An immune-humanized patient-derived xenograft model of estrogen-independent, hormone receptor positive metastatic breast cancer.

BACKGROUND: Metastatic breast cancer (MBC) is incurable, with a 5-year survival rate of 28%. In the USA, more than 42,000 patients die from MBC every year. The most common type of breast cancer is estrogen receptor-positive (ER+), and more patients die from ER+ breast cancer than from any other subtype. ER+ tumors can be successfully treated with hormone therapy, but many tumors acquire endocrine resistance, at which point treatment options are limited. There is an urgent need for model systems that better represent human ER+ MBC in vivo, where tumors can metastasize. Patient-derived xenografts (PDX) made from MBC spontaneously metastasize, but the immunodeficient host is a caveat, given the known role of the immune system in tumor progression and response to therapy. Thus, we attempted to develop an immune-humanized PDX model of ER+ MBC. METHODS: NSG-SGM3 mice were immune-humanized with CD34+ hematopoietic stem cells, followed by engraftment of human ER+ endocrine resistant MBC tumor fragments. Strategies for exogenous estrogen supplementation were compared, and immune-humanization in blood, bone marrow, spleen, and tumors was assessed by flow cytometry and tissue immunostaining. Characterization of the new model includes assessment of the human tumor microenvironment performed by immunostaining. RESULTS: We describe the development of an immune-humanized PDX model of estrogen-independent endocrine resistant ER+ MBC. Importantly, our model harbors a naturally occurring ESR1 mutation, and immune-humanization recapitulates the lymphocyte-excluded and myeloid-rich tumor microenvironment of human ER+ breast tumors. CONCLUSION: This model sets the stage for development of other clinically relevant models of human breast cancer and should allow future studies on mechanisms of endocrine resistance and tumor-immune interactions in an immune-humanized in vivo setting.

MICAL1 regulates actin cytoskeleton organization, directional cell migration and the growth of human breast cancer cells as orthotopic xenograft tumours.

The Molecule Interacting with CasL 1 (MICAL1) monooxygenase has emerged as an important regulator of cytoskeleton organization via actin oxidation. Although filamentous actin (F-actin) increases MICAL1 monooxygenase activity, hydrogen peroxide (H2O2) is also generated in the absence of F-actin, suggesting that diffusible H2O2 might have additional functions. MICAL1 gene disruption by CRISPR/Cas9 in MDA MB 231 human breast cancer cells knocked out (KO) protein expression, which affected F-actin organization, cell size and motility. Transcriptomic profiling revealed that MICAL1 deletion significantly affected the expression of over 700 genes, with the majority being reduced in their expression levels. In addition, the absolute magnitudes of reduced gene expression were significantly greater than the magnitudes of increased gene expression. Gene set enrichment analysis (GSEA) identified receptor regulator activity as the most significant negatively enriched molecular function gene set. The prominent influence exerted by MICAL1 on F-actin structures was also associated with changes in the expression of several serum-response factor (SRF) regulated genes in KO cells. Moreover, MICAL1 disruption attenuated breast cancer tumour growth in vivo. Elevated MICAL1 gene expression was observed in invasive breast cancer samples from human patients relative to normal tissue, while MICAL1 amplification or point mutations were associated with reduced progression free survival. Collectively, these results demonstrate that MICAL1 gene disruption altered cytoskeleton organization, cell morphology and migration, gene expression, and impaired tumour growth in an orthotopic in vivo breast cancer model, suggesting that pharmacological MICAL1 inhibition could have therapeutic benefits for cancer patients.

NF-κB and neutrophil extracellular traps cooperate to promote breast cancer progression and metastasis.

Aberrant NF-κB activation and neutrophil extracellular traps (NETs) are associated with breast cancer progression. How NF-κB and NETs modulate each other in breast cancer development remains unclear. Here, we found that NETs induced by phorbol 12-myristate 13-acetate promote breast cancer cell progression. In turn, cancer cells-derived factors, such as IL-8 and granulocyte colony-stimulating factor, stimulate neutrophils to form NETs. Mechanistically, NETs increased the interaction of NF-κB essential modifier (NEMO) with IκB kinase (IKK)α/ß and enhanced NF-κB activation. We then employed a cell-permeable peptide corresponding to the NEMO-binding domain (NBD) of IKKα/ß, termed NBD peptide, which disrupts NETs-mediated NEMO interaction with IKKα/ß and abolished NF-κB activation in vitro. NBD peptide also reduced IL-8 level and NETs formation, and suppressed primary tumor growth and/or lung metastasis in human breast cancer mouse xenograft models and mouse spontaneous breast cancer model. Blockade of NET formation using a peptidylarginine deiminase 4 (PAD4) pharmacologic inhibitor decreased NF-κB activation and tumor metastasis. Collectively, these data suggest that NF-κB associates with NETs to form a positive loop facilitating breast tumor progression and metastasis, and that selective inhibition of NF-κB and PAD4-dependent NETs provides an effective therapeutic approach for treating breast cancer.

CPSF4 regulates circRNA formation and microRNA mediated gene silencing in hepatocellular carcinoma.

CircRNAs play essential roles in various physiological processes and involves in many diseases, in particular cancer. Global downregulation of circRNA expression has been observed in hepatocellular carcinoma (HCC) in many studies. Previous studies revealed that the pre-mRNA 3' end processing complex participates in circRNA cyclization and plays an important role in HCC tumorigenesis. Therefore, we explored the role of CPSF4, for 3' end formation and cleavage, in circRNA formation. Clinical research has shown that CPSF4 expression is upregulated in HCC and that high expression of CPSF4 is associated with poor prognosis in HCC patients. Mechanistic studies have demonstrated that CPSF4 reduces the levels of circRNAs, which possess a polyadenylation signal sequence and this decrease in circRNAs reduces the accumulation of miRNA and disrupts the miRNA-mediated gene silencing in HCC. Experiments in cell culture and xenograft mouse models showed that CPSF4 promotes the proliferation of HCC cells and enhances tumorigenicity. Moreover, CPSF4 antagonizes the tumor suppressor effect of its downstream circRNA in HCC. In summary, CPSF4 acts as an oncogene in HCC through circRNA inhibition and disruption of miRNA-mediated gene silencing.

Natural killer cells and cytotoxic T lymphocytes are required to clear solid tumor in a patient-derived xenograft.

Existing patient-derived xenograft (PDX) mouse models of solid tumors lack a fully tumor donor-matched, syngeneic, and functional immune system. We developed a model that overcomes these limitations by engrafting lymphopenic recipient mice with a fresh, undisrupted piece of solid tumor, whereby tumor-infiltrating lymphocytes (TILs) persisted in the recipient mice for several weeks. Successful tumor engraftment was achieved in 83% to 89% of TIL-PDX mice, and these were seen to harbor exhausted immuno-effector as well as functional immunoregulatory cells persisting for at least 6 months postengraftment. Combined treatment with interleukin-15 stimulation and immune checkpoint inhibition resulted in complete or partial tumor response in this model. Further, depletion of cytotoxic T lymphocytes and/or natural killer cells before combined immunotherapy revealed that both cell types were required for maximal tumor regression. Our TIL-PDX model provides a valuable resource for powerful mechanistic and therapeutic studies in solid tumors.

Human stem cells harboring a suicide gene improve the safety and standardisation of neural transplants in Parkinsonian rats.

Despite advancements in human pluripotent stem cells (hPSCs) differentiation protocols to generate appropriate neuronal progenitors suitable for transplantation in Parkinson's disease, resultant grafts contain low proportions of dopamine neurons. Added to this is the tumorigenic risk associated with the potential presence of incompletely patterned, proliferative cells within grafts. Here, we utilised a hPSC line carrying a FailSafeTM suicide gene (thymidine kinase linked to cyclinD1) to selectively ablate proliferative cells in order to improve safety and purity of neural transplantation in a Parkinsonian model. The engineered FailSafeTM hPSCs demonstrated robust ventral midbrain specification in vitro, capable of forming neural grafts upon transplantation. Activation of the suicide gene within weeks after transplantation, by ganciclovir administration, resulted in significantly smaller grafts without affecting the total yield of dopamine neurons, their capacity to innervate the host brain or reverse motor deficits at six months in a rat Parkinsonian model. Within ganciclovir-treated grafts, other neuronal, glial and non-neural populations (including proliferative cells), were significantly reduced-cell types that may pose adverse or unknown influences on graft and host function. These findings demonstrate the capacity of a suicide gene-based system to improve both the standardisation and safety of hPSC-derived grafts in a rat model of Parkinsonism.

Deep Learning Enables Individual Xenograft Cell Classification in Histological Images by Analysis of Contextual Features.

Patient-Derived Xenografts (PDXs) are the preclinical models which best recapitulate inter- and intra-patient complexity of human breast malignancies, and are also emerging as useful tools to study the normal breast epithelium. However, data analysis generated with such models is often confounded by the presence of host cells and can give rise to data misinterpretation. For instance, it is important to discriminate between xenografted and host cells in histological sections prior to performing immunostainings. We developed Single Cell Classifier (SCC), a data-driven deep learning-based computational tool that provides an innovative approach for automated cell species discrimination based on a multi-step process entailing nuclei segmentation and single cell classification. We show that human and murine cell contextual features, more than cell-intrinsic ones, can be exploited to discriminate between cell species in both normal and malignant tissues, yielding up to 96% classification accuracy. SCC will facilitate the interpretation of H&E- and DAPI-stained histological sections of xenografted human-in-mouse tissues and it is open to new in-house built models for further applications. SCC is released as an open-source plugin in ImageJ/Fiji available at the following link: https://github.com/Biomedical-Imaging-Group/SingleCellClassifier .

Surgical Transplantation of Human RPE Stem Cell-Derived RPE Monolayers into Non-Human Primates with Immunosuppression.

Recent trials of retinal pigment epithelium (RPE) transplantation for the treatment of disorders such as age-related macular degeneration have been promising. However, limitations of existing strategies include the uncertain survival of RPE cells delivered by cell suspension and the inherent risk of uncontrolled cell proliferation in the vitreous cavity. Human RPE stem cell-derived RPE (hRPESC-RPE) transplantation can rescue vision in a rat model of retinal dystrophy and survive in the rabbit retina for at least 1 month. The present study placed hRPESC-RPE monolayers under the macula of a non-human primate model for 3 months. The transplant was able to recover in vivo and maintained healthy photoreceptors. Importantly, there was no evidence that subretinally transplanted monolayers underwent an epithelial-mesenchymal transition. Neither gliosis in adjacent retina nor epiretinal membranes were observed. These findings suggest that hRPESC-RPE monolayers are safe and may be a useful source for RPE cell replacement therapy.

Establishment and histopathological study of patient-derived xenograft models and primary cell lines of epithelioid malignant peritoneal mesothelioma.

Malignant peritoneal mesothelioma (MPM) is a rare malignancy with few experimental models. This study used the human surgical specimen to establish MPM patient-derived xenograft (PDX) models and primary cell lines to provide a study platform for MPM in vitro and in vivo, and conducted histopathological analysis. Our study used the experimental peritoneal cancer index (ePCI) score to evaluate gross pathology, and the results showed that the ePCI score of the female and male nude mice were 8.80 ± 1.75 and 9.20 ± 1.81 (P=0.6219), respectively. The Hematoxylin and eosin (HE) staining of animal models showed that the tumor was epithelioid mesothelioma and invaded multiple organs. Immunohistochemistry (IHC) staining showed that Calretinin, Cytokeratin 5/6, WT-1 and Ki-67 were all positive. The Swiss-Giemsa and Immunofluorescence (IF) staining of primary cell lines were also consistent with the pathological characteristics of mesothelioma. We also performed the whole-exome sequencing (WES) to identify the mutant genes between models and the patient. And the results showed that 21 mutant genes were shared between the two groups, and the genes related to tumorigenesis and development including BAP1, NF2, MTBP, NECTIN2, CDC23, LRPPRC, TRIM25, and DHRS2. In conclusion, the PDX models and primary cell lines of MPM were successfully established with the epithelioid mesothelioma identity confirmed by histopathological evidence. Moreover, our study has also illustrated the shared genomic profile between models and the patient.

Paediatric Burkitt lymphoma patient-derived xenografts capture disease characteristics over time and are a model for therapy.

Burkitt lymphoma (BL) accounts for almost two-thirds of all B-cell non-Hodgkin lymphoma (B-NHL) in children and adolescents and is characterised by a MYC translocation and rapid cell turnover. Intensive chemotherapeutic regimens have been developed in recent decades, including the lymphomes malins B (LMB) protocol, which have resulted in a survival rate in excess of 90%. Recent clinical trials have focused on immunochemotherapy, with the addition of rituximab to chemotherapeutic backbones, showing encouraging results. Despite these advances, relapse and refractory disease occurs in up to 10% of patients and salvage options for these carry a dismal prognosis. Efforts to better understand the molecular and functional characteristics driving relapse and refractory disease may help improve this prognosis. This study has established a paediatric BL patient-derived xenograft (PDX) resource which captures and maintains tumour heterogeneity, may be used to better characterise tumours and identify cell populations responsible for therapy resistance.

Quantification of Tumor Vasculature by Analysis of Amount and Spatial Dispersion of Caliber-Classified Vessels.

This protocol focuses on the quantitative description of the angioarchitecture of experimental tumor xenografts. This semiautomatic analysis is carried out on functional vessels and microvessels acquired by confocal imaging and processed into progressively reconstructed angioarchitectures following a caliber-classification step. The protocol can be applied also to the quantification of pathological angioarchitectures other than tumor grafts as well as to the microvasculature of physiological tissue samples.

A Xenograft Model for Venous Malformation.

Xenograft models allow for an in vivo approach to monitor cellular functions within the context of a host microenvironment. Here we describe a protocol to generate a xenograft model of venous malformation (VM) based on the use of human umbilical vein endothelial cells (HUVEC) expressing a constitutive active form of the endothelial tyrosine kinase receptor TEK (TIE2 p.L914F) or patient-derived EC containing TIE2 and/or PIK3CA gene mutations. Hyperactive somatic TIE2 and PIK3CA mutations are a common hallmark of VM in patient lesions. The EC are injected subcutaneously on the back of athymic nude mice to generate ectatic vascular channels and recapitulate histopathological features of VM patient tissue histology. Lesion plugs with TIE2/PIK3CA-mutant EC are visibly vascularized within 7-9 days of subcutaneous injection, making this a great tool to study venous malformation.

A Universal Gelfoam 3-D Histoculture Method to Establish Patient-derived Cancer Cells (3D-PDCC) Without Fibroblasts from Patient-derived Xenografts.

BACKGROUND/AIM: The direct placement of patient tumors in 2-D culture on plastic or glass surfaces has inhibited the establishment of patient-derived cancer cells (PDCCs). The aim of the present study was to develop universal and efficient methods to prepare PDCCs. MATERIALS AND METHODS: Fragments of patient-derived xenograft (PDX) tumors established form colon cancer liver metastasis (1 mm3) were placed on Gelfoam and cultured in DMEM. RESULTS: PDX tumor fragments were cultured on Gelfoam. Cancer cells migrated from the explant and formed distinct 3-D structures in the Gelfoam. Each of the three PDCCs showed a distinct morphology. The cultures were essentially all cancer cells without fibroblasts, the opposite of what usually occurs in 2-D culture on plastic or glass. Gelfoam cultures could be readily passaged from one Gelfoam cube to anothers suggesting indefinite culture potential. CONCLUSION: A potentially universal method to establish PDCC using PDX tumors and 3-D Gelfoam histoculture was developed.

Clinical and morphological results of xenografts to use in myringoplasty.

The main aim of our study was to study morphological state of the autograft from the fascia of the temporal muscle in myringoplasty. Until now, there is no consensus on issue of which fabrics are more suitable for use in the eardrum. We decided to study of use of an autograft from the fascia of the temporal muscle for myringoplasty in rabbits in the experiment, and in patients with chronic dry mesotympanitis. An electron microscopic examination of the fascia taken immediately, after 10, 20 min and 1 h after sampling. It was found that there are no gross destructive changes in the fascia structure. Minor changes are detected in the form of a light disorganization of the collagen complex, granular dystrophy with an increase in cell. Inflammatory diseases of the middle ear are widespread among population of all age groups. In the experiment, a positive result was obtained in 29 (82.8%) rabbits. In our research we performed morphological features of xenograft engraftment in an experimental animal on 3, 7, 14, 21 days and 1-3 months. In period from 3 days to 3 months after operation, the animals were euthanized by an air embolism and subjected to pathological examination. Then recovered xenograft, was examined macro and microscopically. Pieces were fixed in a 10% solution of neutral formalin. After washing with water, dehydration was carried out in alcohol and chloroform, and after, waxed with paraffin. Histological sections were stained with hematoxylin-eosin. Collagen fibers were detected by method of Vann-Gieson.

ARG1 functions as a tumor suppressor in breast cancer.

Arginase I (ARG1) is a cytosolic enzyme that catalyzes the hydrolysis of L-arginine to L-ornithine and urea. The association of ARG1 with cancer has mostly been focused on the ARG1 released by tumor-associated myeloid cells in tumor microenvironment. However, the role of ARG1 expressed in cancer cells is unclear. Here, we showed that the expression of ARG1 in human breast cancer (BC) is related to a good prognosis in BC patients. Overexpression of ARG1 suppresses BC cell proliferation and migration in vitro and xenograft tumor growth and development in mouse models. Furthermore, ARG1 expression down-regulates the expression of p-AKT, leading to the de-activation of AKT signal pathway in BC cells. Thus, our results established that in contrast to the role of ARG1 released from tumor-associated myeloid cells in tumor microenvironment that promotes tumor immune escape, ARG1 expressed in BC cells suppresses AKT signaling pathway and functions as a tumor suppressor.

125I seeds irradiation inhibits tumor growth and induces apoptosis by Ki-67, P21, survivin, livin and caspase-9 expression in lung carcinoma xenografts.

BACKGROUND: Lung cancer is a fatal disease and a serious health problem worldwide. Patients are usually diagnosed at an advanced stage, and the effectiveness of chemotherapy for such patients is very limited. Iodine 125 seed (125I) irradiation can be used as an important adjuvant treatment for lung carcinoma. The purpose of this study was to examine the role of irradiation by 125I seeds in human lung cancer xenograft model and to determine the underlying mechanisms involved, with a focus on apoptosis. METHODS: 40 mice with A549 lung adenocarcinoma xenografts were randomly divided into 4 groups: control group (n = 10), sham seed (0 mCi) implant group (n = 10), 125I seed (0.6 mCi) implant group (n = 10) and 125I seed (0.8 mCi) implant group (n = 10), respectively. The body weight and tumor volume, were recorded every 4 days until the end of the study. Apoptotic cells were checked by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and activities of caspase-3 and caspase-8 enzyme were tested. Expression of P21, survivin, livin, caspase-9 and proliferating cell nuclear antigen (Ki-67) was detected with immunohistochemical staining. RESULTS: The results of TUNEL staining assays showed that 125I seed irradiation suppresses the growth of lung cancer xenografts in nude mice and induced apoptosis. The activity of caspase-3 and caspase-8 was significantly higher. The expression levels Ki67, survivin and livin were substantially downregulated, while P21 and caspase-9 protein expression were significantly increased following 125I seed irradiation. This study revealed that 125I seed irradiation could significantly change apoptosis-related protein in human lung cancer xenografts. CONCLUSIONS: Overall, our study demonstrates that radiation exposure by 125I seeds could be a new treatment option for lung cancer.

Quantitative in vivo bioluminescence imaging of orthotopic patient-derived glioblastoma xenografts.

Despite extensive research, little progress has been made in glioblastoma therapy, owing in part to a lack of adequate preclinical in vivo models to study this disease. To mitigate this, primary patient-derived cell lines, which maintain their specific stem-like phenotypes, have replaced established glioblastoma cell lines. However, due to heterogenous tumour growth inherent in glioblastoma, the use of primary cells for orthotopic in vivo studies often requires large experimental group sizes. Therefore, when using intracranial patient-derived xenograft (PDX) approaches, it is advantageous to deploy imaging techniques to monitor tumour growth and allow stratification of mice. Here we show that stable expression of near-infrared fluorescent protein (iRFP) in patient-derived glioblastoma cells enables rapid, direct non-invasive monitoring of tumour development without compromising tumour stemness or tumorigenicity. Moreover, as this approach does not depend on the use of agents like luciferin, which can cause variability due to changing bioavailability, it can be used for quantitative longitudinal monitoring of tumour growth. Notably, we show that this technique also allows quantitative assessment of tumour burden in highly invasive models spreading throughout the brain. Thus, iRFP transduction of primary patient-derived glioblastoma cells is a reliable, cost- and time-effective way to monitor heterogenous orthotopic PDX growth.

Oncogenic properties and signaling basis of the PAX8-GLIS3 fusion gene.

Hyalinizing trabecular tumors of the thyroid are rare and mostly benign epithelial neoplasms of follicular cell origin, which have recently been shown to be underpinned by the PAX8-GLIS3 fusion gene. In our study, we sought to investigate the potential oncogenic mechanisms of the PAX8-GLIS3 fusion gene. Forced expression of PAX8-GLIS3 was found to increase proliferation, clonogenic potential and migration of human nonmalignant thyroid (Nthy-ori 3-1) and embryonic kidney (HEK-293) cells. Moreover, in xenografts, Nthy-ori 3-1 PAX8-GLIS3 expressing cells generated significantly larger and more proliferative tumors compared to controls. These oncogenic effects were found to be mediated through activation of the Sonic Hedgehog (SHH) pathway. Targeting of smoothened (SMO), a key protein in the SHH pathway, using the small molecule inhibitor Cyclopamine partially reversed the increased proliferation, colony formation and migration in PAX8-GLIS3 expressing cells. Our data demonstrate that the oncogenic effects of the PAX8-GLIS3 fusion gene are, at least in part, due to an increased activation of the SHH pathway.

Podoplanin promotes cancer-associated thrombosis and contributes to the unfavorable overall survival in an ectopic xenograft mouse model of oral cancer.

BACKGROUND: Podoplanin (PDPN) is a transmembrane glycoprotein that mediates tumor cell-induced platelets aggregation in different cancer types. Emerging data indicate that PDPN is a marker for poor prognosis of human oral squamous cell carcinoma (OSCC). However, the functional impacts of PDPN on cancer formation and disease progression of OSCC remain to be elucidated. METHODS: The sublines of the OECM-1 oral cancer cells with PDPN knockdown or overexpression were established. The cellular characteristics and the ability to induce platelet aggregation of these cells lines were analyzed. An ectopic xenograft animal model by inoculating cancer cells into the anterior neck region of nude mice was established to investigate the functional impact of PDPN on disease progression and cancer-associated thrombosis of OSCC. RESULTS: PDPN promoted OSCC cell migration and invasion, but had no effect on cell proliferation in vitro and tumor growth in vivo. Co-incubation of PDPN-positive (PDPN+) OSCC cells with platelets induced platelet activation and aggregation. The mice bearing PDPN+ tumor had a decrease in overall survival despite that there was no gross appearance of distant metastasis. A speckled immunofluorescence staining pattern of platelet marker mCD41 was defined in the PDPN+ tumor sections and the intensity was greater than in the PDPN-low or negative tumor sections. Co-immunofluorescence staining of the tumor sections with mCD41 and the endothelial cell marker mCD31 further demonstrated that platelet aggregates were located in the lumen of blood vessel and were also distributed intratumorally in the mice bearing PDPN+ tumors. CONCLUSIONS: These data demonstrated that PDPN expression in the cancer cells is associated with high risk of thrombosis, leading to unfavorable overall survival of the mice. This study provides new insights into the functions of PDPN in cancer-associated thrombosis and in the pathophysiology of OSCC.

Evaluation of anticancer activities of Poria cocos ethanol extract in breast cancer: In vivo and in vitro, identification and mechanism.

ETHNOPHARMACOLOGICAL RELEVANCE: Poria cocos Wolf (P. cocos), a well-known traditional East-Asian medicinal and edible fungus, is one of the most important components in Chinese medicine formulas like "Guizhi fuling wan" to treat hyperplasia of mammary glands and breast cancer. AIMING OF STUDY: In this study, we attempted to verify the anticancer efficacy of the ethanol extract of P. cocos (PC) on the breast cancer as well as to investigate its most active compound and its underlying molecular mechanism in vivo and in vitro. MATERIALS AND METHODS: The key anti-cancer components were separated and purified through chromatography and identified by spectral analyses. The in vivo anti-breast cancer efficacy and side effects of PC were evaluated in BALB/c nude mice that have been subcutaneously injected with breast cancer cells MDA-MB-231. Cytotoxicity, apoptosis and cell cycle arrest of PC were evaluated in vitro by cell viability assays and flow cytometry. The protein levels were examined via western blotting. RESULTS: Pachymic acid (PA), separated and identified as the most active compound, induced the significant cytotoxicity on breast cancer cells MDA-MB-231(IC50 value, 2.13 ± 0.24 µg/mL) and was not active against the normal breast epithelium cells MCF-10A. The in vivo experiment revealed that PC could significantly inhibit the tumor development and the final mean tumor weight of the mice in the PC group (0.51 ± 0.12g) was significantly lower than that in the model group (1.22 ± 0.45g). Notably, compared to the first-line anticancer drug cisplatin, PC showed less side effects on the function of the vital organs and the muscle strength of the mice. Among in vitro study, PC significantly inhibited the cell growth of MDA-MB-231 by inducing cell apoptosis and cell cycle arrested at G0/G1 phase in a dose-dependent manner. The expression of cell cycle-associated cyclin D1, cyclin E, CDK2, and CDK4 were downregulated, while p53 and p21 expression were upregulated following the PA treatment. In addition, PA downregulated the apoptotic regulator Bcl-2, increased the expression of pro-apoptotic protein Bax, and promoted the release of cytochrome c and the activation of cleaved caspase-3, -9 and caspase -8 via mitochondria-mediated and death receptor-mediated signaling pathways. CONCLUSION: This study verified the anticancer efficacy of PC on breast cancer in vivo and in vitro through induction of cell apoptosis and G0/G1 cell cycle arrest. The data also suggested that PA could be developed as an efficacious agent for breast cancer treatment with less side effects.