CTBP1 ve NF2 Gen Ekspresyonlarının Prostat Kanser Hücrelerinde KDM6A/B Aracılı Epigenetik Regülasyonunun İncelenmesi

Amaç: Çalışmanın amacı, epigenetik regülatörlerden lizin demetilaz 6A (KDM6A) ve/veya lizin demetilaz 6B’nin (KDM6B) prostat kanseri oluşumu ve progresyonunda rol oynayan C Terminal bağlayıcı protein1 (CTBP1) ve Nörofibromatozis Tip2 (NF2) ekspresyonlarının regülasyonu üzerindeki modüle edici rolünün aydınlatılmasıdır. Yöntemler: Prostat kanseri metastatik hücreleri LNCaP’lere KDM6 ailesi selektif inhibitörü GSK-J4 uygulanıp, total RNA izolasyonu ve cDNA sentezi yapılarak; CTBP1 ve NF2 mRNA düzeylerindeki değişiklikler reverse transcription-quantitative real time PCR (RT-qPCR) ile gösterilmiştir. Saptanan değişikliklerin transkripsiyonel inhibisyondan kaynaklanıp kaynaklanmadığını araştırmak üzere CTBP1 ve NF2 pre-spliced mRNA düzeylerindeki değişiklikler RT-qPCR ile ölçülmüştür. KDM6A ve/veya KDM6B’nin (KDM6A/B) small interfering RNA (siRNA) aracılı susturulduğu hücrelerde CTBP1 ve NF2 mRNA düzeylerindeki değişiklikler RT-qPCR ile gösterilerek bu genlerin KDM6 enzimlerinin hangisi tarafından regüle edildiği belirlenmiştir. Bulgular: CTBP1 ve NF2 rölatif mRNA düzeyleri GSK-J4 ile %45 ve %49 azalmıştır. Saptanan azalmaların CTBP1 ve NF2 mRNA ekspresyonlarının GSK-J4 tarafından transkripsiyonel baskılanmasıyla ilişkili olup olmadığını belirlemek üzere yapılan RT-qPCR deneylerinde CTBP1 ve NF2 pre-spliced mRNA düzeyleri de %43, %29 azalmıştır. GSK-J4’ün KDM6 ailesine selektif bir inhibitör olması nedeniyle CTBP1 ve NF2 mRNA ekspresyonlarının hangi KDM6 enzimi tarafından kontrol edildiğini belirlemek amacıyla KDM6A ve/veya KDM6B susturulduğunda; KDM6A ve KDM6B dual inhibisyonunda CTBP1 ve NF2 mRNA düzeyleri %56, %39 azalmıştır. Sonuç: Özetle; GSK-J4 uygulanmasıyla CTBP1 ve NF2 mRNA düzeylerinde saptadığımız azalmaların transkripsiyondaki değişiklikten kaynaklandığı pre-spliced mRNA verilerimizle güçlü bir şekilde desteklenmiştir. KDM6A ve KDM6B’nin her ikisinin de CTBP1 ve NF2 ekspresyonlarının prostat kanserindeki regülasyonunda kontrol edici rollerinin olduğunun gösterilmesi; terapötik olarak hedeflenebilecek, KDM6A ve KDM6B aracılığıyla CTBP1 ve NF2 ekspresyonlarını modüle eden yeni bir mekanizmanın aydınlatılmasına katkı sağlayabilir.

Anahtar Kelimeler:

CTBP1, NF2, prostat kanseri, epigenetik, KDM6A/B

The Measurement of Neutrophil Gelatinase Associated Lipocalin in Umbilical Cord Blood and the Assessment of Its Relationship with Neonatal Results

Objectives: In this study, the relationship of cord blood Neutrophil Gelatinase-Associated Lipocalin (NGAL) with neonatal diseases was investigated. Methods: NGAL levels were measured in the cord blood of 180 babies born between 2015 and 2016. Patients were classified according to maternal diseases, neonatal diseases and demographic characteristics. Obtained data were compared with cord blood NGAL levels. Results: In our study, the mean NGAL levels were 1283.99 ng/mL in boys and 1306.52 ng/mL in girls. Umbilical cord blood NGAL levels of infants diagnosed with intrauterine growth retardation (1913.4±2833.5 ng/mL) and prolonged premature rupture of membranes (2594.2±2037.1 ng/mL) were found to be statistically high (p0.05). Conclusions: Neutrophil Gelatinase-Associated Lipocalin, may be useful as a diagnostic biomarker in the evaluation of maternal and neonatal diseases. However, studies on larger patient populations are needed.

Keywords:

Neutrophil Gelatinase Associated Lipocalin, umbilical cord blood, newborn,

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1. Jerónimo C, Bastian PJ, Bjartell A, et al. Epigenetics in prostate cancer: biologic and clinical relevance. Eur Urol. 2011; 60: 753-66.
2. Vieira FQ, Costa-Pinheiro P, Ramalho-Carvalho J, et al. Deregulated expression of selected histone methylases and demethylases in prostate carcinoma. Endocr Relat Cancer. 2014; 21: 51-61.
3. Graça I, Pereira-Silva E, Henrique R, et al. Epigenetic modulators as therapeutic targets in prostate cancer. Clin Epigenetics. 2016; 8: 1-24.
4. Jones PA, Baylin SB. The epigenomics of cancer. Cell. 2007; 128: 683-92.
5. Sharma S, Kelly TK, Jones PA. Epigenetics in cancer. Carcinogenesis. 2010; 31: 27-36.
6. He W, Zhang MG, Wang XJ, et al. KAT5 and KAT6B are in positive regulation on cell proliferation of prostate cancer through PI3K-AKT signaling. Int J Clin Exp Pathol. 2013; 6: 2864-71.
7. Chinaranagari S, Sharma P, Bowen NJ, Chaudhary J. Prostate cancer epigenome. Methods Mol Biol. 2015; 1238: 125-40.
8. Agger K, Cloos PA, Christensen J, et al. UTX and JMJD3 are histone H3K27 demethylases involved in HOX gene regulation and development. Nature. 2007; 449: 731-4.
9. Deng Y, Deng H, Liu J, et al. Transcriptional down regulation of Brca1 and E-cadherin by CtBP1 in breast cancer. Mol Carcinog. 2012; 51: 500-7.
10. Dorman K, Shen Z, Yang C, Ezzat S, Asa SL. CtBP1 interacts with Ikaros and modulates pituitary tumor cell survival and response to hypoxia. Mol Endocrinol. 2012; 26: 447-57.
11. Kuppuswamy M, Vijayalingam S, Zhao LJ, et al. Role of the PLDLS-binding cleft region of CtBP1 in recruitment of core and auxiliary components of the corepressor complex. Mol Cell Biol. 2008; 28: 269- 81.
12. Wang R, Asangani IA, Chakravarthi BV, et al. Role of transcriptional corepressor CtBP1 in prostate cancer progression. Neoplasia. 2012; 14: 905-14.
13. Petrilli AM, Fernández-Valle C. Role of Merlin/NF2 inactivation in tumor biology. Oncogene. 2016; 35: 537-48.
14. Malhotra A, Shibata Y, Hall IM, Dutta A. Chromosomal structural variations during progression of a prostate epithelial cell line to a malignant metastatic state inactivate the NF2, NIPSNAP1, UGT2B17, and LPIN2 genes. Cancer Biol Ther. 2013; 14: 840-52.
15. Erzurumlu Y, Ballar P. Androgen Mediated Regulation of Endoplasmic Reticulum-Associated Degradation and its Effects on Prostate Cancer. Sci Rep. 2017; 7: 1-12.
16. Kruidenier L, Chung C, Cheng Z, et al. A selective jumonji H3K27 demethylase inhibitor modulates the proinflammatory macrophage response. Nature. 2012;488:404-408.
17. Yildirim-Buharalioglu G, Bond M, Sala-Newby GB, Hindmarch CC, Newby AC. Regulation of Epigenetic Modifiers, Including KDM6B, by Interferon-gamma and Interleukin-4 in Human Macrophages. Front Immunol. 2017; 8: 1-18.
18. Elferink CJ, Reiners JJ, Jr. Quantitative RT-PCR on CYP1A1 heterogeneous nuclear RNA: a surrogate for the in vitro transcription run-on assay. Biotechniques. 1996; 20: 470-7.
19. Ghasemi S. Cancer's epigenetic drugs: where are they in the cancer medicines? Pharmacogenomics J. 2020; 20: 367-79.
20. Blevins MA, Huang M, Zhao R. The Role of CtBP1 in Oncogenic Processes and Its Potential as a Therapeutic Target. Mol Cancer Ther. 2017; 16: 981- 90.
21. Moiola CP, De Luca P, Zalazar F, et al. Prostate tumor growth is impaired by CtBP1 depletion in high-fat diet-fed mice. Clin Cancer Res. 2014; 20: 4086-95.
22. Dalton GN, Massillo C, Scalise GD, et al. CTBP1 depletion on prostate tumors deregulates miRNA/mRNA expression and impairs cancer progression in metabolic syndrome mice. Cell Death Dis. 2019; 10: 1-12.
23. Zhou J, Su M, Zhang H, Wang J, Chen Y. miR-539- 3P inhibits proliferation and invasion of gastric cancer cells by targeting CTBP1. Int J Clin Exp Pathol. 2019; 12: 1618-25.
24. Horiguchi A, Zheng R, Shen R, Nanus DM. Inactivation of the NF2 tumor suppressor protein merlin in DU145 prostate cancer cells. Prostate. 2008; 68: 975-84.
25. Gonzalez-Gomez P, Bello MJ, Alonso ME, et al. CpG island methylation in sporadic and neurofibromatis type 2-associated schwannomas. Clin Cancer Res. 2003; 9: 5601-6.
26. Kino T, Takeshima H, Nakao M, et al. Identification of the cis-acting region in the NF2 gene promoter as a potential target for mutation and methylation-dependent silencing in schwannoma. Genes Cells. 2001; 6: 441-54.
27. Sato T, Sekido Y. NF2/Merlin Inactivation and Potential Therapeutic Targets in Mesothelioma. Int J Mol Sci. 2018; 19: 1-18.
28. Wu X, Mao F, Li N, et al. NF2/Merlin suppresses proliferation and induces apoptosis in colorectal cancer cells. Front Biosci (Landmark Ed). 2020; 25: 513-25.
29. Hao S, Baltimore D. The stability of mRNA influences the temporal order of the induction of genes encoding inflammatory molecules. Nat Immunol. 2009; 10: 281-8.
30. De Santa F, Narang V, Yap ZH, et al. Jmjd3 contributes to the control of gene expression in LPS activated macrophages. Embo j. 2009; 28: 3341-52