterized demonstrated all the biochemical hallmarks of this disease, including loss of intracellular glycosidase activity due to impaired mannose 6-phosphate-dependent lysosomal sorting, lysosomal cholesterol accumulation, and increased markers of autophagic dysfunction. RNA sequencing identified altered transcript abundance of several neuronal markers and genes involved in drug metabolism and transport, and neurodegeneration-related pathways. Using selective exo-enzymatic labeling (SEEL) coupled with proteomics to profile cell surface glycoproteins, we demonstrated altered abundance of several glycoproteins in the knockout cells. Most striking was increased abundance of the amyloid precursor protein and apolipoprotein B, indicating that loss of GNPTAB function in these cells corresponds with elevation in proteins associated with neurodegeneration. The implication of these findings on lysosomal disease pathogenesis and the emerging neurological manifestations of GNPTAB-related disorders is discussed.

Identification of CDH4 Gene Copy Number Alteration and Its Association with Clinical Profile of Colorectal Cancer Patient

Background: The genes cadherin 4 (CDH4), Kirsten rat sarcoma viral oncogene homolog (KRAS), V-Raf murine sarcoma viral oncogene homolog B (BRAF), and microsatellite instability (MSI) each play a role in the development of colorectal cancer (CRC). CDH4 gene is located in chromosome 20q13.33 and its amplification or gain is the earliest mutational event found in the majority of CRC and colon polyps. This study aimed to identify copy number alteration in the CDH4 gene and its association with the clinical profile of CRC patients, including gender, age, tumor location and differentiation, frequency of BRAF and KRAS mutations, and MSI status. Methods: The DNA was extracted from 50 tumors and adjacent normal tissues based on the manufacturer’s instructions. Detection of MSI status was carried out by pentaplex polymerase chain reaction (PCR) and PCR products were size separated by capillary electrophoresis (CE) using an ABI 310 Genetic Analyzer. Both KRAS and BRAF mutations were identified by PCR and sequencing while CDH4 copy measurement was measured using TaqManTM Copy Number Assay. Results: Our findings showed 22 (44%) samples with no changes in the copy number of CDH4 gene. Interestingly, 21 (42%) cases had an amplification of the CDH4 gene or CDH4 gene gain, and seven (14%) cases decreased in CDH4 gene expression or CDH4 gene loss. We found an association between changes in the CDH4 gene and gender (p=0.001). However, there was no association between changes in the CDH4 gene and age (p=0.979), tumor location (p=0.145) and differentiation (p=0.648), the frequency of BRAF (p=0.171) and KRAS mutations (p=0.184) and MSI status (p=0.923). Conclusions: Copy number alteration in CDH4 gene in CRC patients and this alteration is significantly associated with gender. Further studies with a larger number of samples are essential to confirm this result and to identify the cause of CDH4 copy number alteration and its biological significance.

TMPRSS2:ERG Gene Fusion Might Predict Resistance to PARP Inhibitors in Metastatic Castration-resistant Prostate Cancer.

The emergence of novel DNA damage repair (DDR) pathways in molecular-target therapy drugs (MTTD) has shown promising outcomes in treating patients with metastatic castration-resistant prostate cancer (mCRPC). About 25% of mCRPC patients have actionable deleterious aberrations in DDR genes, primarily in the homologous recombination (HR) pathway. However, the response rate in patients with BRCA1/2 or mutations in HRR-related genes is only 45%-55%, when exposed to poly ADP ribose polymerase (PARP) inhibitor-based therapy (PARPi). A frequent characteristic feature of prostate cancer (PC) is the occurrence of genomic rearrangement that affects the transmembrane protease serine 2 (TMPRSS2) and E26 transformation-specific (ETS)- transcription factor-related gene (ERG). In this study, a total of 114 patients with mCRPC had their RNA and DNA sequenced using next-generation sequencing. Based on their genetic profile of deleterious gene alterations of BRCA1/2 or ATM, six patients were selected for PARPi. Patients with TMPRSS2:ERG gene fusion and homozygous alteration in ATM or BRCA2 (n=2) or heterozygous alterations (BRCA1 or BRCA2) and lack of TMPRSS2:ERG gene fusion (n=2) did not show clinical benefit from PARPi (treatment duration <16 weeks). In contrast, patients (n=2) without TMPRSS2:ERG gene fusion and homozygous deleterious alterations in ATM or BRCA2 all had clinical benefit from PARPi (treatment duration ≥16 weeks). The TMPRSS2:ERG transcript product might be used as a PARPi resistance biomarker.

Paternal preconception donepezil exposure enhances learning in offspring

BackgroundRecent research has indicated that parental use of central nervous system-targeting medications during periconceptional periods may affect offspring across various developmental and behavioral domains. The present study sought to investigate the potential influence of paternal use of donepezil, a specific reversible central acetylcholinesterase inhibitor that activates the cholinergic system to promote cognition, on offspring.ResultsIn this study, male rats were bred after 21 days of oral donepezil administration at a dose of 4 mg/kg to generate F1 offspring. Both male and female F₁ offspring displayed enhanced performance in learning and short-term memory tests, including novel object recognition, Y maze, and operant learning. Transcriptomic analysis revealed notable alterations in genes associated with the extracellular matrix in the hippocampal tissue of the F1 generation. Integration with genes related to intelligence identified potential core genes that may be involved in the observed behavioral enhancements.ConclusionsThese findings indicate that prolonged paternal exposure to donepezil may enhance the learning and memory abilities of offspring, possibly by targeting nonneural, extracellular regions. Further research is required to fully elucidate any potential transgenerational effects.