Protein phosphatase 2A activators under investigation for smoking-related chronic obstructive pulmonary disease and related disorders.

INTRODUCTION: Chronic obstructive pulmonary disease (COPD) is characterized by progressive inflammation during therapy. Cystic fibrosis (CF), alpha-one antitrypsin deficiency (AATD), and non-CF bronchiectasis are also chronic respiratory disorders with inflammation and progression that share many similarities with COPD. Therefore, various anti-inflammatory approaches are currently being investigated, and protein phosphatase 2A (PP2A) activators may represent one such approach. AREAS COVERED: Systematic review of papers published from 2000-to date on the anti-inflammatory role of endogenous PP2A, the consequences of its inhibition by smoking, and the beneficial effects of its activation in COPD. EXPERT OPINION: PP2A activation is a plausible therapeutic approach in COPD and related disorders, such as CF, AATD, and non-CF bronchiectasis, although the available evidence is still mostly experimental. Metformin repurposing and consideration of inhalation for some of the molecules discussed in this study are promising approaches.

KRAS-mediated upregulation of CIP2A promotes suppression of PP2A-B56α to initiate pancreatic cancer development.

Oncogenic mutations in KRAS are present in approximately 95% of patients diagnosed with pancreatic ductal adenocarcinoma (PDAC) and are considered the initiating event of pancreatic intraepithelial neoplasia (PanIN) precursor lesions. While it is well established that KRAS mutations drive the activation of oncogenic kinase cascades during pancreatic oncogenesis, the effects of oncogenic KRAS signaling on regulation of phosphatases during this process is not fully appreciated. Protein Phosphatase 2A (PP2A) has been implicated in suppressing KRAS-driven cellular transformation. However, low PP2A activity is observed in PDAC cells compared to non-transformed cells, suggesting that suppression of PP2A activity is an important step in the overall development of PDAC. In the current study, we demonstrate that KRASG12D induces the expression of both an endogenous inhibitor of PP2A activity, Cancerous Inhibitor of PP2A (CIP2A), and the PP2A substrate, c-MYC. Consistent with these findings, KRASG12D sequestered the specific PP2A subunit responsible for c-MYC degradation, B56α, away from the active PP2A holoenzyme in a CIP2A-dependent manner. During PDAC initiation in vivo, knockout of B56α promoted KRASG12D tumorigenesis by accelerating acinar-to-ductal metaplasia (ADM) and the formation of PanIN lesions. The process of ADM was attenuated ex vivo in response to pharmacological re-activation of PP2A utilizing direct small molecule activators of PP2A (SMAPs). Together, our results suggest that suppression of PP2A-B56α through KRAS signaling can promote the MYC-driven initiation of pancreatic tumorigenesis.

Sideroflexin-1 promotes progression and sensitivity to lapatinib in triple-negative breast cancer by inhibiting TOLLIP-mediated autophagic degradation of CIP2A.

Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer and it lacks specific therapeutic targets and effective treatment protocols. By analyzing a proteomic TNBC dataset, we found significant upregulation of sideroflexin 1 (SFXN1) in tumor tissues. However, the precise function of SFXN1 in TNBC remains unclear. Immunoblotting was performed to determine SFXN1 expression levels. Label-free quantitative proteomics and liquid chromatography-tandem mass spectrometry were used to identify the downstream targets of SFXN1. Mechanistic studies of SFXN1 and cellular inhibitor of PP2A (CIP2A) were performed using immunoblotting, immunofluorescence staining, and reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Functional experiments were used to investigate the role of SFXN1 in TNBC cells. SFXN1 was significantly overexpressed in TNBC tumor tissues and was associated with unfavorable outcomes in patients with TNBC. Functional experiments demonstrated that SFXN1 promoted TNBC growth and metastasis in vitro and in vivo. Mechanistic studies revealed that SFXN1 promoted TNBC progression by inhibiting the autophagy receptor TOLLIP (toll interacting protein)-mediated autophagic degradation of CIP2A. The pro-tumorigenic effect of SFXN1 overexpression was partially prevented by lapatinib-mediated inhibition of the CIP2A/PP2A/p-AKT pathway. These findings may provide a new targeted therapy for patients with TNBC.

Loss of function variant in CIP2A associated with female infertility with early embryonic arrest and fragmentation.

BACKGROUND: Early embryonic arrest and fragmentation (EEAF) is a common cause of female infertility, but the genetic causes remain to be largely unknown. CIP2A encodes the cellular inhibitor of PP2A, playing a crucial role in mitosis and mouse oocyte meiosis. METHODS: Exome sequencing and Sanger sequencing were performed to identify candidate causative genes in patients with EEAF. The pathogenicity of the CIP2A variant was assessed and confirmed in cultured cell lines and human oocytes through Western blotting, semi-quantitative RT-PCR, TUNEL staining, and fluorescence localization analysis. FINDINGS: We identified CIP2A (c.1510C > T, p.L504F) as a novel disease-causing gene in human EEAF from a consanguineous family. L504 is highly conserved throughout evolution. The CIP2A variant (c.1510C > T, p.L504F) reduced the expression level of the mutant CIP2A protein, leading to the abnormal aggregation of mutant CIP2A protein and cell apoptosis. Abnormal aggregation of CIP2A protein and chromosomal dispersion occurred in the patient's oocytes and early embryos. We further replicated the patient phenotype by knockdown CIP2A in human oocytes. Additionally, CIP2A deficiency resulted in decreased levels of phosphorylated ERK1/2. INTERPRETATION: We first found that the CIP2A loss-of-function variant associate with female infertility characterized by EEAF. Our findings suggest the uniqueness and importance of CIP2A gene in human oocyte and early embryo development. FUNDING: This work was supported by National Key Research and Development Program of China (2023YFC2706302), the National Natural Science Foundation of China (81000079, 81170165, and 81870959), the HUST Academic Frontier Youth Team (2016QYTD02), and the Key Research of Huazhong University of Science and Technology, Tongji Hospital (2022A20).

Chronic stress induces Alzheimer's disease-like pathologies through DNA damage-Chk1-CIP2A signaling.

Stress is an important initiating factor in promoting Alzheimer's disease (AD) pathogenesis. However, the mechanism by which stress induces AD-like cognitive impairment remains to be clarified. Here, we demonstrate that DNA damage is increased in stress hormone Corticotropin-releasing factor (CRF)-treated cells and in brains of mice exposed to chronic restraint stress. Accumulation of DNA damage drives activation of cell cycle checkpoint protein kinase 1 (Chk1), upregulation of cancerous inhibitor of PP2A (CIP2A), tau hyperphosphorylation, and Aß overproduction, eventually resulting in synaptic impairment and cognitive deficits. Pharmacological intervention targeting Chk1 by specific inhibitor and DNA damage by vitamin C, suppress DNA damage-Chk1-CIP2A signaling pathway in chronic stress animal model, which in turn attenuate AD-like pathologies, synaptic impairments and cognitive deficits. Our study uncovers a novel molecular mechanism of stress-induced AD-like pathologies and provides effective preventive and therapeutic strategies targeting this signaling pathway.

ent-8(14),15-Pimaradiene-2ß,19-diol, a diterpene from Aleuritopteris albofusca, inhibits growth and induces protective autophagy in hepatocellular carcinoma cells.

A new pimarane-type diterpene, ent-8(14),15-pimaradiene-2ß,19-diol (JXE-23), was isolated from the fern plant Aleuritopteris albofusca by our previous work; however, the biological activity of this diterpene remains unclear. In the present study, the anti-cancer potential of JXE-23 in various cancer cells was investigated. Among MCF-7 breast cancer cells, A549 lung cancer cells, and HepG2 liver cancer cells, JXE-23 displayed significant cytotoxicity to HepG2 cells with an IC50 value of 17.20 ± 1.73 µM, while showing no obvious toxicity in normal hepatocytes HL7702. JXE-23 inhibited cell growth and colony formation in HepG2 cells. A cell cycle distribution analysis showed that JXE-23 caused G2/M cell cycle arrest. Besides, JXE-23 also suppressed the migration of HepG2 cells. Interestingly, an increase of light chain 3 II (LC3II) and Beclin 1 and a decrease of P62 have occurred in JXE-23-treated cells, as well as the formation of GFP-LC3 dots, indicative of autophagy induction by JXE-23. When combined with autophagy inhibitor 3-methyladenine and chloroquine, the cell viability was significantly reduced, suggesting that JXE-23 triggered protective autophagy in hepatoma cells. Further study showed that JXE-23 inactivated the CIP2A/p-AKT/c-Myc signaling axis in HepG2 cells. Our data provided evidence that JXE-23 inhibited cell growth, arrested cells at the G2/M phase, and induced protective autophagy in HepG2 hepatocellular carcinoma cells. JXE-23 may be a potential lead compound for anti-cancer drug development, and autophagy inhibitor treatment may provide an effective strategy for improving its anti-cancer effect.

Celastrol impairs tumor growth by modulating the CIP2A-GSK3ß-MCL-1 axis in gastric cancer cells.

BACKGROUND/AIM: High Cancerous Inhibitor of PP2A (CIP2A) expression has been reported in solid and hematologic malignancies and is inversely associated with prognosis in Gastric Cancer, the non-small cell lung cancer, et al. CIP2A can be a drug target for the development of novel anti-gastric cancer agent. Our study was designed to explore the anti-cancer effect of celastrol, a small natural compound, and whether it has an anti-proliferative effect through inducing CIP2A degradation against gastric cancer cells. MATERIALS AND METHODS: Employing human gastric cancer cells AGS and BCG-823 cells, the effects of celastrol on cell proliferation, apoptosis and cell cycle was specifically investigated via Annexin V-FITC/PI staining and CCK8 assay. The functional association between celastrol and CIP2A was evaluated by using CIP2A knockdown and overexpression technique. The mechanism of underlying celastrol-triggering anti-gastric cancer effect was detected by real-time PCR and western blot analysis. RESULTS: Celastrol concentration- and time-dependently induced CIP2A degradation and led to gastric cancer cell apoptosis. More in depth studies revealed specific activation of Protein phosphatase 2A (PP2A)-GSK3ß-MCL-1 signaling pathway was involved in pro-apoptosis effect of celastrol, due to celastrol-triggering degradation of CIP2A, which mainly suppressed PP2A activity. CONCLUSION: Our findings highlight that celastrol has therapeutic potential via inducing apoptosis of gastric cancer cells.

Endogenous PP2A inhibitor CIP2A degradation by chaperone-mediated autophagy contributes to the antitumor effect of mitochondrial complex I inhibition.

Combined inhibition of oxidative phosphorylation (OXPHOS) and glycolysis has been shown to activate a PP2A-dependent signaling pathway, leading to tumor cell death. Here, we analyze highly selective mitochondrial complex I or III inhibitors in vitro and in vivo to elucidate the molecular mechanisms leading to cell death following OXPHOS inhibition. We show that IACS-010759 treatment (complex I inhibitor) induces a reactive oxygen species (ROS)-dependent dissociation of CIP2A from PP2A, leading to its destabilization and degradation through chaperone-mediated autophagy. Mitochondrial complex III inhibition has analogous effects. We establish that activation of the PP2A holoenzyme containing B56δ regulatory subunit selectively mediates tumor cell death, while the arrest in proliferation that is observed upon IACS-010759 treatment does not depend on the PP2A-B56δ complex. These studies provide a molecular characterization of the events subsequent to the alteration of critical bioenergetic pathways and help to refine clinical studies aimed to exploit metabolic vulnerabilities of tumor cells.

cAMP/PKA signaling promotes AKT deactivation by reducing CIP2A expression, thereby facilitating decidualization.

cAMP signaling is widely known to be indispensable for decidualization, but the details are not fully understood. Here, we show that cAMP signaling promotes AKT deactivation in endometrial stromal cells, which favors their decidualization. The deactivation of AKT is found to be a consequence of the reduced expression of several inhibitors of PP2A, the major phosphatase of AKT, with CIP2A being the most prominent. CIP2A reduction is obligatory for decidualization, as persistent CIP2A expression impairs chromatin remodeling and the expression of several decidualization markers (IGFBP1, PRL, MAOA, and IL-15). Furthermore, analyses of the responsiveness of the CIP2A promoter to cAMP signaling suggest the ETS family to be a bridge between cAMP signaling and CIP2A reduction. Our results provide novel insights into the role of cAMP signaling in decidualization and might benefit the development of novel therapies for decidualization deficiency, AKT-driven tumors, and the reverse, insulin resistance.

Cancer progression by the okadaic acid class of tumor promoters and endogenous protein inhibitors of PP2A, SET and CIP2A.

PURPOSE: Okadaic acid class of tumor promoters are transformed into endogenous protein inhibitors of PP2A, SET, and CIP2A in human cancers. This indicates that inhibition of PP2A activity is a common mechanism of cancer progression in humans. It is important to study the roles of SET and CIP2A vis-à-vis their clinical significance on the basis of new information gathered from a search of PubMed. RESULTS AND DISCUSSION: The first part of this review introduces the carcinogenic roles of TNF-α and IL-1, which are induced by the okadaic acid class of compounds. The second part describes unique features of SET and CIP2A in cancer progression for several types of human cancer: (1) SET-expressing circulating tumor cells (SET-CTCs) in breast cancer, (2) knockdown of CIP2A and increased PP2A activity in chronic myeloid leukemia, (3) CIP2A and epidermal growth factor receptor (EGFR) activity in erlotinib sensitive- and resistant-non-small cell lung cancer, (4) SET antagonist EMQA plus radiation therapy against hepatocellular carcinoma, (5) PP2A inactivation as a common event in colorectal cancer, (6) prostate cancer susceptibility variants, homeobox transcription factor (HOXB13 T) and CIP2A T, and (7) SET inhibitor OP449 for pre-clinical investigation of pancreatic cancer. In the Discussion, the binding complex of SET is briefly introduced, and overexpression of SET and CIP2A proteins is discussed in relation to age-associated chronic inflammation (inflammaging). CONCLUSION: This review establishes the concept that inhibition of PP2A activity is a common mechanism of human cancer progression and activation of PP2A activity leads to effective anticancer therapy.

The TT Genotype of the KIAA1524 rs2278911 Polymorphism Is Associated with Poor Prognosis in Multiple Myeloma.

BACKGROUND: The KIAA1524 gene encodes an oncoprotein, CIP2A, which inhibits the phosphorylation of the Akt kinase B, stabilizes the c-Myc protein, and, through that, promotes cancerogenesis. An increase in CIP2A expression has been observed in numerous solid tumors and hematologic malignancies, including multiple myeloma (MM). The aim of our study was to evaluate the clinical impact of the functional single nucleotide polymorphisms (SNP) of the KIAA1524 gene (rs2278911, 686C > T) in MM patients. METHODS: The study group consisted of 128 patients with de novo MM. EDTA venous blood samples were collected prior to the treatment. The SNPs were analyzed by Real-Time PCR with the use of specific Taqman probes. RESULTS: Multivariable analysis revealed that variables independently associated with shorter progression-free survival (PFS) included thrombocytopenia, delTP53 and IGH/CCND1 translocation and the TT genotype of the KIAA1524 gene (686C > T) (median PFS: 6 vs. 25 months; HR = 7.18). On the other hand, autologous haematopoietic stem cell transplantation (AHSCT) was related to a lower risk of early disease progression. Moreover, light chain disease, International Staging System (ISS) 3, poor performance status, hypoalbuminemia, IGH/FGFR3 translocation and the TT genotype of the KIAA1524 gene (686C > T) were independent prognostic factors associated with shorter overall survival (OS) (median OS: 8 vs. 45 months; HR = 7.08). CONCLUSION: The evaluation of the SNP 686C > T of the KIAA1524 gene could be used as a diagnostic tool in MM patients at risk of early disease progression and death.

From Basic Science to Clinical Practice: The Role of Cancerous Inhibitor of Protein Phosphatase 2A (CIP2A)/p90 in Cancer.

Cancerous inhibitor of protein phosphatase 2A (CIP2A), initially reported as a tumor-associated antigen (known as p90), is highly expressed in most solid and hematological tumors. The interaction of CIP2A/p90, protein phosphatase 2A (PP2A), and c-Myc can hinder the function of PP2A toward c-Myc S62 induction, thus stabilizing c-Myc protein, which represents a potential role of CIP2A/p90 in tumorigeneses such as cell proliferation, invasion, and migration, as well as cancer drug resistance. The signaling pathways and regulation networks of CIP2A/p90 are complex and not yet fully understood. Many previous studies have also demonstrated that CIP2A/p90 can be used as a potential therapeutic cancer target. In addition, the autoantibody against CIP2A/p90 in sera may be used as a promising biomarker in the diagnosis of certain types of cancer. In this Review, we focus on recent advances relating to CIP2A/p90 and their implications for future research.

Oct4 reduction contributes to testicular injury of unilateral testicular torsion in mice model and apoptotic death of Sertoli cells through mediating CIP2A expression.

This study explored the mechanism of ipsilateral testis injury after ipsilateral testicular torsion detorsion (T/D) and the potential testis-protective part of the octamer-binding transcription factor 4 (Oct4)-cancerous inhibitors of protein phosphatase 2A (CIP2A) axis in a T/D animal model and in ischemia-reperfusion (IR)-treated testicular Sertoli TM4 cells. Quantitative Polymerase chain reaction (PCR) and western blot (WB) confirmed the downregulation of both CIP2A and Oct4 expression in the testicular tissue from T/D mice compared with sham-operated mice. T/D model was then established in mice with upregulated Oct4 expression in the testis. Oct4 elevation restored CIP2A expression in testes after T/D treatment. Furthermore, we observed that an increase in Oct4 ameliorated the testicular damage caused by torsion in the testis. Biochemical analysis indicated that T/D treatment increased serum anti-sperm antibody levels, but reduced testosterone levels. Meanwhile, in testicular tissue, reactive oxygen species (ROS), malondialdehyde (MDA), and activity of testicular myeloperoxidase (MPO) enzymes were promoted, while glutathione peroxidase activity (GPx) was decreased by T/D injury. Notably, testicular Oct4 restoration partially counteracted the effect of T/D treatment on these biochemical indices. Hypoxia/reoxygenation (HR) treatment was applied to TM4 cells to mimic TT injury in vitro. A gain-of-function study showed that Oct4 overexpression partly counteracted the promoting role of HR in cell damage, apoptosis, and oxidative stress in TM4 cells. These observations provide novel insights into the possible biochemical mechanism underlying the mediation of the Oct4-CIP2A axis in T/D injury.

Cip2a induces arginine biosynthesis and promotes tumor progression in non-small cell lung cancer.

Cancerous inhibitor of protein phosphatase 2A (Cip2a) is an oncoprotein, playing important roles in tumor progression. However, the underlying mechanisms by which Cip2a promotes tumor aggressiveness in NSCLC remain to be further investigated. In this study, we found that Cip2a expression is elevated in NSCLC and correlates with poor prognosis. Knockdown of Cip2a significantly reduced the ability of cell proliferation, invasion, and metastasis of NSCLC both in vitro and in vivo. Furthermore, we found that Cip2a promotes tumor progression partly by inducing arginine biosynthesis, and knockdown of Cip2a exhibited a significantly increased sensitivity to arginine deprivation and mTOR inhibition. In addition, we found that p53 mutants in NSCLC cells increased Cip2a expression by inhibiting the activity of wild-type p53. Our findings provide new insights into the mechanisms of Cip2a in promoting tumor progression and suggest that Cip2a represents a potential therapeutic target for treating NSCLC.

Age-related alterations in protein phosphatase 2A methylation levels in brains of cynomolgus monkeys: a pilot study.

The abnormal activity of PP2A, a dominant member of type 2A serine/threonine protein phosphatase, has been implicated in the development of Alzheimer's disease (AD) and dementia with Lewy bodies (DLB). PP2A is a holoenzyme, and protein methylation of the catalytic subunit, PP2Ac, alters the complex composition. A decrease in PP2Ac methylation levels has been reported in AD and DLB. Aging is the most common risk factor for AD and DLB, but the relationship between aging and PP2A has not been studied in detail. Cynomolgus monkey show increased phosphorylation levels of tau and α-synuclein with aging. In this study, we investigated the alterations in the PP2A activity regulation with aging in monkey brains from 2 to 43 years of age using fractionated proteins. We found that type 2A protein phosphatase activity decreased with aging in cytoplasmic and nuclear-soluble fractions. PP2Ac methylation level was decreased in cytoplasmic and sarkosyl-insoluble fractions. A principal component analysis using PP2Ac, demethylated PP2Ac and PP2A methylesterase PME-1 levels in cytoplasmic and nuclear-soluble fractions as attributes showed that aged monkeys were in the same cluster. Our results show that brain aging in cynomolgus monkeys is closely related to changes in PP2A methylation.

Oncogenic RAS promotes MYC protein stability by upregulating the expression of the inhibitor of apoptosis protein family member Survivin.

The small GTPase KRAS is frequently mutated in pancreatic cancer and its cooperation with the transcription factor MYC is essential for malignant transformation. The key to oncogenic KRAS and MYC working together is the stabilization of MYC expression due to KRAS activating the extracellular signal-regulated kinase 1/2, which phosphorylates MYC at serine 62 (Ser 62). This prevents the proteasomal degradation of MYC while enhancing its transcriptional activity. Here, we identify how this essential signaling connection between oncogenic KRAS and MYC expression is mediated by the inhibitor of apoptosis protein family member Survivin. This discovery stemmed from our finding that Survivin expression is downregulated upon treatment of pancreatic cancer cells with the KRASG12C inhibitor Sotorasib. We went on to show that oncogenic KRAS increases Survivin expression by activating extracellular signal-regulated kinase 1/2 in pancreatic cancer cells and that treating the cells either with siRNAs targeting Survivin or with YM155, a small molecule that potently blocks Survivin expression, downregulates MYC and strongly inhibited their growth. We further determined that Survivin protects MYC from degradation by blocking autophagy, which then prevents cellular inhibitor of protein phosphatase 2A from undergoing autophagic degradation. Cellular inhibitor of protein phosphatase 2A, by inhibiting protein phosphatase 2A, helps to maintain MYC phosphorylation at Ser 62, thereby ensuring its cooperation with oncogenic KRAS in driving cancer progression. Overall, these findings highlight a novel role for Survivin in mediating the cooperative actions of KRAS and MYC during malignant transformation and raise the possibility that targeting Survivin may offer therapeutic benefits against KRAS-driven cancers.

Protein Phosphatase 2a Inhibits Gastric Cancer Cell Glycolysis by Reducing MYC Signaling.

Aerobic glycolysis, also known as the Warburg effect, has emerged as a hallmark of cancer and is associated with tumor progression and unfavorable clinical outcomes in cancer patients. PP2A is a highly conserved eukaryotic serine/threonine protein phosphatase that functions as a tumor suppressor in a variety of human cancers. However, the relationship between PP2A and the Warburg effect in gastric cancer has yet to be fully understood. In this study, the expression profile of two endogenous inhibitors of PP2A, SET and CIP2A, in gastric cancer, were analyzed by real-time quantitative polymerase chain reaction. Loss-of-function and gain-of-function studies were performed to investigate the roles of PP2A in gastric cancer cell proliferation and glycolysis. Cell biological, molecular, and biochemical approaches were employed to uncover the underlying mechanisms. The results showed that SET and CIP2A were overexpressed in gastric cancer and associated with a decreased PP2A activity. Pharmacological activation of PP2A with FTY-720 and DT-061 in two gastric cancer cell lines significantly reduced gastric cancer cell proliferation and glycolytic ability. Importantly, inhibition of PP2A activity by genetic silencing of PPP2R5A resulted in a growth advantage, which can be largely compromised by the addition of the glycolysis inhibitor 2-Deoxy-D-glucose, suggesting a glycolysis-dependent effect of PP2A in gastric cancer. Mechanistically, the well-known transcription factor and glycolysis regulator c-Myc was discovered as the functional mediator of PP2A in regulating cell glycolysis. Ectopic expression of a phosphorylation-mutant c-Myc resistant to PP2A (MycT58A) restored the inhibitory effect of FTY-720 and DT-061 on lactate production and glucose uptake. Furthermore, there was a close association between SET and CIP2A expression and c-Myc gene signatures in gastric cancer samples. Collectively, this study provides strong evidence of the involvement of PP2A in the Warburg effect and indicates that it could be a novel antitumor strategy to target tumor metabolism in gastric cancer.

Exploring the composition of protein-ligand binding sites for cancerous inhibitor of PP2A (CIP2A) by inhibitor guided binding analysis: paving a new way for the Discovery of drug candidates against triple negative breast cancer (TNBC).

Triple-negative breast cancer (TNBC) is associated with high-grade invasive carcinoma leading to a 10% to 15% death rate in younger premenopausal women. Targeting cancerous inhibitors of protein phosphatase (CIP2A) has been a highly effective approach for exploring therapeutic drug candidates. Lapatinib, a dual tyrosine kinase inhibitor, has shown promising inhibition properties by inducing apoptosis in TNBC carcinogenesis in vivo. Despite knowledge of the 3D structure of CIP2A, no reports provide insight into CIP2A ligand binding sites. To this effect, we conducted in silico site identification guided by lapatinib binding. Four of the five sites identified were cross-validated, and the stem domain revealed more excellent ligand binding affinity. The binding affinity of lapatinib in these sites was further computed using the Molecular Mechanics/Poisson-Boltzmann Surface Area (MM/PBSA) approach. According to MM/PBSA//200 ns MD simulations, lapatinib exhibited a higher binding affinity against CIP2A in site 2 with ΔG critical values of -37.1 kcal/mol. The steadiness and tightness of lapatinib with CIP2A inside the stem domain disclosed glutamic acid-318 as the culprit amino acid with the highest electrostatic energy. These results provide clear information on the CIP2A domain capable of ligand binding and validate lapatinib as a promising CIP2A inhibitor in TNBC carcinogenesis.

The Pivotal Role of Protein Phosphatase 2A (PP2A) in Brain Tumors.

Protein phosphatase 2A (PP2A) is a highly complex heterotrimeric Ser/Thr phosphatase that regulates many cellular processes. PP2A is dysregulated in several human diseases, including oncological pathology; interestingly, PP2A appears to be essential for controlling cell growth and may be involved in cancer development. The role of PP2A as a tumor suppressor has been extensively studied and reviewed. To leverage the potential clinical utility of combination PP2A inhibition and radiotherapy treatment, it is vital that novel highly specific PP2A inhibitors be developed. In this review, the existing literature on the role of PP2A in brain tumors, especially in gliomas and glioblastoma (GBM), was analyzed. Interestingly, the review focused on the role of PP2A inhibitors, focusing on CIP2A inhibition, as CIP2A participated in tumor cell growth by stimulating cell-renewal survival, cellular proliferation, evasion of senescence and inhibition of apoptosis. This review suggested CIP2A inhibition as a promising strategy in oncology target therapy.

[Ethoxysanguinarine directly targets CIP2A to inhibit proliferation and induce autophagy of SGC7901/DDP cells].

This study aims to investigate the effect of ethoxysanguinarine(Eth) on cisplatin(DDP)-resistant human gastric cancer cells and decipher the underlying mechanism. The human gastric cancer cell line SGC7901 and the DDP-resistant cell line SGC7901/DDP were used as the cell models. Western blot was employed to determine the expression levels of multidrug resistance-related proteins, and methyl thiazolyl tetrazolium(MTT) assay to detect the proliferation of SGC7901 and SGC7901/DDP cells exposed to DDP. After treatment with different concentrations of Eth, the proliferation of SGC7901 and SGC7901/DDP cells was detected by MTT assay, trypan blue exclusion assay, colony formation assay, and high-content imaging and analysis system. The apoptosis of SGC7901/DDP cells was detected by flow cytometry with Annexin V-FITC/PI staining. GFP-LC3 transfection was carried out to detect the effect of Eth on the autophagy of SGC7901/DDP cells. The expression levels of the multidrug resistance-related protein P-glycoprotein(P-gp), the apoptosis-related proteins [caspase-9, caspase-3, and poly(ADP-ribose) polymerase(PARP)], the autophagy-related protein light chain 3-Ⅱ(LC3-Ⅱ), the key effectors [mammalian target of rapamycin(mTOR), 70 kDa ribosomal protein S6 kinase(P70 S6 K), and 4 E binding protein 1(4 E-BP1)] of the mammalian target of rapamycin complex 1(mTORC1) signaling pathway, cancerous inhibitor of protein phosphatase 2A(CIP2A), and protein kinase B(Akt) were measured by Western blot. The mRNA level of CIP2A in the SGC7901/DDP cells exposed to Eth for 24 h was analyzed by RT-qPCR. After SGC7901/DDP cells were transfected with CIP2A expression vector pcDNA3.1-HA-CIP2A and treated with different concentrations of Eth, MTT assay was used to determine the prolife-ration of SGC7901/DDP cells and Western blot to detect the expression levels of related proteins. The interaction sites of Eth and CIP2A were predicted by molecular docking. The affinity between Eth and CIP2A was determined by drug affinity responsive target stability(DARTS) assay. The pharmacokinetic properties and drug-like activity of Eth were predicted by SwissADME. The results indicated that SGC7901/DDP cells were more sensitive to Eth than SGC7901 cells. Eth significantly inhibited proliferation and colony formation and changed the morphology, roundness, and area of SGC7901/DDP cells. Eth treatment caused the nucleus shrinking and significantly increased the apoptosis rate of the cells. Furthermore, Eth down-regulated the expression of caspase-9 and caspase-3 precursors and promoted the cleavage of PARP, which suggested that Eth induced the apoptosis of SGC7901/DDP cells. The GFP-LC3 in Eth-treated cells showed speckled aggregation. The up-regulated expression of LC3-Ⅱ by Eth indicated that Eth activated the autophagy of SGC7901/DDP cells. Eth down-regulated the expression of P-gp, the phosphorylation of mTOR, P70 S6K, and 4E-BP1, the expression of CIP2A, and the phosphorylation of Akt. Additionally, it increased the activity of PP2A, and had no significant effect on the expression of CIP2A in SGC7901/DDP cells. CIP2A overexpression antagonized the inhibition of cell proliferation and the activation of autophagy by Eth. Molecular docking suggested that Eth bound to CIP2A. The results of DARTS assay further proved the above binding effect. Eth has potential drug-like activity. The above results demonstrated that Eth inhibited the proliferation, induced the apoptosis, and activated the autophagy of SGC7901/DDP cells by targeting CIP2A and then down-regulating PP2A/mTORC1 signaling pathway. This study provided a new target for the treatment of cisplatin-resistant gastric cancer.