Sulindac sulfide as a non-immune suppressive γ-secretase modulator to target triple-negative breast cancer.

Introduction: Triple-negative breast cancer (TNBC) comprises a heterogeneous group of clinically aggressive tumors with high risk of recurrence and metastasis. Current pharmacological treatment options remain largely limited to chemotherapy. Despite promising results, the efficacy of immunotherapy and chemo-immunotherapy in TNBC remains limited. There is strong evidence supporting the involvement of Notch signaling in TNBC progression. Expression of Notch1 and its ligand Jagged1 correlate with poor prognosis. Notch inhibitors, including g-secretase inhibitors (GSIs), are quite effective in preclinical models of TNBC. However, the success of GSIs in clinical trials has been limited by their intestinal toxicity and potential for adverse immunological effects, since Notch plays key roles in T-cell activation, including CD8 T-cells in tumors. Our overarching goal is to replace GSIs with agents that lack their systemic toxicity and ideally, do not affect tumor immunity. We identified sulindac sulfide (SS), the active metabolite of FDA-approved NSAID sulindac, as a potential candidate to replace GSIs. Methods: We investigated the pharmacological and immunotherapeutic properties of SS in TNBC models in vitro, ex-vivo and in vivo. Results: We confirmed that SS, a known γ-secretase modulator (GSM), inhibits Notch1 cleavage in TNBC cells. SS significantly inhibited mammosphere growth in all human and murine TNBC models tested. In a transplantable mouse TNBC tumor model (C0321), SS had remarkable single-agent anti-tumor activity and eliminated Notch1 protein expression in tumors. Importantly, SS did not inhibit Notch cleavage in T- cells, and the anti-tumor effects of SS were significantly enhanced when combined with a-PD1 immunotherapy in our TNBC organoids and in vivo. Discussion: Our data support further investigation of SS for the treatment of TNBC, in conjunction with chemo- or -chemo-immunotherapy. Repurposing an FDA-approved, safe agent for the treatment of TNBC may be a cost-effective, rapidly deployable therapeutic option for a patient population in need of more effective therapies.

The Efficacy of CB-103, a First-in-Class Transcriptional Notch Inhibitor, in Preclinical Models of Breast Cancer.

BACKGROUND: The efficacy of CB-103 was evaluated in preclinical models of both ER+ and TNBC. Furthermore, the therapeutic efficacy of combining CB-103 with fulvestrant in ER+ BC and paclitaxel in TNBC was determined. METHODS: CB-103 was screened in combination with a panel of anti-neoplastic drugs. We evaluated the anti-tumor activity of CB-103 with fulvestrant in ESR1-mutant (Y537S), endocrine-resistant BC xenografts. In the same model, we examined anti-CSC activity in mammosphere formation assays for CB-103 alone or in combination with fulvestrant or palbociclib. We also evaluated the effect of CB-103 plus paclitaxel on primary tumors and CSC in a GSI-resistant TNBC model HCC1187. Comparisons between groups were performed with a two-sided unpaired Students' t-test. A one-way or two-way ANOVA followed by Tukey's post-analysis was performed to analyze the in vivo efficacy study results. THE RESULTS: CB-103 showed synergism with fulvestrant in ER+ cells and paclitaxel in TNBC cells. CB-103 combined with fulvestrant or paclitaxel potently inhibited mammosphere formation in both models. Combination of CB-103 and fulvestrant significantly reduced tumor volume in an ESR1-mutant, the endocrine-resistant BC model. In a GSI-resistant TNBC model, CB-103 plus paclitaxel significantly delayed tumor growth compared to paclitaxel alone. CONCLUSION: our data indicate that CB-103 is an attractive candidate for clinical investigation in endocrine-resistant, recurrent breast cancers with biomarker-confirmed Notch activity in combination with SERDs and/or CDKis and in TNBCs with biomarker-confirmed Notch activity in combination with taxane-containing chemotherapy regimens.

H2S Prodrug, SG-1002, Protects against Myocardial Oxidative Damage and Hypertrophy In Vitro via Induction of Cystathionine ß-Synthase and Antioxidant Proteins.

Endogenously produced hydrogen sulfide (H2S) is critical for cardiovascular homeostasis. Therapeutic strategies aimed at increasing H2S levels have proven cardioprotective in models of acute myocardial infarction (MI) and heart failure (HF). The present study was undertaken to investigate the effects of a novel H2S prodrug, SG-1002, on stress induced hypertrophic signaling in murine HL-1 cardiac muscle cells. Treatment of HL-1 cells with SG-1002 under serum starvation without or with H2O2 increased the levels of H2S, H2S producing enzyme, and cystathionine ß-synthase (CBS), as well as antioxidant protein levels, such as super oxide dismutase1 (SOD1) and catalase, and additionally decreased oxidative stress. SG-1002 also decreased the expression of hypertrophic/HF protein markers such as atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), galectin-3, TIMP1, collagen type III, and TGF-ß1 in stressed HL-1 cells. Treatment with SG-1002 caused a significant induction of cell viability and a marked reduction of cellular cytotoxicity in HL-1 cells under serum starvation incubated without or with H2O2. Experimental results of this study suggest that SG-1002 attenuates myocardial cellular oxidative damage and/or hypertrophic signaling via increasing H2S levels or H2S producing enzymes, CBS, and antioxidant proteins.

Targeting the Cbl-b-Notch1 axis as a novel immunotherapeutic strategy to boost CD8+ T-cell responses.

A critical feature of cancer is the ability to induce immunosuppression and evade immune responses. Tumor-induced immunosuppression diminishes the effectiveness of endogenous immune responses and decreases the efficacy of cancer immunotherapy. In this study, we describe a new immunosuppressive pathway in which adenosine promotes Casitas B-lineage lymphoma b (Cbl-b)-mediated Notch1 degradation, causing suppression of CD8+ T-cells effector functions. Genetic knockout and pharmacological inhibition of Cbl-b prevents Notch1 degradation in response to adenosine and reactivates its signaling. Reactivation of Notch1 results in enhanced CD8+ T-cell effector functions, anti-cancer response and resistance to immunosuppression. Our work provides evidence that targeting the Cbl-b-Notch1 axis is a novel promising strategy for cancer immunotherapy.

Association of Obesity and Diabetes With the Incidence of Breast Cancer in Louisiana.

INTRODUCTION: Breast cancer is a heterogeneous disease, consisting of multiple molecular subtypes. Obesity has been associated with an increased risk for postmenopausal breast cancer, but few studies have examined breast cancer subtypes separately. Obesity is often complicated by type 2 diabetes, but the possible association of diabetes with specific breast cancer subtypes remains poorly understood. METHODS: In this retrospective case-control study, Louisiana Tumor Registry records of primary invasive breast cancer diagnosed in 2010-2015 were linked to electronic health records in the Louisiana Public Health Institute's Research Action for Health Network. Controls were selected from Research Action for Health Network and matched to cases by age and race. Conditional logistic regression was used to identify metabolic risk factors. Data analysis was conducted in 2020‒2021. RESULTS: There was a significant association between diabetes and breast cancer for Luminal A, Triple-Negative Breast Cancer, and human epidermal growth factor 2‒positive subtypes. In multiple logistic regression, including both obesity status and diabetes as independent risk factors, Luminal A breast cancer was also associated with overweight status. Diabetes was associated with increased risk for Luminal A and Triple-Negative Breast Cancer in subgroup analyses, including women aged ≥50 years, Black women, and White women. CONCLUSIONS: Although research has identified obesity and diabetes as risk factors for breast cancer, these results underscore that comorbid risk is complex and may differ by molecular subtype. There was a significant association between diabetes and the incidence of Luminal A, Triple-Negative Breast Cancer, and human epidermal growth factor 2‒positive breast cancer in Louisiana.

Obesity Modulates the Gut Microbiome in Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is an aggressive, molecularly heterogeneous subtype of breast cancer. Obesity is associated with increased incidence and worse prognosis in TNBC through various potential mechanisms. Recent evidence suggests that the gut microbiome plays a central role in the progression of cancer, and that imbalances or dysbiosis in the population of commensal microbiota can lead to inflammation and contribute to tumor progression. Obesity is characterized by low-grade inflammation, and gut dysbiosis is associated with obesity, chronic inflammation, and failure of cancer immunotherapy. However, the debate on what constitutes a "healthy" gut microbiome is ongoing, and the connection among the gut microbiome, obesity, and TNBC has not yet been addressed. This study aims to characterize the role of obesity in modulating the gut microbiome in a syngeneic mouse model of TNBC. 16S rRNA sequencing and metagenomic analyses were performed to analyze and annotate genus and taxonomic profiles. Our results suggest that obesity decreases alpha diversity in the gut microbiome. Metagenomic analysis revealed that obesity was the only significant factor explaining the similarity of the bacterial communities according to their taxonomic profiles. In contrast to the analysis of taxonomic profiles, the analysis of variation of functional profiles suggested that obesity status, tumor presence, and the obesity-tumor interaction were significant in explaining the variation of profiles, with obesity having the strongest correlation. The presence of tumor modified the profiles to a greater extent in obese than in lean animals. Further research is warranted to understand the impact of the gut microbiome on TNBC progression and immunotherapy.

Precision Medicine and Triple-Negative Breast Cancer: Current Landscape and Future Directions.

Triple-negative breast cancer (TNBC) is an aggressive and heterogeneous subtype of breast cancer associated with a high recurrence and metastasis rate that affects African-American women disproportionately. The recent approval of targeted therapies for small subgroups of TNBC patients by the US 'Food and Drug Administration' is a promising development. The advancement of next-generation sequencing, particularly somatic exome panels, has raised hopes for more individualized treatment plans. However, the use of precision medicine for TNBC is a work in progress. This review will discuss the potential benefits and challenges of precision medicine for TNBC. A recent clinical trial designed to target TNBC patients based on their subtype-specific classification shows promise. Yet, tumor heterogeneity and sub-clonal evolution in primary and metastatic TNBC remain a challenge for oncologists to design adaptive precision medicine-based treatment plans.

ERK5 Is Required for Tumor Growth and Maintenance Through Regulation of the Extracellular Matrix in Triple Negative Breast Cancer.

Conventional mitogen-activated protein kinase (MAPK) family members regulate diverse cellular processes involved in tumor initiation and progression, yet the role of ERK5 in cancer biology is not fully understood. Triple-negative breast cancer (TNBC) presents a clinical challenge due to the aggressive nature of the disease and a lack of targeted therapies. ERK5 signaling contributes to drug resistance and metastatic progression through distinct mechanisms, including activation of epithelial-to-mesenchymal transition (EMT). More recently a role for ERK5 in regulation of the extracellular matrix (ECM) has been proposed, and here we investigated the necessity of ERK5 in TNBC tumor formation. Depletion of ERK5 expression using the CRISPR/Cas9 system in MDA-MB-231 and Hs-578T cells resulted in loss of mesenchymal features, as observed through gene expression profile and cell morphology, and suppressed TNBC cell migration. In vivo xenograft experiments revealed ERK5 knockout disrupted tumor growth kinetics, which was restored using high concentration Matrigel™ and ERK5-ko reduced expression of the angiogenesis marker CD31. These findings implicated a role for ERK5 in the extracellular matrix (ECM) and matrix integrity. RNA-sequencing analyses demonstrated downregulation of matrix-associated genes, integrins, and pro-angiogenic factors in ERK5-ko cells. Tissue decellularization combined with cryo-SEM and interrogation of biomechanical properties revealed that ERK5-ko resulted in loss of key ECM fiber alignment and mechanosensing capabilities in breast cancer xenografts compared to parental wild-type cells. In this study, we identified a novel role for ERK5 in tumor growth kinetics through modulation of the ECM and angiogenesis axis in breast cancer.

Obesity-Altered Adipose Stem Cells Promote Radiation Resistance of Estrogen Receptor Positive Breast Cancer through Paracrine Signaling.

Obesity is associated with poorer responses to chemo- and radiation therapy for breast cancer, which leads to higher mortality rates for obese women who develop breast cancer. Adipose stem cells (ASCs) are an integral stromal component of the tumor microenvironment (TME). In this study, the effects of obesity-altered ASCs (obASCs) on estrogen receptor positive breast cancer cell's (ER+BCCs) response to radiotherapy (RT) were evaluated. We determined that BCCs had a decreased apoptotic index and increased surviving fraction following RT when co-cultured with obASCs compared to lnASCs or non-co-cultured cells. Further, obASCs reduced oxidative stress and induced IL-6 expression in co-cultured BCCs after radiation. obASCs produce increased levels of leptin relative to ASCs from normal-weight individuals (lnASCs). obASCs upregulate the expression of IL-6 compared to non-co-cultured BCCs, but BCCs co-cultured with leptin knockdown obASCs did not upregulate IL-6. The impact of shLeptin obASCs on radiation resistance of ER+BCCs demonstrate a decreased radioprotective ability compared to shControl obASCs. Key NOTCH signaling players were enhanced in ER+BBCs following co-culture with shCtrl obASCs but not shLep obASCs. This work demonstrates that obesity-altered ASCs, via enhanced secretion of leptin, promote IL-6 and NOTCH signaling pathways in ER+BCCs leading to radiation resistance.

Racial Disparities in Triple Negative Breast Cancer: A Review of the Role of Biologic and Non-biologic Factors.

Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer that lacks expression of the estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor (HER2). TNBC constitutes about 15-30 percent of all diagnosed invasive breast cancer cases in the United States. African-American (AA) women have high prevalence of TNBC with worse clinical outcomes than European-American (EA) women. The contributing factors underlying racial disparities have been divided into two major categories based on whether they are related to lifestyle (non-biologic) or unrelated to lifestyle (biologic). Our objective in the present review article was to understand the potential interactions by which these risk factors intersect to drive the initiation and development of the disparities resulting in the aggressive TNBC subtypes in AA women more likely than in EA women. To reach our goal, we conducted literature searches using MEDLINE/PubMed to identify relevant articles published from 2005 to 2019 addressing breast cancer disparities primarily among AA and EA women in the United States. We found that disparities in TNBC may be attributed to racial differences in biological factors, such as tumor heterogeneity, population genetics, somatic genomic mutations, and increased expression of genes in AA breast tumors which have direct link to breast cancer. In addition, a large number of non-biologic factors, including socioeconomic deprivation adversities associated with poverty, social stress, unsafe neighborhoods, lack of healthcare access and pattern of reproductive factors, can promote comorbid diseases such as obesity and diabetes which may adversely contribute to the aggression of TNBC biology in AA women. Further, the biological risk factors directly linked to TNBC in AA women may potentially interact with non-biologic factors to promote a higher prevalence of TNBC, more aggressive biology, and poor survival. The relative contributions of the biologic and non-biologic factors and their potential interactions is essential to our understanding of disproportionately high burden and poor survival rates of AA women with TNBC.

Leptin produced by obesity-altered adipose stem cells promotes metastasis but not tumorigenesis of triple-negative breast cancer in orthotopic xenograft and patient-derived xenograft models.

BACKGROUND: Breast cancer is the second leading cause of cancer deaths in the USA. Triple-negative breast cancer (TNBC) is a clinically aggressive subtype of breast cancer with high rates of metastasis, tumor recurrence, and resistance to therapeutics. Obesity, defined by a high body mass index (BMI), is an established risk factor for breast cancer. Women with a high BMI have increased incidence and mortality of breast cancer; however, the mechanisms(s) by which obesity promotes tumor progression are not well understood. METHODS: In this study, obesity-altered adipose stem cells (obASCs) were used to evaluate obesity-mediated effects of TNBC. Both in vitro and in vivo analyses of TNBC cell lines were co-cultured with six pooled donors of obASCs (BMI > 30) or ASCs isolated from lean women (lnASCs) (BMI < 25). RESULTS: We found that obASCs promote a pro-metastatic phenotype by upregulating genes associated with epithelial-to-mesenchymal transition and promoting migration in vitro. We confirmed our findings using a TNBC patient-derived xenograft (PDX) model. PDX tumors grown in the presence of obASCS in SCID/beige mice had increased circulating HLA1+ human cells as well as increased numbers of CD44+CD24- cancer stem cells in the peripheral blood. Exposure of the TNBC PDX to obASCs also increased the formation of metastases. The knockdown of leptin expression in obASCs suppressed the pro-metastatic effects of obASCs. CONCLUSIONS: Leptin signaling is a potential mechanism through which obASCs promote metastasis of TNBC in both in vitro and in vivo analyses.

Gene shaving using a sensitivity analysis of kernel based machine learning approach, with applications to cancer data.

BACKGROUND: Gene shaving (GS) is an essential and challenging tools for biomedical researchers due to the large number of genes in human genome and the complex nature of biological networks. Most GS methods are not applicable to non-linear and multi-view data sets. While the kernel based methods can overcome these problems, a well-founded positive definite kernel based GS method has yet to be proposed for biomedical data analysis. METHODS AND FINDINGS: Since the kernel based methods on genomic information can improve the prediction of diseases, here we proposed a noble method, "kernel based gene shaving" which is based on the influence function of kernel canonical correlation analysis. To investigate the performance of the proposed method in comparison to state-of-the-art-method in gene saving, we analyzed extensive simulated and real microarray gene expression data set. The performance metrics including true positive rate, true negative rate, false positive rate, false negative rate, misclassification error rate, the false discovery rate and area under curves were computed for each methods. In colon cancer data analysis, the proposed method identified a significant subsets of 210 genes out of 2000 genes and suggestive superior performance compared with other methods. The proposed method can be applied to the study of other disease process where two view data is a common task. CONCLUSIONS: We addressed the challenge of finding unique kernel based GS methods by using the influence function of kernel canonical correlation analysis. The proposed method has shown to have better performance than state-of-the-art-methods in gene saving and has identified many more significant gene interactions, suggesting that genes function in a concerted effort in colon cancer. In similar biomedical data analysis, kernel based methods could be applied to select a potential subset of genes. The positive definite kernel based methods can overcome the non-linearity problem and improve the prediction process.

Corrigendum: Adenosine A2A Receptor Stimulation Inhibits TCR-Induced Notch1 Activation in CD8+T-Cells.

[This corrects the article DOI: 10.3389/fimmu.2019.00162.].

Neighborhood Social Determinants of Triple Negative Breast Cancer.

Triple Negative Breast Cancer (TNBC) is an aggressive, heterogeneous subtype of breast cancer, which is more frequently diagnosed in African American (AA) women than in European American (EA) women. The purpose of this study is to investigate the role of social determinants in racial disparities in TNBC. Data on Louisiana TNBC patients diagnosed in 2010-2012 were collected and geocoded to census tract of residence at diagnosis by the Louisiana Tumor Registry. Using multilevel statistical models, we analyzed the role of neighborhood concentrated disadvantage index (CDI), a robust measure of physical and social environment, in racial disparities in TNBC incidence, stage at diagnosis, and stage-specific survival for the study population. Controlling for age, we found that AA women had a 2.21 times the incidence of TNBC incidence compared to EA women. Interestingly, the incidence of TNBC was independent of neighborhood CDI and adjusting for neighborhood environment did not impact the observed racial disparity. AA women were more likely to be diagnosed at later stages and CDI was associated with more advanced stages of TNBC at diagnosis. CDI was also significantly associated with poorer stage-specific survival. Overall, our results suggest that neighborhood disadvantage contributes to racial disparities in stage at diagnosis and survival among TNBC patients, but not to disparities in incidence of the disease. Further research is needed to determine the mechanisms through which social determinants affect the promotion and progression of this disease and guide efforts to improve overall survival.

Adenosine A2A Receptor Stimulation Inhibits TCR-Induced Notch1 Activation in CD8+T-Cells.

Notch receptors signaling is required for optimal T-cell activation and function. T-cell receptor (TCR) engagement can activate Notch receptors in T-cells in a ligand-independent fashion. In this study, we examined the role of adenosine A2A receptor (A2AR) signaling pathway in regulating the activity of Notch1 induced by TCR stimulation in CD8+T-cells. A selective A2AR agonist decreased Notch1 protein expression and Notch1 cleavage, and reduced transcripts of Notch1-target genes Hes1 and Myc in activated CD8+T-cells. Inhibition of TCR-induced Notch1 expression by an A2AR agonist was accompanied by increased cAMP concentration and mimicked by forskolin. This effect was associated with reduced IFN-γ and granzyme B production. The effect of an A2AR agonist was abrogated by a selective A2AR antagonist and absent in CD8+T-cells harvested from A2AR-/- mice. Stimulation of A2AR reduced Notch1 receptor levels by inhibiting upstream TCR signals, including ZAP70 phosphorylation, in turn impairing the generation of the active Notch1 intracellular domain (N1ICD). Direct activation of PKC with PMA and ionomycin bypassed A2AR-induced Notch1 inhibition. Overexpression of N1ICD in CD8+T-cells prevented the suppressive effects of an A2AR agonist on proliferation and cytokine release during activation. Our results identify the A2AR signaling pathway as an important regulator of TCR-induced Notch1 receptor activation in CD8+T-cells, and Notch as an important target of the immune suppressive effects of A2AR. We propose a mechanism whereby A2AR impairs CD8 T-cells function through inhibition of Notch1 receptor activation.

Notch Signaling Regulates Mitochondrial Metabolism and NF-κB Activity in Triple-Negative Breast Cancer Cells via IKKα-Dependent Non-canonical Pathways.

Triple negative breast cancer (TNBC) patients have high risk of recurrence and metastasis, and current treatment options remain limited. Cancer stem-like cells (CSCs) have been linked to cancer initiation, progression and chemotherapy resistance. Notch signaling is a key pathway regulating TNBC CSC survival. Treatment of TNBC with PI3K or mTORC1/2 inhibitors results in drug-resistant, Notch-dependent CSC. However, downstream mechanisms and potentially druggable Notch effectors in TNBC CSCs are largely unknown. We studied the role of the AKT pathway and mitochondrial metabolism downstream of Notch signaling in TNBC CSC from cell lines representative of different TNBC molecular subtypes as well as a novel patient-derived model. We demonstrate that exposure of TNBC cells to recombinant Notch ligand Jagged1 leads to rapid AKT phosphorylation in a Notch1-dependent but RBP-Jκ independent fashion. This requires mTOR and IKKα. Jagged1 also stimulates mitochondrial respiration and fermentation in an AKT- and IKK-dependent fashion. Notch1 co-localizes with mitochondria in TNBC cells. Pharmacological inhibition of Notch cleavage by gamma secretase inhibitor PF-03084014 in combination with AKT inhibitor MK-2206 or IKK-targeted NF-κB inhibitor Bay11-7082 blocks secondary mammosphere formation from sorted CD90hi or CD44+CD24low (CSCs) cells. A TNBC patient-derived model gave comparable results. Besides mitochondrial oxidative metabolism, Jagged1 also triggers nuclear, NF-κB-dependent transcription of anti-apoptotic gene cIAP-2. This requires recruitment of Notch1, IKKα and NF-κB to the cIAP-2 promoter. Our observations support a model where Jagged1 triggers IKKα-dependent, mitochondrial and nuclear Notch1 signals that stimulate AKT phosphorylation, oxidative metabolism and transcription of survival genes in PTEN wild-type TNBC cells. These data suggest that combination treatments targeting the intersection of the Notch, AKT and NF-κB pathways have potential therapeutic applications against CSCs in TNBC cases with Notch1 and wild-type PTEN expression.

Notch Signaling in Myeloid Cells as a Regulator of Tumor Immune Responses.

Cancer immunotherapy, which stimulates or augments host immune responses to treat malignancies, is the latest development in the rapidly advancing field of cancer immunology. The basic principles of immunotherapies are either to enhance the functions of specific components of the immune system or to neutralize immune-suppressive signals produced by cancer cells or tumor microenvironment cells. When successful, these approaches translate into long-term survival for patients. However, durable responses are only seen in a subset of patients and so far, only in some cancer types. As for other cancer treatments, resistance to immunotherapy can also develop. Numerous research groups are trying to understand why immunotherapy is effective in some patients but not others and to develop strategies to enhance the effectiveness of immunotherapy. The Notch signaling pathway is involved in many aspects of tumor biology, from angiogenesis to cancer stem cell maintenance to tumor immunity. The role of Notch in the development and modulation of the immune response is complex, involving an intricate crosstalk between antigen-presenting cells, T-cell subpopulations, cancer cells, and other components of the tumor microenvironment. Elegant studies have shown that Notch is a central mediator of tumor-induced T-cell anergy and that activation of Notch1 in CD8 T-cells enhances cancer immunotherapy. Tumor-infiltrating myeloid cells, including myeloid-derived suppressor cells, altered dendritic cells, and tumor-associated macrophages along with regulatory T cells, are major obstacles to the development of successful cancer immunotherapies. In this article, we focus on the roles of Notch signaling in modulating tumor-infiltrating myeloid cells and discuss implications for therapeutic strategies that modulate Notch signaling to enhance cancer immunotherapy.

A novel patient-derived xenograft model for claudin-low triple-negative breast cancer.

BACKGROUND: Triple-negative breast cancer (TNBC) subtypes are clinically aggressive and cannot be treated with targeted therapeutics commonly used in other breast cancer subtypes. The claudin-low (CL) molecular subtype of TNBC has high rates of metastases, chemoresistance and recurrence. There exists an urgent need to identify novel therapeutic targets in TNBC; however, existing models utilized in target discovery research are limited. Patient-derived xenograft (PDX) models have emerged as superior models for target discovery experiments because they recapitulate features of patient tumors that are limited by cell-line derived xenograft methods. METHODS: We utilize immunohistochemistry, qRT-PCR and Western Blot to visualize tumor architecture, cellular composition, genomic and protein expressions of a new CL-TNBC PDX model (TU-BcX-2O0). We utilize tissue decellularization techniques to examine extracellular matrix composition of TU-BcX-2O0. RESULTS: Our laboratory successfully established a TNBC PDX tumor, TU-BCX-2O0, which represents a CL-TNBC subtype and maintains this phenotype throughout subsequent passaging. We dissected TU-BCx-2O0 to examine aspects of this complex tumor that can be targeted by developing therapeutics, including the whole and intact breast tumor, specific cell populations within the tumor, and the extracellular matrix. CONCLUSIONS: Here, we characterize a claudin-low TNBC patient-derived xenograft model that can be utilized for therapeutic research studies.

Exogenous lipid uptake induces metabolic and functional reprogramming of tumor-associated myeloid-derived suppressor cells.

Myeloid-derived suppressor cells (MDSC) promote tumor growth by blocking anti-tumor T cell responses. Recent reports show that MDSC increase fatty acid uptake and fatty acid oxidation (FAO) to support their immunosuppressive functions. Inhibition of FAO promoted a therapeutic T cell-mediated anti-tumor effect. Here, we sought to determine the mechanisms by which tumor-infiltrating MDSC increase the uptake of exogenous lipids and undergo metabolic and functional reprogramming to become highly immunosuppressive cells. The results showed that tumor-derived cytokines (G-CSF and GM-CSF) and the subsequent signaling through STAT3 and STAT5 induce the expression of lipid transport receptors with the resulting increase in the uptake of lipids present at high concentrations in the tumor microenvironment. The intracellular accumulation of lipids increases the oxidative metabolism and activates the immunosuppressive mechanisms. Inhibition of STAT3 or STAT5 signaling or genetic depletion of the fatty acid translocase CD36 inhibits the activation of oxidative metabolism and the induction of immunosuppressive function in tumor-infiltrating MDSC and results in a CD8+ T cell-dependent delay in tumor growth. Of note, human tumor-infiltrating and peripheral blood MDSC also upregulate the expression of lipid transport proteins, and lipids promote the generation of highly suppressive human MDSC in vitro. Our data therefore provide a mechanism by which tumor-derived factors and the high lipid content in the tumor microenvironment can cause the profound metabolic and functional changes found in MDSC and suggest novel approaches to prevent or reverse these processes. These results could further enhance the efficacy of cancer immunotherapy.

γ-Secretase inhibitors in cancer clinical trials are pharmacologically and functionally distinct.

γ-Secretase inhibitors (GSIs) are being actively repurposed as cancer therapeutics based on the premise that inhibition of NOTCH1 signaling in select cancers is therapeutic. Using novel assays to probe effects of GSIs against a broader panel of substrates, we demonstrate that clinical GSIs are pharmacologically distinct. GSIs show differential profiles of inhibition of the various NOTCH substrates, with some enhancing cleavage of other NOTCH substrates at concentrations where NOTCH1 cleavage is inhibited. Several GSIs are also potent inhibitors of select signal peptide peptidase (SPP/SPPL) family members. Extending these findings to mammosphere inhibition assays in triple-negative breast cancer lines, we establish that these GSIs have different functional effects. We also demonstrate that the processive γ-secretase cleavage pattern established for amyloid precursor protein (APP) occurs in multiple substrates and that potentiation of γ-secretase cleavage is attributable to a direct action of low concentrations of GSIs on γ-secretase. Such data definitively demonstrate that the clinical GSIs are not biological equivalents, and provide an important framework to evaluate results from ongoing and completed human trials with these compounds.