CHEK2 Mutations Research Articles

Abstract PO1-09-02: Universal genetic testing for women with newly diagnosed breast cancer identified many mutations, impacted clinical management and caused no psychological distress to patients

Abstract Background: For newly diagnosed breast cancer (BC) patients, identification of pathogenic germline mutations in hereditary breast cancer (HBC) genes can inform clinical management and risk mitigation strategies and identify patients who would benefit from targeted therapy and clinical trials. Offering germline testing based on NCCN or other guidelines fails to identify all patients with germline HBC gene mutations, potentially with adverse consequences for patients and their families. As an integral part of the study, we have explored the acceptance of universal testing among patients and clinicians. Methods: This is the largest Australian multi-centre prospective study exploring the prevalence of hereditary pathogenic variants in the Australian BC population. 650 newly diagnosed consecutive, consented patients with non-metastatic breast cancer or high-grade DCIS were recruited from May 2020 to March 2023 in 2 phases. Phase 1 (n=157) offered a combination of germline and somatic sequencing. Germline testing was performed by whole genome sequencing on DNA from blood or saliva, and the data analyzed for actionable HBC gene mutations, including large genomic rearrangements. Germline variants were interrogated for pathogenic variants in BRCA1, BRCA2, PALB2, ATM, CHEK2, BARD1, BRIP1, RAD51B, RAD51C, RAD51D, MLH1, MSH2, MSH6, PMS2, CDH1, PTEN, STK11, TP53 and NTHL1.Tumour sequencing will be reported separately. Phase 2 (n=493) included germline exome sequencing with no somatic sequencing. The objectives were to assess the additional actionable HBC gene mutations identified by universal testing compared to algorithmic based (MANCHESTER and BOADICEA scores) and NCCN guidelines, the clinical impact of this identification and the prevalence of germline mutations in the general BC population. A 3-generation pedigree was constructed for every patient for accurate estimation of NCCN, MANCHESTER, and BOADICEA scores. Phase 1 patients completed surveys of perceptions of universal testing, psychological distress and cancer-specific worry pre- and post-testing. Results: 50 carriers of actionable germline mutations were identified: (7.7%). BRCA1 (n=10), BRCA2 (n=12), PALB2 (n=7), CHEK2 (n=10), ATM (n=4), PMS2 (n=2), MSH6 (n=1), RAD51C (n=2), and BARD1 (n=3). One carrier had both PALB2 and CHEK2 mutations. Updated NCCN guidelines (2023 version) identified 44/50 pathogenic germline variant carriers (88%). The BOADICEA and MANCHESTER scores only identified 22 carriers (44%). Immediate clinical management changed due to the discovery of HBC gene in 40/50 (80%). Among 105 participants, cancer-specific distress declined from pre- to post-testing. Anxiety, stress and depression did not change. 90/103 participants agreed that genetic testing should be routine, 100/104 reported their decision to undergo testing as correct. All (n=25) BC clinicians reported that genetic test results were helpful for important treatment decisions and none that testing was distressing to patients. Conclusion: HBC gene mutation prevalence was 7.7%. Universal germline HBC gene testing is the best method for detecting carriers. Current testing guidelines would have missed HBC genes which would have impacted clinical management. Updated NCCN guidelines were sensitive in the detection of HBC gene mutation, but more than half of HBC gene mutations were missed by Australian standards. Discovery of HBC gene mutations led to changed management in most cases. Universal testing is highly acceptable to clinicians and patients who reported no adverse impact on psychological distress or cancer-specific worry and no decision regret. Germline mutations identified and their impact on management Citation Format: Dilanka De Silva, Lesley Stafford, Anita Skandarajah, Michelle Sinclair, Lisa Devereux, Kirsten Hogg, Maira Kentwell, Allan Park, luxi Lal, Magnus Zethoven, Madawa Jayawardena, Fiona Chan, Paul James, Geoffrey Lindeman, Ian Campbell, Bruce Mann. Universal genetic testing for women with newly diagnosed breast cancer identified many mutations, impacted clinical management and caused no psychological distress to patients [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 PO1-09-02.

Abstract PO3-22-08: Impact of Zip Code-Based Income on Reconstructive Method and Outcomes in Mutation-Positive Breast Cancer Patients

Abstract Background: In the United States, 13% of women develop invasive breast cancer, with certain genetic mutations increasing the risk. Genetic screening is costly and burdensome for patients, particularly those from lower socioeconomic backgrounds. Moreover, lower income (LI) patients face barriers in accessing healthcare and have lower rates of breast reconstruction. This study aims to examine the impact of socioeconomic status, as indicated by ZIP code median income, on the type of breast surgery, reconstructive technique, and perioperative outcomes among mutation-positive (M+) patients. Methods: A multicenter retrospective cohort study included patients with a genetic predisposition for breast cancer and undergoing breast and reconstructive surgery between January 2016 and October 2022. Patient demographics, history, operative details, and postoperative outcomes were collected. Patient ZIP codes were cross-referenced with the American Community Survey to determine the median household income, which served as a proxy for the patient-specific household income. Four distinct income quartiles were established: quartile 1 (Q1) = $37,045-$81,586 (“LI”); quartile 2 (Q2) = $81,587-$103,703; quartile 3 (Q3) = $103,704-$129,768; and quartile 4 (Q4) = $129,769-$213,724. Results: Among the 170 patients analyzed, 42 (24.7%) were in Q1, 42 (24.7%) in Q2, 44 (25.9%) in Q3, and 42 (24.7%) in Q4. Most patients had BRCA1 (n=73, 42.9%), BRCA2 (n=68, 40.0%), PALB2 (n=11,6.5%), or CHEK2 (n=8, 64.7%) mutations, and underwent prophylactic mastectomy without evidence of breast cancer (n=119, 70.0%). Mean age and body mass index (BMI) were 41±16 years and 25.0±7.5 kg/m2, respectively. There was a significant association between patient race and income (p< 0.0001), such that 30.1% of White patients were in Q4 (vs. 8.3% of Black patients), compared to 38.9% of Black patients in Q1 (vs.19.0% of White patients). LI was significantly associated with higher BMI (p=0.0083) and obesity rates (p=0.0080). Nipple-sparing mastectomy (NSM) (n=103, 60.6%) was the most frequent surgical approach, followed by skin-sparing mastectomy (SSM) (n=42, 24.7%), simple mastectomy (n=13, 7.6%), lumpectomy (n=4, 2.4%), and radical mastectomy (n=4, 2.4%). Patients in Q3 underwent NSM significantly more often than those in Q1 (p=0.0080). Tissue expander (n=73, 42.9%) was the most common reconstruction method. 34.7% of patients (n=59) experienced postoperative complications. Short-term (< 30 days postoperatively) complications included delayed wound healing (n=22, 12.9%), surgical site infection (n=16, 9.4%), dehiscence (n=12, 7.1%), cellulitis (n=10, 5.9%), seroma (n=9, 5.3%) and hematoma (n=9, 4.7%). Long term (≥30 days postoperatively) complications included mastectomy flap necrosis (n=22, 12.9%), total reconstructive failure (n=8, 4.7%), and red breast syndrome (n=2, 1.2%). There was no significant association between income level and incidence of complications (p >0.05). Income level significantly correlated with cancer recurrence, as patients in Q2 (n=0, 0.0%) and Q4 (n=1, 2.4%) had significantly lower recurrence rates compared to Q1 (n=6, 14.3%) (p=0.0140). Of the 30.0% of patients with diagnosis of cancer at DOS, 66.7% (n=34) of patients had ductal carcinoma in situ (DCIS). Eleven of these patients were in Q1, five in Q2, six in Q3, and twelve in Q4 (p=0.259). Conclusion: LI M+ patients had lower NSM rates than higher income patients, despite similar rates of prophylactic mastectomies, possibly due to perceived costs of increased follow-up care. Moreover, LI patients experienced higher cancer recurrence, despite similar incidences of equivalent cancer subtypes at presentation, likely reflecting differences in post-diagnostic care and morbidity. These findings highlight the need to address financial barriers and ensure equitable access to comprehensive care for M+ patients of all income strata. Citation Format: Jaytin Gupta, Shrayus Sortur, Christian Lava, Margaret Lee, Lauren Berger, Varsha Harish, Daisy Spoer, Lindy Rosal, Ian Greenwalt, Lucy De La Cruz, David Song, Kenneth Fan. Impact of Zip Code-Based Income on Reconstructive Method and Outcomes in Mutation-Positive Breast Cancer Patients [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 PO3-22-08.

Case report: Mutation evolution in a patient with TdT positive high grade B cell lymphoma with MYC and BCL2 rearrangements following the treatment of concurrent follicular lymphoma and diffuse large B-cell lymphoma

BackgroundConcurrent follicular lymphoma (FL) and diffuse large B-cell lymphoma (DLBCL)was reported in some studies, while the diagnosis of TdT (terminal deoxynucleotydil transferase) positive high grade B cell lymphoma (HGBL) with MYC and BCL2 rearrangements (“double hit”) transformed from FL/DLBCL has been rarely reported. Herein, we described the clinical features and mutation profiles of a case diagnosed with TdT positive “double hit” HGBL following the treatment of FL/DLBCL.Case presentationThis is a 43-year-old Chinese man who was diagnosed with low grade FL (account for 80%) combined with DLBCL (20%) at a stage of IVB. The patient presented with BCL2/IGH translocation without MYC rearrangement, as well as the expressions of CD20, CD19, CD10 and BCL2 at the initial diagnosis of FL/DLBCL. MYC rearrangement and TdT expression occurred after the treatment. The targeted sequencing revealed mutations in KMT2D, FOXO1, CREBBP, ATM, STAT6, BCL7A, DDX3X, MUC4, FGFR3, ARID5B, DDX11 and PRKCSH genes were the co-mutations shared by the FL/DLBCL and TdT positive “double hit” HGBL, while CCND3, BIRC6, ROBO1 and CHEK2 mutations specifically occurred after the treatment. The overall survival time was 37.8 and 17.8 months after the initial diagnosis of FL/DLBCL and TdT positive “double hit” HGBL, respectively.ConclusionThis study reports a rare case of TdT positive “double hit” HGBL following the treatment of concurrent FL/DLBCL and highlights the mutation characteristics. Collectively, this study will help enrich the knowledge of TdT positive “double hit” HGBL transformed from FL/DLBCL.

Variation Analysis in Premenopausal and Postmenopausal Breast Cancer Cases.

Menopausal status affects the prognoses and consequences of breast cancer. Therefore, this retrospective study aimed to reveal the molecular variation profile differences in breast cancer patients according to their menopausal status, with the hypothesis that the molecular variation profiles will be different at premenopausal and postmenopausal ages. Breast cancer patients (n = 254) who underwent molecular subtyping and QIAseq Human Breast Cancer NGS Panel screening between 2018 and 2022 were evaluated retrospectively. Their menopausal status was defined by age, and those aged 50 years and above were considered postmenopausal. Of the subjects, 58.66% (n = 149) were premenopausal and 41.34% (n = 105) were postmenopausal. The mean age at the time of diagnosis for all patients was 49.31 ± 11.19 years, with respective values of 42.11 ± 5.51 and 59.54 ± 9.01 years for the premenopausal and postmenopausal groups, respectively (p = 0.000). Among premenopausal patients, the percentages of patients in BCa subtypes (luminal A, luminal B-HER2(-), luminal B-HER2(+), HER2 positive, and triple-negative) were determined to be 34.90%, 8.05%, 26.17%, 10.74%, and 20.13%, respectively, while in the postmenopausal group, these values were 39.05%, 16.19%, 24.76%, 6.67%, and 13.33%, respectively (p > 0.05). Considering menopausal status, the distribution of hormone receptors in premenopausal patients was ER(+)/PgR(+) 63.76%, ER(-)/PgR(-) 23.49%, ER(+)/PgR(-) 10.74%, and ER(-)/PgR(+) 2.01%, respectively, while in postmenopausal women, this distribution was observed to be 74.29%, 23.81%, 1.90% and 0.00% in the same order (p = 0.008). The most frequently mutated gene was TP53 in 130 patients (51.18%), followed by PIK3CA in 85 patients (33.46%), BRCA2 and NF1 in 56 patients (22.05%), PTEN in 54 patients (21.26%), and ATR and CHEK2 in 53 patients (20.87%). TP53, PIK3CA, NF1, BRCA2, PTEN, and CHEK2 mutations were more frequently observed in premenopausal patients, while TP53, PIK3CA, BRCA2, BRCA1, and ATR mutations in postmenopausal patients. These findings contribute to a deeper understanding of the underlying causes of breast cancer with respect to menopausal status. This study is the first from Turkey that reflects the molecular subtyping and somatic mutation profiles of breast cancer patients according to menopausal status.

Immunohistochemical findings and clinicopathological features of breast cancers with pathogenic germline mutations in Non-BRCA genes

Deleterious germline mutations in multiple genes confer an increased breast cancer (BC) risk. Immunohistochemical (IHC) expression of protein products of mutated high-risk genes has not been investigated in BC. We hypothesized that pathogenic mutations may lead to an abnormal IHC expression pattern in the tumor cells. BCs with deleterious germline mutations in CHEK2, ATM, PALB2 &PTEN were identified. Immunohistochemistry was performed using Dako staining platform on formalin fixed paraffin embedded tumor tissue. Primary antibodies for PALB2 (ab202970), ATM [2C1(1A10)}, CHK2 (EPR4325), and PTEN (138G6) proteins were used for BCs with respective deleterious mutations. IHC expression was assessed in tumor and adjacent benign breast tissue. Total 27 BCs with 10 CHEK2, 9 ATM, 6 PALB2 & 2 PTEN deleterious germline mutations were identified. IHC staining was performed on 8 CHEK2, 7 ATM, 6 PALB2 & 2 PTEN cases. Abnormal CHEK2 IHC staining was identified in 7/8(88%) BCs. Three distinct CHK2 IHC patterns were noted: 1) Strong diffuse nuclear positivity (5 BC), 2) Null-pattern (2 BC), & 3) Normal breast–like staining in 1 BC Four of 5 (80%) strong CHK2 staining BC had missense CHEK2 mutations. Null-pattern was present with a missense & a frameshift mutation. Normal breast-like CHEK2 IHC staining pattern was present in 1 BC with CHEK2 frameshift mutation. Loss of nuclear/cytoplasmic PTEN IHC expression was noted in 2 in-situ carcinomas. Abnormal PTEN and CHK2 IHC were present in atypical ductal hyperplasia and flat epithelial atypia. ATM and PALB2 IHC expression patterns were similar in tumor cells and benign breast epithelium: mild to moderate intensity nuclear and cytoplasmic staining. We report abnormal CHEK2 IHC expression in 88% of BCs with pathogenic CHEK2 mutations. With PTEN and CHEK2 pathogenic mutations, abnormal IHC patterns are seen in early atypical proliferative lesions. IHC may be applied to identify CHEK2 &PTEN mutated BCs and precursor lesions.

Effect of inactivating heterozygous mutations in DNA repair genes on experimental lung carcinogenesis in mice

Background. Inactivating mutations in Chek2 and Gprc5a genes are known to be associated with cancer development. Experimental carcinogenesis studies in genetically modified mice generate new data on their influence on pathology development.Aim. In the present study in a model of lung carcinogenesis, survival parameters as well as tumor multiplicity and size in mice with Chek2 and Gprc5a heterozygous inactivating mutations were evaluated.Material and methods. F2 hybrid mice from crosses between CBAB6F1 males heterozygous for the studied mutations and wild-type BALB / c females were used: Chek2dAA-carriers (76 males and 64 females) and Gprc5ainsA-carriers (60 males and 42 females). Starting at four months of age, mice received urethane (ethyl carbamate) intraperitoneally at a dose of 600 mg / kg weekly for 6 weeks. After genotyping by allele-specific PCR, animals were allocated to groups. Carcinogenesis parameters were evaluated 40 weeks after the beginning of the experiment.Results. The proportion of mice with mutations surviving to the age of three months roughly followed the Mendelian distribution (35 / 41 males and 33 / 31 females) for the offspring of males heterozygous for Chek2dAA and was significantly lower in the case of Gprc5ainsA (20 / 40 males and 17 / 25 females, p = 0.043). The death of Gprc5ainsA carriers during the experiment was also higher than in the control group (p = 0.0506 in females). Synchronous lung and thymus neoplasms were found in 2 out of 4 Gprc5ainsA females that died before the end of the experiment, which were not found in other groups. At the end of the experiment, no significant differences in tumor multiplicity, mean linear size, and volume were found between the groups of mice with and without mutations.Conclusion. It was found that heterozygous inactivating mutation Chek2dAA does not affect early age development and does not modify the parameters of induced lung carcinogenesis in mice. Heterozygous carriage of Gprc5ainsA mutation in mice increases the risk of early death and sensitivity to the toxic and carcinogenic effects of urethane.

Genetic mutation patterns among glioblastoma patients in the Taiwanese population – insights from a single institution retrospective study

This study utilized Next-Generation Sequencing (NGS) to explore genetic determinants of survival duration in Glioblastoma Multiforme (GBM) patients. We categorized 30 primary GBM patients into two groups based on their survival periods: extended survival (over two years, N = 17) and abbreviated survival (under two years, N = 13). For identifying pathogenic or likely pathogenic variants, we leveraged the ClinVar database. The cohort, aged 23 to 66 (median: 53), included 17 patients in Group A (survival >2 years, 10 males, 7 females), and 13 patients in Group B (survival <2 years, 8 males, 5 females), with a 60% to 40% male-to-female ratio. Identified mutations included CHEK2 (c.1477 G > A, p.E493K), IDH1 (c.395 G > A, p.R132H), and TP53 mutations. Non-coding regions exhibited variants in the TERT promoter (c.-146C > T, c.-124C > T) and TP53 RNA splicing site (c.376-2 A > C, c.376-2 A > G). While Group A had more mutations, statistical significance wasn’t reached, likely due to sample size. Notably, TP53, and ATR displayed a trend toward significance. Surprisingly, TP53 mutations were more prevalent in Group A, contradicting Western findings on poorer GBM prognosis. In Taiwanese GBM patients, bevacizumab usage is linked to improved survival rates, affirming its safety and effectiveness. EGFR mutations are infrequent, suggesting potential distinctions in carcinogenic pathways. Further research on EGFR mutations and amplifications is essential for refining therapeutic approaches. TP53 mutations are associated with enhanced survival, but their functional implications necessitate detailed exploration. This study pioneers genetic analysis in Taiwanese GBM patients using NGS, advancing our understanding of their genetic landscape.

Association between missense variants of uncertain significance in the CHEK2 gene and hereditary breast cancer: a cosegregation and bioinformatics analysis.

Inherited mutations in the CHEK2 gene have been associated with an increased lifetime risk of developing breast cancer (BC). We aim to identify in the study population the prevalence of mutations in the CHEK2 gene in diagnosed BC patients, evaluate the phenotypic characteristics of the tumor and family history, and predict the deleteriousness of the variants of uncertain significance (VUS). A genetic study was performed, from May 2016 to April 2020, in 396 patients diagnosed with BC at the University Hospital Lozano Blesa of Zaragoza, Spain. Patients with a genetic variant in the CHEK2 gene were selected for the study. We performed a descriptive analysis of the clinical variables, a bibliographic review of the variants, and a cosegregation study when possible. Moreover, an in-depth bioinformatics analysis of CHEK2 VUS was carried out. We identified nine genetic variants in the CHEK2 gene in 10 patients (two pathogenic variants and seven VUS). This supposes a prevalence of 0.75% and 1.77%, respectively. In all cases, there was a family history of BC in first- and/or second-degree relatives. We carried out a cosegregation study in two families, being positive in one of them. The bioinformatics analyses predicted the pathogenicity of six of the VUS. In conclusion, CHEK2 mutations have been associated with an increased risk for BC. This risk is well-established for foundation variants. However, the risk assessment for other variants is unclear. The incorporation of bioinformatics analysis provided supporting evidence of the pathogenicity of VUS.

Relationship between the Expression of CHK2 and p53 in Tumor Tissue and the Course of Papillary Thyroid Cancer in Patients with CHEK2 Germline Mutations.

The aim of this study was to determine whether the expression of CHK2 and p53 in tumor tissue in carriers of germline CHEK2 mutations can serve as a prognostic marker for PTC, and whether CHEK2 and TP53 copy numbers correlates with the course of PTC disease. This study included 156 PTC patients previously tested for the presence of CHEK2. Clinicopathological features, treatment response, disease outcome, and germline mutation status of the CHEK2 gene were assessed with respect to CHK2 and p53 expression, and CHEK2 and TP53 gene copy statuses. In patients with and without a germline mutation in CHEK2 and with higher CHK2 expression, the chances of an excellent treatment response and no evidence of disease were lower than in patients without or with lower CHK2 expression. TP53 deletion was associated with angioinvasion. In patients with a truncating mutation, the chance of a CHEK2 deletion was higher than in patients with WT CHEK2 alone or those with WT CHEK2 and with the missense I157T mutation. Higher CHK2 expression was associated with poorer treatment responses and disease outcomes. Higher CHK2 expression and positive p53 together with a TP53 deletion could be a prognostic marker of unfavorable disease outcomes in patients with germline truncating mutations in CHEK2.

Survival impact of homologous recombination deficiency in veterans with cholangiocarcinoma including mutual exclusivity with pathogenic KRAS and TP53.

546 Background: Cholangiocarcinoma (CCA) has poor prognosis and limited treatment options. The US Veteran Health Administration’s (VHA) National Precision Oncology Program (NPOP) was established to characterize pathogenic drivers across the integrated VA network. Here, we summarize an important subgroup of CCA with homologous recombination deficiency (HRD) and its impact on overall survival (OS). Methods: Next-generation sequencing (NGS) was performed using FoundationOne CDx Analysis included clinicopathological, molecular alterations including, single nucleotide variants, gene amplification/loss, and gene fusion/rearrangements obtained from centralized data of the VHA’s Corporate Data Warehouse. Homologous recombination deficiency was defined by pathogenic variant mutations (MT) in ARID1A, ATM, ATR, BAP1, BRCA1, BRCA2, CDK12, CHEK2, FANC, NBN, PALB2, and RAD51. Statistical analysis was performed using Chi-squared test for concurrent gene enrichment and log-rank Kaplan-Meier analysis. Results: 483 CCA samples underwent tissue NGS as standard of care. Pathogenic variants in HRD genes included ARID1A (15.5%, n=75), BAP1 (9.5%, n=46), ATM (2.7%, n=13), PALB2 (2.3%, n=11) and BRCA (2.0%, n=10), representing 31.7% (n=153) of the population. HRD mutations were found in similar proportions of localized (31.8%) and metastatic (31.5%) clinical presentations at diagnosis. OS was not significantly different between CCA with HRD mutations (12.3 mo WT vs. 13.8 months with HRD MT, p=64). KRAS MT (n=77, 15.9% of population) did not impact OS (WT 12.3 mo v. MT 13.1 mo) including subgroup analysis stratified by HRD status. There was increased frequency in KRAS MT with HRD WT 18.5% versus HRD MT10.4% (p&lt;0.03). Additionally, there was increased frequency in TP53 MT with HRD WT 54.8% versus HRD MT 28.1% (p&lt;0.00001). TP53 MT (n=224, 46.3% of population) conferred inferior OS (WT 12.3 mo v. MT 13.1 mo) including subgroup analysis stratified by HRD status. Subgroup analysis of TP53 MT had worsened OS with concurrent HRD MT (n=39, 4.1 mo) v. HRD WT (n=158, 9.0 mo; p&lt;0.04). Conclusions: Across a diverse integrated healthcare system of Veterans, mutations associated with HRD were found to be common, however, not prognostic in OS across a diverse patient population of CCA. Further work is needed to clarify exposure to platinum containing chemotherapy. TP53 MT CCA remains a critical unmet need that drives worsened outcomes including here in patients with mutations in HRD. Future analysis should consider mutation signatures of HRD, loss in heterozygosity of HRD, and platinum exposure to further define the subgroups of CCA with mutations in HRD. [Table: see text]

CHEK2 knockout is a therapeutic target for TP53-mutated hepatocellular carcinoma

Currently, there is still a lack of novel and effective drug targets to improve the prognosis of hepatocellular carcinoma (HCC). Additionally, the role of CHEK2 in HCC has not been reported yet. The eQTLgen database and two HCC Genome-Wide Association Study (GWAS) datasets (ieu-b-4953, ICD10 C22.0) were used to find the drug target: CHEK2. Next, Colony, Edu, β-gal, and cell cycle analysis were facilitated to evaluate the role of CHEK2 knockout in HCC. In addition, Nultin-3 was added to evaluate the apoptosis of TP53-mutated HCC cells with CHEK2 knockout. Furthermore, MitoSox, electron microscopy, mitochondrial ATP, and NADH+/NADH levels were assessed in the CHEK2 knockout HCC cells with or without Metformin. Finally, cell-derived tumor xenograft was used to evaluate the role of CHEK2 knockout in vivo. We initially identified a potential drug target, CHEK2, through GWAS data analysis. Furthermore, we observed a significant upregulation of CHEK2 expression in HCC, which was found to be correlated with a poor prognosis. Subsequently, the results indicated that knocking out CHEK2 selectively affects the proliferation, cell cycle, senescence, and apoptosis of TP53-mutant HCC cells. Additionally, the introduction of Nultin-3 further intensified the functional impact on TP53-mutant cells. Then ClusterProfiler results showed high CHEK2 and TP53 mutation group was positively enriched in the mitochondrial ATP pathway. Then we used MitoSox, electron microscopy, mitochondrial ATP, and NADH + /NADH assay and found knockout of CHECK could induce the ATP pathway to inhibit the growth of HCC. Our research introduces a novel drug target for TP53-mutant HCC cells via mitochondrial ATP, addressing the limitation of Nultin-3 as a standalone treatment that does not induce tumor cell death.

O-11 A rare cause of thyroid cancer - Li Fraumeni 2 syndrome

Abstract Introduction Thyroid cancer often originates from follicular cells, and papillary thyroid cancer (PTC) is the most common type. There is a familial predisposition in some cases of thyroid cancer. However, a minority of cases have thyroid cancer syndrome (e.g., familial polyposis, MEN 2, or Cowden syndrome). Many known genetic susceptibility genes are RET/PTC, BRAF, RAS, TERT, and TP53. We want to present a case of Li Fraumeni type 2 syndrome as a rare cause of thyroid cancer. Clinical Case A 69-year-old female patient with a history of PTC is admitted to the endocrinology department for follow-up. In 1976, this patient had a thyroid lobectomy, but the pathology result is unknown, and she takes 75 mcg of levothyroxine daily. In 1999, she was diagnosed with rectal cancer. The patient had a completion thyroidectomy in 2002, and the pathology was compatible with PTC. In 2014, she was diagnosed with right breast noninvasive ductal cancer. Cases of cancer were also quite common in her family history. The results of laboratory tests revealed normal TSH levels with negative thyroglobulin and anti-thyroglobulin levels. Neck ultrasonography showed no rest gland and pathological LAP. We requested genetic analysis (55 genes) for familial cancer syndromes. As a result of the analysis, 2 genes were found to be heterozygous. The c.599T&amp;gt;c heterozygous variant in the CHEK 2 gene was found (Figure 1). It was evaluated as pathogenic in the database review. Another heterozygous mutation in the BRIP1 gene, which was evaluated as a VUS also detected. Conclusion The CHEK 2 gene is a DNA-repairing protein kinase at position 22.q12.1. It is responsible for TP53 stabilization, genome maintenance, and apoptosis. CHEK2 phosphorylates p53 in response to DNA damage, leading to cell cycle arrest and stabilization (Figure 2). The mutation(CHEK 2 c.599T&amp;gt;c) we detected in this case has been considered possibly pathogenic in the academic databases. There are reported CHEK 2 c.599T&amp;gt;c cases on the CLINVAR site; the first report is from 2013. Classic Li-Fraumeni syndrome is a rare autosomal dominant syndrome caused by TP53 gene mutation. The chromosome location on classical Li fraumeni is 17p13.1. Li-Fraumeni type 2 develops due to the CHEK 2 gene mutation on chromosome 22q12.1. The results of a mutation of the CHEK2 gene are breast cancer, prostate cancer, PTC, colon cancer, kidney cancer, adrenocortical cancer, lung cancer, and rare cases of myelodysplastic syndromes. There are no data on thyroid-specific mortality in carriers of the CHEK2 mutations, and it is not yet known whether thyroid cancer in this population is more aggressive or has a better outcome.The familial syndromes should also be considered if PTC is detected in breast, colon, and prostate cancer patients. Studies on Li-Fraumeni type 2 syndrome are scarce, and considering the prevalence in European countries, further investigations of this syndrome and screening in patients with multiple cancers in the family should be warranted.Figure 1.Genetic sequencing image of the case

Abstract B025: Exceptional response to the ataxia telangiectasia and Rad3-related inhibitor (ATRi), camonsertib, in a patient with alternative lengthening of telomeres (ALT)-positive metastatic melanoma

Abstract One hallmark of cancer tumorigenesis is the maintenance of telomere length which canonically occurs through the reactivation of telomerase. Alternative lengthening of telomeres (ALT) is a minor telomere maintenance mechanism that uses homologous recombination (HR) to maintain telomere length and is associated with replication stress and defects in genome maintenance. In preclinical models, ALT positivity (ALT+) has been shown to sensitize tumor cells to ataxia telangiectasia and Rad3-related inhibitors (ATRi). Here we describe a confirmed clinical and molecular response to the ATRi, camonsertib, in a patient with ALT+ metastatic melanoma. To our knowledge, this is the first report of clinical experience with ATRi in ALT+ tumors. A 63-year-old male was diagnosed in July 2019 with stage IV (cTX, cNX, pM1c[1]) metastatic melanoma and treated with nivolumab (nivo) + ipilumumab (ipi) with initial partial response (PR) documented after 3 months which was confirmed on subsequent nivo maintenance. Ipi was resumed following suspected pseudoprogression in March 2020, and continued until confirmed progression in June 2020. Genomic profiling of a liver biopsy collected at the time of progression identified somatic alterations in TP53 (p.S127F, p.R65fs), NF1 (p.K1263*), BRCA2 (p.R2482C) and ATRX (p.R418*), a germline CHEK2 mutation (p.T376fs), and high tumor mutational burden. The patient received additional targeted regimens with olaparib (ola): nivo ± ola and later ola ± trametinib; no response was documented, and progression was noted in April 2021. In June 2021, the patient received camonsertib monotherapy at an initial dose of 120 mg QD 3 days on/4 days off in the phase I TRESR trial (NCT04497116). The patient tolerated treatment well and continued at the same dose until dose was reduced via a change in schedule (to 2 weeks on/1 week off) at 9 months due to anemia. The patient had an initial RECIST PR (-36%) at 3 months with circulating tumor DNA (ctDNA) clearance. The target lesions continued to decrease with a best decline of -74% after about 1 year of treatment. The cancer progressed at 19 months (+105% from nadir, -47% from baseline) though due to ongoing clinical benefit treatment was continued for a total duration of 22 months. Interrogation of ctDNA tumor fraction in longitudinal plasma samples is ongoing and will be reported. To understand the long and deep response to camonsertib treatment in this patient, we further characterized the screening biopsy by a custom targeted sequencing panel, SyNthetic lethal Interactions for Precision Diagnostics (SNiPDx), and whole genome sequencing (WGS). Both the BRCA2 and gCHEK2 mutations were monoallelic and genomic signature analysis defined the tumor as HR-proficient, consistent with the lack of benefit from ola. WGS confirmed a biallelic loss of ATRX with a genomic signature consistent with an ALT+ phenotype. The clinical experience of ATRi therapy in patients with ALT+ tumors is limited but further investigation of ATRi such as camonsertib is warranted in this patient population. Citation Format: Timothy A. Yap, Ian M. Silverman, Adrienne Johnson, Chenfeng Meng, Joseph D. Schonhoft, Michal Zimmermann, Danielle Ulanet, Victoria Rimkunas, Maria Koehler. Exceptional response to the ataxia telangiectasia and Rad3-related inhibitor (ATRi), camonsertib, in a patient with alternative lengthening of telomeres (ALT)-positive metastatic melanoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: DNA Damage Repair: From Basic Science to Future Clinical Application; 2024 Jan 9-11; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2024;84(1 Suppl):Abstract nr B025.

Germline DNA Damage Response Gene Mutations in Localized Prostate Cancer.

Background and Objectives: Germline DNA damage response (DDR) gene mutations correlate with increased prostate cancer (PCa) risk and a more aggressive form of the disease. DDR mutation testing is recommended for metastatic PCa cases, while eligible information about the mutations' burden in the early-stage localized PCa is still limited. This study is aimed at the prospective detection of DDR pathway mutations in cases with localized PCa and correlation with clinical, histopathological, and radiological data. A comparison to the previously assessed cohort of the advanced PCa was performed. Materials and Methods: Germline DDR gene mutations were assessed prospectively in DNA samples from 139 patients, using a five-gene panel (BRCA1, BRCA2, ATM, CHEK2, and NBN) targeted next-generation sequencing. Results: This study revealed an almost three-fold higher risk of localized PCa among mutation carriers as compared to non-carriers (OR 2.84 and 95% CI: 0.75-20.23, p = 0.16). The prevalence of germline DDR gene mutations in PCa cases was 16.8% (18/107) and they were detected only in cases with PI-RADS 4/5 lesions. BRCA1/BRCA2/ATM mutation carriers were 2.6 times more likely to have a higher (>1) cISUP grade group compared to those with a CHEK2 mutation (p = 0.27). However, the number of cISUP > 1-grade patients with a CHEK2 mutation was significantly higher in advanced PCa than in localized PCa: 66.67% vs. 23.08% (p = 0.047). Conclusions: The results of our study suggest the potential of genetic screening for selected DDR gene mutations for early identification of cases at risk of aggressive PCa.

Germline mutations in pediatric cancer cohort with mixed-ancestry Mexicans.

Childhood cancer is one of the primary causes of disease-related death in 5- to 14-year-old children and currently no prevention strategies exist to reduce the incidence of this disease. Childhood cancer has a larger hereditary component compared with cancer in adults. Few genetic studies have been conducted on children with cancer. Additionally, Latin American populations are underrepresented in genomic studies compared with other populations. Therefore, the aim of this study is to analyze germline mutations in a group of mixed-ancestry Mexican pediatric patients with solid and hematological cancers. We analyzed genetic variants from 40 Mexican childhood cancer patients and their relatives. DNA from saliva or blood samples was used for whole-exome sequencing. All variants were identified following GATK best practices. We found that six patients (15%) were carriers of germline mutations in CDKN2A, CHEK2, DICER1, FANCA, MSH6, MUTYH, NF1, and SBDS cancer predisposition genes, and additional new variants predicted to be deleterious by in silico algorithms. A population genetics analysis detected five components consistent with the demographic models assumed for modern mixed-ancestry Mexicans. This report identifies potential genetic risk factors and provides a better understanding of the underlying mechanisms of childhood cancer in this population.

Breast cancers in monoallelic MUTYH germline mutation carriers have clinicopathological features overlapping with those in BRCA1 germline mutation carriers.

Breast cancer patients referred to genetic counseling often undergo genetic testing with broad panels that include both breast cancer susceptibility genes as well as genes more specific for extramammary sites. As a result, patients are often incidentally found to have germline mutations in genes that are not necessarily related to breast cancer risk. One such gene is MUTYH. To understand the role MUTYH may play in breast cancer, the clinicopathological features of patients with monoallelic MUTYH germline mutation and breast cancer were examined. The clinicopathological characteristics of the breast cancers from patients with monoallelic MUTYH mutation were compared to breast cancer patients with other germline mutations in known breast cancer susceptibility genes, including ATM, BRCA1/2, CHEK2, and PALB2. The breast cancer patients who received genetic counseling but tested negative for the aforementioned gene mutations were used as a control group. Histologic characteristics of the breast cancers arising in monoallelic MUTYH mutation carriers had significantly larger tumor size, higher tumor grade, and more high-risk biomarker profiles (i.e., Her2-positive and triple-negative) than breast cancer patients with susceptibility genes, except for BRCA1. MUTYH mutation carriers also showed a trend of more frequent intratumoral divergency in terms of tumor grade and biomarker profiles. Although germline monoallelic MUTYH mutation is not thought to confer a meaningfully increased risk of breast cancer development, it may contribute to pathological aggressiveness and diversity of breast cancers when they sporadically arise in MUTYH carriers.

Profiling of the genetic features of patients with breast, ovarian, colorectal and extracolonic cancers: Association to CHEK2 and PALB2 germline mutations

Background and aimsCancer predisposition goes beyond BRCA and DNA Mismatch Repair (MMR) genes since multi-gene panel testing has become the routine diagnostic tool for hereditary cancer suspicion (HCS) cases. CHEK2 and PALB2 are some of the foremost-mutated non-BRCA/MMR actionable genes in families with a significant familial aggregation. Therefore, the purpose of this work is to unravel which tumours other than breast, ovary or colorectal display the patients. Materials and methodsWe have analysed 528 probands that meet the inclusion criteria for Hereditary Breast and Ovarian Cancer and Lynch Syndrome established by our Hereditary Cancer Regional Program with a customized 35 genes-panel by using Ion Torrent™ Technology. ResultsWe have identified pathogenic variants (PVs) in 61 families (1.55%), of which more than half (31 probands) harboured PVs in CHEK2 and PALB2 genes. Ours results reveal that not only were PVs CHEK2 and PALB2 carriers more likely to have family history of cancer not limited to breast, ovarian or colorectal cancers, but also they are prone to other extracolonic cancers, noteworthy endometrial and gastric cancers. ConclusionsMultigene panel testing improves the chance of finding PVs in actionable genes in families with HCS. In addition, the coexistence of variants should be recorded to implement a polygenic risk algorithm that might explain the missing heritability in the aforementioned families.

Mutation in Chek2 triggers von Hippel-Lindau hemangioblastoma growth

PurposeVon Hippel-Lindau (VHL) is a rare inherited disease mainly characterized by the growth of tumours, predominantly hemangioblastomas (Hbs) in the CNS and retina, and renal carcinomas. The natural history of VHL disease is variable, differing in the age of onset and its penetrance, even among relatives. Unfortunately, sometimes VHL shows more severe than average: the onset starts in adolescence, and surgeries are required almost every year. In these cases, the factor that triggers the appearance and growth of Hbs usually remains unknown, although additional mutations are suspected.MethodsWe present the case of a VHL patient whose first surgery was at 13 years of age. Then, along his next 8 years, he has undergone 5 surgeries for resection of 10 CNS Hbs. To clarify this severe VHL condition, DNA from a CNS Hb and white blood cells (WBC) was sequenced using next-generation sequencing technology.ResultsMassive DNA sequencing of the WBC (germ line) revealed a pathogenic mutation in CHEK2 and the complete loss of a VHL allele (both tumour suppressors). Moreover, in the tumour sample, several mutations, in BRAF1 and PTPN11 were found. Familiar segregation studies showed that CHEK2 mutation was in the maternal lineage, while VHL was inherited by paternal lineage.ConclusionsFinally, clinical history correlated to the different genotypes in the family, concluding that the severity of these VHL manifestations are due to both, VHL-and-CHEK2 mutations. This case report aims to notice the importance of deeper genetic analyses, in inherited rare diseases, to uncover non-expected mutations.

Efficacy of olaparib in advanced cancers with germline or somatic mutations in BRCA1, BRCA2, CHEK2 and ATM, a Belgian Precision tumor-agnostic phase II study

The Belgian Precision initiative aims to maximize the implementation of tumor-agnostic next-generation sequencing in patients with advanced cancer and enhance access to molecularly guided treatment options. Academic tumor-agnostic basket phase II studies are part of this initiative. The current investigator-driven trial aimed to investigate the efficacy of olaparib in advanced cancers with a (likely) pathogenic mutation (germline or somatic) in a gene that plays a role in homologous recombination (HR). This open-label, multi-cohort, phase II study examines the efficacy of olaparib in patients with an HR gene mutation in their tumor and disease progression on standard of care. Patients with a somatic or germline mutation in the same gene define a cohort. For each cohort, a Simon minimax two-stage design was used. If a response was observed in the first 13 patients, 14 additional patients were included. Here, we report the results on four completed cohorts: patients with a BRCA1, BRCA2, CHEK2 or ATM mutation. The overall objective response rate across different tumor types was 11% in the BRCA1-mutated (n= 27) and 21% in the BRCA2-mutated (n= 27) cohorts. Partial responses were seen in pancreatic cancer, gallbladder cancer, endocrine carcinoma of the pancreas and parathyroid cancer. One patient with a BRCA2 germline-mutated colon cancer has an ongoing complete response with 19+ months on treatment. Median progression-free survival in responding patients was 14+ months (5-34+ months). The clinical benefit rate was 63% in the BRCA1-mutated and 46% in the BRCA2-mutated cohorts. No clinical activity was observed in the ATM (n= 13) and CHEK2 (n= 14) cohorts. Olaparib showed efficacy in different cancer types harboring somatic or germline mutations in the BRCA1/2 genes but not in ATM and CHEK2. Patients with any cancer type harboring BRCA1/2 mutations should have access to olaparib.

152P Efficacy of olaparib in advanced cancers with germline or somatic tumor mutations in BRCA1, BRCA2, CHEK2 and ATM: A Belgian precision tumor-agnostic phase II study

152P Efficacy of olaparib in advanced cancers with germline or somatic tumor mutations in BRCA1, BRCA2, CHEK2 and ATM: A Belgian precision tumor-agnostic phase II study