Buthionine sulfoximine sensitizes antihormone-resistant human breast cancer cells to estrogen-induced apoptosis

Introduction  

Estrogen deprivation using aromatase inhibitors is one of the standard treatments for postmenopausal women with estrogen receptor (ER)-positive breast cancer. However, one of the consequences of prolonged estrogen suppression is acquired drug resistance. Our group is interested in studying antihormone resistance and has previously reported the development of an estrogen deprived human breast cancer cell line, MCF-7:5C, which undergoes apoptosis in the presence of estradiol. In contrast, another estrogen deprived cell line, MCF-7:2A, appears to have elevated levels of glutathione (GSH) and is resistant to estradiol-induced apoptosis. In the present study, we evaluated whether buthionine sulfoximine (BSO), a potent inhibitor of glutathione (GSH) synthesis, is capable of sensitizing antihormone resistant MCF-7:2A cells to estradiol-induced apoptosis.     

Rad51 inhibition sensitizes breast cancer stem cells to PARP inhibitor in triple-negative breast cancer

Breast cancer susceptibility gene 1 (BRCA1) is a tumorsuppressor gene, and its protein BRCA1 plays a role inDNA repair [1]. BRCA1 is generally expressed in the cellsof mammary glands and other tissues, helping to repairdamaged DNA or disrupting cells when DNA cannot berepaired. When BRCA1 is mutated and cannot functionand therefore the damaged DNA cannot be repaired ontime, the risk of breast cancer will greatly increase [2]. InBRCA1-mutant tumors, the capability of DNA damagerepair is decreased, which makes tumor cells sensitiveto DNA-damaging drugs; however, high BRCA1 activityweakens the effect of these drugs [3].     

Silencing of prion protein sensitizes breast adriamycin-resistant carcinoma cells to TRAIL-mediated cell death

We investigated the relationship between the resistance to the proapoptotic action of tumor necrosis factor-related apoptosis inducing ligand (TRAIL) and cellular prion protein (PrPc) function, using the TRAIL-sensitive MCF-7 human breast adenocarcinoma cell line and two TRAIL-resistant sublines: 2101 and MCF-7/ADR. All of the cell lines tested expressed TRAIL-R1 and TRAIL-R2. TRAIL decoy receptors were not detected, suggesting that the resistance of 2101 and MCF-7/ADR cells, strongly expressing PrPc, to TRAIL-mediated cell death was independent from the expression of TRAIL receptors and death-inducing signaling complex formation. Down-regulation of PrPc by small interfering RNA increased the sensitivity of Adriamycin- and TRAIL-resistant cells to TRAIL, but not to epirubicin/Adriamycin. TRAIL-mediated apoptosis in PrPc knocked-down cells was associated with caspase processing, Bid cleavage, and Mcl-1 degradation. In addition, an increased sensitivity of apoptosis-resistant cells to TRAIL after PrPc silencing was not associated with the increased recruitment of receptors and intracellular signaling molecule to the death-inducing signaling complex. Bcl-2 expression was substantially decreased after PrPc knock-down but the levels of Bcl-X(L) and Mcl-1 were not affected. The down-regulation of Bcl-2 was concomitant with Bax delocalization. Our findings support the notion that silencing of PrPc facilitates the activation of proapoptotic Bax by down-regulation of Bcl-2 expression, thereby abolishing the resistance of breast cancer cells to TRAIL-induced apoptosis.     

Induction of cell death in antiestrogen resistant human breast cancer cells by the protein kinase CK2 inhibitor DMAT

Protein kinase CK2 is involved in cell proliferation and survival, and found overexpressed in virtually all types of human cancer, including breast cancer. We demonstrate that inhibition of CK2 with 2-dimethylamino-4,5,6,7-tetrabromo-benzimidazole (DMAT), a potent and specific CK2 inhibitor, results in caspase-mediated killing of human breast cancer cells with acquired resistance to antiestrogens, while DMAT fails to kill parental MCF-7 cells. The antiestrogen resistant breast cancer cells express reduced levels of Bcl-2 compared to MCF-7 cells. Reduced Bcl-2 protein level is also found in a tamoxifen resistant human breast tumor grown as a xenograft. We show that re-expression of Bcl-2 partially rescues antiestrogen resistant MCF-7 sublines from DMAT-induced cell death. In summary, our data suggest a novel role of CK2 in antiestrogen resistance.     

Letrozole sensitizes breast cancer cells to ionizing radiation

Introduction  

Radiotherapy (RT) is considered a standard treatment option after surgery for breast cancer. Letrozole, an aromatase inhibitor, is being evaluated in the adjuvant setting. We determined the effects of the combination of RT and letrozole in the aromatase-expressing breast tumour cell line MCF-7CA, stably transfected with the CYP19 gene.     

Functional inactivation of Rb sensitizes cancer cells to TSC2 inactivation induced cell death

We showed previously that inactivation of TSC2 induces death in cancer cells lacking the Retinoblastoma (Rb) tumor suppressor under stress conditions, suggesting that inactivation of TSC2 can potentially be used as an approach to specifically kill cancers that have lost WT Rb. As Rb is often inactivated in cancers by overexpression of cyclin D1, loss of p16^ink4a cdk inhibitor, or expression of viral oncoproteins, it will be interesting to determine if such functional inactivation of Rb would similarly sensitize cancer cells to TSC2 inactivation induced cell death. In addition, many cancers lack functional Pten, resulting in increased PI3K/Akt signaling that has been shown to modulate E2F-induced cell death. Therefore it will be interesting to test whether loss of Pten will affect TSC2 inactivation induced killing of Rb mutant cancer cells. Here, we show that overexpression of Cyclin D1 or the viral oncogene E1a sensitizes cancer cells to TSC2 knockdown induced cell death and growth inhibition. On the other hand, knockdown of p16^ink4a sensitizes cancer cells to TSC2 knockdown induced cell death in a manner that is likely dependant on serum induction of Cyclin D1 to inactivate the Rb function. Additionally, we demonstrate that loss of Pten does not interfere with TSC2 knockdown induced cell death in Rb mutant cancer cells. Together, these results suggest that TSC2 is potentially a useful target for a large spectrum of cancer types with an inactivated Rb pathway.     

Restoration of ASC expression sensitizes colorectal cancer cells to genotoxic stress-induced caspase-independent cell death

Apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), an essential component of the inflammasome complex, is frequently silenced by epigenetic methylation in many tumor cells. Here, we demonstrate that restoration of ASC expression in human colorectal cancer DLD-1 cells, in which ASC is silenced by aberrant methylation, potentiated cell death mediated by DNA damaging agent. Contrarily, ASC knockdown in HT-29 cells rendered cells less susceptible to etoposide toxicity. The increased susceptibility of ASC-expressing DLD-1 cells to genotoxic stress was independent of inflammasome or caspase activation, but partially dependent on mitochondrial ROS production and JNK activation. Thus, our data suggest that ASC expression in cancer cells is an important factor in determining their susceptibility to chemotherapy.     

乳腺癌芳香化酶抑制剂耐药的研究进展

乳腺癌是女性常见的恶性肿瘤,约70％的患者为雌激素受体(estrogen receptor,ER)和(或)孕激素受体(progesterone receptor,PR)阳性,内分泌治疗是激素受体(hormone receptor,HR)阳性乳腺癌的主要治疗方式之一.近几十年来,内分泌治疗药物不断发展并应用于临床,乳腺癌...     

Targeting triple-negative breast cancer cells with the histone deacetylase inhibitor panobinostat

Introduction

Of the more than one million global cases of breast cancer diagnosed each year, approximately fifteen percent are characterized as triple-negative, lacking the estrogen, progesterone, and Her2/neu receptors. Lack of effective therapies, younger age at onset, and early metastatic spread have contributed to the poor prognoses and outcomes associated with these malignancies. Here, we investigate the ability of the histone deacetylase inhibitor panobinostat (LBH589) to selectively target triple-negative breast cancer (TNBC) cell proliferation and survival in vitro and tumorigenesis in vivo.

Methods

TNBC cell lines MDA-MB-157, MDA-MB-231, MDA-MB-468, and BT-549 were treated with nanomolar (nM) quantities of panobinostat. Relevant histone acetylation was verified by flow cytometry and immunofluorescent imaging. Assays for trypan blue viability, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) proliferation, and DNA fragmentation were used to evaluate overall cellular toxicity. Changes in cell cycle progression were assessed with propidium iodide flow cytometry. Additionally, qPCR arrays were used to probe MDA-MB-231 cells for panobinostat-induced changes in cancer biomarkers and signaling pathways. Orthotopic MDA-MB-231 and BT-549 mouse xenograft models were used to assess the effects of panobinostat on tumorigenesis. Lastly, flow cytometry, ELISA, and immunohistochemical staining were applied to detect changes in cadherin-1, E-cadherin (CDH1) protein expression and the results paired with confocal microscopy in order to examine changes in cell morphology.

Results

Panobinostat treatment increased histone acetylation, decreased cell proliferation and survival, and blocked cell cycle progression at G2/M with a concurrent decrease in S phase in all TNBC cell lines. Treatment also resulted in apoptosis induction at 24 hours in all lines except the MDA-MB-468 cell line. MDA-MB-231 and BT-549 tumor formation was significantly inhibited by panobinostat (10 mg/kg/day) in mice. Additionally, panobinostat up-regulated CDH1 protein in vitro and in vivo and induced cell morphology changes in MDA-MB-231 cells consistent with reversal of the mesenchymal phenotype.

Conclusions

This study revealed that panobinostat is overtly toxic to TNBC cells in vitro and decreases tumorigenesis in vivo. Additionally, treatment up-regulated anti-proliferative, tumor suppressor, and epithelial marker genes in MDA-MB-231 cells and initiated a partial reversal of the epithelial-to-mesenchymal transition. Our results demonstrate a potential therapeutic role of panobinostat in targeting aggressive triple-negative breast cancer cell types.     

A sphingosine kinase inhibitor induces cell death in temozolomide resistant glioblastoma cells

Purpose  Sphingosine kinase is an oncogene that is up-regulated in several solid tumors. The product of the sphingosine kinase activity, sphingosine-1-phosphate is a potent mitogen involved in diverse cell processes such as cell survival and migration. Current standard therapy in the treatment of glioblastoma multiforme (GBM) is a combination of surgery, radiation, and chemotherapy using the drug temozolomide (TMZ). However, virtually all tumors become resistant to TMZ. Therefore, new drug targets are necessary. In this study, we investigated whether inhibiting sphingosine kinase could induce cell death in TMZ-resistant GBM cells. Methods  To study TMZ resistance in vitro, we have generated TMZ-resistant cell lines from established GBM cells. We used a potent inhibitor of sphingosine kinase to study its effect on colony formation and cell growth in GBM cells with a limited dilution and WST assay. Moreover, cell death was determined by measuring caspase-3 activity using flow cytometry. Results  A sphingosine kinase inhibitor reduced cell colony formation and activated caspase-3 in both TMZ-sensitive and resistant GBM cells. Conclusion  Addition of a sphingosine kinase inhibitor to the standard chemotherapy regimen against GBM may be beneficial.     

Targeting immunogenic cell death in cancer

Immunogenic cell death (ICD) is a type of cancer cell death triggered by certain chemotherapeutic drugs, oncolytic viruses, physicochemical therapies, photodynamic therapy, and radiotherapy. It involves the activation of the immune system against cancer in immunocompetent hosts. ICD comprises the release of damage‐associated molecular patterns (DAMPs) from dying tumor cells that result in the activation of tumor‐specific immune responses, thus eliciting long‐term efficacy of anticancer drugs by combining direct cancer cell killing and antitumor immunity. Remarkably, subcutaneous injection of dying tumor cells undergoing ICD has been shown to provoke anticancer vaccine effects in vivo. DAMPs include the cell surface exposure of calreticulin (CRT) and heat‐shock proteins (HSP70 and HSP90), extracellular release of adenosine triphosphate (ATP), high‐mobility group box‐1 (HMGB1), type I IFNs and members of the IL‐1 cytokine family. In this review, we discuss the cell death modalities connected to ICD, the DAMPs exposed during ICD, and the mechanism by which they activate the immune system. Finally, we discuss the therapeutic potential and challenges of harnessing ICD in cancer immunotherapy.

Abbreviations

ATP

adenosine triphosphate

BAK

BCL‐2 homologous antagonist killer

BAX

BCL‐2‐associated X protein

BCL‐2

B‐cell lymphoma 2

BID

BH3‐interacting domain death agonist

c‐FLIP

cellular FLICE‐like inhibitory protein

cGAMP

cyclic guanosine monophosphate–adenosine monophosphate

cGAS

cyclic GMP‐AMP synthase

CRT

calreticulin

CXCL10

chemokine C‐X‐C motif ligand 10

DAMPs

damage‐associated molecular patterns

DCs

dendritic cells

DISC

death‐inducing signaling complex

ER

endoplasmic reticulum

FADD

FAS‐associated protein with death domain

FASL

FAS ligand

GSDMD

gasdermin D

GSDMD^NT

N‐terminal fragment of gasdermin D

GSDME

gasdermin E

HMGB1

high‐mobility group box‐1

HSP

heat‐shock proteins

Hyp‐PDT

hypericin‐based photodynamic therapy

ICD

immunogenic cell death

IFN

interferon

IFNAR

IFN‐α and IFN‐β receptors

IL

interleukin

IRF3

interferon regulatory factor 3

ISGs

IFN‐stimulated genes

LPS

lipopolysaccharide

MAPK

mitogen‐activated protein kinase

MHC

major histocompatibility complex

MLKL

mixed‐lineage kinase‐like

MOMP

mitochondrial outer membrane permeabilization

mtDNA

mitochondrial DNA

NF‐κB

nuclear factor kappa‐light‐chain‐enhancer of activated B cells

NK cells

natural killer cells

NLR

NOD‐like receptor

NLRP3

NOD‐like receptor family, pyrin domain‐containing 3 protein

P2RX7

purinergic receptor P2X 7

PD‐L1

programmed death ligand

PRRs

pattern recognition receptors

PS

phosphatidyl serine

RCD

regulated cell death

RIPK1

receptor‐interacting serine/threonine protein kinase 1

RIPK3

receptor‐interacting serine/threonine protein kinase 3

ROS

reactive oxygen species

STING

stimulator of interferon genes

tBID

truncated form of BID

TBK1

TANK‐binding kinase 1

TLR

Toll‐like receptor

TNF

tumor necrosis factor

TRAIL

TNF‐related apoptosis‐inducing ligand

ZBP

Z‐DNA‐binding protein

     

Targeting pro-apoptotic trail receptors sensitizes HeLa cervical cancer cells to irradiation-induced apoptosis

PURPOSE: To investigate the potential of irradiation in combination with drugs targeting the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) death receptor (DR)4 and DR5 and their mechanism of action in a cervical cancer cell line. METHODS AND MATERIALS: Recombinant human TRAIL (rhTRAIL) and the agonistic antibodies against DR4 and DR5 were added to irradiated HeLa cells. The effect was evaluated with apoptosis and cytotoxicity assays and at the protein level. Membrane receptor expression was measured with flow cytometry. Small-interfering RNA against p53, DR4, and DR5 was used to investigate their function on the combined effect. RESULTS: rhTRAIL and the agonistic DR4 and DR5 antibodies strongly enhanced 10-Gy-induced apoptosis. This extra effect was 22%, 23%, and 29% for rhTRAIL, DR4, and DR5, respectively. Irradiation increased p53 expression and increased the membrane expression of DR5 and DR4. p53 suppression, as well as small-interfering RNA against DR5, resulted in a significant downregulation of DR5 membrane expression but did not affect apoptosis induced by irradiation and rhTRAIL. After small-interfering RNA against DR4, rhTRAIL-induced apoptosis and the additive effect of irradiation on rhTRAIL-induced apoptosis were abrogated, implicating an important role for DR4 in apoptosis induced through irradiation in combination with rhTRAIL. CONCLUSION: Irradiation-induced apoptosis is strongly enhanced by targeting the pro-apoptotic TRAIL receptors DR4 or DR5. Irradiation results in a p53-dependent increase in DR5 membrane expression. The sensitizing effect of rhTRAIL on irradiation in the HeLa cell line is, however especially mediated through the DR4 receptor.     

Prospects of neoadjuvant aromatase inhibitor therapy in breast cancer

Over 40% of breast cancer patients are older than 70 years of age. In the last three decades, several clinical trials have attempted to determine the most effective and well-tolerated systemic treatments, especially for elderly women. Tamoxifen had been the standard adjuvant endocrine treatment for postmenopausal women with hormone-sensitive breast cancer, but recent trials have shown advantages in using third-generation aromatase inhibitors. Patients with large, operable or locally advanced breast cancer are often treated initially with neoadjuvant therapy to reduce their tumor size and allow for breast-conserving surgery. Neoadjuvant chemotherapy has been widely studied, but little has been published on neoadjuvant endocrine therapy. This article reviews the use of neoadjuvant aromatase inhibitors in postmenopausal women with hormone-sensitive breast cancer.