BRCA1 Expression Research Articles

Enhanced cancer cell proliferation and aggressive phenotype counterbalance in breast cancer with high BRCA1 gene expression.

While comprehensive research exists on the mutation of the DNA repair gene BRCA1, limited information is available regarding the clinical significance of BRCA1 gene expression. Given that cancer cell proliferation is aggrevated by DNA repair, we hypothesized that high BRCA1 gene expression breast cancer (BC) might be linked with aggressive tumor biology and poor clinical outcomes. The cohorts: The Cancer Genome Atlas (TCGA, n = 1069), METABRIC (n = 1903), and SCAN-B (n = 3273) were utilzed to obtain data of 6245 BC patients. BC patients without BRCA1 mutation exhibited higher BRCA1 expression, which was associated with DNA repair functionality. However, no such correlation was observed with BRCA2 expression. The association of high BRCA1 expression with cancer cell proliferation was evidenced by significant enrichment of cell proliferation-related gene sets, higher histological grade, and proliferation score. Furthermore, increased levels of homologous recombination deficiency, intratumoral heterogeneity, and altered fractions were associated with high BRCA1 expression. Moreover, BC with high BRCA1 expression exhibited reduced infiltration of dendritic cells and CD8 T-cells, while showing increased infiltration of Th1 cells. Surprisingly, BRCA1 expression was not associated with the survival of BC irrespective of the subtypes. Conversely, BC with low BRCA1 expression enriched cancer aggravating pathway gene sets, such as Cancer Stem Cell-related signaling (NOTCH and HEDGEHOG), Angiogenesis, Epithelial-Mesenchymal Transition, Inflammatory Response, and TGF-beta signaling. Despite being linked to heightened proliferation of cancer cells and unassertive phenotype, BRCA1 expression did not show any association with survival in BC.

Arsenic and Chromium Induced Toxicity on Zebrafish Kidney: Mixture Effects on Oxidative Stress and Involvement of Nrf2-Keap1-ARE, DNA Repair, and Intrinsic Apoptotic Pathways.

In polluted water, cooccurrences of two carcinogens, arsenic (As) and chromium (Cr), are extensively reported. Individual effects of these heavy metals have been reported in kidney of fishes, but underlying molecular mechanisms are not well established. There is no report on combined exposure of As and Cr in kidney. Thus, the present study investigated and compared individual and combined effects of As and Cr on zebrafish (Danio rerio) kidney treating at their environmentally relevant concentrations for 15, 30, and 60 days. Increased ROS levels, lipid peroxidation, GSH level, and decreased catalase activity implied oxidative stress in treated zebrafish kidney. Damage in histoarchitecture in treated groups was also noticed. The current study involved gene expression study of Nrf2, an important transcription factor of cellular stress responses along with its negative regulator Keap1 and downstream antioxidant genes nqo1 and ho1. Results indicated activation of Nrf2-Keap1 pathway after combined exposure. Expression pattern of ogg1, apex1, polb, and creb1 revealed the inhibition of base excision repair pathway in treatments. mRNA expression of tumor suppressor genes p53 and brca2 was also altered. Expressional alteration in bax, bcl2, caspase9, and caspase 3 indicated apoptosis (intrinsic pathway) induction, which was maximum in combined group. Inhibition of DNA repair and induction of apoptosis indicated that the activated antioxidant system was not enough to overcome the damage caused by As and Cr. Overall, this study revealed additive effects of As and Cr in zebrafish kidney after chronic exposure focusing cellular antioxidant and DNA damage responses.

Modulation of Tumor Suppressor Genes by Histone Deacetylase Inhibitors in Cancer Therapy

Histone deacetylase (HDAC) inhibitors have emerged as promising therapeutics for various cancers due to their ability to modulate gene expression profiles and restore normal cellular functions. Among the key targets of HDAC inhibitors are tumor suppressor genes, which play crucial roles in regulating cell cycle progression, apoptosis, and DNA repair. However, despite their therapeutic potential, HDAC inhibitors come with significant drawbacks. These include off-target effects that can lead to unintended alterations in gene expression, toxicity to normal cells, and the development of drug resistance in some cancer patients. Additionally, the non-specific nature of some HDAC inhibitors can result in side effects such as fatigue, gastrointestinal disturbances, and hematological toxicity, limiting their clinical utility. This review provides a comprehensive overview of the impact of HDAC inhibitors on tumor suppressor genes in cancer therapy. We discussed the mechanisms by which HDAC inhibitors regulate the expression of tumor suppressor genes, including p53, p21, BRCA1, PTEN, and others, through histone hyperacetylation and chromatin remodeling. Furthermore, we described the therapeutic implications of HDAC inhibitor-induced modulation of tumor suppressor genes in various cancer types, highlighting the potential for combination therapies and personalized treatment approaches. Understanding the interplay between HDAC inhibitors and tumor suppressor genes is critical for optimizing the efficacy of HDAC inhibitor-based therapies and advancing precision medicine in oncology.

BRCA1 Promotes Repair of DNA Damage in Cochlear Hair Cells and Prevents Hearing Loss.

Cochlear hair cells (HCs) sense sound waves and allow us to hear. Loss of HCs will cause irreversible sensorineural hearing loss. It is well known that DNA damage repair plays a critical role in protecting cells in many organs. However, how HCs respond to DNA damage and how defective DNA damage repair contributes to hearing loss remain elusive. In this study, we showed that cisplatin induced DNA damage in outer hair cells (OHCs) and promoted OHC loss, leading to hearing loss in mice of either sex. Cisplatin induced the expression of Brca1, a DNA damage repair factor, in OHCs. Deficiency of Brca1 induced OHC and hearing loss, and further promoted cisplatin-induced DNA damage in OHCs, accelerating OHC loss. This study provides the first in vivo evidence demonstrating that cisplatin mainly induces DNA damage in OHCs and that BRCA1 promotes repair of DNA damage in OHCs and prevents hearing loss. Our findings not only demonstrate that DNA damage-inducing agent generates DNA damage in postmitotic HCs but also suggest that DNA repair factors, like BRCA1, protect postmitotic HCs from DNA damage-induced cell death and hearing loss.

Newly synthesized chitosan nanoparticles loaded with caffeine/moringa leaf extracts Halt Her2, BRCA1, and BRCA2 expressions

Breast cancer is among the highest morbidity and mortality rates in women around the world. In the present investigation we aimed to synthesis novel nanosystem combining two naturally important anticancer agents with different mechanism of action namely Moringa oleifera and caffeine. Firstly, chemical analysis of Moringa oleifera extract and caffeine was done by gas chromatography-mass spectroscopy (GC–MS) in order to assess the main chemical compounds present and correlate between them and the possible anticancer effect. The novel nanosystem was characterized through dynamic light scattering techniques which revealed the stability and homogeneity of the prepared M. oleifera leaves extract/Caffeine loaded chitosan nanoparticles, while FTIR and transmission electron microscope (TEM) proved the shape and the successful incorporation of M. oleifera leaves extract/Caffeine onto the nanochitosan carrier. Our initial step was to assess the anticancer effect in vitro in cancer cell line MCF-7 which proved the significant enhanced effect of M. oleifera leaves extract/Caffeine nanosystem compared to M. oleifera leaves extract or caffeine loaded nanoparticles. Further studies were conducted in vivo namely tumor biomarkers, tumor volume, bioluminescence imaging, molecular and histopathological investigations. The present study proved the potent anticancer effect of the synthesized M. oleifera leaves extract/Caffeine loaded chitosan nanoparticles. Mo/Caf/CsNPs exhibited a large number of apoptotic cells within the tumor mass while the adipose tissue regeneration was higher compared to the positive control. The prepared nanoparticles downregulated the expression of Her2, BRCA1 and BRCA2 while mTOR expression was upregulated. The aforementioned data demonstrated the successful synergistic impact of Moringa and caffeine in decreasing the carcinoma grade.

Alterations in the expression of homologous recombination repair (HRR) genes in breast cancer tissues considering germline BRCA1/2 mutation status

IntroductionHomologous recombination (HR) is a crucial DNA-repair mechanism, and its disruption can lead to the accumulation of mutations that initiate and promote cancer formation. The key HR genes, BRCA1 and BRCA2, are particularly significant as their germline pathogenic variants are associated with a hereditary predisposition to breast and/or ovarian cancer.Materials and methodsThe study was performed on 45 FFPE breast cancer tissues obtained from 24 and 21 patients, with and without the germline BRCA1/2 mutation, respectively. The expression of 11 genes: BRCA1, BRCA2, ATM, BARD1, FANCA, FANCB, FANCI, RAD50, RAD51D, BRIP1, and CHEK2 was assessed using Custom RT2 PCR Array (Qiagen), and results were analysed using R.ResultsCancer tissues from patients with BRCA1 or BRCA2 germline mutations displayed no significant differences in the expression of the selected HR genes compared to BRCA1 or BRCA2 wild-type cancer tissues. In BRCA1mut cancer tissues, BRCA1 expression was significantly higher than in BRCA2mut and BRCA wild-type cancer tissues.ConclusionsIn cancer tissues harbouring either BRCA1 or BRCA2 germline mutations, no significant differences in expression were observed at the mRNA level of any tested HR genes, except BRCA1. However, the significant differences observed in BRCA1 expression between germline BRCA1mut, germline BRCA2mut and BRCA1/2wt tissues may indicate a compensatory mechanism at the mRNA level to mitigate the loss of BRCA1 function in the cells.

The combination of temozolomide and perifosine synergistically inhibit glioblastoma by impeding DNA repair and inducing apoptosis

Temozolomide (TMZ) is widely utilized as the primary chemotherapeutic intervention for glioblastoma. However, the clinical use of TMZ is limited by its various side effects and resistance to chemotherapy. The present study revealed the synergistic inhibition of glioblastoma through the combined administration of TMZ and perifosine. This combination therapy markedly diminished BRCA1 expression, resulting in the suppression of DNA repair mechanisms. Furthermore, the combination of TMZ and perifosine elicited caspase-dependent apoptosis, decreasing glioblastoma cell viability and proliferation. The observed synergistic effect of this combination therapy on glioblastoma was validated in vivo, as evidenced by the substantial reduction in glioblastoma xenograft growth following combined treatment with TMZ and perifosine. In recurrent glioma patients, higher BRCA1 expression is associated with worse prognosis, especially the ones that received TMZ-treated. These findings underscore the potent antitumor activity of the AKT inhibitor perifosine when combined with TMZ and suggest that this approach is a promising strategy for clinical glioblastoma treatment.

Type I or Type II Endometrial Carcinoma? Role of BRCA1 Immunohistochemistry.

Investigation of diagnostic and prognostic relevance of BRCA1 immunohistochemistry (IHC) in endometrial carcinoma. Ninty four specimens of endometrial carcinomas were evaluated. Full sections stained with hematoxylin & eosin were revaluated for assessment of tumor type, grade, myometrial, & lympho-vascular invasion (LVI). Tissue microarray blocks were constructed using the pencil tip method and immunostained with Anti-BRCA1 antibody. BRCA1 was correlated with clinicopathological parameters as well as disease free survival and overall survival. There was a statistically significant difference (P=0.001) between serous and endometroid carcinomas regarding BRCA1 expression where most cases of serous carcinoma showed negative expression. No statistically significant difference was found between BRCA1 positive and negative cases regarding disease free survival (DFS) or overall survival. Serous histotype, high grade, advanced stage, and omental deposits were the parameters significantly associated with decreased DFS. Results of this study can support inclusion of BRCA1 IHC in a panel to differentiate both endometroid and serous carcinomas. The current study found no prognostic relevance for BRCA1 in terms of overall survival and disease-free survival.

Evaluation of Tumorigenic Properties of MDA-MB-231 Cancer Stem Cells Cocultured with Telocytes and Telocyte-Derived Mitochondria Following miR-146a Inhibition.

Telocytes have some cytoplasmic extensions called telopodes, which are thought to play a role in mitochondrial transfer in intercellular communication. Besides, it is hypothesized that telocytes establish cell membrane-mediated connections with breast cancer cells in coculture and may contribute to the survival of neoplastic cell clusters together with other stromal cells. The aim of this study is to investigate the contribution of telocytes and telocyte-derived mitochondria, which have also been identified in breast tumors, to the tumor development of breast cancer stem cells (CSCs) via miR-146a-5p. The isolation/characterization of telocytes from bone marrow mononuclear cells and the isolation of mitochondria from these cells were performed, respectively. In the next step, CSCs were isolated from the MDA-MB-231 cell line and were characterized. Then, miR-146a-5p expressions of CSCs were inhibited by anti-miR-146a-5p. The epithelial-mesenchymal transition (EMT) was determined by evaluating changes in vimentin protein levels and was evaluated by analyzing BRCA1, P53, SOX2, E-cadherin, and N-cadherin gene expression changes. Our results showed that miR-146a promoted stemness and oncogenic properties in CSCs. EMT (N-cadherin, vimentin, E-cadherin) and tumorigenic markers (BRCA1, P53, SOX2) of CSCs decreased after miR-146a inhibition. Bone marrow-derived telocytes and mitochondria derived from telocytes favored the reduction of CSC aggressiveness following this inhibition.

Modulation of homologous recombination gene activity in breast tumor cells in an <i>in vitro</i> model

Introduction. It has been established that the presence of homologous recombination deficiency in a breast tumor is associated with the effectiveness of treatment. But despite the high chemosensitivity of the tumor to DNA-damaging agents, complete pathological responses to treatment are very rare. And this process may be based on a change in the somatic status of BRCA1, that is, a reversion and return of the wild-type allele occurs and the DNA repair function is restored.Aim. To evaluate changes in the presence of chromosomal aberrations and the expression profile of the main genes of homologous recombination in cell models of breast cancer under the influence of cisplatin and docetaxel.Materials and methods. The study was conducted on breast cancer tumor cell cultures: MCF-7, MDA-MB-231 and MDA-MB-468. A cell model of drug resistance was obtained for two drugs: cisplatin and docetaxel. RNA and DNA were isolated from cell suspension using the RNeasy Plus Mini Kit and QIAamp DNA Mini Kit (Qiagen, Germany), respectively. The expression level of homologous recombination genes was assessed using reverse transcription polymerase chain reaction. To assess the presence of chromosomal aberrations, microarray analysis was performed on DNA chips.Results. Restoration of normal copy number for the BRCA1, CDK12, CHEK1 and RAD51D genes in MCF-7 under the influence of cisplatin was shown. For BRCA2 and PALB2, amplifications were detected. A statistically significant increase in the expression of the BRCA1 (p = 0.04), BRCA2 (p = 0.02), PALB2 (p = 0.01) and RAD51D (p = 0.05) genes was also shown. MDAMB-231 shows that all identified loci with deletions, where the BRCA2, BARD1, CHEK2, PALB2 and RAD54L genes are localized, are restored to normal copy number by cisplatin. The appearance of amplifications was registered for BRCA1, BRIP1, FANCL, RAD51B, PARP1. A similar result was shown for docetaxel. An increase in the expression level is typical for the genes BRCA1 (p = 0.02), BRCA2 (p = 0.02), CHEK2 (p = 0.05), FANCL (p = 0.04), PALB2 (p = 0.05), RAD51C (p = 0.02), PARP1 (p = 0.02), which corresponds to the appearance of amplifications. In the MDA-MB-468 cell culture, an increase in the copy number of only the BRCA1 gene is observed. The effect of docetaxel has no effect on this cell culture. The level of BRCA1 expression increases in direct proportion to the duration of drug action.Conclusion. Thus, the study showed that under the influence of cisplatin, reversion of not only homologous recombination gene mutations, but also other disorders can occur.

The change of albumen quality during the laying cycle and its potential physiological and molecular basis of laying hens

To elucidate the regulatory mechanisms that impact variability in albumen quality of laying hens from the peak of lay to the late production phase. A 60-week study was conducted on a cohort of 20,000 Hy-Line Brown laying hens from 20 to 80 weeks old. Before commencement at 20 weeks, the 10-week-old hens were acclimatized for ten weeks. This study examined changes in albumen quality, serum, and liver antioxidant capacity, magnum morphology, and expression of albumen-protein-related genes in the magnum. To reduce sampling error, we collected eggs (n=90) from pre-determined cages at every sampling point (5-week intervals), and 8 hens were selected at 10-week intervals for blood and tissue collection. Our findings revealed that age significantly affected most evaluated parameters. Albumen gel properties, including hardness, gumminess, and chewiness, increased significantly with age (P < 0.05). With the increasing of hens' age from 20 to 80 weeks, the albumen proportion of eggs was decreased, but eggshell proportion, yolk proportion, thick albumen proportion, thick to thin ratio, thick albumen solid content, albumen height, Haugh units, and yolk color were increased and then decreased (P < 0.05). Compared to hens aged 20-60 weeks, the hens (70-80 weeks) had significantly reduced total antioxidant capacity (TAC) and glutathione levels (GSH) in the liver and lower serum TAC and superoxide dismutase levels (SOD) (P < 0.05). The magnum mucosal folds were highest in 40-60 weeks-old hens, and the luminal diameter increased with age (P < 0.05). In the magnum, the mRNA expression levels for OVA, CPE, and NUP205 increased significantly between 30 and 40 weeks, while FBN1 expression was higher between 30 and 50 weeks (P < 0.05). At 70-80 weeks, the expression of BRCA2 was significantly downregulated (P < 0.05). Albumen height, thick albumen proportion with protein secretion-related genes, enhanced antioxidant function, and luminal diameter correlated positively. However, the thick-to-thin albumen ratio negatively correlated with BRCA2, downregulated in aged laying hens. We used principal component and cluster analysis to deduce albumen quality changes during three phases: 25-35, 40-55, and 60-80 weeks. The decline in albumen quality in aging hens is linked with decreased antioxidant capacity, magnum health, and downregulation of key genes involved in protein synthesis and secretion. These findings emphasize critical albumen quality changes in laying hens and suggest molecular pathways underlying age-related albumen quality alterations.

Epstein-Barr virus deubiquitinating enzyme BPLF1 is involved in EBV carcinogenesis by affecting cellular genomic stability

Increased mutational burden and EBV load have been revealed from normal tissues to Epstein-Barr virus (EBV)-associated gastric carcinomas (EBVaGCs). BPLF1, encoded by EBV, is a lytic cycle protein with deubiquitinating activity has been found to participate in disrupting repair of DNA damage. We first confirmed that BPLF1 gene in gastric cancer (GC) significantly increased the DNA double strand breaks (DSBs). Ubiquitination mass spectrometry identified histones as BPLF1 interactors and potential substrates, and co-immunoprecipitation and in vitro experiments verified that BPLF1 regulates H2Bub by targeting Rad6. Over-expressing Rad6 restored H2Bub but partially reduced γ-H2AX, suggesting that other downstream DNA repair processes were affected. mRNA expression of BRCA2 were significantly down-regulated by next-generation sequencing after over-expression of BPLF1, and over-expression of p65 facilitated the repair of DSBs. We demonstrated BPLF1 may lead to the accumulation of DSBs by two pathways, reducing H2B ubiquitination (H2Bub) and blocking homologous recombination which may provide new ideas for the treatment of gastric cancer.

DNA damage and repair in patients undergoing myocardial perfusion single-photon emission computed tomography

As patient exposure to ionizing radiation from medical imaging and its risks are continuing issues, this study aimed to evaluate DNA damage and repair markers after myocardial perfusion single-photon emission computed tomography (MPS). Thirty-two patients undergoing Tc-99m sestamibi MPS were studied. Peripheral blood was collected before radiotracer injection at rest and 60–90 min after injection. The comet assay (single-cell gel electrophoresis) was performed with peripheral blood cells to detect DNA strand breaks. Three descriptors were evaluated: the percentage of DNA in the comet tail, tail length, and tail moment (the product of DNA tail percentage and tail length). Quantitative PCR (qPCR) was performed to evaluate the expression of five genes related to signaling pathways in response to DNA damage and repair (ATM, ATR, BRCA1, CDKN1A, and XPC). Mann–Whitney’s test was employed for statistical analysis; p < 0.05 was considered significant. Mean Tc-99m sestamibi dose was 15.1 mCi. After radiotracer injection, comparing post-exposure to pre-exposure samples of each of the 32 patients, no statistically significant differences of the DNA percentage in the tail, tail length or tail moment were found. qPCR revealed increased expression of BRCA1 and XPC, without any significant difference regarding the other genes. No significant increase in DNA strand breaks was detected after a single radiotracer injection for MPS. There was activation of only two repair genes, which may indicate that, in the current patient sample, the effects of ionizing radiation on the DNA were not large enough to trigger intense repair responses, suggesting the absence of significant DNA damage.

Higher expression of BRCA1, and CHUK and lower expression of NFKBIA, ESR1, PIK3R1, and PPARG in HAM/TSP compared to ATLL, a diverse pathological consequence

Higher expression of BRCA1, and CHUK and lower expression of NFKBIA, ESR1, PIK3R1, and PPARG in HAM/TSP compared to ATLL, a diverse pathological consequence

Epigenetic editing of BRCA1 promoter increases cisplatin and olaparib sensitivity of ovarian cancer cells

ABSTRACT Drug resistance is the primary contributor to the high mortality rate of ovarian cancer (OC). The loss of BRCA1/2 function is linked to drug sensitivity in OC cells. The aim of this study is to enhance the drug sensitivity of OC cells by inducing BRCA1 dysfunction through promoter epigenetic editing. Epigenetic regulatory regions within the BRCA1 promoter, affecting gene expression, were initially discerned through analysis of clinical samples. Subsequently, we designed and rigorously validated epigenetic editing tools. Ultimately, we evaluated the cisplatin and olaparib sensitivity of the OC cells after editing. The BRCA1 promoter contains two CpG-rich regions, with methylation of the region covering the transcription start site (TSS) strongly correlating with transcription and influencing OC development, prognosis, and homologous recombination (HR) defects. Targeting this region in OC cells using our designed epigenetic editing tools led to substantial and persistent DNA methylation changes, accompanied by significant reductions in H3K27ac histone modifications. This resulted in a notable suppression of BRCA1 expression and a decrease in HR repair capacity. Consequently, edited OC cells exhibited heightened sensitivity to cisplatin and olaparib, leading to increased apoptosis rates. Epigenetic inactivation of the BRCA1 promoter can enhance cisplatin and olaparib sensitivity of OC cells through a reduction in HR repair capacity, indicating the potential utility of epigenetic editing technology in sensitization therapy for OC.

ADAR1 promotes cisplatin resistance in intrahepatic cholangiocarcinoma by regulating BRCA2 expression through A-to-I editing manner.

Aberrant A-to-I RNA editing, mediated by ADAR1 has been found to be associated with increased tumourigenesis and the development of chemotherapy resistance in various types of cancer. Intrahepatic cholangiocarcinoma (iCCA) is a highly aggressive malignancy with a poor prognosis, and overcoming chemotherapy resistance poses a significant clinical challenge. This study aimed to clarify the roles of ADAR1 in tumour resistance to cisplatin in iCCA. We discovered that ADAR1 expression is elevated in iCCA patients, particularly in those resistant to cisplatin, and associated with poor clinical outcomes. Downregulation of ADAR1 can increase the sensitivity of iCCA cells to cisplatin treatment, whereas its overexpression has the inverse effect. By integrating RNA sequencing and Sanger sequencing, we identified BRCA2, a critical DNA damage repair gene, as a downstream target of ADAR1 in iCCA. ADAR1 mediates the A-to-I editing in BRCA2 3'UTR, inhibiting miR-3157-5p binding, consequently increasing BRCA2 mRNA and protein levels. Furthermore, ADAR1 enhances cellular DNA damage repair ability and facilitates cisplatin resistance in iCCA cells. Combining ADAR1 targeting with cisplatin treatment markedly enhances the anticancer efficacy of cisplatin. In conclusion, ADAR1 promotes tumour progression and cisplatin resistance of iCCA. ADAR1 targeting could inform the development of innovative combination therapies for iCCA.

PARP inhibitor synthetic lethality in ATM biallelic mutant cancer cell lines is associated with BRCA1/2 and RAD51 downregulation.

Ataxia telangiectasia-mutated (ATM) kinase is a central regulator of the DNA damage response (DDR) signaling pathway, and its function is critical for the maintenance of genomic stability in cells that coordinate a network of cellular processes, including DNA replication, DNA repair, and cell cycle progression. ATM is frequently mutated in human cancers, and approximately 3% of lung cancers have biallelic mutations in ATM, i.e., including 3.5% of lung adenocarcinomas (LUAD) and 1.4% of lung squamous cell carcinomas (LUSC). We investigated the potential of targeting the DDR pathway in lung cancer as a potential therapeutic approach. In this context, we examined whether ATM loss is synthetically lethal with niraparib monotherapy. This exploration involved the use of hATM knockout (KO) isogenic cell lines containing hATM homozygous (-/-) and heterozygous (+/-) generated via CRISPR/Cas9 gene knockout technology in DLD-1, a human colorectal adenocarcinoma cell line. Subsequently, we extended our investigation to non-small cell lung cancer (NSCLC) patient derived xenograft (PDX) models for further validation of poly ADP-ribose polymerase inhibitor (PARPi) synthetic lethality in ATM mutant NSCLC models. Here, we demonstared that biallelic hATM deletion (-/-) in DLD-1 impairs homologous recombination (HR) repair function and sensitizes cells to the PARPi, niraparib. Niraparib also caused significant tumor regression in one-third of the NSCLC PDX models harboring deleterious biallelic ATM mutations. Loss of hATM (-/-) was concomitantly associated with low BRCA1 and BRCA2 protein expression in both the hATM (-/-) DLD-1 cell line and PARPi-sensitive ATM mutant NSCLC PDX models, suggesting a downstream effect on the impairment of HR-mediated DNA checkpoint signaling. Further analysis revealed that loss of ATM led to inhibition of phosphorylation of MRN (Mre11-Rad50-NBS1) complex proteins, which are required for ATM-mediated downstream phosphorylation of p53, BRCA1, and CHK2. Taken together, our findings highlight that the synthetic lethality of niraparib in ATM-deficient tumors can be regulated through a subsequent effect on the modulation of BRCA1/2 expression and its effect on HR function.

Abstract PO4-23-10: Restoring the efficacy of standard of care in treatment-refractory ER+ breast cancer by re-activation of cAMP-ROS-DNA damage axis

Abstract Estrogen receptor (ER)-positive (ER+) breast cancer has the highest incidence rate and accounts for around 75% of all cases. Endocrine therapy has been the mainstay therapy for the treatment of both early and late-stage ER+ breast cancer for decades. Although most ER+ breast cancer patients initially respond well to endocrine therapy, resistance is common. The addition of CDK4/6 inhibitors to endocrine therapy led to significant improvements in clinical outcome, and they are now considered one of the standard of care (SOC) therapies. However, a significant proportion of patients still suffer from disease relapse upon prolonged use of endocrine therapy in combination with CDK4/6 inhibitors, representing a major clinical challenge that reduces the long-term benefit and patient survival. Therefore, elucidating the mechanisms of sensitivity and resistance to SOC therapy and identifying novel actionable targets are urgently needed. Here, we show that endocrine therapies and CDK4/6 inhibitors cause toxic PARP1 trapping and generation of a functional BRCAness phenotype by downregulating key DNA repair proteins that ultimately result in increased histone parylation and reduced H3K9 acetylation, leading to transcriptional blockage and cell death. Mechanistically, we found that SOC therapy downregulates phosphodiesterase 4D (PDE4D), resulting in increased cAMP levels, PKA-dependent phosphorylation of mitochondrial COXIV-I, generation of mitochondrial reactive oxygen species (ROS), and DNA damage. Importantly, we identified PDE4D as a novel ER target gene that in turn stimulates ER activity in a feedforward loop in endocrine-responsive models and regulates BRCA1 expression. However, during SOC resistance, an ER-to-EGFR switch induces PDE4D overexpression via c-Jun. Inhibition of PDE4D using BPN14770, the first-in-class PDE4D allosteric inhibitor that has successfully completed Phase II trials in Fragile X syndrome, or its upstream EGFR in combination with SOC therapies in drug-resistant settings, including multiple acquired SOC resistant cell line models, primary cultures and organoids of endocrine resistant ER+ PDXs reinstates PARP1 trapping and BRCAness, leading to drug sensitization in vitro and in vivo. Notably, we demonstrated that high PDE4D mRNA and protein expression is associated with dramatically worse disease-free survival and overall survival in endocrine therapy-treated ER+ breast cancer. Considering the availability of potent and non-toxic PDE4D inhibitors, clinically approved EGFR and PARP1 inhibitors, our findings have great translational potential. Citation Format: Ozge Saatci, Metin Cetin, Meral Uner, Unal Metin Tokat, Ioulia Chatzistamou, Pelin Gulizar Ersan, Elodie Montaudon, Aytekin Akyol, Sercan Aksoy, Aysegul Uner, Elisabetta Marangoni, Sajish Mathew, Ozgur Sahin. Restoring the efficacy of standard of care in treatment-refractory ER+ breast cancer by re-activation of cAMP-ROS-DNA damage axis [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-23-10.

Abstract PO3-15-10: Enhanced Cancer Cell Proliferation and Aggressive Phenotype Counterbalance in Breast Cancer with High BRCA1 Gene Expression

Abstract Introduction: It is estimated that 5–10% of all breast cancer cases have a hereditary component with BRCA1 mutation being one of the most frequently observed gene mutations in breast cancer patients. BRCA1 plays a crucial role in DNA repair, and its mutation has been extensively studied. However, the clinical significance of BRCA1 gene expression remains largely unexplored. Given that heightened DNA repair mechanisms can potentially increase cancer cell proliferation, we hypothesized that breast cancer with high BRCA1 gene expression might be associated with aggressive tumor biology and worse survival. Methods: The clinical, pathological and gene expression levels of a total of 6,245 breast cancer patients were analyzed from three large independent cohorts: METABRIC (n = 1,903), GSE96058 (n = 3,273), and The Cancer Genome Atlas (TCGA, n = 1,069). The data was retrieved through cBioPortal. The xCell algorithm was used to correlate the BRCA1 gene expression with the stromal and immune cell fractions infiltrating the tumor microenvironment (TME). Results: BRCA1 gene expression was higher in breast cancer patients without BRCA1 mutation, and it correlated with the DNA repair activity (p = 0.026 and p &amp;lt; 0.01 respectively), however, it did not correlate with BRCA2 gene expression (p = 0.38). Breast cancer with high BRCA1 gene expression was associated with cancer cell proliferation as evidenced by higher Nottingham histological grade (p &amp;lt; 0.001), Ki67 expression (p &amp;lt; 0.001), significant enrichment of cell proliferation-related gene sets such as E2F Targets, G2M Checkpoint, MYC Targets V1 and V2, Mitotic Spindle (all FDR &amp;lt; 0.25), and significantly less infiltration of stromal cells such as adipocytes, fibroblasts, microvascular and lymphatic endothelial cells, and pericytes (all p &amp;lt; 0.05). High BRCA1 gene expression was associated with higher homologous recombination deficiency, intratumor genomic heterogeneity, and fraction altered (all p &amp;lt; 0.001); however, there was no correlation observed with mutation rates or neoantigens. It was observed that breast cancer with high BRCA1 gene expression had less infiltration of CD8 T cells, dendritic cells, regulatory T cells, and B cells and had higher infiltration of Th1 cells (all p &amp;lt; 0.05). To our surprise, there was no significant difference in overall survival regardless of the subtypes (except for ER+/HER2- subtype in METABRIC cohort, p = 0.03). Breast cancer with low BRCA1 gene expression enriched cancer aggravating pathway gene sets such as Cancer Stem Cell-related signaling pathways (NOTCH and HEDGEHOG), Angiogenesis, Epithelial Mesenchymal Transition, Inflammatory Response, and TGF-beta signaling (all FDR &amp;lt; 0.25). Conclusion: Contrary to our initial hypothesis, we did not find any significant association between BRCA1 gene expression and survival outcomes in breast cancer patients. Based on the results of our study, we speculate that the interplay between enhanced cancer cell proliferation and aggressive tumor characteristics may counterbalance the effect of high BRCA1 expression in breast cancer. Citation Format: Kohei Chida, Masanori Oshi, Arya Mariam Roy, Itaru Endo, Kazuaki Takabe. Enhanced Cancer Cell Proliferation and Aggressive Phenotype Counterbalance in Breast Cancer with High BRCA1 Gene Expression [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO3-15-10.

Functional Analysis of BRCA1 3'UTR Variants Predisposing to Breast Cancer.

Breast Cancer (BC) is the main female cancer diagnosed worldwide, and it has been described that few genes, such as BRCA1, have a high penetrance for this type of cancer. In this manuscript, we were interested in evaluating the effect of 3'UTR variants on BRCA1 expression. To accomplish this objective, Whole Exome Sequencing (WES) data of 400 patients with unselected BC was used to filter variants located in the region of interest of BRCA1 gene, finding two of them (c.*36C>G and c.*369_373del). miRGate and miRanda in silico tools were used to predict microRNA (miRNA) interaction. The two variants (c.*36C>G, c.*369_373del) were predicted to affect miRNA interaction. After cloning of BRCA1 3'UTR into pMIR-Report vector, the construct was transfected into two BC cell lines (MDA-MB-231 and MCF-7), and the variant c.*36C>G evidenced overexpression of reporter gene luciferase, showing that the transcript was not being degraded by the miRNA in MDA-MB-231 cells. The variant seems to protect against Triple Negative BC probably due to the expression level of miRNA in this particular cell line (MDA-MB-231). This is consistent with the clinical history of the patients who harbor BC Hormone Receptors positive (HR+).