Basal Type Research Articles

Abstract PS18-03: Differential gene expression in luminal-type invasive lobular carcinoma and invasive ductal carcinoma by MammaPrint risk stratification

Abstract Background: Invasive lobular carcinoma (ILC) comprises 10-15% of breast tumors and is the second most common histological type after invasive ductal carcinoma (IDC). Patients with ILC are often diagnosed at an older age and more advanced stage than those with IDC. Late recurrences and worse long-term survival suggest the need for improved approaches to treatment optimization and exploration of molecular pathways unique to ILC. Although previous reports have described comprehensive transcriptomic profiling of ILC, these were limited by small sample sizes. Furthermore, differential gene expression between ILC and IDC within genomic risk groups and molecular subtypes has yet to be explored. Here we characterize differential gene expression between ILC and IDC in a large, age-matched patient subset categorized by 70-gene signature/MammaPrint (MP) risk and 80-gene signature/BluePrint (BP) subtype. Methods: The prospective FLEX Registry (NCT03053193) includes stage I-III primary invasive breast cancer patients who receive MP/BP testing and consent to full transcriptome and clinical data collection. This sub-analysis included 450 ILC patients enrolled from 2017 to present. Compared with a random selection of IDC patients (n=450, mean age, 60 years), ILC patients were older (mean, 63 years, p<0.001). Thus, we selected an age-matched subset for differential gene expression analysis. There were few non-Luminal ILCs; thus, gene expression analyses were limited to BP Luminal tumors. A subset of 413 age-matched pairs (n=826) of ILC and IDC were used for analysis. Gene expression data were quantile normalized using R limma package, and differentially expressed genes (DEGs) were compared between groups. DEGs with an adjusted p<0.05 and log2 fold change > ± 1.0 were considered significant. Results: ILC represented 13% of FLEX cases (n=450/3562), and were 81% lymph node-negative, 99% ER+, 94% HER2-negative, and 68% MP Low Risk (LR). By BP, ILC were 99% Luminal, 1% HER2, and <1% Basal type. BP Luminal ILC were predominantly grade 2 (63%), T1 (61%), node-negative (84%), and MP LR (69%). Menopausal status, nodal status, ethnicity, BMI distribution, and frequency of type 2 diabetes mellitus were similar between ILC and IDC. However, IDC were more likely to be MP HR (46% IDC vs. 31% ILC, p<0.001) and grade 3 (15% IDC vs. 4% ILC, p<0.001). ILC were more likely to be T3 (10% ILC vs. 1% IDC, p<0.001). We found 4 DEGs common to all comparisons: all Luminal ILC vs. IDC, MP LR ILC vs. IDC, and MP HR ILC vs. IDC. ILC had lower expression of CDH1 (E-cadherin) than IDC, regardless of MP risk. Including CDH1, 6 unique genes were differentially expressed in LR ILC compared with IDC, and 21 genes were differentially expressed in HR ILC compared with IDC. Genes with increased expression in HR ILC were related to immune cell migration/chemotaxis, hormone signaling, and growth factor signaling. HR ILCs were also enriched for TGFβ signaling and angiogenesis pathway genes. Conclusions: Here we report differential clinical and molecular characteristics between ILC and IDC in a large, age-matched patient subset. Regardless of MP risk, expression of CDH1 was lower in ILC compared with IDC. Approximately one-third of ILCs were MP HR, and we report a greater number and diversity of DEGs between HR ILC and HR IDC compared with LR tumors, in particular genes related to TGFβ signaling. TGFβ pathway genes play a variety of roles in the tumor microenvironment, including induction of angiogenesis, fibroblast growth factor stimulation, and inhibition and/or exclusion of an immune response. These results suggest that therapeutic strategies targeting the TGFβ pathway may be future avenues of exploration in ILC, although further studies are warranted to characterize underlying molecular mechanisms. Citation Format: Beth-Ann Lesnikoski, Jennifer A. Crozier, Gordan Srkalovic, Patricia Robinson, Clodia Osipo, Kaylan Banda, Heather M. Kling, Josien Haan, William Audeh, FLEX Investigators Group. Differential gene expression in luminal-type invasive lobular carcinoma and invasive ductal carcinoma by MammaPrint risk stratification [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS18-03.

Enteric methane emission and growth rate of three different breeds of beef cattle fed on oil palm frond or grass basal diet

Enteric methane emission in ruminants is affected by the feed quality. The study aimed to evaluate the performance and enteric methane emission of three beef cattle breeds fed on grass or oil palm frond basal diet. The study used three cattle breed, namely Bali, Ongole crossbred, and PO crossbred. The cattle were fed on grass or oil palm frond (OPF) basal diet and supplemented with concentrate based on palm kernel cake (PKC). Diet was fed in total mixed ration contained 12% crude protein. The study was conducted in a randomized block design in a 3x2 factorial arrangement. Results showed that the growth rate, dry matter digestibility, and enteric methane emission were not affected (P>0.05) by the interaction of cattle breed and basal diet type. No main factor affected (P>0.05) on the parameter observed. The average values of dry matter digestibility, average daily gain (ADG), methane emission, and methane intensity of the cattle were 63.38%, 359 g day−1, 91. L day-1, and 177.32 gCH4 kg ADG−1, respectively. The study indicates that OPF could be used as a basal diet of beef cattle without affecting its performance. Cattle breeds had similar performance either fed on grass or OPF basal diet.

Comparative stemness and differentiation of luminal and basal breast cancer stem cell type under glutamine-deprivation.

Glutamine (gln) metabolism has emerged as a cancer therapeutic target in past few years, however, the effect of gln-deprivation of bCSCs remains elusive in breast cancer. In this study, effect of glutamine on stemness and differentiation potential of bCSCs isolated from MCF-7 and MDAMB-231 were studied. We have shown that bCSCs differentiate into CD24+ epithelial population under gln-deprivation and demonstrated increased expression of epithelial markers such as e-cadherin, claudin-1 and decreased expression of mesenchymal protein n-cadherin. MCF-7-bCSCs showed a decrease in EpCAMhigh population whereas MDAMB-231-bCSCs increased CD44high population in response to gln-deprivation. The expression of intracellular stem cell markers such sox-2, oct-4 and nanog showed a drastic decrease in gene expression under gln-deprived MDAMB-231-bCSCs. Finally, localization of β-catenin in MCF-7 and MDAMB-231 cells showed its accumulation in cytosol or perinuclear space reducing its efficiency to transcribe downstream genes. Conclusively, our study demonstrated that gln-deprivation induces differentiation of bCSCs into epithelial subtypes and also reduces stemness of bCSCs mediated by reduced nuclear localization of β-catenin. It also suggests that basal and luminal bCSCs respond differentially towards changes in extracellular and intracellular gln. This study could significantly affect the gln targeting regimen of breast cancer therapeutics.

P63 expression in human tumors and normal tissues: a tissue microarray study on 10,200 tumors

BackgroundTumor protein 63 (p63) is a transcription factor of the p53 gene family involved in differentiation of several tissues including squamous epithelium. p63 immunohistochemistry is broadly used for tumor classification but published data on its expression in cancer is conflicting.MethodsTo comprehensively catalogue p63 expression, tissue microarrays (TMAs) containing 12,620 tissue samples from 115 tumor entities and 76 normal tissue types were analyzed.Resultsp63 expression was seen in various normal tissues including squamous epithelium and urothelium. At least occasional weak p63 positivity could be detected in 61 (53%) of 115 different tumor types. The frequencies of p63 positivity was highest in squamous cell carcinomas irrespective of their origin (96–100%), thymic tumors (100%), urothelial carcinomas (81–100%), basal type tumors such as basal cell carcinomas (100%), and various salivary gland neoplasias (81–100%). As a rule, p63 was mostly expressed in cancers derived from p63 positive normal tissues and mostly not detectable in tumors derived from p63 negative cancers. However, exceptions from this rule occurred. A positive p63 immunostaining in cancers derived from p63 negative tissues was unrelated to aggressive phenotype in 422 pancreatic cancers, 160 endometrium cancers and 374 ovarian cancers and might be caused by aberrant squamous differentiation or represent stem cell properties. In 355 gastric cancers, aberrant p63 expression occurred in 4% and was linked to lymph node metastasis (p = 0.0208). Loss of p63 in urothelial carcinomas - derived from p63 positive urothelium - was significantly linked to advanced stage, high grade (p < 0.0001 each) and poor survival (p < 0.0001) and might reflect clinically relevant tumor dedifferentiation.ConclusionThe high prevalence of p63 expression in specific tumor types makes p63 immunohistochemistry a suitable diagnostic tool. Loss of p63 expression might constitute a feature of aggressive cancers.

Stem Cell-derived Respiratory Epithelial Cell Cultures as Human Disease Models.

Advances in stem cell biology and the understanding of factors that determine lung stem cell self-renewal have enabled long-term in vitro culture of human lung cells derived from airway basal and alveolar type II cells. Improved capability to expand and study primary cells long term, including in clonal cultures that are recently derived from a single cell, will allow experiments that address fundamental questions about lung homeostasis and repair, as well as translational questions in asthma, chronic obstructive pulmonary disease, pulmonary fibrosis, and lung cancer research. Here, we provide a brief history of postnatal lung epithelial cell culture and describe recent methodological advances. We further discuss the applications of primary cultures in defining "normal" epithelium, in modeling lung disease, and in future cell therapies.

Basal type ‘broken heart’ in a postpartum woman: Utility of speckle tracking echocardiography

Basal type ‘broken heart’ in a postpartum woman: Utility of speckle tracking echocardiography

Generation of ACE2-Expressing Airway and Alveolar Epithelial Cells from Human Induced Pluripotent Stem Cells

SARS-CoV-2 is known to bind ACE2 as initial action of infection and cause COVID-19. ACE2 is known to express in various types of cells, such as, heart, liver, cornea, ileum, and respiratory cells are considered important in terms of clinical outcome. Supplying ACE2-expressing respiratory cells such as airway basal, club, ciliated and alveolar epithelial type II (ATII) cells are essential for COVID-19 research. However, differentiation of human ACE2-expressing respiratory cells is not fully understood. In the present study, we have generated ATII and club cells from human iPSCs using two-dimensional culture method. We differentiated human iPSCs using a stepwise protocol. After differentiation of lung progenitor cells by flow cytometry, airway and alveolar cells were induced. We found that the differentiated cells contained SFTPB-positive ATII cells and SCGB3A2-positive club cells. In addition, we found that these cells express ACE2 by RT-qPCR and immunofluorescence analysis. These data suggest that the iPSC-derived airway and alveolar cells can be applied as in vitro models for COVID-19 research. We are planning to perform infection of SARS-CoV-2 in the models.

Investigation of Candidate Genes and Pathways in Basal/TNBC Patients by Integrated Analysis.

Purpose:This study aims to identify the key pathway and related genes and to further explore the potential molecular mechanisms of triple negative breast cancer (TNBC).Methods:The transcriptome data and clinical information of breast cancer patients were downloaded from the TCGA database, including 94 cases of paracancerous tissue, 225 cases of Basal like type, 151 cases of Her2 type, 318 cases of Luminal type A, 281 cases of Luminal type B, and 89 cases of Normal Like type. The differentially expressed genes (DEGs) were identified based on the criteria of |logFC|≥1.5 and adjust P < 0.001.Their functions were annotated by gene ontology (GO) analysis and Kyoto Encyclopedia of differentially expressed genes & Genomes (KEGG) pathway analysis. Cox regression univariate analysis and Kaplan-Meier survival curves (Log-rank method) were used for survival analysis. FOXD1, DLL3 and LY6D were silenced in breast cancer cell lines, and cell viability was assessed by CCK-8 assay. Further, the expression of FOXD1, DLL3 and LY6D were explored by immunohistochemistry on triple negative breast tumor tissue and normal breast tissue.Results:A total of 533 DEGs were identified. Functional annotation showed that DEGs were significantly enriched in intermediate filament cytoskeleton, DNA−binding transcription activator activity, epidermis development, and Neuroactive ligand−receptor interaction. Survival analysis found that FOXD1, DLL3, and LY6D showed significant correlation with the prognosis of patients with the Basal-like type (P < 0.05). CCK-8 assay showed that compared with Doxorubicin alone group, the cytotoxicity of Doxorubicin combined with siRNA-knockdown of FOXD1, DLL3, or LY6D was much significant.Conclusion:The DEGs and their enriched functions and pathways identified in this study contribute to the understanding of the molecular mechanisms of TNBC. In addition, FOXD1, DLL3, and LY6D may be defined as the prognostic markers and potential therapeutic targets for TNBC patients.

On the activation energy for plastic flow during tension test at room temperature and the anisotropy behaviour of Ti-6Al-4V alloy

The movement of dislocations inside of crystals during the tension test at constant crosshead velocity is related to activation energy for plastic flow in spatially extended polycrystalline systems. In this work, the main objectives are to establish the influence of the slip systems for α hexagonal compact packaging (hcp) titanium phase on the activation energy for plastic flow and the anisotropy influence on the plastic flow behaviour of Ti-6Al-4V alloy in sheet form under tension test at constant crosshead velocity of 1 mm/min in three in-plane orientations, namely in the rolling 0° (RD), 45° (DD) and transverse (TD) directions, respectively. By applying the quantum mechanics and relativistic model (QM-RM) proposed by Muñoz-Andrade, it is shown that for a <c+a> pyramidal type slip with a Burgers vector b=0.553 nm, the activation energy barrier for plastic flow at maximum stress in uniaxial tension in the sheet rolling direction, Q=115.115 kJ/mol is lower in comparison with Q=116,629 kJ/mol obtained by the <a> basal type slip with a Burgers vector b=0.295nm. These results are in accordance with the reduced cross-slip activation energy barrier for cross-slip onto pyramidal planes in an a hexagonal crystalline structure with compact packaging (hcp) titanium phase. Additionally, it was shown that transverse direction (TD) of Ti-6Al-4V alloy in sheet form has developed the maximum stress in uniaxial tension test and ductility.

A Correlation of Pathological Characteristics and Immuno Histo Chemistry (IHC) Based Biomarker Expression in Triple Negative Breast Carcinomas

Background and aim: Triple-negative breast cancer (TNBC) is defined as a group of breast carcinomas that are negative for expression of hormone receptors estrogen receptor (ER) or progesterone receptor (PR) and human epidermal growth factor receptor 2 (Her2). In India, several reports have suggested that TNBC incidence is higher and up to 31%. Histological features of triple-negative breast cancer are reported to be common with those of basallike subtype, comprising of high-grade invasive ductal carcinoma, no special type, invasive ductal carcinoma with a large central acellular zone, typical medullary carcinoma, and metaplastic carcinomas. In the present study, we aimed to correlate the pathological characteristics and evaluate IHC based expression of basal type biomarkers in triple negative breast carcinomas. Our study showed heterogeneity in histology, most of the TNBC cases in our study were IDC, NOS and there were a slightly higher percentage of atypical medullary cases as compared to other studies. Most of the cases were high grade based on modified NBR grading and majority of them turned out to be basal like on IHC. Distinctly found in our cases which had metastatic axillary lymph node was, basal immune phenotype unlike the case in other studies which demonstrated a higher proportion of lymph node negativity. As expected a high proportion of triple negative tumors showed a consistent basal cytokeratin expression (CK5/6-66%, CK14-72%, CK17-68%). EGFR and p53 positivity did not show statistical significance between basal and non-basal groups. Proliferation marker Ki67 was statistically significant in basal like groups. Gene expression profiling is the gold standard for TNBC molecular subtype classification, however, IHC is an accepted 'surrogate marker' for identifying and classifying the 'basal like group'. Our findings suggest that pathologic characteristics cannot be used to accurately classify triple-negative breast cancer into basal and non-basal groups. Relevance to patients: DNA microarray based molecular profiling may not be accessible always in clinical settings, this study has emphasized that identification of basal like subtype of breast cancers can be done by doing easily available surrogate Immunohistochemical biomarkers on TNBC, and thus help to provide the patient with relevant target based therapeutic benefit.

Microstructure and Tensile Properties of Cost-Efficient Thermally Hardenable α + β Alloys of Ti–Al–Mo–Fe and Ti–Al–Mo–Cr Systems

Four new compositions of titanium alloys of Ti–Al–Mo–Fe and Ti–Al–Mo–Cr systems with content of β-phase stabilizing elements in a range of 3.8–9.8 wt.% (in terms of “molybdenum equivalent” CMo) were studied. The alloys were melted by single-melt electron-beam cold hearth technique, thermomechanically processed, and subjected to conventional (in furnace) or continuous rapid (resistant heating by electric current) heat treatments with different regimes. The microstructure, phase composition, and mechanical properties both in as-deformed state and after heat treatments were studied in detail. It is found that the proposed thermomechanical processing allows to obtain a transformed microstructure characterized by nearly equiaxed α-phase particles with relatively small aspect ratio and weak crystallographic texture of a basal type. Subsequent annealing in the α + β field led to completion of the α + β microstructure transformation; a microstructure of globular type was obtained in the low-alloyed (CMo = 3.8%) material. The phase transformations at various stages of strengthening heat treatments and their influence on the final microstructure and properties were studied, depending on the content of β-stabilizing elements and processing route. Particular attention was paid to the influence of the content of β-stabilizing element on the mechanical properties, as well as to the difference between the alloying systems with iron or chromium. It is shown that the alloys of proposed compositions provide high strength and reliability; they are competitive with conventional commercial alloys in terms of the balance of mechanical properties in hot deformed, annealed, and thermally strengthened states. Strengthening treatment based on rapid heating allows to achieve strength above 1500 MPa with sufficient ductility.

Basal type ‘broken heart’ in a postpartum woman: Utility of speckle tracking echocardiography

Basal type ‘broken heart’ in a postpartum woman: Utility of speckle tracking echocardiography

HER3 Differentiates Basal From Claudin Type Triple Negative Breast Cancer and Contributes to Drug and Microenvironmental Induced Resistance.

Triple Negative Breast Cancer (TNBC) is an aggressive form of Breast Cancer (BC). Numerous kinase inhibitors (KI) targeting different pathway nodes have shown limited benefit in the clinical setting. In this study, we aim to characterize the extent of HER3 reliance and to define the effect of Neuregulin (NRG) isoforms in TNBCs. Basal and Claudin type TNBC cell lines were treated with a range of small molecule inhibitors, in the presence or absence of the HER3 ligand NRG. Single agent and combination therapy was also evaluated in human cancer cell lines through viability and biochemical assessment of the AKT/MAPK signaling pathway. We show that Basal (BT20, HCC-70, and MDA-MB-468) and Claudin type (MDA-MB-231, BT-549) TNBC cell lines displayed differential reliance on the HER family of receptors. Expression and dynamic HER3 upregulation was predominant in the Basal TNBC subtype. Furthermore, the presence of the natural ligand NRG showed potent signaling through the HER3-AKT pathway, significantly diminishing the efficacy of the AKT and PI3K inhibitors tested. We report that NRG augments the HER3 feedback mechanism for continued cell survival in TNBC. We demonstrate that combination strategies to effectively block the EGFR-HER3-AKT pathway are necessary to overcome compensatory mechanisms to NRG dependent and independent resistance mechanisms. Our findings suggests that the EGFR-HER3 heterodimer forms a major signaling hub and is a key player in tumorigenesis in Basal but not Claudin type TNBC tested. Thus, HER3 could potentially serve as a biomarker for identifying patients in which targeted therapy against the EGFR-HER3-AKT axis would be most valuable.

Breast carcinoma histopathological correlation with molecular classification: A comparative study

Breast cancer is the most common malignancy in females. It isroutinely classified according to the WHO classification. However, molecular classification can be more powerful than histopathology as a predictive factor for the different treatment strategies. This study gives an insight to overall prognosis, role of molecular markers, various molecular subtypes and better categorization of triple negative breast cancer cases.This was a cross sectional study conducted in the department of pathology over a period of 2 years.In this study, 500 cases of breast carcinoma were included. Molecular phenotype was determined using expression of estrogen receptor, progesterone receptor, HER2/neu, Ki67, epithelial growth factor receptor (EGFR), and cytokeratin 5/6. Correlation of molecular subtypes with histological grade, prognostic and predictive factors done by using chi square test. Using software Medcalc version 12.2.1.0.Of the 500 cases, maximum number of cases 38.20% were luminal A. Most common histological subtype in all category were Infiltrating duct carcinoma (Not Otherwise Specified) {IDC (NOS)} with higher grade of IDC (NOS) in her2neu and basal type. Unclassified category includes both low grade tumors and high grade tumors. Statistically significant association of molecular subtype was found with histological subtype, tumor necrosis, lymphocytic response and lymphovascular invasion. (P ≤ 0.001) Molecular classification would result in less frequent use of chemotherapy in breast carcinoma and have its considerable advantages in reducing toxicity and costs.In view of increasing awareness, younger age of presentation and occasional family history various screening program should be popularized.

Study of 5-Fluorouracil Loaded Chitosan Nanoparticles for Treatment of Skin Cancer.

The purpose of this study was to formulate, characterize and in-vitro cytotoxicity of 5-Fluorouracil loaded controlled release nanoparticles for the treatment of skin cancer. The patents on nanoparticles (US8414926B1), (US61654404A), (WO2007150075A3) etc. helped in the selection polymers and method for the preparation of nanoparticles. In the present study nanoparticles were prepared by simple ionic gelation method using various concentrations of chitosan and sodium tripolyphosphate (TPP). Several process and formulation parameters were screened and optimized using 25-2 fractional factorial design. The prepared nanoparticles were evaluated for particle size, shape, charge, entrapment efficiency, crosslinking mechanism and drug release study. The optimized 5-Fluorouracil loaded nanoparticle were found with particle size of of 320±2.1 nm, entrapment efficiency of 85.12%± 1.1% and Zeta potential of 29mv±1mv. Scanning electron microscopy and dynamic light scattering technique revealed spherical particles with uniform size. The invitro release profile showed controlled release up to 24 hr. Further study was carried using A375 basal cell carcinoma cell-line to elucidate the mechanism of its cytotoxicity by MTT assay. These results demonstrate that the possibility of delivering 5-Fluorouracil to skin with enhanced encapsulation efficiency indicating effectiveness of the formulation for treatment of basal cell carcinoma type of skin cancer.

Efficacy of neoadjuvant atezolizumab treatment in patients with advanced urothelial bladder cancer according to the BASQ classification: a study protocol for an open-label, two-cohort, phase II trial

IntroductionAtezolizumab is a programmed death ligand-1 inhibitor for urothelial bladder cancer treatment. Atezolizumab has become the standard therapy for patients with urothelial bladder cancer who are not responding to cisplatin-based...

Cholesterol Pathway Inhibition Induces TGF-β Signaling to Promote Basal Differentiation in Pancreatic Cancer

Oncogenic transformation alters lipid metabolism to sustain tumor growth. We define a mechanism by which cholesterol metabolism controls the development and differentiation of pancreatic ductal adenocarcinoma (PDAC). Disruption of distal cholesterol biosynthesis by conditional inactivation of the rate-limiting enzyme Nsdhl or treatment with cholesterol-lowering statins switches glandular pancreatic carcinomas to a basal (mesenchymal) phenotype in mouse models driven by KrasG12D expression and homozygous Trp53 loss. Consistently, PDACs in patients receiving statins show enhanced mesenchymal features. Mechanistically, statins and NSDHL loss induce SREBP1 activation, which promotes the expression of Tgfb1, enabling epithelial-mesenchymal transition. Evidence from patient samples in this study suggests that activation of transforming growth factor β signaling and epithelial-mesenchymal transition by cholesterol-lowering statins may promote the basal type of PDAC, conferring poor outcomes in patients.

Letter by Finsterer Regarding Article, “COVID-19-Associated Stress (Takotsubo) Cardiomyopathy”

HomeCirculation: Cardiovascular ImagingVol. 13, No. 10Letter by Finsterer Regarding Article, “COVID-19-Associated Stress (Takotsubo) Cardiomyopathy” Free AccessLetterPDF/EPUBAboutView PDFView EPUBSections ToolsAdd to favoritesDownload citationsTrack citationsPermissions ShareShare onFacebookTwitterLinked InMendeleyReddit Jump toFree AccessLetterPDF/EPUBLetter by Finsterer Regarding Article, “COVID-19-Associated Stress (Takotsubo) Cardiomyopathy” Josef Finsterer, MD, PhD Josef FinstererJosef Finsterer https://orcid.org/0000-0003-2839-7305 Klinikum Landstrasse, Messerli Institute, Vienna, Austria. Search for more papers by this author Originally published23 Sep 2020https://doi.org/10.1161/CIRCIMAGING.120.011577Circulation: Cardiovascular Imaging. 2020;13:e011577To the Editor:With interest we read the article by Tsao et al1 about a 59-year-old obese but otherwise healthy woman who experienced a severe infection with severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) manifesting as coronavirus disease 2019 (COVID-19) and complicated by the development of Takotsubo syndrome (TTS), also known as stress cardiomyopathy or broken-heart syndrome. The patient recovered from TTS despite the severe COVID-19 infection.1 We have the following comments and concerns.We do not agree with the notion that “TTS associated with COVID-19 has not been well described.”1 A literature search brought to light that at least 37 cases with TTS in the setting of an acute SARS-CoV-2 infection, including the index case, have been reported. The first patient with TTS triggered by a SARS-CoV-2 infection was an 83-year-old Swiss woman described by Meyer et al.2 She had a history of arterial hypertension and developed acute chest pain together with dry cough 3 days before admission.2 Transthoracic echocardiography revealed apical ballooning with hyperkinetic basal segments.2 On hospital day 3, she was tested positive for SARS-CoV-2. She recovered completely within 10 days upon conventional heart failure therapy.2 The most recently presented case was published by Sattar et al.3In the vast majority of the patients with SARS-CoV-2 associated TTS, TTS developed after onset of the SARS-CoV-2 infection. Age ranged between 39 and 87 years. The distribution between genders was almost equal, and the majority of patients presented with the classical type of TTS. Only 2 patients each presented with the midventricular, respectively, basal type of TTS. Treatment for TTS was quite variable, but most patients received β-blockers, diuretics, heparin, or acetyl-salicylic acid. Ten patients recovered; 3 patients died; and in 20 patients, the outcome was not reported.Since the infection with SARS-CoV-2 may go along with myocarditis4 and myocarditis may be misinterpreted as TTS,4 we should know how myocarditis was ruled in the index patient. Particularly, we should know whether the patient underwent cardiac magnetic resonance imaging with contrast medium or endomyocardial biopsy to see whether there was myocardial inflammation or not. Myocarditis on cardiac magnetic resonance imaging may show up as edema (pseudohypertrophy)4 or enhancement with contrast medium.4 Acute myocarditis on endomyocardial biopsy may show up as diffuse infiltration of the myocardium with T lymphocytes in the absence of any scars.4 Myocarditis in SARS-CoV-2–infected patients is usually virus negative but characterized by T-cell infiltration.4Interestingly, the prevalence of TTS generally increased during the COVID-19 crisis.5 The median length of hospitalization of these patients was longer than that during the non-COVID period.5 However, mortality and rehospitalization rate were the same as that before the crisis.5In summary, TTS is a well-recognized complication of COVID-19. Clinical presentation, epidemiology, treatment, and outcome of SARS-CoV-2–associated TTS are similar to COVID-19–unrelated TTS. Workup for TTS in patients with COVID-19 is indicated if they develop symptoms and signs of a myocardial infarction or heart failure in addition to the pulmonary compromise.AcknowledgmentsDr Finsterer contributed to the design, literature search, discussion, first draft, and critical comments.DisclosuresNone.FootnotesThe opinions expressed in this article are not necessarily those of the editors or of the American Heart Association.

Abstract 1424: Her3 up regulation in response to AKT and PI3K inhibition causes differential responses dependent on the presence of neuregulin isoforms in the microenvironmental milieu

Abstract Introduction: Triple Negative Breast Cancer (TNBC) is an aggressive form of Breast Cancer (BC) affecting ~20% of BC patients. TNBC is a highly heterogenous disease and numerous tyrosine kinase inhibitors (TKI) targeting different pathways including the PI3K, AKT and MAPK pathway have shown limited benefit in the clinical setting. Intrinsic and extrinsic resistance mechanisms to TKI are the main reason of therapeutic failure. Signalling from the human epidermal growth factor receptor (HER) family of proteins is dependent on a well-orchestrated series of interactions between family members to form either homo- or heterodimers. The heterodimeric complex formed with HER3 is a particularly potent oncogenic signalling unit that can act as a driver of cancer growth. HER3 upregulation in response to small molecule inhibitors has been shown to rescue a large number of cancers including TNBC. In this study we aim to characterize the extent of HER3 reliance in TNBC and to define the effect of neuregulin isoforms in TNBC cancers. Method and Materials: Basal and Claudin type Breast cancer cell lines were treated with a range of small molecule inhibitors, known to exhibit limited efficacy as single agents, including the AKT inhibitor (GDC0068), the PI3K inhibitor (GDC0077) and the EGFR inhibitors (Gefitinib and Lapatinib) or a combination of these drugs in the presence or absence of the HER3 ligand neuregulin. The effect these inhibitors on Akt/ MAPK signalling was characterized using specific biomarkers of the respective pathways. Combination therapy was evaluated in human cancer cell lines through Annexin V staining. Results: Basal (HCC-70 and MDA-MB-468) and Caudin type (MDA-MB-231) TNBC cell lines displayed differential reliance on the HER family of receptors. Dynamic HER3 upregulation was predominant in the TNBC cells that show HER signaling reliance coinciding with the basal subtype. Furthermore, the presence of the natural ligand neuregulin showed potent signaling through the HER3-AKT pathway and significantly diminished the efficacy of the inhibitors tested. We report that neuregulin augments the HER3 feedback mechanism for continued proliferation in TNBC. Interestingly, we show that neuregulin mediated resistance is dependent on the type and dose of the neuregulin isoform. Neuregulin 1β showed potent proliferative effects and overcoming this effect required a 10 fold increase in the drug concentration. In contrast, the presence of neuregulin 1α isoform may paradoxically confer sensitivity to small molecule inhibitors. We show that cellular proliferation is maintained through both the PI3K and MAPK pathway and single targeting is inefficient in overcoming resistance. We demonstrate that combination strategies that block compensatory mechanisms are effective strategies to neuregulin dependent and independent resistance mechanisms. Conclusion: The highly dynamic nature of HER3 expression and signalling combined with redundant HER signalling and downstream mechanisms allow for continued signalling in a subtype of TNBC patients. Targeted therapy to ensure complete abrogation of HER signalling is essential for effective therapeutic response. In conclusion, TNBC cancers display differential mechanism of resistance that should be adequately addressed for therapeutic strategies. Citation Format: Nicoleta Sinevici, Pedram Heidari, Umar Mahmood. Her3 up regulation in response to AKT and PI3K inhibition causes differential responses dependent on the presence of neuregulin isoforms in the microenvironmental milieu [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 1424.

Abstract 1714A: TGF-beta signaling on myeloid cells regulates ECM deposition in mammary carcinoma via adenosine dependent mechanisms

Abstract TGF-beta plays a crucial role in tumor microenvironment by regulating cell-cell and cell-stroma interactions. We previously demonstrated that TGF-beta signaling on myeloid cells regulates expression of CD73, a key enzyme for production of adenosine, a pro-tumorigenic metabolite implicated in regulation of tumor cell behaviors, immune response and angiogenesis. Using MMTV-PyMT mouse mammary tumor model, we discovered that deletion of TGF-beta signaling on myeloid cells (PyMT/TGFbRII-LysM) affects ECM formation in tumor tissue, specifically increases collagen and decreases fibronectin deposition, and these changes associate with mitigated tumor growth and reduced metastases. Using multiplexed five-color immunofluorescent staining and spatial analysis on a single-cell level, we discovered that reduced TGF-beta signaling on fibroblasts associates with their proximity to CD73 positive myeloid cells in tumor tissue. Consistent with these findings, in vitro gel contraction assay and Western blotting for Collagen I and pSMAD proteins confirmed that adenosine significantly downregulates TGF-beta signaling on fibroblasts. Using in vitro pharmacological approach, we found that this effect is regulated by A2a and A2b adenosine receptors. TCGA data base analysis revealed that patients with triple negative breast cancer and basal type have similar signature of adenosine and ECM profiles: high expression of A2b adenosine receptors correlates with decreased expression of Col1 and is associates with poor outcome. Taken together, our studies reveal a new role for TGF-beta signaling on myeloid cells in tumorigenesis. Discovered crosstalk between TGF-beta/CD73 on myeloid cells and TGF-beta signaling on fibroblasts can contribute to ECM remodeling and pro-tumorigenic actions of CAFs. Citation Format: Georgii Vasiukov, Tatiana Novitskaya, Samantha Schwager, Harold Moses, Timothy Blackwell, Igor Feoktistov, Sergey Novitskiy. TGF-beta signaling on myeloid cells regulates ECM deposition in mammary carcinoma via adenosine dependent mechanisms [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 1714A.