Inhibitors of apoptosis proteins (IAPs) as potential molecular targets for therapy of hematological malignancies

Apoptosis, a programmed cell death, plays a key role in the regulation of tissue homeostasis. However, impairment of its regulation may promote formation and progression of malignancy. An important part of the apoptotic machinery are the inhibitor of apoptosis protein (IAP) family, regulating caspase activity, cell division or cell survival pathways through binding to their baculovirus AIP repeat (BIR) domains and/or by their ubiquitin-ligase RING zinc finger (RZF) activity. The following IAPs have been described so far: NAIP (neuronal apoptosis inhibitory protein; BIRC1), cIAP1 and cIAP2 (cellular inhibitor of apoptosis 1 and 2; BIRC2 and BIRC3, respectively), XIAP (X-chromosome binding IAP; BIRC4), survivin (BIRC5), BRUCE (Apollon; BIRC6), livin (BIRC7) and Ts-IAP (testis-specific IAP; BIRC8). Several studies suggested a potential contribution of IAPs to oncogenesis and resistance to anti-tumor treatment. Increased IAP expression was found in variety of human cancers, including hematological malignancies, such as leukemias and B-cell lymphomas. A correlation between the progression of those diseases and high levels of survivin or XIAP has been reported. Overexpression of XIAP in acute myeloid leukemia or survivin in acute lymphoblastic leukemia and diffuse large B-cell lymphoma have been indicated as an unfavorable prognostic factors. Elevated cellular levels of cIAP1, cIAP2, XIAP and survivin correlated with a progressive course of chronic lymphocytic leukemia. Thus, targeting IAPs with small-molecule inhibitors by their antisense approaches or natural IAP antagonist mimetics, may be an attractive strategy of anti-cancer treatment. Such agents can either directly induce apoptosis of tumor cells or sensitize them to other cytotoxic agents, hence overcoming drug-resistance. This review demonstrates the current knowledge on IAP molecular biology, as well as the mechanisms of action and the development of IAP-targeting agents for treatment of hematological malignancies.     

SRSF1和BIRC5在胃癌组织中的表达及临床意义

目的通过对丝氨酸/精氨酸富有剪接因子1(SRSF1)和凋亡抑制因子(BIRC5)在胃癌组织中表达的检测及两者之间相关性的研究,来探讨两者与胃癌的发生、发展之间的关系;为胃癌的转移及复发预警,以及预后评估提供新靶点。方法应用逆转录-聚合酶链式反应(RT-PCR)和蛋白质印迹(Western blotting)检测60例术中切除的胃癌肿组织以及对应的癌旁组织中SRSF1与BIRC5的表达水平,分析这两种蛋白的表达水平与临床病理学特征的关系。结果 (1)胃癌肿组织中SRSF1 mRNA和BIRC5mRNA表达情况:SRSF1阳性表达率(75.0%,45/60)高于癌旁组织中(8.3%,5/60);BIRC5阳性表达率(81.7%,49/60)高于癌旁组织中(11.7%,7/60)。胃癌肿组织中SRSF1mRNA与BIRC5mRNA的表达与患者的年龄、性别、肿瘤大小等因素无关,与胃癌肿组织的分化程度、淋巴结转移以及TNM分期有关;SRSF1和BIRC5 mRNA在胃癌肿组织中的阳性表达呈正相关(r=0.3496,P=0.0062)。(2)胃癌肿组织中SRSF1蛋白与BIRC5蛋白表达情况:胃癌肿组织中SRSF1蛋白表达量相较于其在癌旁组织中的表达量明显增多(t=44.87,P0.01),BIRC5蛋白表达量较癌旁组织同样异常增高(t=69.84,P0.01)。结论在胃癌肿组织中SRSF1和BIRC5表达均存在异常增高,与胃癌的发生发展存在正相关,这可能与SRSF1选择性剪接BIRC5片段,促进凋亡抑制基因BIRC5的高表达有关,具体机制还需进一步探讨。     

Polo-like kinase and survivin are esophageal tumor-specific promoters

For developing successful cancer gene therapy strategies, tumor-specific gene delivery is essential. In this study, we used esophageal cancer (EC) cells to identify and evaluate esophageal tumor-specific gene promoters. Four genes (polo-like kinase-1/PLK, survivin/BIRC5, karyopherin alpha 2/KPNA2, and pituitary tumor transforming gene protein 1/PTTG1) were identified by a microarray analysis as highly expressed in EC cell lines vs. five normal organ tissues (liver, lung, kidney, brain, and heart). By quantitative RT-PCR, the average mRNA expression levels of these four genes in 20 primary ECs were 2.7-fold (PLK), 6.1-fold (survivin), 2.6-fold (KPNA2), and 2.4-fold (PTTG1) higher than that of each gene in 24 different normal organs. By dual luciferase assay, the promoter activity of PLK and survivin in EC cell lines was 18.9-fold and 28.5-fold higher, respectively, than in normal lung and renal cells. The promoters of PLK and survivin could be useful tools for developing EC-specific gene therapy vectors.     

High-Throughput Screening of Myxoid Liposarcoma Cell Lines: Survivin Is Essential for Tumor Growth

Myxoid liposarcoma (MLS) is a soft tissue sarcoma characterized by a recurrent t(12;16) translocation. Although tumors are initially radio- and chemosensitive, the management of inoperable or metastatic MLS can be challenging. Therefore, our aim was to identify novel targets for systemic therapy. We performed an in vitro high-throughput drug screen using three MLS cell lines (402091, 1765092, DL-221), which were treated with 273 different drugs at four different concentrations. Cell lines and tissue microarrays were used for validation. As expected, all cell lines revealed a strong growth inhibition to conventional chemotherapeutic agents, such as anthracyclines and taxanes. A good response was observed to compounds interfering with Src and the mTOR pathway, which are known to be affected in these tumors. Moreover, BIRC5 was important for MLS survival because a strong inhibitory effect was seen at low concentration using the survivin inhibitor YM155, and siRNA for BIRC5 decreased cell viability. Immunohistochemistry revealed abundant expression of survivin restricted to the nucleus in all 32 tested primary tumor specimens. Inhibition of survivin in 402-91 and 1765-92 by YM155 increased the percentage S-phase but did not induce apoptosis, which warrants further investigation before application in the treatment of metastatic MLS. Thus, using a 273-compound drug screen, we confirmed previously identified targets (mTOR, Src) in MLS and demonstrate survivin as essential for MLS survival.     

Survivin is regulated by interleukin‐4 in colon cancer stem cells

Colorectal cancer has provided an important model to test the stem cell hypothesis of cancer origin, which implies that cancer arises as a result of genetic aberrations in stem cells leading to deregulation of the proliferation/differentiation balance. We and others have demonstrated that, similarly to other solid tumors, colon carcinogenesis and progression are dictated by highly apoptosis‐resistant stem‐like cells. Our data have suggested that protection from apoptosis is achieved by autocrine production of interleukin‐4 (IL‐4) through up‐regulation of anti‐apoptotic mediators. In this study, we extend our analysis to another apoptosis inhibitor widely expressed in tumors, namely survivin (also known as BIRC‐5, baculoviral IAP repeat‐containing protein 5). We show that this protein, with important roles in cell death counteraction and mitotic progression control, is regulated by the IL‐4 pathway in colon rectal cancer stem cells (CR‐CSC). Hence, the presence of IL‐4 increases survivin levels in our model while cytokine neutralization has opposing effects. Treatment with cytokine neutralizing agent or with leflunomide, Stat6 inhibitor, have similar consequences on survivin localization, increasing its nuclear pool, an observation known to be correlated with a good prognosis in colon cancer patients. These results demonstrate that IL‐4, through activation of the STAT‐6 signaling pathway, is involved in survivin expression levels as well as its localization. These findings shed more light on the molecular mechanisms involved in IL‐4‐mediated chemoresistance. J. Cell. Physiol. 225: 555–561, 2010. © 2010 Wiley‐Liss, Inc.     

Functional polymorphism -31C/G in the promoter of BIRC5 gene and risk of nasopharyngeal carcinoma among chinese

Background

Baculoviral inhibitor of apoptosis repeat-containing 5 (BIRC5, also called as survivin) is a member of the inhibitor of apoptosis protein (IAP) family, which plays an important role in the occurrence and progression of cancer. Recently, a polymorphism in the promoter of BIRC5, -31C/G (rs9904341), was shown to influence BIRC5 expression.

Methods

We examined whether the -31C/G was related to the risk of developing nasopharyngeal carcinoma (NPC) in a case-control population from Guangxi province in southern China, which consists of 855 patients with NPC and 1036 controls. This polymorphism was genotyped by TaqMan assay. The genetic associations with the occurrence and progression of NPC were estimated by logistic regression.

Results

We observed a statistically significant increased occurrence of NPC associated with the CC genotype (odds ratio [OR], 1.40; 95% confidence interval [CI], 1.13–1.73; P = 0.0020) compared with the genotypes containing G allele (CG + GG genotype). However, no significant association was observed for the -31C/G with the severity of NPC (as measured by tumor-node-metastasis staging system).

Conclusion

Our findings suggest that the functional polymorphism -31C/G in the promoter of BIRC5 gene may play a role in mediating the susceptibility to NPC among Chinese.     

Survivin contributes to the anti-apoptotic activities of transforming growth factor alpha in mouse blastocysts through phosphatidylinositol 3'-kinase pathway

Transforming growth factor alpha (TGFA) is produced by epithelial cells in the oviducts and uteri and has the potential to act as an anti-apoptotic factor on preimplantation embryos expressing its receptor. Previously, we demonstrated that survivin (also known as BIRC5), an anti-apoptotic gene expressed in mouse preimplantation embryos, protects embryos from apoptosis. In this study, we investigated the role of survivin on TGFA-mediated inhibition of apoptosis in mouse blastocysts. Under the suboptimal conditions produced by single embryo culture, blastocysts showed an increase of apoptosis that correlated with a decrease of survivin expression. TGFA treatment significantly decreased apoptosis and increased the levels of survivin mRNA in a dose-dependent manner in blastocyst, and conversely, these activities were neutralized by an anti-TGFA antibody. Antibody treatment alone exerted little effect on either the occurrence of apoptosis or the levels of survivin mRNA. Upregulation of survivin expression by TGFA treatment was insignificant before the blastocyst stage. Using an antisense approach, we examined whether upregulation of survivin is responsible for the anti-apoptotic effect of TGFA in blastocysts. Apoptosis was inhibited by TGFA treatment in blastocysts, but the effect was abrogated by cotreatment with antisense oligonucleotides directed against survivin. These data suggest that survivin contributes to the anti-apoptotic activities of TGFA in blastocysts. We also found that the upregulation of survivin expression was mediated by activation of the phosphatidylinositol 3-kinase (PI3K) signaling pathway. Thus, TGFA inhibits apoptosis in mouse blastocysts through upregulation of survivin expression via the PI3K pathway.     

Association of a Polymorphism in the BIRC6 Gene with Pseudoexfoliative Glaucoma

Recently an association was observed between alleles in genes of the unfolded protein response pathway and primary open angle glaucoma (POAG). The goal of the current study is to investigate the role of these two genes, protein disulphide isomerase A member 5 (PDIA5) and baculoviral IAP repeat containing 6 (BIRC6), in different forms of glaucoma. 278 patients with POAG, 132 patients with primary angle closure glaucoma (PACG) and 135 patients with pseudoexfoliative glaucoma (PEXG) were genotyped for single nucleotide polymorphisms (SNPs) rs11720822 in PDIA5 and 471 POAG, 184 PACG and 218 PEXG patients were genotyped for rs2754511 in BIRC6. Genotyping was done by allelic discrimination PCR, and genotype and allele frequencies were calculated. Logistic regression analyses were performed using R software to determine the association of these SNPs with glaucoma. The allele and genotype frequencies of rs11720822 in PDIA5 were not associated with POAG, PACG or PEXG. The TT genotype of rs2754511 in BIRC6 was found to be protective for PEXG (p = 0.05, OR 0.42 [0.22–0.81]) in the Pakistani population, but not for POAG or PACG. This study did not confirm a previously reported association of risk alleles in PDIA5 and BIRC6 with POAG, but did demonstrate a protective role of the T allele of rs2754511 in the BIRC6 gene in PEXG. This supports a role for the unfolded protein response pathway and regulation of apoptotic cell death in the pathogenesis of PEXG.     

Survivin’s Dual Role: An Export’s View

Survivin is proposed to function as a mitotic regulator and an apoptosis inhibitor duringdevelopment and pathogenesis. As such, survivin has aroused keen interest in disparateareas of basic and translational research. Survivin acts as a subunit of the chromosomalpassenger complex (CPC), composed of the mitotic kinase Aurora-B, Borealin 5 and INCENP,and is essential for proper chromosome segregation and cytokinesis. Our recent findingsindicate that the nuclear export receptor Crm1 is critically involved in tethering the CPC to thecentromere by interacting with a leucine-rich nuclear export signal (NES), evolutionaryconserved in all mammalian survivin proteins. In addition, the survivin/Crm1 interaction10 seems to be required for the cytoprotective activity of survivin, because export deficientsurvivin fails to protect tumor cells against cancer therapy-induced apoptosis. These findingsappear of clinical relevance since preferential nuclear localization of survivin turned out to bea favorable prognostic factor in cancer patients. Besides emphasizing the functionalsignificance of the Crm1/survivin interface, we suggest to exploit the pharmacogenetic15 interference with survivin’s export as a novel strategy to antagonize survivin’s activity.