Allele Frequencies Research Articles

An overview of autosomal STRs and identity SNPs in a Norwegian population using massively parallel sequencing

In recent years, probabilistic genotyping software has been adapted for the analysis of massively parallel sequencing (MPS) forensic data. Likelihood ratios (LR) are based on allele frequencies selected from populations of interest. This study provides an outline of sequence-based (SB) allele frequencies for autosomal short tandem repeats (aSTRs) and identity single nucleotide polymorphisms (iSNPs) in 371 individuals from Southern Norway. 27 aSTRs and 94 iSNPs were previously analysed with the ForenSeq™ DNA Signature Prep Kit (Verogen). The number of alleles with frequencies less than 0.05 for sequenced-based alleles was 4.6 times higher than for length-based alleles. Consistent with previous studies, it was observed that sequence-based data (both with and without flanks) exhibited higher allele diversity compared to length-based (LB) data; random match probabilities were lower for SB alleles confirming their advantage to discriminate between individuals. Two alleles in markers D22S1045 and Penta D were observed with SNPs in the 3´ flanking region, which have not been reported before. Also, a novel SNP with a minor allele frequency (MAF) of 0.001, was found in marker TH01. The impact of the sample size on minor allele frequency (MAF) values was studied in 88 iSNPs from Southern Norway (n = 371). The findings were then compared to a larger Norwegian population dataset (n = 15,769). The results showed that the smaller Southern Norway dataset provided similar results, and it was a representative sample. Population structure was analyzed for regions within Southern Norway; FST estimates for aSTR and iSNPs did not indicate any genetic structure. Finally, we investigated the genetic differences between Southern Norway and two other populations: Northern Norway and Denmark. Allele frequencies between these populations were compared, and we found no significant frequency differences (p-values > 0.0001). We also calculated the pairwise FST values per marker and comparisons between Southern and Northern Norway showed small differences. In contrast, the comparisons between Southern Norway and Denmark showed higher FST values for some markers, possibly driven by distinct alleles that were present in only one of the populations. In summary, we propose that allele frequencies from each population considered in this study could be used interchangeably to calculate genotype probabilities.

Trans-Atlantic genomic differentiation and parallel environmental and allelic variation in Lumpfish (Cyclopterus lumpus )

Abstract In marine species with large populations and high dispersal potential, large-scale genetic differences and clinal trends in allele frequency can provide insight into the evolutionary processes that shape diversity. Lumpfish, Cyclopterus lumpus, is found throughout the North Atlantic and has traditionally been harvested for roe and more recently used as a cleaner fish in salmon aquaculture. We used a 70 K SNP array to evaluate trans-Atlantic differentiation, genetic structuring, and clinal variation across the North Atlantic. Basin-scale structuring between the Northeast and Northwest Atlantic was significant, with enrichment for loci associated with developmental/mitochondrial function. We identified a putative structural variant on chromosome 2, likely contributing to differentiation between Northeast and Northwest Atlantic Lumpfish, and consistent with post-glacial trans-Atlantic secondary contact. Redundancy analysis identified climate associations both in the Northeast (N = 1269 loci) and Northwest (N = 1637 loci), with 103 shared loci between them. Clinal patterns in allele frequencies were observed in some loci (15%—Northwest and 5%—Northeast) of which 708 loci were shared and involved with growth, developmental processes, and locomotion. The combined evidence of trans-Atlantic differentiation, environmental associations, and clinal loci, suggests that both regional and large-scale potentially-adaptive population structuring is present across the North Atlantic.

Phenomic landscape and pharmacogenomic implications for HLA region in a Taiwan Han Chinese population

BackgroundThe human leukocyte antigen (HLA) genes, exhibiting significant genetic diversity, are associated with susceptibility to various clinical diseases and diverse in drug responses. High costs of HLA sequencing and the population-specific architecture of this genetic region necessitate the establishment of a population-specific HLA imputation reference panel. Moreover, there is a lack of understanding about the genetic and phenotypic landscape of HLA variations within the Taiwanese population.MethodsWe created models for a Taiwanese-specific HLA imputation reference panel. These models were trained with the array genotype data and HLA sequencing data from 845 Taiwanese subjects. HLA imputation was applied for 59,448 Taiwanese subjects to characterize the HLA allele and haplotype frequencies. Additionally, a phenome-wide association study (PheWAS) was conducted to identify the phenotypes associated with HLA variations. The association of the biallelic HLA variants with the binary and quantitative traits were evaluated with additive logistic and linear regression models, respectively. Furthermore, an omnibus test with likelihood-ratio test was applied for each HLA amino acid position in the multiallelic HLA amino acid polymorphisms to compare the difference between a fitted model and a null model following a χ2 distribution of n-1 degree of freedom at a position with n residues. Finally, we estimated the prevalence of adverse drug reactions (ADR)-related HLA alleles in the Taiwanese population.ResultsIn this study, the reference panel models displayed remarkable accuracy, with averages of 99.3%, 98.9%, and 99.1% for 2-, 4-, 6-digit alleles of the eight classical HLA genes, respectively. For PheWAS, a total of 18,136 significant associations with HLA variants across 26 phenotypes are identified (p < 5×10-8), highlighting the pleiotropy feature of the HLA region. Among the independent signals, 15 are novel, including the association of HLA-B pos 138 variation with ankylosing spondylitis (AS), and rs9266290 and rs9266292 with allergy. Through an analysis spanning the entire HLA region, we identified clusters of phenotype correlations. Finally, the carriers of pharmacogenomic related HLA alleles, including HLA-C*01:02 (35.86%), HLA-B*58:01 (20.9%), and HLA-B*15:02 (8.38%), were characterized in the Taiwanese general population.ConclusionsWe successfully delivered the HLA imputation for 59,448 Taiwanese subjects and characterized the genetic and phenotypic landscapes of the HLA variations. In addition, we quantified the estimated prevalence of the ADR-related HLA alleles in the Taiwanese population. The developed HLA imputation reference panel could be used for estimation of population HLA allele frequencies, which can facilitate further studies in the role of HLA variants in a wider range of phenotypes in the population.

Comparing vitamin D receptor gene polymorphisms in rs11568820, rs7970314, rs4334089 between COVID-19 patients with mild and severe symptoms: a case control study

The associations of vitamin D receptor (VDR)- single nucleotide polymorphisms (SNPs) with the symptoms of COVID-19 may vary between patients with different severities of COVID-19. Therefore, in the present study, we aim to compare VDR polymorphisms in severe and mild COVID-19 patients. In this study, a total number of 85 hospitalized patients and 91 mild/moderate patients with COVID-19 were recruited. SNPs in VDR genes were determined using ARMS and then confirmed by sanger sequencing. The mean (SD) age of participants in hospitalized and non-hospitalized group was 59.0 (12.4) and 47.8 (14.8) years, respectively. Almost 46% of participants in hospitalized and 48% of participant in non-hospitalized group were male. The frequency of TT genotype of SNP rs11568820 was significantly lower in hospitalized than non-hospitalized group (3.5% vs. 17.6%; P = 0.018). However, there was no significant differences between genotypes of SNPs rs7970314 and rs4334089 and also alleles frequencies in all SNPs of two groups. The genotype of rs11568820 SNP had an inverse association with hospitalization of patients with COVID-19 after adjustment for comorbidities [OR 0.18, 95% CI 0.04, 0.88; P = 0.034]. While, there was no relationship between genotypes of SNPs rs7970314 and rs4334089 and hospitalization. The TT genotype of rs11568820 plays protective role in sever COVID-19 and hospitalization. Further studies with a large sample size which consider various confounding factors are warranted to confirm our results.

Genetic variation in two candidate genes against gastrointestinal parasites in Colombian Hair Sheep

Objective. To characterize two SNP-type genetic polymorphisms in the GLI1 (rs411868094) and IL20RA (rs419463995) genes, candidates for resistance against gastrointestinal parasites in two biotypes of Colombian hair sheep. Materials and methods. From the DNA bank of the Animal Genetics Laboratory of the University of Sucre, 167 samples, belonging to the Ethiopian (n=94) and Sudan (n=73) biotypes, were analyzed by PCR and subsequent bidirectional sequencing of two SNPs in the GLI1 (T&gt;G) and IL20RA (G&gt;A) genes. Allelic and genotypic frequencies, observed (Ho) and expected (He) heterozygosity, F index, and Hardy-Weinberg equilibrium deviations (EHW) were calculated using the GENALEX version 6.5 program. Results. For the GLI1 locus, the mean genotypic frequencies were 0.155±0.07, 0.370±0.07, and 0.475±0.07 for GG, GT, and TT, respectively. In the Ethiopian biotype, the highest frequencies of the genotype of interest (GG) were found. For the IL20RA locus, the AA and AG genotypes had similar and the highest frequency (0.465±0.03) compared to the GG genotype (0.110±0.01). The genotype of interest at this locus (AA) was the most frequent in both OPC biotypes. Conclusions. The alleles of interest associated with low FEC had a low frequency for the GLI1 gene, but a high frequency for IL20RA. The Ethiopian OPC biotype showed the highest frequencies of the genotypes of interest.

Selection leads to remarkable variability in the outcomes of hybridisation across replicate hybrid zones.

Hybrid zones have been viewed as an opportunity to see speciation in action. When hybrid zones are replicated, it is assumed that if the same genetic incompatibilities are maintaining reproductive isolation across all instances of secondary contact, those incompatibilities should be identifiable by consistent patterns in the genome. In contrast, changes in allele frequencies due to genetic drift should be idiosyncratic for each hybrid zone. To test this assumption, we simulated 20 replicates of each of 12 hybrid zone scenarios with varied genetic incompatibilities, rates of migration, selection and different starting population size ratios of parental species. We found remarkable variability in the outcomes of hybridisation in replicate hybrid zones, particularly with Bateson-Dobzhansky-Muller incompatibilities and strong selection. We found substantial differences among replicates in the overall genomic composition of individuals, including admixture proportions, inter-specific ancestry complement and number of ancestry junctions. Additionally, we found substantial variation in genomic clines among replicates at focal loci, regardless of locus-specific selection. We conclude that processes other than selection are responsible for some consistent outcomes of hybridisation, whereas selection on incompatibilities can lead to genomically widespread and highly variable outcomes. We highlight the challenge of mapping between pattern and process in hybrid zones and call attention to how selection against incompatibilities will commonly lead to variable outcomes. We hope that this study informs future research on replicate hybrid zones and encourages further development of statistical techniques, theoretical models and exploration of additional axes of variation to understand reproductive isolation.

Influencing risk factors of voriconazole-induced liver injury in Uygur pediatric patients undergoing allogeneic hematopoietic stem cell transplantation

PurposeWe aimed to investigated the influencing risk factors of voriconazole-induced liver injury in Uygur pediatric patients undergoing allogeneic hematopoietic stem cell transplantation (HSCT).MethodsThis was a prospective cohort design study. High-performance liquid chromatography-mass spectrometry was employed to monitor voriconazole concentration. First-generation sequencing was performed to detect gene polymorphisms. Indicators of liver function were detected at least once before and after voriconazole therapy.ResultsForty-one patients were included in this study, among which, 15 patients (36.6%) had voriconazole-induced liver injury. The proportion of voriconazole trough concentration > 5.5 μg·mL−1 patients within the DILI group (40.0%) was significantly higher compared to the control group (15.4%) (p < 0.05). After administration of voriconazole, the values of ALT (103.3 ± 80.3 U/L) and AST (79.9 ± 60.6 U/L) in the DILI group were higher than that in the control group (24.3 ± 24.8 and 30.4 ± 8.6 U/L) (p < 0.05). There was no significant difference between the two groups in genotype and allele frequencies of CYP2C19*2, CYP2C19*3, CYP2C19*17, and UGT1A4 (rs2011425) (p > 0.05).ConclusionThere was a significant correlation between voriconazole-induced liver injury and voriconazole trough concentration in high-risk Uygur pediatric patients with allogeneic HSCT.

Modified Unit-Mediated Strand Displacement Reactions for Direct Detection of Single Nucleotide Variants in Active Double-Stranded DNA.

Accurate identification of single nucleotide variants (SNVs) in key driver genes holds a significant value for disease diagnosis and treatment. Fluorescent probes exhibit tremendous potential in specific, high-resolution, and rapid detection of SNVs. However, additional steps are required in most post-PCR assays to convert double-stranded DNA (dsDNA) products into single-stranded DNA (ssDNA), enabling them to possess hybridization activity to trigger subsequent reactions. This process not only prolongs the complexity of the experiment but also introduces the risk of losing target information. In this study, we proposed two strategies for enriching active double-stranded DNA, involving PCR based on obstructive groups and cleavable units. Building upon this, we explored the impact of modified units on the strand displacement reaction (SDR) and assessed their discriminatory efficacy for mutations. The results showed that detection of low variant allele frequencies (VAF) as low as 0.1% can be achieved. The proposed strategy allowed orthogonal identification of 45 clinical colorectal cancer tissue samples with 100% specificity, and the results were generally consistent with sequencing results. Compared to existing methods for enriching active targets, our approach offers a more diverse set of enrichment strategies, characterized by the advantage of being simple and fast and preserving original information to the maximum extent. The objective of this study is to offer an effective solution for the swift and facile acquisition of active double-stranded DNA. We anticipate that our work will facilitate the practical applications of SDR based on dsDNA.

Abstract PO1-14-01: Clonal Architecture of circulating tumor DNA predicts Early Progression on Tamoxifen With or Without Palbociclib in Hormone Receptor-Positive Advanced Breast Cancer: NCCH1607/PATHWAY trial

Abstract Objective: Some patients with poor prognosis hormone receptor-positive, human epidermal growth factor receptor 2-negative (HR+/HER2-) advanced breast cancer (ABC) experience early disease progression on the combination of CDK4/6 inhibitors and endocrine therapy. We aimed to determine whether clonal architecture assessed by circulating tumor DNA (ctDNA) could identify patients at higher risk of early progression on tamoxifen therapy with or without palbociclib. Methods: PATHWAY trial is a randomized, placebo-controlled, Asian international, double-blind, phase 3 study evaluating palbociclib plus tamoxifen (PAL+TAM) versus placebo plus tamoxifen (PLB+TAM) in pre/perimenopausal and postmenopausal women with HR+/HER2- ABC (NCT03423199). The study was conducted as a Clinical Research Collaboration with the National Cancer Center Hospital as the regulatory study sponsor and Pfizer providing drug and financial support. The primary endpoint was investigator assessed progression-free survival (PFS). Genomic profiling by next-generation sequencing was an exploratory endpoint to characterize molecular alterations in ctDNA and correlate these with clinical outcomes. Plasma samples were collected at cycle 1-day 1 (pre-treatment) and cycle 2-day 15 and sequenced with the Guardant360 assay (Guardant Health). Patients were included if they had a minimum of 14 days of treatment in the first cycle. The variant allele frequency (VAF) was defined as number of mutant molecules at a specific nucleotide location over the total number of molecules present in the background at a specific given genomic location. We assessed the gene with highest VAF as the dominant clone allele frequency (DCAF). The association of changes in ctDNA alterations with PFS was evaluated to identify biomarkers of early progression. Results: Pre-treatment plasma was successfully sequenced in 177 of the 184 patients; 88 patients treated with PAL+TAM, and 89 with PLB+TAM. ctDNA was detected in 144 (81.4%) patients; the median number of genetic abnormalities was 2 (range, 0-19). PIK3CA, TP53, ERBB2, ESR1, AKT1, EGFR and PTEN were the genes with highest VAF as the dominant clone (24.3%, 11.9%, 4.5%, 4.5%, 3.4%, 3.4%, 3.4%, respectively). The median DCAF was 1.79 (range, 0.1-81.23); PAL+TAM, 2.53 (range, 0.1-63.5) and PLB+TAM, 1.65 (range, 0.08-81.23). Patients with ≥ 2 detectable genomic alterations and DCAF ≥ 1.8% had worse PFS (hazard ratio (HR) 1.93 [95% confidence interval (CI): 1.375 to 2.721], p = 0.0003 and HR 1.95 [95% CI: 1.345 to 2.834], p &amp;lt; 0.0001, respectively) compared to all other patients. In addition, we evaluated whether changes of clonal architecture in ctDNA at early treatment time point (ie. Cycle 2 Day 15) are associated with PFS. Patients with increased DCAF had worse PFS on both PAL+TAM (HR 2.80 [95% CI: 0.552 to 14.15], p = 0.04) and PLB+TAM (HR 2.02 [95% CI: 1.022 to 3.990], p = 0.01). All patients with high pretreatment DCAF (≥ 1.8%) and increase in DCAF progressed within 1 year (n=5, 2 patients treated with PAL+TAM, and 3 with PLB+TAM). Conclusion: Evaluation of clonal architecture in ctDNA could support identification of HR+/HER2- ABC patients with poorer prognosis, who may be at higher risk of early progression, regardless of CDK4/6 inhibitor use. Citation Format: Yoon-Sim Yap, Yuki Kojima, Akinobu Hamada, Hirofumi Mukai, Yen-Shen Lu, Joo Hyuk Sohn, Yoshiko Umeyama, Emi Noguchi, Kazuki Sudo, Tomomi Hata, Aya Kuchiba, Taro Shibata, Shigehiro Yagishita, Masayuki Yoshida, Shinji Kohsaka, Kouya Shiraishi, Kenichi Nakamura, Kenji Tamura, Kan Yonemori. Clonal Architecture of circulating tumor DNA predicts Early Progression on Tamoxifen With or Without Palbociclib in Hormone Receptor-Positive Advanced Breast Cancer: NCCH1607/PATHWAY trial [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 PO1-14-01.

Cardiac arrhythmia and epilepsy genetic variants in sudden unexpected death in epilepsy.

Sudden Unexpected Death in Epilepsy (SUDEP) is the leading epilepsy-related cause of death, affecting approximately 1 per 1,000 individuals with epilepsy per year. Genetic variants that affect autonomic function, such as genes associated with cardiac arrhythmias, may predispose people with epilepsy to greater risk of both sudden cardiac death and SUDEP. Advances in next generation sequencing allow for the exploration of gene variants as potential biomarkers. Genetic testing for the presence of cardiac arrhythmia and epilepsy gene variants was performed via genetic panels in 39 cases of SUDEP identified via autopsy by the Ontario Forensic Pathology Service. Variants were summarized by in-silico evidence for pathogenicity from 4 algorithms (SIFT, PolyPhen-2, PROVEAN, Mutation Taster) and allele frequencies in the general population (GnomAD). A maximum credible population allele frequency of 0.00004 was calculated based on epilepsy prevalence and SUDEP incidence to assess whether a variant was compatible with a pathogenic interpretation. Median age at the time of death was 33.3 years (range: 2, 60). Fifty-nine percent (n=23) were male. Gene panels detected 62 unique variants in 45 genes: 19 on the arrhythmia panel and 26 on the epilepsy panel. At least one variant was identified in 28 (72%) of decedents. Missense mutations comprised 57 (92%) of the observed variants. At least three in silico models predicted 12 (46%) cardiac arrhythmia panel missense variants and 20 (65%) epilepsy panel missense variants were pathogenic. Population allele frequencies were <0.00004 for 11 (42%) of the cardiac variants and 10 (32%) of the epilepsy variants. Together, these metrics identified 13 SUDEP variants of interest. Nearly three-quarters of decedents in this SUDEP cohort carried variants in comprehensive epilepsy or cardiac arrhythmia gene panels, with more than a third having variants in both panels. The proportion of decedents with cardiac variants aligns with recent studies of the disproportionate cardiac burden the epilepsy community faces compared to the general population and suggests a possible cardiac contribution to epilepsy mortality. These results identified 13 priority targets for future functional studies of these genes potential role in sudden death and demonstrates the necessity for further exploration of potential genetic contributions to SUDEP.

Abstract PO3-16-01: Personalized circulating tumor DNA assay precisely predicts the response of neoadjuvant chemotherapy in breast cancer patients

Abstract Introduction Circulating tumor DNA (ctDNA), which is detected in the blood as cell free DNA fragment from tumor cells, could indicates genomic landscape, tumor burden and treatment response during chemotherapy as a non-invasive method. Especially, personized ctDNA assay by next-generation sequencing(NGS) is able to detect a trace amount of ctDNA and precisely figure out the change of the amount of ctDNA during NAC and follow up period after curative surgery. Here, we performed serial ctDNA evaluations in EBC patients who diagnosed as TNBC or HER2-positive BC and received NAC followed by curative surgery. We aimed to predict NAC response and detect minimal residual disease (MRD) using personalized ctDNA assay. Methods CtDNA was detected by AlphaLiquid®Detect, a tumor-informed personalized MRD detection assay exploring most of the mutations in tumor. Whole exome sequencing (WES) of tumor tissue and peripheral blood mononuclear cells (PBMCs) was performed. Patient-specific somatic mutations were selected using a proprietary algorithm. In brief, clonal variants were prioritized using integrated information including variant allele frequency, population allele frequency databases, somatic variant databases, variant pathogenicity, and genomic context. Up to 100 variants were selected for patient monitoring. A hybridization capture panel consisting of a pool of 4 patients’ selected variants was synthesized. These bespoke panels (BSPs) were used for ctDNA detection. Patients with more than 2 tumor-derived mutations detected in plasma were considered as ctDNA positive. Inclusion criteria included patient who diagnosed as stage IIA-IIIC BC planned to NAC followed by curative surgery. Among BC subtypes, TNBC and HER2-positive BC were allowed. Collection of specimens and associated clinical data used in this study was approved by the Institutional Review Board of Samsung Medical Center (IRB File No.2021-02-033), and we received informed consents for this study. Results From May 2021 to Sep 2022, 158 patients has been enrolled. Archival tissues were not available in 47 patients and tissue WES had failed in six patients. Therefore, 105 patients were enrolled and analyzed their ctDNA at diagnosis based on tissue WES data. Of 105 patients, Median age of BC patients was 49.3 years of age (range: 26.1, 67.8). Thirty-six patients (34.3%) were post-menopausal status and others (65.7%) were pre-menopausal status. In BC pathology at diagnosis, 56 (53.3%) were TNBC and other 49 (46.7%) were HER2-positive BC. Among HER2-positive BC, hormone receptor (HR)-positive were in 21 (20.0%). In clinical stage at diagnosis, stage II were 46 (43.8%) and stage III in 59 (56.2%). In 105 patients, median number of somatic mutations was 60 (interquartile range [IQR]: 38.5, 91). CtDNA detection rate at BC diagnosis was 90.5% and all of BC which not be detected ctDNA was HER2-positive BC with clinical stage II. Median amount of ctDNA at diagnosis was 20 (IQR: 6, 44). In the number of somatic mutations, there was no difference according to BC subtypes (P=0.121) and clinical stage (P=0.700) but the amounts of ctDNA at diagnosis was different. CtDNA was much more detected in TNBC compared to HER2-positive BC (Median: 34.5 vs. 15.9; P&amp;lt; 0.001) and clinical stage III compared to clinical stage II (Median 33.2 vs. 19.5; P=0.002). After NAC, ctDNA at curative surgery was tested in 70 patients. In 70 patients, pathologic complete response(pCR) was observed in 44 patients. CtDNA was detected in ten patients (14.3%) and eight did not achieve pCR and two with pCR (Specificity of assay: 95.5%, positive predictive value: 0.80, P=0.032). Conclusions Personalized ctDNA assay can precisely detected ctDNA at diagnosis and their detection rate was associated with BC subtypes and clinical stage. In addition, ctDNA at curative surgery also can predict NAC response in EBC patients. Long term ctDNA follow up of MRD would be warranted. Citation Format: Ji-Yeon Kim, Hae Hyun Jung, Jeong Eon Lee, Seok Won Kim, Hwang-Phill Kim, Sunghoon Heo, Jongseong Ahn, Tae-You Kim, Young-Hyuck Im. Personalized circulating tumor DNA assay precisely predicts the response of neoadjuvant chemotherapy in 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 PO3-16-01.

Abstract PO2-15-02: Personalized circulating tumor DNA (ctDNA) monitoring can early identify disease progression and predict prognosis in patients with breast cancer undergoing neoadjuvant therapy

Abstract Background: Circulating tumor DNA (ctDNA) analysis offers a non-invasive method to assess tumor burden during treatment. Although some data have been reported on the use of ctDNA in neoadjuvant therapy (NAT) for breast cancer (BC), there is a lack of data on neoadjuvant efficacy and prognosis in China specifically using tumor-informed personalized ctDNA monitoring panel. Methods: We performed personalized ctDNA analyses at different time of NAT (before, during, and after) in 19 patients with Stage IIA-IIIB BC. WES was conducted on tumor tissues, and single nucleotide variants (SNVs) (20-40) were selected for each patient to ensure accurate and sensitive detection of individual ctDNA(Shanghai OrigiMed Co., Ltd.). Results: A total of 19 BC patients, 6 HER2-negative and 13 HER2-positive, were enrolled at the Breast Center, Fourth Hospital of Hebei Medical University (China) from June 2022 to May 2023. The median age of the patients was 53 (min-max: 28-64) years old. Through WES, it was discovered that 78.5% (3377/4303) of the mutated genes were unique to each patient. Additionally, 96.1% (637/663) of the selected tumor-informed SNVs were VUS variants. These findings suggest that tumor-informed ctDNA monitoring is more effective than panel-based ctDNA monitoring in BC. Among the patients, 84% (16/19) completed baseline blood collection, 63% (12/19) completed blood collection after NAT and 42% (8/12) patients completed blood collection after the surgery. The baseline positivity rate was 100%, and the baseline ctDNA variant allele frequency was significantly associated with T stage and N stage (both p &amp;lt; 0.01). This suggests that designing a panel based on puncture samples is feasible for ctDNA analysis. The detection of ctDNA before surgery after NAT correlated with the postoperative ctDNA status. Patients with persistently negative ctDNA during NAT were associated with pathologic complete response (pCR), although the difference was not statistically significant due to the small sample size (n=7). In 84% of patients, dynamic changes in ctDNA aligned with imaging findings, indicating a correlation between ctDNA and disease progression. A 56-year-old patient with HER2-negative stage IIIC BC had persistent ctDNA positivity during NAT. Imaging evaluation showed disease progression, with ctDNA allele frequency increasing 18 days before detection on imaging. After surgical tumor removal, the pathology assessment revealed a change in HER2 status from negative to positive. The patient received adjuvant chemotherapy combined with HER2-targeted therapy, resulting in a transition from positive to negative ctDNA status. The disease has remained stable for nearly 9 months. A 29-year-old patient with HER2-positive stage IIIA BC underwent NAT for 8 cycles. Due to drug intolerance, the treatment was interrupted multiple times during the first 4 cycles, and ctDNA remained positive. After modifying the treatment regimen in the 5th cycle, ctDNA became negative until the patient underwent surgery. The postoperative pathology evaluation revealed a pCR. Conclusions: Personalized ctDNA monitoring is more effective than panel-based ctDNA (a predefined set of cancer-associated genes) monitoring in BC. The status of ctDNA before and after surgery following NAT may be correlated, suggesting that patients with positive ctDNA before surgery may benefit from intensive therapy. More importantly, dynamic changes in ctDNA closely align with radiological findings, enabling early detection of disease progression and guiding treatment modifications. Citation Format: Yunjiang Liu, Chao Shi, Xiangmei Zhang, Xiaofei Ren, Shang Wu, Yaxian Wang, Yue Che, Zanmei Xu, Fei Pang. Personalized circulating tumor DNA (ctDNA) monitoring can early identify disease progression and predict prognosis in patients with breast cancer undergoing neoadjuvant therapy [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 PO2-15-02.

Abstract PO2-13-10: Predicting outcomes in locally advanced breast cancer through profiling post-neoadjuvant tissue-based minimal residual disease

Abstract Background: Neoadjuvant chemotherapy (NAC) for locally advanced breast cancer can down-stage disease prior to surgery, monitor chemo-sensitivity, and provide key prognostic information via pathologic response that guides adjuvant treatment. Greater residual disease as assessed by the residual cancer burden (RCB) index is associated with inferior outcomes but imperfectly predicts recurrence, particularly in triple negative breast cancer (TNBC) where there is heterogeneity of outcomes across RCB scores. As an alternative to RCB and PCR, blood-based minimal residual disease (MRD) from circulating tumor DNA (ctDNA) has been explored. However, poor sensitivity limits ctDNA, which is typically negative post-NAC when treatment decisions are made in patients who later recur. Methods: To address this lack of sensitivity, we developed tissue-based MRD (t-MRD) to assess molecular disease burden in post-NAC breast and lymph node resection tissue. We performed whole exome sequencing on diagnostic tumor and matched germline tissues from five pilot patients with early-stage TNBC with a range of responses to NAC. For each patient, we designed a personalized assay to track tumor-specific SNVs, combined with limited panel of genes recurrently mutated in breast cancer, to enable MRD detection. We then applied this assay to DNA from individual pathology blocks from the tumor resection via ultra-deep targeted hybrid capture. We assessed each sample for the presence of MRD using a Monte Carlo statistical framework to integrate all tumor mutations as previously described (Newman et al, Nature Biotechnology 2016). We aggregated results across all pathology blocks from each case to define each patient’s t-MRD profile and assessed the performance of t-MRD versus gold standard pathologic assessment. Results: We tracked a mean of 136 MRD reporters per case across 119 post-NAC blocks derived from five patients. Twenty-five samples had pathologic residual disease and 28 had detectable t-MRD. There was strong concordance between t-MRD mean allele frequency (AF) and tumor cellularity as assessed by pathology, defined as percent cellularity within the region containing viable tumor (Spearman r = 0.9052, p&amp;lt; 0.0001), and excellent classification of true positive pathology samples by genomic sequencing (AUC = 0.9675, p &amp;lt; 0.0001). Samples (n = 6) where t-MRD but not pathologic residual disease was detected had a lower mean AF than those where there was also histologic residual disease (p=0.0001). Among the five studied cases, we confirmed a molecular complete response with no t-MRD detected in the RCB-0 case (0/44 t-MRD positive blocks). In the two RCB-III cases, we observed more diffuse t-MRD than that assessed by pathology (n = 4 additional blocks with t-MRD but not pathologic disease detected), and in the RCB-II case, we detected minimal t-MRD in only a single block where pathology did not detect residual disease. Significance: By applying blood-based MRD assessment tools to tumor tissue, we developed t-MRD to profile molecular residual disease at the landmark post-NAC timepoint when disease burden is low and adjuvant treatment decisions are made. We demonstrated high concordance with gold standard pathologic assessment, with increased sensitivity and minimal false positive signal. Further studies assessing the additional prognostic value of t-MRD beyond stage-based prognostication and post-NAC RCB scores are ongoing. Citation Format: Julia Ransohoff, Mia Carleton, Alisha Birk, Gregory Bean, Melinda Telli, Ash Alizadeh, James Ford, George Duran, Michael Khodadoust, David Kurtz. Predicting outcomes in locally advanced breast cancer through profiling post-neoadjuvant tissue-based minimal residual disease [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 PO2-13-10.

Abstract PO4-14-10: Circulating Tumor DNA as a Biomarker for ADCs in Metastatic Breast Cancer

Abstract Background: Breast cancer is a complex disease characterized by heterogeneity, and the analysis of circulating tumor DNA (ctDNA) holds promise for capturing this heterogeneity. Recently, ctDNA testing has been employed in clinical practice to guide targeted therapies for metastatic breast cancer, particularly in cases of hormone receptor-positive disease with mutations in PIK3CA and ESR1. This approach becomes necessary when traditional tumor biopsies are inadequate for next-generation sequencing (NGS) testing or fail to capture the full extent of the cancer's heterogeneity. Moreover, the utilization of antibody-drug conjugates (ADC) in addressing tumor heterogeneity continues to expand. However, there has been no biomarker, outside of HER2 protein expression, to predict ADC response. In this project, we test the hypothesis that ctDNA can be utilized as a biomarker to predict response to novel therapies such as ADCs. Methods: We analyzed a subset of patients from the Dallas Metastatic Breast Cancer Study comprised of patients with metastatic breast cancer (n= 109) who underwent ctDNA testing using the Tempus xF liquid biopsy ctDNA sequencing panel. This panel detects 105-genes that are known oncogenic drivers and resistance mutations. The data was collected from a single academic medical center between the initial year of ctDNA collection in 2019 and 2023. Results: Among these patients, 85 (77.9%) had hormone receptor-positive disease, 13 (11.9%) had triple-negative breast cancer, and 11 (10.0%) had HER2-positive disease (including 3 (2.75%) patients with triple-positive disease). Only 22 of these patients had tissue biopsies that underwent NGS testing. Analysis of all patients revealed that the most common gene alteration was PIK3CA, which was identified in 70 patients. Specifically, the most frequently observed alteration was PIK3CA p.545K, found in 25 patients. Table 1 further details the frequency of each gene in this subset of patients. Among the patients, 34 out of 109 (31%) received an ADC, while only 8 out of 109 (7%) received immunotherapy. Patients with CDKN2A achieved the longest progression-free survival (PFS) on an ADC, with a median PFS of 7 months, whereas patients carrying mutations in TP53, PIK3R1, PIK3CA, PTEN, NF1, BRAF, RHOA, FGFR4, KRAS, TERT, GATA3, HNF1A, FB1, STOP had median PFSs of 2 months or less. Table 2 provides additional information on the response of patients with specific mutations to ADCs. Notably, only one patient received sequential ctDNA testing, and in that case, there was a gain of TP53 p.N239S following sacituzumab govitecan administration compared to the baseline ctDNA test, followed by a gain of PIK3CA p.P449_L455del after trastuzumab deruxtecan administration. Conclusions: As we observe changes in subtypes following treatment progression, researchers are actively seeking biomarkers to better characterize real-time changes in tumor biology that can predict response and resistance to treatments. Further studies are needed to identify additional genes present in ctDNA that can provide mechanistic insights into why certain patients respond favorably to ADCs while others do not. We are currently enrolling patients for a prospective study that aims to assess baseline ctDNA levels and monitor changes in variant allele frequency following treatment with ADCs and immunotherapy. Table 2 Median PFS on ADC by Gene Citation Format: Hannah Chang, Isabela Anawate, Alyssa Low, Shao-Po Huang, Julia Maues, Christine Hodgdon, Isaac Chan. Circulating Tumor DNA as a Biomarker for ADCs in Metastatic Breast Cancer [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 PO4-14-10.

Abstract PS02-05: Trajectories and predictors of peripheral neuropathy after neoadjuvant chemotherapy in a prospective cohort of 11,014 patients with early breast cancer

Abstract Background Peripheral neuropathy (PN) is a debilitating adverse event in patients with early breast cancer (BC) receiving (neo)adjuvant chemotherapy (CT). We harnessed the CANTO cohort study to detail clinical trajectories and explore clinical and genetic predictors of PN. Methods CANTO (CANcer TOxicities - NCT01993498) prospectively enrolled invasive stage I-III BC patients (pts) of 26 French comprehensive cancer centers. Pts were assessed at diagnosis, 3-6 (M6), 12 (M12), 36 (M36), and 60 (M60) months after treatment, defined as completion of surgery, CT or radiotherapy, whichever comes last. At each time point, PN events including paresthesia, sensory and motor neuropathy were collected according to NCI-CTC v4.0 criteria. A genome-wide association study (GWAS) was conducted to identify genetic predictors of PN, using Illumina GSA BeadChips. Minimac4/1000G was used to impute additional single nucleotide polymorphisms (SNPs). After stringent quality control measures, 1,894,475 SNPs with a minor allele frequency (MAF) &amp;gt;0.05 were evaluated. Longitudinal trajectories of PN events were descriptively examined. Statistical associations between each SNP and PN events measured at different time points, and between SNPs and trajectories, were performed with logistic regression assuming a log-additive genetic model. All analyses were adjusted for key clinical parameters and the first ten axes of principal component analysis of the genetic data to control for population stratification. Results Of 12,012 included pts (data lock Aug. 2022), 11,014 (91.7%) were analyzed. Age was &amp;lt; 50 and &amp;gt;65 in 3407 and 2759 pts (31% and 25%, respectively). Overweight/obesity and diabetes were recorded in 5328 (48%) and 458 pts (4%). A neurologic history was observed in 1360 pts (13%). Stage 0-I and II-III were observed in 5356 (49%) and 5512 pts (51%), and 4011 pts had an axillary dissection (37%). CT was administered in 5790 pts (53%), including a taxane (tax) in 5542 pts (96%). Shortly, all grade and grade 2+ PN was observed in 29% and 12%; 27% and 10%; 20% and 10%; 13% and 10% of pts at M6, M12, M36 and M60, respectively. We derived 5 trajectories of overall PN, deemed “never”, “always”, “appears”, “transient” and “disappears”, in respectively 54%, 10%, 9%, 7% and 19% of the pts. Similar trajectories were built for the 3 categories of PN events. We built 9 predictive nomograms including key clinical parameters and time of analysis. E.g., grade 2+ PN at M6 was independently predicted by a medical history of carpal tunnel syndrome, past neurologic history and exposure to tax. Grade 2+ PN at M36 was predicted by past neurologic history and exposure to tax, and largely by previous PN at M6 or M12. The GWAS analysis included 7633 pts (84%). Four independent SNPs adding predictive value to the clinical nomograms were identified at a suggestive level of association (p &amp;lt; 1e-6), including one SNP in the NCAM1 gene (involved in neurogenesis) for M6 PN (OR=1.40, 95%CI 1.23-1.60, p=4.8e-7, see table) and one SNP in the CLDN11 gene (regulates oligodendrocytes) for M12 sensory PN (OR=1.32; 95%CI 1.17-1.49, p=7.5e-6). For rare ( &amp;lt; 5%) toxicities, 10 independent suggestive SNPs were identified, including one SNP in the NELL1 gene (neural cell growth regulation and differentiation) for the “always” trajectory (OR=1.61, 95%CI 1.31-1.99, p=7.6e-6) and one SNP in the KCNIP1 gene (neuronal sensor) for M36 motor PN (OR=1.5, 95%CI 1.7-3.5, p=3.1e-07). Conclusions Risk of early peripheral neuropathy is associated with previous neurological history and taxane exposure. Patients with PN at M6 or M12 are highly exposed to long-term PN. Some key SNPs may add independent predictive value for specific PN endpoints. Exploratory results of interest were produced for rarer toxicities and typical clinical trajectories. Detailed results will be presented at the meeting. All PN, M6 Multivariate analyses of the risk of peripheral neuropathy, all grades, at M6, without and with GWAS input. Citation Format: Youenn Drouet, Emilie Thomas, Florence Lerebours, Barbara Pistilli, Olivier Trédan, Christelle Jouannaud, Marion Fournier, Philippe Rouanet, Laurence Vanlemmens, CHARLES COUTANT, Asma Dhaini Merimeche, Baptiste Sauterey, Christelle Levy, Mario Campone, Carole Tarpin, Marie-Ange Mouret-Reynier, Olivier Rigal, Thierry Petit, Sophie Guillermet, Antoine Arnaud, Mahmoud Ibrahim, Sylvie Giacchetti, Florence Dalenc, Johanna Wassermann, Olivier Arsène, Ariane Darut-Jouve, Sibille Everhard, Ines Vaz Luis, Anne-Laure Martin, Fabrice André, Jean-François Deleuze, Alain Viari, Matthieu Carton, Paul Cottu. Trajectories and predictors of peripheral neuropathy after neoadjuvant chemotherapy in a prospective cohort of 11,014 patients with early breast cancer [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 PS02-05.

Abstract PO4-13-07: Association of clonal hematopoiesis (CH) with clinical outcomes among survivors of breast cancer (BC)

Abstract Background: Clonal hematopoiesis (CH) refers to the identification of clonal expansion of hematopoietic cells due to somatic mutations in leukemia-associated genes without evidence of hematologic anomalies. CH, particularly with variant allele frequency (VAF) ≥ 10%, has been associated with adverse outcomes, including reduced survival in advanced malignancies and higher risk of cardiovascular events. The aim of this study was to evaluate the biological and prognostic relevance of CH in survivors of breast cancer (BC). Methods: CANTO (NCT01993498) is a prospective, multicenter cohort enrolling patients diagnosed with stage I-III BC. CANTO collects clinical, tumor and treatment-related characteristics at diagnosis (dx). Follow-up visits are scheduled at 1, 2, 4 and 6 years after dx and data on disease status, type of recurrence, and survival status are collected. We assessed CH among pts ≥ 40 years, without previous solid or hematological cancer, with blood samples available at dx and year 4 or experiencing death or disease recurrence before year 4. CH was determined using Next-Generation Sequencing with unique molecular identifiers (HaloPlexHS, Agilent Technologies) for 17 genes (including DNMT3A, TET2, ASXL1, PPM1D, ATM, JAK2 and TP53) on blood samples obtained at dx. Outcomes of interest included invasive disease-free survival (iDFS), distant disease-free survival (DDFS) and overall survival (OS) defined according to STEEP criteria. Kaplan-Meier estimator, log-rank test and multivariate Cox models assessed prognostic role of CH. Results: In the cohort with available CH data (N=1219) mean age (SD) was 57.5 (10.1) years, 62% of pts were postmenopausal, 40.9% were current or former smokers, 46.2% had stage I BC, 77.7% had HR+/HER2- BC, 53.5% received chemotherapy and 80.8% hormonal therapy. CH was detected with a VAF ≥ 1% in 306 pts (25.1%) including 207 pts (67.6%) with DNMT3A mutation, ≥ 2% in 186 (15.3%) and ≥ 10% in 45 (3.7%) pts. Pts with CH ≥ 10% were more frequently older (65.3 vs 57.2 years, p&amp;lt; .0001) and with a previous history of cardiovascular disease (53.3% vs 37.3%, p&amp;lt; 0.02). At a median (IQR) follow-up of 7.0 (6.2-7.8) years 246 iDFS, 161 DDFS and 118 OS events were observed. Only 8 cases of new hematological malignancies were observed. Pts harboring CH with a VAF ≥ 10% had worse OS (log-rank p=0.04) and CH with a VAF ≥ 10% was associated with higher risk of death (HR= 2.4, 95%CI 1.04-5.6); presence of CH with lower VAF was not associated with worse OS. Table displays complete results for all tested associations. Conclusions: CH at dx was identified in 25% of pts with stage I-III BC aged ≥ 40 years in the CANTO cohort, including 4% with a VAF ≥ 10%. In a large, prospective cohort of survivors of BC, we confirmed known associations of CH with older age and cardiovascular history. We observed few secondary hematological malignancies. CH with a VAF ≥ 10% was significantly associated with higher risk of death while adjusting for other relevant prognostic factors. Furthermore, a trend towards higher risk of DDFS events was observed among pts harboring CH with a VAF ≥ 10%. Table. Citation Format: Davide Soldato, Antonin Della Noce, Antonio Di Meglio, Christophe Marzac, Barbara Pistilli, Anne-Laure Martin, Sibille Everhard, Marie Breckler, Sandrine Boyault, Marina Rousseau, Olivier Trédan, Charles Coutant, Paul Cottu, Olivier Rigal, Christelle Levy, Christelle Jouannaud, Baptiste Sauterey, Nathalie Droin, Bastien Job, Aurélie Bertaut, Fabrice André, Ines Vaz Luis, Jean-Baptiste Micol. Association of clonal hematopoiesis (CH) with clinical outcomes among survivors of breast cancer (BC) [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 PO4-13-07.

Abstract PO2-16-03: ESR1 genomic alterations (GAs) and coexistent putative resistance alterations in comprehensive genomic profiling (CGP) of metastatic breast cancer (MBC)

Abstract Background: ESR1 mutations (ESR1mut) are an established biomarker of endocrine therapy (ET) resistance in patients (pts) with hormone receptor positive (HR+) MBC. Moreover, ESR1mut acquired in response to standard-of-care ET now confer access to novel ET recently approved by health authorities. However, the nuanced genomic landscape beyond the simple presence or absence of ESR1mut may be critically important. We report on detailed ESR1 GAs detected by tissue and liquid biopsies (TBx, LBx) and examine the prevalence of co-occurring GAs in other genes reported to confer intrinsic or acquired resistance to ET. Methods: CGP results from MBC biopsies were retrospectively analyzed (TBx 33,653 pts; LBx 7,134 pts) using FoundationOne® CDx and FoundationOne® Liquid CDx, hybrid-capture based sequencing platforms profiling 324 cancer-related genes. Foundation Medicine’s ctDNA tumor fraction (TF) is a composite algorithm prioritizing aneuploidy at higher levels to avoid germline signal and prioritizing variant allele frequency (VAF) of canonical alterations at lower levels to maximize dynamic range. Results: ESR1mut were detected in 3,755 (11.2%) TBx and 1,513 (21.2%) LBx. Among samples with ESR1mut, multiple ESR1mut were more common in LBx (19% 2, 12% 3+) than in TBx (6% 2, 0.5% 3+), with up to 10 different ESR1mut seen in one LBx. The most common mutations were D538G (39% and 52% of TBx and LBx with ESR1mut), Y537S (27%, 38%), Y537N (10%, 15%), and E380Q (10%, 14%). Activating ESR1 insertion-deletion mutations (indels) were detected in 81 TBx and 62 LBx (2% and 4%), representing 23 distinct variants at the protein level, including V422del (107), indels affecting codons 532-538 (29), G344_L345insC (7), and Y328_S329del (6). 28 fusions of the ESR1 DNA binding domain to 13 unique gene partners were detected; 10/28 (36%) were ESR1-CCDC170 fusions. ESR1 amplifications (ESR1amp) were detected in 704 (2.1%) TBx; 62 (8.8%) also harbored ESR1mut. ESR1amp were detected among 34 (0.5%) LBx; 8 (24%) also had ESR1mut. In TBx with ESR1mut and LBx collected from the same pt within 8 months (43 pairs), LBx sensitivity of detection of ESR1mut was 96% (26/27, 95% confidence interval CI [95%CI]: 79%-99%) when TF was ≥1%, but only 6% (1/16, 95%CI: 0.3%-32%) when LBx had TF &amp;lt; 1%. In 988/1,513 (65%) of LBx with ESR1mut, ≥1 mutations or fusions in genes associated with alternative ET resistance pathways were detected, including PIK3CA (46% of ESR1mut LBx), RB1 (12%), PTEN (10%), NF1 (7%), ARID1A (7%), AKT1 (5%), FGFR2 (4%), and KRAS (4%) and ERBB2 (3%). Prevalence of co-occurring GA associated with ET resistance was higher in LBx than in TBx (RB1, NF1, FGFR2, PTEN, EGFR, KRAS, PIK3CA, FDR &amp;lt; 0.001). Moreover, prevalence was higher in 240 LBx where ESR1 was a minor allele (VAF/TF &amp;lt; 10%) compared to 1,273 LBx were ESR1 was a major allele (FGFR2, PIK3CA, FGFR3, BRAF, ERBB2, RB1, FDR &amp;lt; 0.05). In pts with ESR1mut LBx and a historical TBx collected &amp;lt; 5 years earlier (182 pairs), GAs in RB1, FGFR2, KRAS, EGFR, and BRAF were detected primarily on the LBx but not the preceding TBx. Conclusions: CGP detects a wide spectrum of mutations in ESR1, including missense and indel mutations and fusions. When LBx TF is &amp;lt; 1%, sensitivity for ESR1mut is reduced and repeat testing should be considered. ESR1mut can coexist with complementary or competing resistance mechanisms, particularly when ESR1 is a minor allele, which could impact benefit from novel ET approved for patients with ESR1 mutations. Currently, CGP of LBx informs therapeutic recommendations for pts with HR+ MBC based on the binary presence or absence of ESR1mut; however, the clinical implications of the ESR1 mutation spectrum described herein and the potential impact of co-expression of other key pathway signals warrant further investigation. Citation Format: Heather McArthur, Hanna Tukachinsky, Alexa Schrock, Russell Madison, Oliver Holmes, Smruthy Sivakumar, Ethan Sokol, Ryon Graf, Julia Quintanilha, Kali Dougherty, Lincoln Pasquina, Geoffrey Oxnard, Mia Levy, Eric Winer. ESR1 genomic alterations (GAs) and coexistent putative resistance alterations in comprehensive genomic profiling (CGP) of metastatic breast cancer (MBC) [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 PO2-16-03.

Abstract PO3-14-11: Association between BRCA alterations detected by circulating tumor DNA and germline mutations in breast cancer patients: a retrospective mono-institutional analysis

Abstract Background: PARP inhibitors (PARPi) are a standard of care for breast cancer (BC) patients (pts) with germline BRCA (gBRCA) mutation, with benefit demonstrated also in patients with somatic (s) BRCA1/2 mutations. The use of circulating tumor (ct) DNA testing allows to detect resistant and actionable alterations in BC, but it could be associated with the incidental identification of germinal mutations. The aim of this study was to describe the incidence of both, somatic and germinal BRCA1/2 alterations, and the impact on the therapeutic outcomes in a retrospective BC cohort with clinical ctDNA testing for their BC. Materials and methods: A retrospective cohort of 245 BC pts that underwent ctDNA analysis (Guardant 360) between August 2014 and May 2023 at Weill Cornell Medical College was identified. Pts with at least a BRCA1/2 alteration, including variant of uncertain significant (VUS) and synonymous, were enrolled in the study. A descriptive analysis was performed. Results: Among 35 patients included, 34 had metastatic disease, the median age was 61 years old (IQR 40-67), and 65.7% were post-menopausal at diagnosis. 57.1% of pts had hormone receptor positive/HER2 negative (HR+/HER2-) BC, 20% had HER2 positive disease and 22.9% were triple-negative (TN) BC. Ten (28.6%) and 20 (57.1%) pts had a sBRCA1 or a sBRCA2 alteration on ctDNA, respectively; 5 pts (14.3%) had a coexisting sBRCA1/2 mutation. The median variant allele frequency (VAF) was 1.2% for BRCA1 and 2.7% for BRCA2 alterations. Furthermore, 53.3% and 6.7% of sBRCA1 and 40% and 8% of sBRCA2 mutations were identified as VUS and synonymous, respectively. The most common associated genomic alterations included TP53 (62.9%), ESR1 (20%) and PI3KCA (52%) with a median number of 4 variants (IQR 2-7). Next generation sequencing (NGS) testing on tissue was available for 3 patients: two demonstrated concordance with BRCA alterations detected by ctDNA, whereas no alteration in BRCA1/2 genes was found for the third patient despite the tissue analysis was performed at the same time of the ctDNA. Germline testing was available for 24 (69%) pts; a corresponding gBRCA1 mutation was found for 3 pts (VAF 48.6%-84.3%), and a gBRCA2 for 8 pts (VAF 26.7%-49.5%). Due to the small sample size, a univocal VAF cut-off to detect gBRCA1/2 mutations could not be calculated. Twenty-nine pts had a sBRCA1/2 mutation detected by ctDNA test before starting a new therapy; of these 55.2% had HR+/HER2- disease, while 17.2% and 27.6% were HER2 positive and TN BC, respectively. The median progression free survival (mPFS) was 10.3 (3.2-17.4) months (ms) and the 1-year and 2-year overall survival (OS) rates were 82.3% and 70.5%, respectively. The mPFS was 10.3 ms (0-25.0) for patients (n=16) that underwent chemotherapy, 5.6 ms (0-15.1) for those who had received endocrine therapy (n=9), and not reached for patients treated with PARPi (n=4). Conclusions: ctDNA analysis allows the detection of sBRCA1/2 mutations that could be missed by tissue-based testing. The identification of BRCA alterations on ctDNA could guide the use of germline testing even when these are present at a lower VAF than expected, with relevant impact on the therapeutic choices and family screening. Further evaluation to establish the impact of sBRCA1/2 detected by ctDNA on the therapeutic algorithm decision are needed. Citation Format: Letizia Pontolillo, Eleonora Nicolò, Laura Munoz Arcos, Carolina Reduzzi, Mara Serena Serafini, Amanda Kaylan Strickland, Nadia Bayou, Jeannine Donahue, Elisabetta Molteni, Lorenzo Gerratana, Diana Giannarelli, Eleni Andreopoulou, Emilio Bria, Massimo Cristofanilli. Association between BRCA alterations detected by circulating tumor DNA and germline mutations in breast cancer patients: a retrospective mono-institutional analysis [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 PO3-14-11.

Abstract PO5-06-14: Genomic Landscape and Clinical Outcomes of Triple-Negative Invasive Lobular Carcinoma

Abstract Background: Triple-negative (TN) invasive lobular carcinoma (ILC) of the breast is a rare entity, comprising roughly 2% of primary ILC. Clinicopathologic studies have shown that TN ILC is associated with more pleomorphism, higher nuclear grade, older patient age, and worse disease-related outcomes compared to estrogen receptor-positive (ER+) ILC. Intrinsic subtyping indicates that TN ILC less frequently expresses basal markers than TN invasive ductal carcinoma (IDC). This prompted investigation of the genomic landscape and clinical management of TN ILC to provide insight into potential therapeutic approaches. Methods: Twenty patients with TN ILC were included in this study. Three patients were treated at a single academic center and underwent comprehensive genomic profiling (CGP) of breast or metastatic tissue; clinicopathologic details were obtained from their electronic health records. Seventeen patients with TN ILC who underwent CGP of breast or metastatic biopsies were included from the Foundation Medicine, Inc. database. PD-L1 testing for all patients was determined by immunohistochemistry (IHC) using the VENTANA SP142 or Dako 22C3 commercial assays. HER2 expression was determined by IHC. Negative HER2 expression was defined as an IHC score of 0, or IHC 1+ or 2+ with non-amplified fluorescence in situ hybridization (FISH). Results: In the Foundation Medicine database of patients with TN ILC, the most frequent genomic alterations were in CDH1 (15/17 patients [88.2%]), TP53 (10/17 patients [58.8%]), and PIK3CA (10/17 patients [58.8%]). Short variant mutations in ERBB2 were found in 3/17 patients (17.6%). Fourteen patients were HER2-low (HER2 IHC 1+ or 2+ with non-amplified FISH). No cases were MSI high or expressed PD-L1 (8 of 17 cases underwent PD-L1 assessment by SP142). Median tumor mutational burden (TMB) was 5 muts/Mb. Among the three patients with TN ILC treated at a single center, the most common somatic mutations were in CDH1 and TP53, and no cases were MSI high or expressed PD-L1 (by SP142 or 22C3). All three patients had pleomorphic ILC, histologic grade 2 or 3, and HER2 IHC 0. One patient with mpT1cN1a(sn) ER+/HER2- ILC of the right breast in 2011 received adjuvant TAC (docetaxel, doxorubicin, cyclophosphamide), breast/axillary radiation, and ten years of adjuvant endocrine therapy. She was diagnosed with TN ILC of the left supraclavicular nodes in 2022; she did not exhibit a response to docetaxel and carboplatin with pembrolizumab, and she subsequently received palliative radiation followed by capecitabine. Another patient with locally advanced TN ILC progressed on the KEYNOTE-522 regimen, immediately developed metastases, and is currently receiving palliative sacituzumab govitecan. The third patient with cT3N1 TN ILC progressed on neoadjuvant therapy (through the I-SPY 2 trial) with oral paclitaxel, encequidar, and dostarlimab, followed by dostarlimab with dose-dense doxorubicin and cyclophosphamide. Her surgical pathology demonstrated ypT3N3a disease with 10 × 7 cm of tumor, and she developed skin metastases on the chest wall within weeks post-operatively. She received comprehensive chest wall and nodal radiation and is currently receiving palliative capecitabine with neratinib given a high variant allele frequency driver somatic ERBB2 mutation. Conclusions: The most common somatic mutations among TN ILC patients included in this study were in CDH1, TP53, and PIK3CA. Currently the management of TN ILC is extrapolated from that of TN IDC, however our patients did not respond to chemoimmunotherapy. The presence of HER2-low status or somatic ERBB2 mutations may provide opportunities for treatment with an anti-HER2 antibody-drug conjugate or tyrosine kinase inhibitor. Studies with larger sample sizes of TN ILC are needed for molecular profiling and assessment of outcomes with specific therapeutic approaches. Citation Format: Hemali Batra-Sharma, Smruthy Sivakumar, Prashanth Ashok Kumar, Ethan Sokol, Rebecca Shatsky, Jeffrey Ross. Genomic Landscape and Clinical Outcomes of Triple-Negative Invasive Lobular Carcinoma [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 PO5-06-14.

Gastric cancer actionable genomic alterations across diverse populations worldwide and pharmacogenomics strategies based on precision oncology.

Introduction: Gastric cancer is one of the most prevalent types of cancer worldwide. The World Health Organization (WHO), the International Agency for Research on Cancer (IARC), and the Global Cancer Statistics (GLOBOCAN) reported an age standardized global incidence rate of 9.2 per 100,000 individuals for gastric cancer in 2022, with a mortality rate of 6.1. Despite considerable progress in precision oncology through the efforts of international consortia, understanding the genomic features and their influence on the effectiveness of anti-cancer treatments across diverse ethnic groups remains essential. Methods: Our study aimed to address this need by conducting integrated in silico analyses to identify actionable genomic alterations in gastric cancer driver genes, assess their impact using deleteriousness scores, and determine allele frequencies across nine global populations: European Finnish, European non-Finnish, Latino, East Asian, South Asian, African, Middle Eastern, Ashkenazi Jewish, and Amish. Furthermore, our goal was to prioritize targeted therapeutic strategies based on pharmacogenomics clinical guidelines, in silico drug prescriptions, and clinical trial data. Results: Our comprehensive analysis examined 275,634 variants within 60 gastric cancer driver genes from 730,947 exome sequences and 76,215 whole-genome sequences from unrelated individuals, identifying 13,542 annotated and predicted oncogenic variants. We prioritized the most prevalent and deleterious oncogenic variants for subsequent pharmacogenomics testing. Additionally, we discovered actionable genomic alterations in the ARID1A, ATM, BCOR, ERBB2, ERBB3, CDKN2A, KIT, PIK3CA, PTEN, NTRK3, TP53, and CDKN2A genes that could enhance the efficacy of anti-cancer therapies, as suggested by in silico drug prescription analyses, reviews of current pharmacogenomics clinical guidelines, and evaluations of phase III and IV clinical trials targeting gastric cancer driver proteins. Discussion: These findings underline the urgency of consolidating efforts to devise effective prevention measures, invest in genomic profiling for underrepresented populations, and ensure the inclusion of ethnic minorities in future clinical trials and cancer research in developed countries.