Vpliv celic žilja na odpornost celic glioblastoma na kemoterapijo

Kolenc Milavec, Tina (author), Breznik, Barbara (mentor)

Glioblastom kljub agresivnemu zdravljenju in številnim raziskavam v smeri novih načinov spopadanja s to boleznijo ostaja eden izmed najsmrtonosnejših primarnih rakov. Danes vemo, da so za razvoj tumorja in odpornost na klasično terapijo odgovorne rakave matične celice (GSC). Pri ohranjanju lastnosti GSC ima ključno vlogo tumorsko mikrookolje, zato se te v tkivu nahajajo na točno določenih področjih ob žilah in jih imenujemo perivaskularne niše glioblastomskih matičnih celic, ki zagotavljajo ustrezne pogoje za preživetje. V mikrookolju glioblastoma so za rakave matične celice pomembne celice žilja, ki preko izločanja molekul, kot so kemokini in dušikov oksid, spodbujajo rast tumorja in povečujejo odpornost GSC na terapijo. Za iskanje novih tarč za zdravljenje glioblastoma, ki dandanes še vedno temelji na klasični terapiji, je ključno poznavanje biologije niše in interakcij med GSC in celicami žilja. V okviru te magistrske naloge smo zato želeli preučiti interakcije med GSC in celicami žilja v perivaskularni niši in vpliv kemoterapije na te interakcije, pa tudi na lastnosti samih GSC. V ta namen smo postavili in vitro model perivaskularne niše in z uporabo pleriksaforja, ki zavira kemokinsko signalno pot SDF-1α/CXCR4, ter L-NAME, ki inhibira signalno pot sintaze NO, skušali odkriti dejavnike, ki prispevajo k invaziji GSC v perivaskularno nišo, ter ugotoviti, ali te signalne poti vplivajo na fenotip GSC. Ugotovili smo, da imajo tako endotelijske kot tudi gladkomišične celice močan kemotaktični učinek na GSC. Pokazali smo, da je za kemotakso GSC k endotelijskim celicam in v periarteriolarno nišo ključna signalna pot SDF-1α/CXCR4, saj je antagonist CXCR4 pleriksafor zmanjšal invazijo GSC v nišo. Po drugi strani inhibitor sintaze NO (L-NAME) na invazijo GSC v perivaskularno nišo ni imel vpliva. Kemoterapevtik temozolomid ne vpliva na invazijo samih GSC, inhibira pa invazijo GSC v perivaskularno nišo. Dodatno smo ugotovili, da blokada poti SDF-1α/CXCR4 sproži diferenciacijo GSC in da začnejo GSC pod vplivom gladkomišičnih celic pridobivati astrocitom podoben fenotip. Rezultati magistrske naloge dajejo nov vpogled v razumevanje odpornosti glioblastoma na zdravljenje in izpostavijo signalno pot SDF-1α/CXCR4 kot tarčo za zaviranje prehoda GSC v perivaskularne niše. Z uporabo inhibitorjev te signalne poti, kamor sodi tudi pleriksafor, bi lahko zmanjšali delež glioblastomskih celic, ki se nahajajo v nišah in s tem povečali dovzetnost GSC na terapijo.

• Language: Slovenian
• Keywords: glioblastomske matične celice, invazija, celice žilja, pleriksafor, temozolomid
• Work type: Master's thesis/paper
• Typology: 2.09 - Master's Thesis
• Organization: FKKT - Faculty of Chemistry and Chemical Technology
• Year: 2023
• PID: 20.500.12556/RUL-147037
• COBISS.SI-ID: 159783171
• Publication date in RUL: 21.06.2023

Secondary language

• Language: English
• Title: Effect of vascular cells on resistance of glioblastoma cells to chemotherapy

Abstract

Despite aggressive treatment and an array of research, trying to find new approaches to tackle glioblastoma, it remains one of the deadliest primary cancers. Main culprits for resistance of glioblastoma to classical therapy are cancer stem cells (CSCs). Because tumour microenvironment is crucial for survival of GSCs and maintenance of GSC phenotype, those cells can be found in special areas near brain vasculature, known as perivascular GSC niches, which provide perfect environment for GSC survival. An important part of tumour microenvironment represent blood vessels that excrete molecules, such as chemokines and nitric oxide, and thereby promote growth of the tumour and lead to increased resistance of glioblastoma to therapy. Finding new targets to treat glioblastoma, which is nowadays still tackled with classical therapies, requires understanding the biology of GSC niches and interactions between GSCs and vascular cells. In this work we investigated interactions between GSCs and vascular cells in perivascular niche and aimed to find out how chemotherapy affects those interactions as well as features of GSCs. We developed in vitro model of perivascular niche and tried to discover factors that contribute to invasion of GSCs into perivascular niche by using plerixafor, which inhibits SDF-1α/CXCR4 signalling pathway and L-NAME, inhibitor of NO synthase. We discovered that endothelial as well as smooth muscle cells have a profound chemotactic effect on GSCs. We showed that signalling pathway SDF-1α/CXCR4 is crucial for chemotaxis of GSCs into periarteriolar niche, since CXCR4 inhibitor plerixafor significantly reduced invasion of GSCs in the niche. NO synthase inhibitor L-NAME on the other hand did not show any effect on the invasion of GSCs into perivascular niche. Chemotherapeutic temozolomide did not affect invasion of GSCs on their own, but it did inhibit invasion of GSCs into perivascular niche. Additionally, we discovered that inhibition of SDF-1α/CXCR4 pathway leads to GSC differentiation and that in presence of smooth muscle cells, GSCs start to acquire astrocyte-like phenotype. The results of this work give new insight into understanding the resistance of glioblastoma to therapy and expose signalling pathway SDF-1α/CXCR4 as a possible target to inhibit GSC invasion into the perivascular niches. By using inhibitors of this pathway, such as plerixafor, we could lower the amount of GSCs in the perivascular niches and thereby sensitize GSCs to therapy.

• Keywords: glioblastoma stem cells, invasion, vascular cells, plerixafor, temozolomide