Efficacy and mechanism of the combination of PARP and CDK4/6 inhibitors in the treatment of triple-negative breast cancer

Abstract Background PARP inhibitors (PARPi) benefit only a fraction of breast cancer patients with BRCA mutations, and their efficacy is even more limited in triple-negative breast cancer (TNBC) due to clinical primary and acquired resistance. Here, we found that the efficacy of the PARPi olaparib in TNBC can be improved by combination with the CDK4/6 inhibitor (CDK4/6i) palbociclib. Methods We screened primary olaparib-sensitive and olaparib-resistant cell lines from existing BRCAmut/TNBC cell lines and generated cells with acquired olaparib resistance by gradually increasing the concentration. The effects of the PARPi olaparib and the CDK4/6i palbociclib on BRCAmut/TNBC cell lines were examined in both sensitive and resistant cells in vitro and in vivo. Pathway and gene alterations were assessed mechanistically and pharmacologically. Results We demonstrated for the first time that the combination of olaparib and palbociclib has synergistic effects against BRCAmut/TNBC both in vitro and in vivo. In olaparib-sensitive MDA-MB-436 cells, the single agent olaparib significantly inhibited cell viability and affected cell growth due to severe DNA damage. In olaparib-resistant HCC1937 and SUM149 cells, single-agent olaparib was ineffective due to potential homologous recombination (HR) repair, and the combination of olaparib and palbociclib greatly inhibited HR during the G2 phase, increased DNA damage and inhibited tumour growth. Inadequate DNA damage caused by olaparib activated the Wnt signalling pathway and upregulated MYC. Further experiments indicated that the overexpression of β-catenin, especially its hyperphosphorylation at the Ser675 site, activated the Wnt signalling pathway and mediated olaparib resistance, which could be strongly inhibited by combined treatment with palbociclib. Conclusions Our data provide a rationale for clinical evaluation of the therapeutic synergy of the PARPi olaparib and CDK4/6i palbociclib in BRCAmut/TNBCs with high Wnt signalling activation and high MYC expression that do not respond to PARPi monotherapy.

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Efficacy and Mechanism of the Combination of PARP and CDK4/6 Inhibitors in the Treatment of Triple-negative Breast Cancer

10.21203/rs.3.rs-125331/v1 ◽

2020 ◽

Author(s):

XIUZHI ZHU ◽

LI CHEN ◽

Binhao Huang ◽

Xiaoguang Li ◽

Yang Liu ◽

...

Keyword(s):

Breast Cancer ◽

Dna Damage ◽

Wnt Signaling ◽

Cell Lines ◽

Triple Negative ◽

Single Agent ◽

Wnt Signaling Pathway ◽

Tnbc Cell

Abstract Background: PARP inhibitors (PARPi) benefit only a fraction of breast cancer patients with BRCA mutations and their efficacy is even more limited in triple-negative breast cancer (TNBC) due to clinical primary and acquired resistance. Here, we found that the efficacy of PARPi in TNBC can be improved with CDK4/6 inhibitors (CDK4/6i).Methods: We screened primary PARPi-sensitive and resistant cell lines from existing BRCAmut/TNBC cell lines and generated cells with acquired PARPi resistance by gradually increasing the concentration. The effects of the PARPi olaparib and the CDK4/6i palbociclib on BRCAmut/TNBC cell lines were examined in both sensitive and resistant cells in vitro and in vivo. Pathway and gene alterations were assessed mechanistically and pharmacologically.Results: We demonstrated for the first time that the combination of PARPi and CDK4/6i has synergistic effects against BRCAmut/TNBC both in vitro and in vivo. In the PARPi-sensitive MB436 cells, the single agent olaparib significantly inhibited cell viability and affected cell growth due to severe DNA damage. In the PARPi-resistant HCC1937 and SUM149 cells, single-agent olaparib was ineffective due to potential homologous recombination (HR) repair, and the combination of PARPi and CDK4/6i greatly inhibited HR during the G2 phase, increased DNA damage and inhibited tumor growth. Inadequate DNA damage caused by PARPi activated the Wnt signaling pathway and upregulated MYC. Further experiments indicated that the overexpression of β-catenin, especially its hyperphosphorylation at the Ser675 site activated the Wnt signaling pathway and mediated PARPi resistance, which could be strongly inhibited by the combined treatment with CDK4/6i.Conclusions: Our data provide a rationale for the clinical evaluation of the therapeutic synergy of PARPi and CDK4/6i in BRCAmut/TNBCs with high Wnt signaling activation, high MYC expression and do not respond to PARPi monotherapy.

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Phytochemicals inhibit migration of triple negative breast cancer cells by targeting kinase signaling

BMC Cancer ◽

10.1186/s12885-019-6479-2 ◽

2020 ◽

Vol 20 (1) ◽

Cited By ~ 2

Author(s):

Pradip Shahi Thakuri ◽

Megha Gupta ◽

Sunil Singh ◽

Ramila Joshi ◽

Eric Glasgow ◽

...

Keyword(s):

Breast Cancer ◽

Cell Migration ◽

Cancer Cells ◽

Protein Kinases ◽

Cell Lines ◽

Triple Negative Breast Cancer ◽

Triple Negative ◽

Migration And Invasion ◽

Tnbc Cell

Abstract Background Cell migration and invasion are essential processes for metastatic dissemination of cancer cells. Significant progress has been made in developing new therapies against oncogenic signaling to eliminate cancer cells and shrink tumors. However, inherent heterogeneity and treatment-induced adaptation to drugs commonly enable subsets of cancer cells to survive therapy. In addition to local recurrence, these cells escape a primary tumor and migrate through the stroma to access the circulation and metastasize to different organs, leading to an incurable disease. As such, therapeutics that block migration and invasion of cancer cells may inhibit or reduce metastasis and significantly improve cancer therapy. This is particularly more important for cancers, such as triple negative breast cancer, that currently lack targeted drugs. Methods We used cell migration, 3D invasion, zebrafish metastasis model, and phosphorylation analysis of 43 protein kinases in nine triple negative breast cancer (TNBC) cell lines to study effects of fisetin and quercetin on inhibition of TNBC cell migration, invasion, and metastasis. Results Fisetin and quercetin were highly effective against migration of all nine TNBC cell lines with up to 76 and 74% inhibitory effects, respectively. In addition, treatments significantly reduced 3D invasion of highly motile TNBC cells from spheroids into a collagen matrix and their metastasis in vivo. Fisetin and quercetin commonly targeted different components and substrates of the oncogenic PI3K/AKT pathway and significantly reduced their activities. Additionally, both compounds disrupted activities of several protein kinases in MAPK and STAT pathways. We used molecular inhibitors specific to these signaling proteins to establish the migration-inhibitory role of the two phytochemicals against TNBC cells. Conclusions We established that fisetin and quercetin potently inhibit migration of metastatic TNBC cells by interfering with activities of oncogenic protein kinases in multiple pathways.

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Rad51 Silencing with siRNA Delivered by Porous Silicon-Based Microparticle Enhances the Anti-Cancer Effect of Doxorubicin in Triple-Negative Breast Cancer

Journal of Biomedical Nanotechnology ◽

10.1166/jbn.2021.3198 ◽

2021 ◽

Vol 17 (12) ◽

pp. 2351-2363

Author(s):

Zeliang Wu ◽

Lin Zhu ◽

Junhua Mai ◽

Haifa Shen ◽

Rong Xu

Keyword(s):

Breast Cancer ◽

Dna Damage ◽

Porous Silicon ◽

Lung Metastasis ◽

Triple Negative Breast Cancer ◽

Triple Negative ◽

Antitumor Effects ◽

Mesenchymal Transition ◽

Tnbc Cell

Due to its high heterogeneity and aggressiveness, cytotoxic chemotherapy is still a mainstay treatment for triple negative breast cancer. Unfortunately, the above mentioned has not significantly ameliorated TNBC patients and induces drug resistance. Exploring the mechanisms underlying the chemotherapy sensitivity of TNBC and developing novel sensitization strategies are promising approaches for improving the prognosis of patients. Rad51, a key regulator of DNA damage response pathway, repairs DNA damage caused by genotoxic agents through “homologous recombination repair.” Therefore, Rad51 inhibition may increase TNBC cell sensitivity to anticancer agents. Based on these findings, we first designed Rad51 siRNA to inhibit the Rad51 protein expression in vitro and evaluated the sensitivity of TNBC cells to doxorubicin. Subsequently, we constructed discoidal porous silicon microparticles (pSi) and encapsulated discoidal 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) liposomes/siRad51 (PS-DOPC/siRad51) to explore the synergistic antitumor effects of siRad51 and doxorubicin on two mouse models of TNBC in vivo. Our in vitro studies indicated that siRad51 enhanced the efficacy of DOX chemotherapy and significantly suppressed TNBC cell proliferation and metastasis. This effect was related to apoptosis induction and epithelial to mesenchymal transition (EMT) inhibition. siRad51 altered the expression of apoptosis- and EMT-related proteins. In orthotopic and lung metastasis xenograft models, the administration of PS-DOPC/siRad51 in combination with DOX significantly alleviated the primary tumor burden and lung metastasis, respectively. Our current studies present an efficient strategy to surmount chemotherapy resistance in TNBC through microvector delivery of siRad51.

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Co-loading of an anthracycline analogue Pirarubicin and Salinomycin in a 1:3 ratio into Quatramerbiodegradable polymeric nanoparticles synergistically inhibit growth of triple-negative breast cancer.

Journal of Clinical Oncology ◽

10.1200/jco.2021.39.15_suppl.e15047 ◽

2021 ◽

Vol 39 (15_suppl) ◽

pp. e15047-e15047

Author(s):

Surender Kharbanda ◽

Anees Mohammad ◽

Sachchidanand Tiwari ◽

Neha Mehrotra ◽

Sireesh Appajosyula ◽

...

Keyword(s):

Breast Cancer ◽

Triple Negative Breast Cancer ◽

Triple Negative ◽

Breast Cancers ◽

Mice Model ◽

Single Drug ◽

Tnbc Cell ◽

Dual Drug

e15047 Background: Triple negative breast cancer (TNBC) accounts for about 10-15% of all breast cancers and differ from other types of invasive breast cancers in that they grow and spread faster. TNBCs have limited treatment options and a worse prognosis. Therapy with anthracyclines considered to be one of the most effective agents in the treatment. Unfortunately, resistance to anthracycline therapy is very common due to drug efflux mediated by overexpression of ABC transporter. Pirarubicin (PIRA), an analogue of doxorubicin (DOX), is approved in Japan, Korea and China and is shown to be less cardiotoxic than DOX. Recent studies suggest that cancer stem cells (CSCs) play an important role in tumorigenesis and biology of TNBC. Targeting CSCs may be a promising, novel strategy for the treatment of this aggressive disease. Recent studies have shown that salinomycin (SAL) preferentially targets the viability of CSCs. Methods: SAL and PIRA were co-encapsulated in polylactic acid (PLA)-based block copolymeric nanoparticles (NPs) to efficiently co-deliver these agents to treat TNBC cells. Results: Generated SAL-PIRA co-encapsulated dual drug-loaded NPs showed an average diameter of 110 ± 7 nm, zeta potential of -12.5 mV and PDI of less than 0.25. Both of these anti-cancer agents showed slow and sustained release profile in non-physiological buffer (PBS, pH 7.4) from these dual drug-encapsulated NPs. Additionally, multiple ratios (PIRA:SAL = 3:1, 1:1, 1:3) were encapsulated to generate diverse dual drug-loaded NPs. The results demonstrate that, in contrast to 1:1 and 3:1, treatment of TNBC cells with 1:3 ratio of PIRA:SAL dual drug-loaded NPs, was associated with significant inhibition of growth in vitro in multiple TNBC cell lines. Interestingly, PIRA:SAL (1:3) was synergistic as compared to either SAL- or PIRA single drug-loaded NPs. The IC50 of PIRA and SAL in single drug-encapsulated NPs is 150 nM and 700 nM respectively in MDA-MB-468. Importantly, the IC50 of PIRA in dual drug-encapsulated NPs dropped down to 30 nM (5-fold). Similar results were obtained in SUM-149 TNBC cell line. Studies are underway to evaluate in vivo biological activity of PIRA:SAL (1:3) on tumor growth in a TNBC xenograft mice model. Conclusions: These results demonstrate that a novel dual drug-loaded NP formulation of PIRA and SAL in a unique ratio of 1:3 represents an approach for successful targeting of CSCs and bulk tumor cells in TNBC and potentially other cancer types.

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PFKFB4 Overexpression Facilitates Proliferation by Promoting the G1/S Transition and Is Associated with a Poor Prognosis in Triple-Negative Breast Cancer

Disease Markers ◽

10.1155/2021/8824589 ◽

2021 ◽

Vol 2021 ◽

pp. 1-10

Author(s):

Yu-Chen Cai ◽

Hang Yang ◽

Hong-Bo Shan ◽

Hui-Fang Su ◽

Wen-Qi Jiang ◽

...

Keyword(s):

Breast Cancer ◽

Triple Negative Breast Cancer ◽

Triple Negative ◽

Signaling Networks ◽

Oncogenic Signaling ◽

Tnbc Cell ◽

Key Factor ◽

Tissue Specimens

Background. 6-Phosphofructo-2-kinase/fructose-2,6-biphosphate-4 (PFKFB4) is a key factor that plays an important role in tumorigenesis. However, its role in triple-negative breast cancer (TNBC) progression needs to be further validated. We investigated whether PFKFB4 is directly involved in the oncogenic signaling networks of TNBC. Methods. First, we assessed the expression level of PFKFB4 in tumor tissue specimens by immunohistochemistry and evaluated its prognostic value. Next, the effect of PFKFB4 on TNBC cell growth and associated mechanisms were investigated. Finally, the results were further verified in vivo. Results. We found that PFKFB4 overexpression was associated with an unfavorable prognosis in TNBC patients. PFKFB4 was overexpressed in TNBC cell lines in hypoxic environments, and its overexpression promoted tumor progression in vitro and in vivo. Further analyses demonstrated that the possible mechanism might be that PFKFB4 overexpression facilitates TNBC progression by enhancing the G1/S phase transition by increasing the protein level of CDK6 and phosphorylation of Rb. Conclusions. These data suggest that PFKFB4 plays significant roles in the tumorigenesis and development of TNBC.

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Synergistic antitumor effects of S-1 with eribulin in vitro and in vivo for triple-negative breast cancer cell lines

SpringerPlus ◽

10.1186/2193-1801-3-417 ◽

2014 ◽

Vol 3 (1) ◽

pp. 417 ◽

Cited By ~ 15

Author(s):

Masato Terashima ◽

Kazuko Sakai ◽

Yosuke Togashi ◽

Hidetoshi Hayashi ◽

Marco A De Velasco ◽

...

Keyword(s):

Breast Cancer ◽

Cell Lines ◽

Cancer Cell ◽

Breast Cancer Cell ◽

Triple Negative Breast Cancer ◽

Triple Negative ◽

Breast Cancer Cell Lines ◽

Antitumor Effects

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Targeting MYCN-expressing triple-negative breast cancer with BET and MEK inhibitors

Science Translational Medicine ◽

10.1126/scitranslmed.aaw8275 ◽

2020 ◽

Vol 12 (534) ◽

pp. eaaw8275 ◽

Cited By ~ 8

Author(s):

Johanna M. Schafer ◽

Brian D. Lehmann ◽

Paula I. Gonzalez-Ericsson ◽

Clayton B. Marshall ◽

J. Scott Beeler ◽

...

Keyword(s):

Breast Cancer ◽

Cell Lines ◽

Triple Negative Breast Cancer ◽

Triple Negative ◽

Clinical Investigation ◽

Mek Inhibitors ◽

Cancer Subtypes ◽

Bet Inhibitors

Triple-negative breast cancer (TNBC) is an aggressive form of breast cancer that does not respond to endocrine therapy or human epidermal growth factor receptor 2 (HER2)–targeted therapies. Individuals with TNBC experience higher rates of relapse and shorter overall survival compared to patients with receptor-positive breast cancer subtypes. Preclinical discoveries are needed to identify, develop, and advance new drug targets to improve outcomes for patients with TNBC. Here, we report that MYCN, an oncogene typically overexpressed in tumors of the nervous system or with neuroendocrine features, is heterogeneously expressed within a substantial fraction of primary and recurrent TNBC and is expressed in an even higher fraction of TNBCs that do not display a pathological complete response after neoadjuvant chemotherapy. We performed high-throughput chemical screens on TNBC cell lines with varying amounts of MYCN expression and determined that cells with higher expression of MYCN were more sensitive to bromodomain and extraterminal motif (BET) inhibitors. Combined BET and MEK inhibition resulted in a synergistic decrease in viability, both in vitro and in vivo, using cell lines and patient-derived xenograft (PDX) models. Our preclinical data provide a rationale to advance a combination of BET and MEK inhibitors to clinical investigation for patients with advanced MYCN-expressing TNBC.

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ZHX2 Promotes HIF1α Oncogenic Signaling in Triple-Negative Breast Cancer

10.1101/2021.05.27.445959 ◽

2021 ◽

Author(s):

Wentong Fang ◽

Chengheng Liao ◽

Rachel Shi ◽

Jeremy Simon ◽

Travis Ptacek ◽

...

Keyword(s):

Breast Cancer ◽

Gene Expression ◽

Triple Negative Breast Cancer ◽

Triple Negative ◽

Hypoxia Inducible Factor ◽

Oncogenic Signaling ◽

Potential Therapeutic Target ◽

Tnbc Cell

Triple-negative breast cancer (TNBC) is an aggressive and highly lethal disease, which warrants the critical need to identify new therapeutic targets. We show that Zinc Fingers And Homeoboxes 2 (ZHX2) is amplified or overexpressed in TNBC cell lines and patients. Functionally, depletion of ZHX2 inhibited TNBC cell growth and invasion in vitro, orthotopic tumor growth and spontaneous lung metastasis in vivo. Mechanistically, ZHX2 bound with hypoxia inducible factor (HIF) family members and positively regulated HIF1α activity in TNBC. Integrated ChIP-Seq and gene expression profiling demonstrated that ZHX2 co-occupied with HIF1α on transcriptionally active promoters marked by H3K4me3 and H3K27ac, thereby promoting gene expression. Furthermore, multiple residues (R491, R581 and R674) on ZHX2 are important in regulating its phenotype, which correspond with their roles on controlling HIF1α activity in TNBC cells. These studies establish that ZHX2 activates oncogenic HIF1α signaling, therefore serving as a potential therapeutic target for TNBC.

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MiR-664b-3p Inhibited Triple-Negative Breast Cancer Cell Growth Via Targeting BRIP1

10.21203/rs.3.rs-798916/v1 ◽

2021 ◽

Author(s):

Qiu Jin ◽

Bo Lin ◽

Wenhui Zhao ◽

Runyuan Ji

Keyword(s):

Breast Cancer ◽

Cell Proliferation ◽

Cell Lines ◽

Triple Negative Breast Cancer ◽

Triple Negative ◽

Luciferase Reporter ◽

Breast Cancer Cell Growth ◽

Cell Functions

Abstract BackgroundMany studies indicate that microRNAs (miRNAs) play a crucial role in modulating the development and progression of triple-negative breast cancer (TNBC). However, miR-664b-3p affections on the TNBC functions and mechanisms are still unknown. The purpose of our study was to clarify the effects of miR-664b-3p in cellular TNBC development and progression.MethodsIn our study, the expressions of miR-664b-3p in cell lines and tissueswere tested by real-time PCR (RT-PCR), immunofluorescence, H&E and immunohistochemistry staining. CCK-8 assay, colony formation, EdU, flow cytometry apoptosis, wound scratch, Transwell assays were applied to explore the cell functions. The targeted relationship between miR-664b-3p and its target BRIP1 was determined by dual-luciferase reporter assay and rescue experiments. ResultsWe observed that miR-664b-3p was significantly decreased in TNBC cell lines. Overexpression of miR-664b-3p could observably inhibit cell proliferation, migration, invasion and induced apoptosis in vitro. Meanwhile, miR-664-3p suppressed TNBC tumor growth in vivo. Furthermore, luciferase reporter assays identified the interaction between 3’UTR of BRIP1 and miR-664b-3p. Moreover, we investigated the mechanisms underlying the effect of miR-664b-3p on cell functions, and the result showed that miR-664b-3p inhibited cell proliferation, invasion and accelerated apoptosis by targeting BRIP1.ConclusionFrom the above, our findings indicated that miR-664b-3p played a significant role in TNBC progression by targeting BRIP1, providing new therapeutic targets for diagnostic in TNBC.

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3174 Tumor suppressors p53 and ARF control oncogenic potential of triple-negative breast cancer cells by regulating RNA editing enzyme ADAR1

Journal of Clinical and Translational Science ◽

10.1017/cts.2019.61 ◽

2019 ◽

Vol 3 (s1) ◽

pp. 25-26

Author(s):

Che-Pei Kung ◽

Emily Bross ◽

Emily Bramel ◽

Eric Freeman ◽

Thwisha Sabloak ◽

...

Keyword(s):

Breast Cancer ◽

Rna Editing ◽

Cell Lines ◽

Triple Negative Breast Cancer ◽

Tumor Suppressors ◽

Triple Negative ◽

Oncogenic Potential ◽

Tnbc Cell ◽

Editing Enzyme

OBJECTIVES/SPECIFIC AIMS: Triple-negative breast cancer (TNBC) accounts for one-fifth of the breast cancer patient population. The heterogeneous nature of TNBC and lack of options for targeted therapy make its treatment a constant challenge. The co-deficiency of tumor suppressors p53 and ARF is a significant genetic signature enriched in TNBC, but it is not yet clear how TNBC is regulated by this genetic alteration. METHODS/STUDY POPULATION: To answer this question, we established p53/ARF-defective murine embryonic fibroblast (MEF) to study the molecular and phenotypic consequences in vitro. Moreover, transgenic mice were generated to investigate the effect of p53/ARF deficiency on mammary tumor development in vivo. RESULTS/ANTICIPATED RESULTS: Increased transformation capability was observed in p53/ARF-defective cells, and formation of aggressive mammary tumors was also seen in p53-/-ARF-/- mice. RNA-editing enzyme ADAR1 was identified as a potential mediator for the elevated oncogenic potential. Interestingly, we found that the overexpression of ADAR1 is also prevalent in human TNBC cell lines and patient specimen. Using short hairpin RNA (shRNA) to reduce ADAR1 expression abrogated the oncogenic potential of human TNBC cell lines, while non-TNBC cells are less susceptible. Different levels of RNA editing of known ADAR1 targets were detected in shRNA-treated human TNBC cell lines, suggesting that ADAR1-mediated RNA editing contributes to TNBC pathogenesis. DISCUSSION/SIGNIFICANCE OF IMPACT: These results indicate critical roles played by the tumor suppressors p53 and ARF in the pathogenesis of TNBC, partially through affecting ADAR1-mediated RNA editing. Further understanding of this pathway could shed light on potential vulnerabilities of TNBC and inform the development of personalized therapies based on patients’ genetic signiatures.

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