Molecular Profiling Of Breast Cancer Research Articles

Association Between Smoking with a Molecular Profile of Breast Cancer

Background: Female breast cancer is one of the causes of the highest cancer mortality and morbidity in the world. It is already known that there is a strong association between smoking and breast cancer. However, the association between smoking and tumor severity is not very clear. The objective of this study was to assess the severity of the breast tumor, using the tumor's molecular classification as a tool according to its immunohistochemical profile in smoking and non-smoking women. Methods: This is a longitudinal study in which 208 women with a diagnosis of breast cancer were followed for 17 months, 80 of whom were smokers and all underwent anatomopathological diagnosis by core biopsy and subsequent immunohistochemistry, followed by treatment indicated according to the type and the clinical staging of the tumor. The severity of the tumor was assessed by its molecular classification according to its immunohistochemical profile. Results: Smoking was associated with higher mortality. The tumor with the most severe immunohistochemical profile was found in younger smokers. 19.7% of smokers had a triple negative tumor and 10% of non-smokers. The age of female smokers with triple negative was 48.2 years and of non-smoking women was 52.6 years (p = 0.005). In 17 months of follow-up, mortality among smokers was 39.5% and for nonsmokers, 20%. Survival was statistically significantly lower among the group of smokers (p=0.01). Conclusion: smoking is associated with greater breast cancer severity, risk for cancer severity was OR 5.5 times higher for the smoking group survival was statistically significantly lower among the group of smokers

Abstract PO1-02-08: Comprehensive Molecular Profiling of Breast Cancer: A real-time PCR assay for identifying ESR1, PGR, ERBB2, MKI67 and a novel proliferative signature in core needle biopsies and resected invasive breast cancer tissues

Abstract Background: Accurate status determination of breast cancer (BC) biomarkers, including estrogen receptor (ER/ESR1), progesterone receptor (PR/PGR), human epidermal growth factor receptor 2 (HER2/ERBB2), and marker of proliferation Ki67 (MKI67), is crucial for guiding therapeutic decisions. However, current “gold standard” methods such as immunohistochemistry (IHC) for assessing these biomarkers in formalin-fixed paraffin-embedded (FFPE) tissues face challenges in standardization and exhibit substantial inter- and intra-laboratory variability, particularly for Ki67. To address these limitations, APIS Breast Cancer Subtyping Kit has been developed, utilizing reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) to evaluate the status of ER, PR, HER2, KI67, and a novel proliferative signature in breast cancer core needle biopsy (CNB) and resected specimens. The APIS Breast Cancer Subtyping Kit offers enhanced accuracy, improved reproducibility, and the ability to provide results in a timely manner. Herein we report the development and validation of the APIS Breast Cancer Subtyping Kit, assessing clinical performance using retrospective and prospective sample collection. Furthermore, reproducibility and reliability across three testing sites were evaluated. By comparing the results of the kit with those obtained through standard IHC, we demonstrate the reliability and utility of the APIS Breast Cancer Subtyping Kit for precise breast cancer subtyping and guiding personalized patient management decisions. Methods: FFPE tissue sections (n=652) taken by CNB or resection underwent histological analysis in accordance with laboratory’s standard of care methods. Samples with a HER2 score of 2+ were referred to fluorescence in situ hybridization (FISH) to determine ERBB2 amplification, with the FISH result replacing the IHC result in determining the HER2 status (positive/negative) of the sample. Tumors with greater than 1% positive stained nuclei for ER/PR were scored positive. A cut-off of 20% Ki67 staining was used for positive/negative status. The performance was assessed as the agreement with IHC/ISH status and reported as OPA (overall percent agreement) with corresponding 95% confidence intervals (95% CI). To assess the precision of the assay, a panel of RNA pools (samples) was repeatedly measured on different days across three independent laboratories. Reproducibility was evaluated using variance components analysis. Results: The APIS Breast Cancer Subtyping Kit showed high concordance (OPA) with IHC: 93.1% (95% CI: 90.9-94.8) for ER, 86.8% (95% CI: 84.0-89.2) for PR, 94.2% (95% CI: 92.2-95.8) for HER2 (IHC/FISH), 78.3% (95% CI: 75.0-81.3) for Ki67, and 80.1% (95% CI: 76.8-83.1) for proliferative signature (assessed against MKI67 IHC status). Inter-site reproducibility demonstrated excellent agreement in quantitative measurements, with a total SD ranging from 0.00 to 0.22. Binary single-marker status (positive/negative) calling achieved 100% reproducibility for ER, PR, and HER2 for all tested samples (negative/low positive/medium positive), as well as for most Ki67 samples (87% agreement for negative samples). Conclusions: APIS Breast Cancer Subtyping Kit demonstrated a high level of concordance with standard of care IHC/FISH in assessing breast cancer biomarker status. These findings suggest that the APIS Breast Cancer Subtyping Kit provides highly precise and reproducible quantitative assessment of BC biomarkers and molecular subtypes. Inclusion and exclusion criteria Citation Format: Anna Gasior, Joanna Gorniak, Sara Rollinson, Leanne Gough, Anne-Sophie Wegscheider, Axel Niendorf. Comprehensive Molecular Profiling of Breast Cancer: A real-time PCR assay for identifying ESR1, PGR, ERBB2, MKI67 and a novel proliferative signature in core needle biopsies and resected invasive breast cancer tissues [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 PO1-02-08.

Ultrasound-based radiomics model for predicting molecular biomarkers in breast cancer.

Breast cancer (BC) is the most common cancer in women and is highly heterogeneous. BC can be classified into four molecular subtypes based on the status of estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) and proliferation marker protein Ki-67. However, they can only be obtained by biopsy or surgery, which is invasive. Radiomics can noninvasively predict molecular expression via extracting the image features. Nevertheless, there is a scarcity of data available regarding the prediction of molecular biomarker expression using ultrasound (US) images in BC. To investigate the prediction performance of US radiomics for the assessment of molecular profiling in BC. A total of 342 patients with BC who underwent preoperative US examination between January 2013 and December 2021 were retrospectively included. They were confirmed by pathology and molecular subtype analysis of ER, PR, HER2 and Ki-67. The radiomics features were extracted and four molecular models were constructed through support vector machine (SVM). Pearson correlation coefficient heatmaps are employed to analyze the relationship between selected features and their predictive power on molecular expression. The receiver operating characteristic curve was used for the prediction performance of US radiomics in the assessment of molecular profiling. 359 lesions with 129 ER- and 230 ER+, 163 PR- and 196 PR+, 265 HER2- and 94 HER2+, 114 Ki-67- and 245 Ki-67+ expression were included. 1314 features were extracted from each ultrasound image. And there was a significant difference of some specific radiomics features between the molecule positive and negative groups. Multiple features demonstrated significant association with molecular biomarkers. The area under curves (AUCs) were 0.917, 0.835, 0.771, and 0.896 in the training set, while 0.868, 0.811, 0.722, and 0.706 in the validation set to predict ER, PR, HER2, and Ki-67 expression respectively. Ultrasound-based radiomics provides a promising method for predicting molecular biomarker expression of ER, PR, HER2, and Ki-67 in BC.

The molecular profile of breast cancer: primary tumor versus corresponding lymph node metastases.

Breast cancer (BrCa) is the most frequent malignancy in female, and lymph node metastases (LNM) is an important prognostic and therapeutic parameter. The molecular classification is nowadays largely applied to characterize the primary tumors, but few studies focused on the comparison between the molecular profiles of the primary with corresponding LNM. In the current work, we investigated the expression of conventional markers used by molecular classification in both primary tumors and axillary LNM. A series of 156 patients with BrCa was investigated, and from these 80 cases showed LNM. After routine pathological investigation, including the histopathological form and grade, we performed additional step sections from the primary and lymph nodes for immunohistochemistry. All procedures for hormone receptors, human epidermal growth factor receptor 2 (HER2), Ki67, cytokeratin 5 (CK5), epidermal growth factor receptor (EGFR), p53, E-cadherin, and B-cell leukemia∕lymphoma-2 (Bcl-2) were performed using the standard automated procedures. We found significant differences between the primary tumors and corresponding LNM in luminal A, luminal B, and basal-like carcinoma. No phenotypical interconversions were noticed in HER2 and unclassified BrCa. Our data demonstrate that in almost 20% of the cases the molecular profile of the primary does not overlap with aspects found in the lymph nodes. Our results strongly suggest performing the molecular classification in both primary tumors and in LNM. Current data suggest that the application of this diagnostic procedure will significantly influence the therapeutic strategy.

The Present and Future of Clinical Management in Metastatic Breast Cancer.

Regardless of the advances in our ability to detect early and treat breast cancer, it is still one of the common types of malignancy worldwide, with the majority of patients decease upon metastatic disease. Nevertheless, due to these advances, we have extensively characterized the drivers and molecular profiling of breast cancer and further dividing it into subtypes. These subgroups are based on immunohistological markers (Estrogen Receptor-ER; Progesterone Receptor-PR and Human Epidermal Growth Factor Receptor 2-HER-2) and transcriptomic signatures with distinct therapeutic approaches and regiments. These therapeutic approaches include targeted therapy (HER-2+), endocrine therapy (HR+) or chemotherapy (TNBC) with optional combination radiotherapy, depending on clinical stage. Technological and scientific advances in the identification of molecular pathways that contribute to therapy-resistance and establishment of metastatic disease, have provided the rationale for revolutionary targeted approaches against Cyclin-Dependent Kinases 4/6 (CDK4/6), PI3 Kinase (PI3K), Poly ADP Ribose Polymerase (PARP) and Programmed Death-Ligand 1 (PD-L1), among others. In this review, we focus on the comprehensive overview of epidemiology and current standard of care treatment of metastatic breast cancer, along with ongoing clinical trials. Towards this goal, we utilized available literature from PubMed and ongoing clinical trial information from clinicaltrials.gov to reflect the up to date and future treatment options for metastatic breast cancer.

Application of Fluorescence Dye in Combination with Methylene Blue for Axillary Reverse Mapping in Patients with Modified Radical Mastectomy for Breast Cancer.

Background Axillary reverse mapping (ARM) is a novel intraoperative technique developed in recent years. This study was aimed at determining the effect of combined use of fluorescence dye and methylene blue, as well as its feasibility of ARM in patients with breast cancers who undergo modified radical mastectomy. Method From January 2016 to June 2017, 46 patients with primary breast cancer at stage I-IV (Tis-T3, N0-N3, and M0) who received modified radical mastectomy were enrolled in this study. The exclusion criteria included preoperative radiotherapy/chemotherapy or bilateral disease. Patients were divided into 2 groups: methylene blue group (22 patients, 47.8%) and methylene blue+indocyanine green (ICG) group (24 patients, 52.2%). ARM was performed before surgery. Results The overall visualization rate of ARM nodes was 80.4% (37/46). The visualization rate was significantly increased in methylene blue+ICG group (91.67%, 22/24) than that in methylene blue group (63.64%, 14/22) (P = 0.032). There was a statistical difference of visualization rate between clinical groups of N0-N1 and N3-N4 (t = 2.431, P = 0.19 < 0.05). Although there was no significant difference found in the total drainage volume and arm perimeter of patients between the two groups (P > 0.05), the harden diameter of the injection site in methylene blue+ICG group was significantly longer than that in methylene blue group (P < 0.05). Among the 44 patients with different molecular profiling of breast cancer, there was no significant difference of visualization rate of ARM nodes in luminal A group (100%,5/5), luminal B group (75.0%, 18/24, P = 0.21), HER2 group (75.0%, 6/8, P = 0.22), and basal-like group (85.7%, 6/7, P = 0.37). Conclusions ARM using methylene blue+ICG presented greater identification rate than that with methylene blue alone, especially in patients with more invasive breast cancer. Our finding offers support for the improvement of ARM in future breast cancer management.

Socioeconomic, Clinical, and Molecular Features of Breast Cancer Influence Overall Survival of Latin American Women.

Molecular profile of breast cancer in Latin-American women was studied in five countries: Argentina, Brazil, Chile, Mexico, and Uruguay. Data about socioeconomic characteristics, risk factors, prognostic factors, and molecular subtypes were described, and the 60-month overall cumulative survival probabilities (OS) were estimated. From 2011 to 2013, 1,300 eligible Latin-American women 18 years or older, with a diagnosis of breast cancer in clinical stage II or III, and performance status ≦̸1 were invited to participate in a prospective cohort study. Face-to-face interviews were conducted, and clinical and outcome data, including death, were extracted from medical records. Unadjusted associations were evaluated by Chi-squared and Fisher’s exact tests and the OS by Kaplan–Meier method. Log-rank test was used to determine differences between cumulative probability curves. Multivariable adjustment was carried out by entering potential confounders in the Cox regression model. The OS at 60 months was 83.9%. Multivariable-adjusted death hazard differences were found for women living in Argentina (2.27), Chile (1.95), and Uruguay (2.42) compared with Mexican women, for older (≥60 years) (1.84) compared with younger (≤40 years) women, for basal-like subtype (5.8), luminal B (2.43), and HER2-enriched (2.52) compared with luminal A subtype, and for tumor clinical stages IIB (1.91), IIIA (3.54), and IIIB (3.94) compared with stage IIA women. OS was associated with country of residence, PAM50 intrinsic subtype, age, and tumor stage at diagnosis. While the latter is known to be influenced by access to care, including cancer screening, timely diagnosis and treatment, including access to more effective treatment protocols, it may also influence epigenetic changes that, potentially, impact molecular subtypes. Data derived from heretofore understudied populations with unique geographic ancestry and sociocultural experiences are critical to furthering our understanding of this complexity.

Is There a Correlation between Multiparametric Assessment in Ultrasound and Intrinsic Subtype of Breast Cancer?

Molecular profile of breast cancer provides information about its biological activity, prognosis and treatment strategies. The purpose of our study was to investigate the correlation between ultrasound features and molecular subtypes of breast cancer. From June 2019 to December 2019, 86 patients (median age 57 years; range 32–88) with 102 breast cancer tumors were included in the study. The molecular subtypes were classified into five types: luminal A (LA), luminal B without HER2 overexpression (LB HER2−), luminal B with HER2 overexpression (LB HER2+), human epidermal growth factor receptor 2 positive (HER2+) and triple negative breast cancer (TNBC). Histopathological verification was obtained in core biopsy or/and post-surgery specimens in all cases. Univariate logistic regression analysis was performed to assess the association between the subtypes and ultrasound imaging features. Experienced radiologists assessed lesions according to the BIRADS-US lexicon. The ultrasound scans were performed with a Supersonic Aixplorer and Supersonix. Based on histopathological verification, the rates of LA, LB HER2−, LB HER2+, HER2+, and TNBC were 33, 17, 17, 16, 19, respectively. Both LB HER2+ and HER2+ subtypes presented higher incidence of calcification (OR = 3.125, p = 0.02, CI 0.0917–5.87) and HER2+ subtype presented a higher incidence of posterior enhancement (OR = 5.75, p = 0.03, CI 1.2257–32.8005), compared to other subtypes. The calcifications were less common in TNBC (OR = 0.176, p = 0.0041, CI 0.0469–0.5335) compared to other subtypes. There were no differences with regard to margin, shape, orientation, elasticity values and vascularity among five molecular subtypes. Our results suggest that there is a correlation between ultrasonographic features assessed according to BIRADS-US lexicon and BC subtypes with HER2 overexpression (both LB HER2+ and HER2+). It may be useful for identification of these aggressive subtypes of breast cancer.

P–446 Controlled ovarian stimulation protocols for oocyte vitrification induce differential gene expression profiles in primary tumours of breast cancer

Abstract Study question Could controlled ovarian stimulation (COS) protocols used in fertility preservation (FP) impact on malignant cell proliferation and tumour molecular profiling of breast cancer (BC) patients? Summary answer Letrozole supplementation during ovarian stimulation for oocyte vitrification could be considered as a safe procedure in estrogen-dependent BC patients undergoing FP. What is known already High estradiol levels associated to COS could promote changes in gene expression in estrogen-positive BC tumors. Estradiol levels reached during the ovarian stimulation could aggressively promote malignant cell proliferation and cell migration to adjacent organs. Aromatase inhibitors such as letrozole, are added to standard stimulation protocols to avoid this undesirable potential side effect. Despite the reassuring clinical results achieved by using letrozole for FP in BC patients, there is still a lack of evidence regarding its impact on malignant cell behaviour. For this reason, specific molecular studies to properly evaluate safety of letrozole in this specific population are still required. Study design, size, duration Experimental in vivo study. Thirty 5-week-old Nude-nu female mice were divided into three different groups: BC (n = 10), BC and FSH stimulation (BC-FSH, n = 10), or BC and letrozole stimulation (BC-LTZ, n = 10). BC was considered the control group, whereas BC-FSH and BC-LTZ represented distinct COS protocols. Hormone-dependent BC was induced in all mice. Animals were followed-up for 5 months and then euthanized to collect kidney, ovary, spleen, and liver tissues for gene expression and immunohistochemistry (IHC) analysis. Participants/materials, setting, methods One million of human MCF–7 BC cells were injected into the mouse left kidney capsule. Two days after xenograft, COS was induced by 10IU FSH or 1mg/ml letrozole + 10IU FSH, followed by ovarian triggering with 10IU hCG at 48h. Human BC RT2 Profiler PCR Arrays were performed to evaluate the impact of COS on tumour behaviour. BC biomarkers (Ki67, Erα, PR and HER–2) were also analyzed by IHC to validate gene expression results. Main results and the role of chance The differential gene expression was firstly assessed in kidney samples, as they represent the xenograft site, and different expression profiles were obtained depending on the COS protocol used. The BC-FSH group showed a global over-expression pattern of all genes of the array when compared to BC and BC-LTZ. Further gene ontology analysis revealed that cellular process, biological regulation, metabolic process, and proteases were the most over-represented biological terms, with a 20.5-fold over-expression for MMP2 compared to the other groups. On the other hand, BC-LTZ mice presented gene expression profiles similar to that of controls. When other tissues were analysed to detect malignant cell presence, our results revealed a significant up-regulation of matrix-proteases, cell cycle and proliferation related-genes, in liver samples from the BC-FSH group, but no amplification of any of the studied genes was detected in ovarian tissue or spleen. IHC findings confirmed the presence of human BC cells in 100% of samples from kidney tissue and in 30% of samples from liver tissue in the BC-FSH group. No human cells were detected by IHC in the BC and BC-LTZ groups. Limitations, reasons for caution Since this is an animal model of estrogen-dependent BC induced through a cell line, further validation with human tumour breast cancer samples would be required. Wider implications of the findings: Adjuvant letrozole in COS protocols prevents BC cell migration. The present study suggests that this protective effect could be mediated by interfering ER-pathway downstream genes involved in cell proliferation and matrix digestion. Altogether, letrozole could safely be used as a supplement during COS procedures for oocyte vitrification in BC women. Trial registration number Not applicable

Concordance of the molecular subtype classification between core needle biopsy and surgical specimen in primary breast cancer.

Molecular profiling of breast cancer (BC) classifies several intrinsic subtypes based on different patterns of gene expression. Multigene assays estimate the risk of recurrence and help to select high-risk patients for adjuvant chemotherapy. However, these tests are associated with significant costs. Immunohistochemistry (IHC) offers a surrogate classification for molecular subtypes by determining estrogen (ER) and progesterone receptors (PR), human epidermal growth factor (Her2neu), as well as the proliferation marker Ki67. Core needle biopsy (CNB) is well established in BC diagnosis and allows a pre-operative assessment of biomarkers. The aim of this study was to analyze the concordance of these markers between CNB and surgical specimens to assess whether re-testing of the surgical specimen is mandatory. Within a 3-year period, patients with primary BC and paired samples of CNB and surgical specimens were analyzed retrospectively. Concordance rates of ER, PR, Her2neu, Ki67, and the surrogate classification for molecular subtypes were calculated using the Landis and Koch agreement grades. Out of 2254 patients with primary breast cancer, 1307 paired specimens without pre-operative treatment were available for analysis Concordance rates for ER, PR, Her2neu, and Ki67 status showed substantial-to-almost perfect agreement grades (κ = 0.91, 0.75, 0.89, and 0.61, respectively). Though substantial concordance was also found for the subtype classification (κ = 0.70), the molecular subtype changed in 18.5% of patients based on the testing of the surgical specimen, mainly from luminal A-like to luminal B-like. Though the concordance rates for single markers were convincing, a significant proportion of the molecular subtypes differed between CNB and the surgical specimen. Re-testing of PR and Ki67 is mandatory to ensure optimal treatment decisions. Further research is necessary to define safe, efficient, and cost-effective predictive models in adjuvant breast cancer therapy.

Impact of Molecular Profiling of Breast Cancer on the Rate of Locoregional Recurrence in Young Versus Old Female Patients.

BackgroundBreast cancer (BC) is diverse regarding its natural history and treatment responses. The traditional histopathological classification is unable to confine this diverse clinical heterogeneity. Classically, prognosis and treatment response are influenced by factors including histological grade, lymph node status, and tumour size. Recently, research has diverted from histological classification towards molecular classification.We aim to analyse the locoregional recurrence of breast cancer incidence following surgery across the different molecular subtypes as well as relation to age.Materials and methodsFemale patients diagnosed with a locoregional recurrence of breast carcinoma in 2012-2014 were identified from our centre histology department. We only included stage I-III patients who were previously treated with surgery achieving negative surgical margins and later developed locoregional recurrence during our study period. These patients were subdivided by age into old (≥40 years old) and young (<40 years old) groups according to their initial diagnosis age. Furthermore, they were categorised according to the molecular subtype of their primary tumour.ResultsOur study included 184 patients (124 designated to the old age group, 60 to the young age group). In the young group, recurrence occurred after a mean of 4.3 years and the range was one to 23 years, while in the old group, the mean was 3.8 years, and the range was one to 14 years. The most primary cancer subtype recorded was triple-negative (41.85%): 50 old patients and 27 young. Next was the Her-2/neu enriched subtype (27.72%): 35 old patients and 16 young, following this was luminal A subtype (21.19%): 27 old and 12 young. Last was the luminal B subtype (9.24%): 12 old patients and five young.ConclusionsTo conclude, in our series, the most common molecular subtype found in the recurrent cases was the luminal negative subtypes, with a relatively similar pattern across both age groups. The results of this study can be used as a basis for large prospective studies in our centre to further analyse the effect of molecular subtyping on the recurrence rates of BC.

The molecular landscape of Asian breast cancers reveals clinically relevant population-specific differences

Molecular profiling of breast cancer has enabled the development of more robust molecular prognostic signatures and therapeutic options for breast cancer patients. However, non-Caucasian populations remain understudied. Here, we present the mutational, transcriptional, and copy number profiles of 560 Malaysian breast tumours and a comparative analysis of breast cancers arising in Asian and Caucasian women. Compared to breast tumours in Caucasian women, we show an increased prevalence of HER2-enriched molecular subtypes and higher prevalence of TP53 somatic mutations in ER+ Asian breast tumours. We also observe elevated immune scores in Asian breast tumours, suggesting potential clinical response to immune checkpoint inhibitors. Whilst HER2-subtype and enriched immune score are associated with improved survival, presence of TP53 somatic mutations is associated with poorer survival in ER+ tumours. Taken together, these population differences unveil opportunities to improve the understanding of this disease and lay the foundation for precision medicine in different populations.

Ultradeep targeted sequencing of circulating tumor DNA in plasma of early and advanced breast cancer.

We present a study to evaluate the feasibility and clinical utility of amplicon‐based Oncomine Pan‐Cancer cell‐free assay to detect circulating tumor DNA (ctDNA) in patients with early or advanced breast cancer. In this study, 109 early and metastatic breast cancer patients were recruited before the initiation of treatment. ctDNA mutation profiles were assessed through unique molecular tagging (UMT) and ultradeep next generation sequencing (NGS). For patients with mutations, DNA from corresponding white blood cells (WBC) was sequenced to exclude variants of clonal‐hematopoietic (CH) origin. UMT targeted sequencing from plasma of 109 patients achieved a median total coverage of 55 498X and a median molecular coverage of 4187X. Among 53 ctDNA positive samples, 38% were mutation positive by WBC sequencing, indicating potentially false‐positive results contributed by CH origin. Prevalence of CH‐related mutations was associated with age (P = 7.51 × 10−4). After exclusion of CH mutations, ctDNA detection rates were 37% for local or locally advanced breast cancer (stage I‐III) and 81% for metastatic or recurrent breast cancer. The ctDNA detection rate correlated with disease stage (P = 2.60 × 10−4), nodal spread (P = 6.49 × 10−3) and the status of distant metastases (P = 5.00 × 10−4). ctDNA variants were detected mostly in TP53, PIK3CA and AKT1 genes, with variants showing therapeutic relevance. This pilot study endorses the use of targeted NGS for non‐invasive molecular profiling of breast cancer. Paired sequencing of plasma ctDNA and WBC should be implemented to improve accurate interpretation of liquid biopsy.

Abstract 2117: Immune contexture of breast cancer subtypes in real-world molecular data

Abstract Background: Breast cancer has been extensively characterized molecularly with cohorts such as The Cancer Genome Atlas (TCGA). However, as the standard of care evolves, there is a need for contemporaneous datasets with more detailed treatment and outcomes data. The ORIEN Avatar database (M2Gen, Tampa, FL) consists of clinical and tumor sequencing data from treatment-naive or on-treatment biopsies. This study evaluates the quality of ORIEN Avatar's RNA-sequencing (RNASeq) data, interrogates the impact of prior treatment on PAM50 subtyping, and reports associations between breast cancer subtypes and estimated immune cell populations. Methods: In the ORIEN Avatar dataset, RNASeq data were prepared using a multistep normalization process. We assessed the quality of normalized RNASeq data using 3 modalities. Using a principal-component (PC) analysis, we interrogated the association of gene expression with potential technical and biological factors, including processing batch effects, preservation methods, and treatment status. We evaluated concordance between immunohistochemistry (IHC)-based subtyping and intrinsic subtyping using the PAM50 gene expression signature. Prevalence of in silico-derived tumor-infiltrating immune cell compositions in ORIEN Avatar compared with TCGA breast cancer samples was investigated. Results: The ORIEN Avatar breast cancer cohort comprised 560 patients with RNASeq data. The first 3 PCs (accounting for 11%, 9%, and 7% of total explained variance, respectively) were associated with biological factors, including PAM50 subtyping and HER2 receptor and hormone receptor (HR) status. PAM50 subtype was largely concordant with IHC subtype, per clinical record of biomarker testing, and prior treatment status did not impact PAM50 subtyping. We observed that 68% of basal samples were triple negative by IHC, 92% of luminal samples were HER2-negative and HR-positive by IHC, and 62% of HER2-enriched samples per RNASeq were HER2 receptor positive by IHC. Several in silico-derived tumor-infiltrating immune cell compositions, including activated CD4 memory T cells, M0 macrophages, and activated dendritic cells, were enriched in triple-negative breast cancer samples, consistent with TCGA. Correlative analysis of RNASeq data with other biomarker data and clinical outcome is planned. Conclusions: Our results using real-world (RW) ORIEN Avatar RNASeq data were consistent with molecular profiling of breast cancer assessed using IHC and TCGA. This proof-of-concept study found no association between prior treatment status and PAM50 intrinsic subtyping. Our analysis provides a framework to assess the quality of RW molecular profiling and highlights the feasibility of leveraging harmonized molecular data to replicate and discover novel biological evidence. Citation Format: Xuya Wang, Bin Li, Peter M. Szabo, Han Chang, Mustimbo Roberts, Alice M. Walsh. Immune contexture of breast cancer subtypes in real-world molecular data [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2117.

Molecular Profiles of Breast Cancer in a Single Institution.

Historically, breast cancer has been treated according to an evaluation of biomarkers, such as the estrogen receptor and HER2 status. Recently, molecular profiling has been used to detect driver mutations and select anti-cancer treatment strategies. In addition to detecting pathogenic mutations, the total mutation count (tumor mutation burden) has been considered as another biomarker. We performed molecular profiling of 143 breast cancer tissues obtained from resected tissues via surgical operation. Suspected germline mutations were detected in 10% of the patients with a higher somatic mutation ratio. As hypermutated breast cancers are more likely to benefit from certain anti-cancer treatment strategies, molecular profiling can be used as a biomarker.

The US–Latin America Cancer Research Network

PURPOSE The National Cancer Institute (NCI) Center for Global Health promotes global oncology research to reduce cancer burden worldwide. In 2009, NCI launched the Latin American Cancer Research Network (LACRN) to support a clinical cancer research network in Latin America. LACRN was started by a coalition of research institutions through bilateral collaborative agreements between the US Department of Health and Human Services and the governments of Argentina, Brazil, Chile, Mexico, and Uruguay. The LACRN is supported through a research contract to a study coordination center and subcontracts to 6 low- and middle-income country sites. The participating countries have a shared goal that meets the specific research needs of the regions. The overarching purpose of this endeavor is to implement high-quality standards for conducting clinical research studies and developing collaborative cancer research projects. METHODS NCI supported a clinical breast cancer project for LACRN, “Molecular profiling of breast cancer (MPBC) in Latin American women with stage II and III breast cancer receiving standard neo-adjuvant chemotherapy.” The molecular profiling of breast cancer study was conducted in 40 hospitals and research institutions across 5 countries with a study population of approximately 1,400 patients. RESULTS AND CONCLUSION Establishing a comprehensive network in Latin America and their research institutions yielded an incredible research resource that can be used in future studies, driven by the network. Throughout the process of developing and implementing studies, LACRN helped identify key elements of the functionality of research networks, such as the pivotal role of institutional and government commitment for sustainability; the importance of building multidisciplinary teams, transparent communications, and training; the ability to combine translational, epidemiology, and clinical research to close research gaps; and the application of new technologies to standard cancer clinical care.

Abstract IA41: Precision medicine in cancer: Opportunities and challenges in Latin America

Abstract In the last few years, the importance of diversity in precision medicine has been highlighted. However, in many cases, the scientific focus has been put on the need of genetic studies in diverse populations, even when the very definition of precision medicine has included both genetic and environmental aspects that affect the development of a certain disease in an individual. In particular, cancer, like other complex diseases, is considered a result of how chronic exposure to etiologically relevant environmental factors (the “exposome”) impacts on the more or less susceptible genetic information of a subject. Studies on cohorts other than White have highlighted variations on cancer incidence, prognosis, and response to treatment that reinforce the need to establish the unique genetic and environmental characteristics that are present in geographically diverse populations. In particular, Hispanic/Latinos have been the subject of several initiatives in the US that try to discriminate the impact of the variations on genome and exposome in the incidence and severity of cancers. Among other findings, it is fairly clear now that Latinos tend to have overall better health indicators than those of other ethnic groups with whom they share demographic and socioeconomic characteristics; however, cancer incidence rates tend to increase in US Latinos with respect to those reported in Latin America. Registries of cancer are scarcely populated in Latin America and for this reason, differences in completeness of registries cannot be ruled out as a confounding factor in this generalization. Unfortunately, there are still very little data publicly available in Latin America to address the causality of these differences. Moreover, Latin America is in itself a complex mixture of genetic variation and geographical features, and in particular the South American admixture, which represents less than 15% of US Latinos, is hardly represented in the current precision medicine landscape. Thus, there is a compelling need to establish Latin American case and control cohorts that may help to discern the relevance of genetic and environmental factors in the characteristics of cancer in this population. A few regional initiatives try to address, at least partially, this lack of knowledge. An example of these initiatives is the Latin American Cancer Research Network (LACRN), which comprises a coalition of hospitals (mostly public); basic, translational, and clinical science investigators and institutions; and government officials from Argentina, Brazil, Chile, Mexico, and Uruguay. Its first study, sponsored by NCI's Center for Global Health (CGH) and regional partners, focuses in the molecular and epidemiologic characterization of a real-world cohort of breast cancer patients (the MPBC study, for Molecular Profile of Breast Cancer) and has currently 1,314 eligible patients who have been enrolled from 2011 to 2013 and have been studied and followed for &amp;gt;5 years under GCP quality standards. As well, associated biobanks have been established to keep TCGA-quality biospecimens in optimal conditions for molecular studies. In my talk, I am going to address some of the difficulties that arise when these projects are developed in Latin America, especially when developed at the level of public health services. The success in these projects depends upon applying specific strategies to overcome the aforementioned difficulties so as to contribute to the knowledge of diversity in precision medicine in Latin America. Citation Format: Andrea S. Llera. Precision medicine in cancer: Opportunities and challenges in Latin America [abstract]. In: Proceedings of the Eleventh AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2018 Nov 2-5; New Orleans, LA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2020;29(6 Suppl):Abstract nr IA41.

Molecular profiling in breast cancer—ready for clinical routine?

SummaryThe herald of genomic testing opened novel diagnostic and therapeutic possibilities for many tumor entities. For breast cancer, molecular profiling has become an integral part of disease management on multiple levels. Genetic testing allows for the identification of hereditary cancer syndromes in patients with a family history of malignancies and contributes to the successful prevention of breast cancer. In early breast cancer, several prospective randomized trials demonstrated the prognostic significance of commercially available mRNA-based gene expression analyses, which now have become part of standard of care in the adjuvant setting. In advanced breast cancer, testing for targetable mutations ensures personalized cancer treatment. Poly-ADP-ribose polymerase (PARP) inhibitors provide the first targeted alternative for patients with BRCA 1/2-associated breast cancer. In advanced breast cancer of luminal type, the detection of Phosphatidylinositol-4,5-Bisphosphate 3-Kinase Catalytic Subunit Alpha (PIK3CA) mutations provides a novel treatment option with alpelisib, a PIK3CA inhibitor. Further targetable mutations include NTRK3 in rare cases of secretory breast carcinoma and human epidermal growth factor receptor 2 (HER2). Recent data support the importance of the analysis of circulating tumor cells and cell-free DNA. These “liquid biopsies” open novel possibilities of molecular profiling. However, clinical benefit of such analyses remains to be confirmed.

Genetic Profiling of Breast Cancer with and Without Preexisting Metabolic Disease

Breast cancer is the most commonly diagnosed cancer and the leading cause of cancer death among women. Various mechanisms are involved in the initiation and progression of breast cancer. Metabolic dysregulation has been associated with increasing breast cancer incidence and mortality. However, little is known about how metabolic disease regulates the development and progression of breast cancer at the molecular level. Here, using a hybridization capture-based panel including 124 cancer-associated genes, we performed targeted next-generation sequencing of tumor tissues and matched blood samples from 20 postmenopausal patients with primary breast cancer, in which 6 cases suffered from preexisting metabolic disorders including hypertension, type 2 diabetes, and coronary heart disease. We took only the protein-altering variants and identified 170 somatic mutations of 59 genes. Among these, 40 mutated genes were found in the metabolic disease group, and 33 mutated genes were found in the non–metabolic disease group. Importantly, nonsynonymous mutations of 26 genes (MSH3, BRAF, MLH3, MTOR, DDR2, ALK, etc.) were uniquely present in the metabolic disease group. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes enrichment analysis were performed to investigate biological functions and key pathways of somatic mutations. TP53, PIK3CA, and PTEN were the top three commonly mutated genes at a higher frequency compared with the Cancer Genome Atlas (TCGA) data, and several novel but infrequent mutations in other genes were also found. Although further studies are required to validate these variants, our results are the first to suggest a specific molecular profile of breast cancer with preexisting metabolic disease.

Abstract 1731: Functional genomics approach reveals distinct neo-morphic activities and co-driver mutations of different missense mutant TP53 that promote breast cancer heterogeneity and progression

Abstract Phenotypic and molecular profiling of breast cancer demonstrates a high degree of heterogeneity in the breast tumors. TP53 tumor suppressor is mutated in 30% of all breast tumors and 80% of the basal-like subtype with numerous co-existing somatic mutations in other genes. We hypothesize that tumor heterogeneity is a result of combination of neo-morphic functions of specific TP53 driver mutations and functionally important but distinct co-mutations (or ‘co-drivers’) for each type of TP53 mutation. We thus ectopically expressed 10 most common missense p53 mutant proteins found in breast cancer patients in normal-like mammary epithelial cells and examined their phenotypes associated with various hallmarks of cancer. Supporting our hypothesis, a wide spectrum of phenotypic changes in cell survival, resistance to apoptosis and anoikis, cell invasion and cell polarity across the 10 mutant cell lines were observed compared to those of WT p53 expressing cells. Integrated analysis of ChIP-seq and RNA-seq showed distinct promoter binding profiles of different p53 mutant proteins, implying that non-canonical transcriptional activity of mutant p53 proteins may contribute to the phenotypic heterogeneity in TP53-mutated tumor. Further, enrichment and model-based pathway analyses revealed the key biological pathways associated with invasion of cells expressing different p53 mutant proteins. As a proof-of-concept for screening co-drivers of mutant p53, when PTEN was deleted in non-invasive cells expressing p53-Y234C mutant using CRISPR-Cas9 system, an increase in cell invasion was observed. A genome-wide CRISPR library-based screen on p53-Y234C and p53-R273C cells identified completely different candidate co-driver mutations that promoted cell invasion. The top candidates included several known mutated genes in breast cancer patients harboring TP53 mutations and were associated with cytoskeletal and apoptosis resistance pathways. Overall, our combined approach of molecular profiling and functional genomics screens highlighted distinct sets of genes and co-driver mutations that can lead to heterogeneous phenotypes and promote aggressive behaviors of cells with different TP53 mutant backgrounds, which can guide development of novel targeted therapies. Citation Format: Anasuya Pal, Anasuya Pal. Functional genomics approach reveals distinct neo-morphic activities and co-driver mutations of different missense mutant TP53 that promote breast cancer heterogeneity and progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1731.