Browsing by Author "Yerlikaya, Azmi"
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Item Additive enhancement of apoptosis by TRAIL and fenretinide in metastatic breast cancer cells in vitro(Elsevier , 2014-03-13) Dimas, K.; Yerlikaya, Azmi; Ulukaya, Engin; Sarımahmut, Mehmet; Cevatemre, Buse; Arı, Ferda; Uludağ Üniversitesi/Tıp Fakültesi/Tıbbi Biyokimya Anabilim Dalı.; Uludağ Üniversitesi/Fen Edebiyat Fakültesi/Biyoloji Bölümü.; 0000-0003-2647-5875; 0000-0002-6729-7908; 0000-0003-4875-5472; K-5792-2018; AAG-8288-2021; AAG-7012-2021; AHD-2050-2022; 6602927353; 44661687400; 55693788600; 24376085300Item İnsan prostat kanseri PC3 hücre hattında proteozom inhibitörlerine karşı geliştirilen direnç mekanizmalarının araştırılması(Bursa Uludağ Üniversitesi, 2022-02-23) Kanbur, Ertan; Budak, Ferah; Yerlikaya, Azmi; Bursa Uludağ Üniversitesi/Sağlık Bilimleri Enstitüsü/Tıp Fakültesi/İmmünoloji Anabilim Dalı; 0000-0001-8399-8942Günümüzde kanser tedavisi için en umut vaat eden uygulama halen kemoterapi olmasına rağmen kanser ilaçlarına karşı ilaç direnci, medikal onkolojideki en büyük engel teşkil etmeye devam etmektedir. Kemoterapide meydana gelen başarısızlık sebeplerinin %90’ı, kanserde olan veya oluşan ilaç direnci ile meydana gelen invazyon ve metastazdan kaynaklanmaktadır. Tüm dünyada prostat kanseri, erkeklerde akciğer kanserinden (%14,3) sonra ikinci (%14,1) en sık görülen kanser tipidir. Ubiquitin ve proteozom yolağı (UPY) proteinler için bir kalite kontrol sistemidir. Hasarlı ve yanlış katlanmış proteinlerin yıkımından sorumludur. UPY; transkripsiyonda, hücre döngüsünde, hücre içi sinyallerde, antijen sunumunda ve epigenetik mekanizmalarda görev almaktadır. Proteozom inhibitörleri hematolojik malignitelerde kullanılmaktadır. Bortezomib bir proteozom inhibitörüdür. Proteozom inhibitörlerinin solid tümörlerde kullanılması umut vadetmektedir. Proteozom inhibitörlerinin otoimmün hastalıklarda da kullanılabileceği düşünülmektedir. Hücrelerin stres kaynaklarına verdiği yanıtlardan biri, hücrelerdeki moleküler değişiklikler ile kalıcı bir şekilde hücre döngüsü tutuklanması olan senesenstir. Senesent hücreler genellikle biyoaktif proteinler salgılarlar ve bunlar senesent olmayan komşu hücreleri uyararak tümör gelişimine sebep de olabilmektedir ve aynı zamanda yaşlanma sürecindeki kronik enflamasyondan sorumlu olduğu düşünülmektedir. Bu tezde; bir prostat kanseri hücre hattı olan PC3 hücrelerinde [PC3-P, parental; PC3-R, dirençli], klinikte prostat tedavi protokollerinde henüz kullanılmayan ve solid tümörlerde kullanılması umut vadeden bortezomib ilacına karşı oluşmuş ilaç direnci mekanizmaları ile bortezomib’in PC3 hücreleri üzerindeki senesens rolü araştırılmıştır. Dirençli PC3-R ve parental PC3-P hücrelerinde yaptığımız bortezomib sitotoksisitesi, hücre ölüm modu, AO/EB çift boyaması ve 3 boyutlu hücre kültürü çalışmalarında; PC3-R hücrelerinin PC3-P hücrelerine göre ilaca çok daha dirençli oldukları, apoptoza daha dirençli oldukları ve ilaç varlığında bile 3 boyutlu kompleks yapıları oluşturarak kendilerini destekleyebildikleri ve direnç mekanizmasının geri dönüşümlü olmadığı gösterilmiştir. Direnç mekanizmasını aydınlatmaya yönelik MAPK ve otofajik aktivasyon, ısı şok proteinlerin ekspresyonu gibi incelenen stratejik yolaklarda; stres koşullarında aktive olan ve dirence sebebiyet veren bu yolakların ve ilgili moleküllerin dirençli PC3-R hücrelerinde değil, parental PC3-P hücrelerinde bir sağkalım mekanizması olarak aktive oldukları gözlenmiştir. PC3-R hücrelerindeki bortezomib direncinin ilacın bağlandığı PSMB5 enziminde meydana gelen bir mutasyondan kaynaklanabileceği düşünülmektedir. PC3-P ve PC3-R hücrelerinde yapılan senesens deneyleri sonucunda, senesens markerlerinin bortezomib muamelesi sonucunda ekseriyetle azaldığı gözlenmiştir ve bu sebeple bortezomibin bir senolitik etkisinin olabileceği öne sürülmektedir. Buna ek olarak PC3-R hücrelerinde senesens, PC3-P hücrelerine göre daha az görülmektedir ve bunun ilaç direncine bir katkısı olabilir. Son olarak, literatüre göre ERK1 ve ERK2 proteinlerinin fonksiyonel olarak işlevlerinin aynı olup olmadığı hakkında tartışmalar vardır. Tez çalışmamızda bortezomib muamelesinden sonra ERK1 fosforilasyonlarının artarken ERK2 fosforilasyonlarının azaldığı görülmüştür. Bu olay, ERK1 ve ERK2’nin farklı regüle edilebildiğini ve bu sebeple farklı işlevler görebileceklerine işaret etmektedir.Item The p53-independent induction of apoptosis in breast cancer cells in response to proteasome inhibitor bortezomib(Sage Publications, 2012-10) Yerlikaya, Azmi; Okur, Emrah; Ulukaya, Engin; Uludağ Üniversitesi/Tıp Fakültesi/Tıbbi Biyokimya Anabilim Dalı.; K-5792-2018; 6602927353An important hallmark of cancer cells is acquired resistance toward apoptosis. The apoptotic pathway is the most well-defined cell death program and is characterized by several morphological and biochemical features. The tumor suppressor protein p53 is a critical regulator of apoptosis in many cell types. p53 stimulates a wide network of signals that act through either extrinsic or intrinsic pathways of apoptosis. However, a number of studies have shown that apoptosis can be induced in a p53-independent manner as well. In this study, we examined the mechanism of apoptosis in p53-null breast cancer cells in response to the proteasome inhibitor bortezomib. Initially, we determined the p53 status of 4T1 breast carcinoma and 4THMpc (a highly mestatic derivative of 4T1) cells and verified that both cells are p53 deficient. It was subsequently shown that apoptosis can be induced in both cells in a dose-dependent manner in response to bortezomib treatment, based on DNA fragmentation evidence. Western blot analyses of ubiquitin-protein conjugates additionally showed that the proteasome is potently inhibited by bortezomib in p53-null 4T1 and 4THMpc cells. The results presented in the current study also show that caspase-3 is significantly activated in response to the treatment with bortezomib, implying that induction of apoptosis in these p53-deficient cells is occuring via caspase-3. The additional results presented here suggest that the pro-apoptotic proteins Bad, Noxa, and Puma are not critical regulators of apoptosis induction in p53-null 4T1 and 4THMpc cells. Similarly, there was no difference in the expression level of Mcl-1 in treated cells, suggesting that this anti-apoptotic protein is also uninvolved in the apoptotic response resulting from bortezomib treatment. In contrast, a very significant upregulation of the anti-apoptotic protein Hsp25/27 was detected in these p53-deficient cells after treatment with bortezomib. If the increased expression of Hsp25/27 protein levels are muting the apoptotic effects of the bortezomib treatment, then the apoptosis-inducing effects of such proteasome inhibitors might be increased with approaches simultaneously inhibiting Hsp25/27 protein in p53-deficient cells.Item Protein yıkımının önemi(Uludağ Üniversitesi, 2009-07-14) Yerlikaya, Azmi; Dokudur, HarunYaşayan bir organizmanın birçok hücresel proteini sürekli olarak yıkılmakta ve tekrar sentezlenmektedir. Proteinlerin bu yaşam süreleri birkaç dakika ile birkaç hafta veya daha uzun süreler arasında değişmektedir. Hücre içindeki protein yıkım mekanizmalarının bazı işlevleri şunlardır: 1) Normal şartlar altında, bazı önemli metabolik kontrol noktalarında görev alan enzimlerin yıkılarak ve aktivitelerinin düzenlenerek hücrenin değişen çevre şartlarına ve metabolik ihtiyaçlarına verimli bir şekilde cevap vermesini sağlamak. 2) Proteinlerin besin yetersizliği durumunda enerji ve amino asit kaynağı olarak kullanılmasını sağlamak. 3) Transkripsiyonel veya translasyonel hatalardan kaynaklanan hasarlı veya işlevsiz proteinlerin uzaklaştırılması. Bu derlemede hücre içindeki bazı önemli proteolitik mekanizmaların genel özellikleri, yapıları ve işlevleri hakkında bilgi vermeye çalıştık.Item A proteomic analysis of p53-independent induction of apoptosis by bortezomib in 4T1 breast cancer cell line(Elsevier, 2015-01-15) Yerlikaya, Azmi; Okur, Emrah; Baykal, Ahmet Tarik; Acilan, Ceyda; Boyaci, Ihsan; Ulukaya, Engin; Uludağ Üniversitesi/Tıp Fakültesi/Tıbbi Biyokimya Anabilim Dalı.; 0000-0003-4875-5472; K-5792-2018; 6602927353The 26S proteasome is a proteolytic enzyme found in both cytoplasm and nucleus. In this study, we examined the differential expression of proteasome inhibitor bortezomib-induced proteins in p53-deficient 4T1 cells. It was found that GRP78 and TCEB2 were over-expressed in response to treatment with bortezomib for 24 h. Next, we analyzed the expression of intracellular proteins in response to treatment with 100 nM bortezomib for 24 h by label-free LC-MS/MS. These analyses showed that Hsp70, the 26S proteasome non-ATPase regulatory subunit 14 and sequestosome 1 were increased at least 2 fold in p53-deficient 4T1 cells. The proteins identified by label-free LC-MS/MS were then analyzed by Ingenuity Pathway Analysis (IPA) Tool to determine biological networks affected by inhibition of the 26S proteasome. The analysis results showed that post-translational modifications, protein folding, DNA replication, energy production and nucleic acid metabolism were found to be among the top functions affected by the 26S proteasome inhibition. The biological network analysis indicated that ubiquitin may be the central regulator of the pathways modulated after bortezomib-treatment. Further investigation of the mechanism of the proteins modulated in response to the proteasomal inhibition may lead to the design of more effective and novel therapeutic strategies for cancer. Biological significance Although the proteasome inhibitor bortezomib is approved and used for the treatment of human cancer (multiple myeloma), the mechanism of action is not entirely understood. A number of studies showed that proteasome inhibitors induced apoptosis through upregulation of tumor suppressor protein p53. However, the role of tumor suppressor protein p53 in bortezomib-induced apoptosis is controversial and not well-understood. The tumor suppressor p53 is mutated in at least 50% of human cancers and is strongly induced by proteasomal inhibition. Some also reported that the proteasome inhibitor can induce apoptosis in a p53-independent manner. Also, it is reported that Noxa, a target of p53, is induced in response to proteasomal inhibition in a p53-independent manner. However, we have also previously reported that neither Puma nor Noxa are induced by proteasomal inhibition in p53-null 4T1 breast cancer cells, which is commonly used for in vivo breast cancer tumor models. The current results provided additional targets of proteasome inhibitor bortezomib and may therefore help in understanding the p53-independent mechanism of apoptosis induction by proteasome inhibitors. In addition, the results presented in this current study report for the first time that proteasomal subunit Psmd14, anti-apoptotic GRP78, anti apoptotic protein Card10, Dffb, Traf3 and Trp53bp2 are regulated and overexpressed in response to proteasome inhibitor bortezomib in p53-deficient 4T1 cells. Therefore, novel therapeutic strategies targeting these anti-apoptotic or pro-apoptotic proteins as well as inhibiting the proteasome simultaneously may be more effective against cancer cells. The proteins identified here present new avenues for the development of anti-cancer drugs.Publication The ubiquitin-proteasome pathway and epigenetic modifications in cancer(Bentham Science Publ Ltd, 2021-01-01) Yerlikaya, Azmi; Kanbur, Ertan; Stanley, Bruce A.; Tümer, Emrah; Kanbur, Ertan; Bursa Uludağ Üniversitesi/Tıp Fakültesi/İmmünoloji Ana Bilim Dalı; 0000-0001-8399-8942; AAW-6971-2021Background: The ubiquitin-proteasome pathway is involved in almost all cellular processes (cell cycle, gene transcription and translation, cell survival and apoptosis, cell metabolism and protein quality control) mainly through the specific degradation of the majority of intracellular proteins (>80%) or partial processing of transcription factors (e.g., NF-kappa B). A growing amount of evidence now indicates that epigenetic changes are also regulated by the ubiquitin-proteasome pathway. Recent studies indicate that epigenetic regulations are equally crucial for almost all biological processes as well as for pathological conditions such as tumorigenesis, as compared to non-epigenetic control mechanisms (i.e., genetic alterations or classical signal transduction pathways).Objective: Here, we reviewed the recent work highlighting the interaction of the ubiquitin-proteasome pathway components (e.g., ubiquitin, E1, E2 and E3 enzymes and 26S proteasome) with epigenetic regulators (histone deacetylases, histone acetyltransferases and DNA methyltransferases).Results: Alterations in the regulation of the ubiquitin-proteasome pathway have been discovered in many pathological conditions. For example, a 2- to 32-fold increase in proteasomal activity and/or subunits has been noted in primary breast cancer cells. Although proteasome inhibitors have been successfully applied in the treatment of hematological malignancies (e.g., multiple myeloma), the clinical efficacy of the proteasomal inhibition is limited in solid cancers. Interestingly, recent studies show that the ubiquitin-proteasome and epigenetic pathways intersect in a number of ways through the regulation of epigenetic marks (i.e., acetylation, methylation and ubiquitylation).Conclusion: It is therefore believed that novel treatment strategies involving new generation ubiquitin-proteasome pathway inhibitors combined with DNA methyltransferase, histone deacetylase or histone acetyltransferase inhibitors may produce more effective results with fewer adverse effects in cancer treatment as compared to standard chemotherapeutics in hematological as well as solid cancers.Item The ubiquitin-proteasome pathway and resistance mechanisms developed against the proteasomal inhibitors in cancer cells(Bentham Science Publications, 2020) Yerlikaya, Azmi; Kanbur, Ertan; Bursa Uludağ Üniversitesi/Tıp Fakültesi/İmmünoloji Anabilim Dalı.; 0000-0001-8399-8942; AAW-6971-2021; 57218292727Background: The ubiquitin-proteasome pathway is crucial for all cellular processes and is, therefore, a critical target for the investigation and development of novel strategies for cancer treatment. In addition, approximately 30% of newly synthesized proteins never attain their final conformations due to translational errors or defects in post-translational modifications; therefore, they are also rapidly eliminated by the ubiquitin-proteasome pathway. Objective: Here, an effort was made to outline the recent findings deciphering the new molecular mechanisms involved in the regulation of ubiquitin-proteasome pathway as well as the resistance mechanisms developed against proteasome inhibitors in cell culture experiments and in the clinical trials. Results: Since cancer cells have higher proliferation rates and are more prone to translational errors, they require the ubiquitin-proteasome pathway for selective advantage and sustained proliferation. Therefore, drugs targeting the ubiquitin-proteasome pathway are promising agents for the treatment of both hematological and solid cancers. Conclusion: A number of proteasome inhibitors are approved and used for the treatment of advanced and relapsed multiple myeloma. Unfortunately, drug resistance mechanisms may develop very fast within days of the start of the proteasome inhibitor-treatment either due to the inherent or acquired resistance mechanisms under selective drug pressure. However, a comprehensive understanding of the mechanisms leading to the proteasome inhibitor-resistance will eventually help the design and development of novel strategies involving new drugs and/or drug combinations for the treatment of a number of cancers.