Münster (Westfalen), Universiẗat, Diss., 2001.

No description available.

Universiẗat, Diss., 2001--Münster (Westfalen).

One group of chemotherapeutics that are used successfully to treat breast cancer, alone or in combination with other agents, are the taxanes; paclitaxel and docetaxel. They act by interfering with the spindle microtubule dynamics of the cell causing cell cycle arrest. However, the complexities underlying the mechanism of action are yet to be fully elucidated. Arguably, one of the most significant problems with taxanes is chemoresistance. Unfortunately, some patients are intrinsically resistant to taxanes and others acquire resistance to taxanes as treatment advances. This problem is exacerbated by a lack of understanding of the mechanisms underlying taxane resistance. Isogenic breast cancer cell lines that were taxane resistant were generated to use as an experimental model. Paclitaxel resistant (PACR) MDA-MB-231, paclitaxel resistant ZR75-1 and docetaxel resistant (DOCR) ZR75-1 cell lines were successfully generated by incrementally increasing taxane dose in respective native cell lines in vitro. An extensive characterisation of each of the resistant cell lines was conducted, focussing primarily on the 25nM resistant cells which were determined to be the most clinically relevant dose of taxane. A suboptimal dose of 5nM, a “superoptimal” dose of 50nM and the native, taxane sensitive cells was included. Dose response cell count experiments were performed that confirmed taxane resistant cells had been generated. It was shown that MDA-MB-231 native cells were more sensitive to paclitaxel than the ZR75-1 native cells, suggesting that ZR75-1 cells may already have low level inherent resistance. The MDA-MB-231 25nM PACR cells were tested to determine whether they retained PACR when maintained in media containing no paclitaxel. MDA-MB-231 25nM PACR cells were maintained in a taxane free environment for six months and then rechallenged with taxane. When rechallenged, the PACR cells previously maintained in the absence of paclitaxel mirrored the pattern of growth of corresponding PACR cells that had been maintained in the presence of paclitaxel. This proved that in the absence of paclitaxel, PACR cells did not revert to parent phenotype. This meant that experiments could be designed to grow cell lines as xenografts in mice, (in the absence of paclitaxel) & bring in vitro experiments into an in vivo setting. Effects of taxane treatment on both native and resistant cells were analysed using flow cytometry. Paclitaxel treatment exerted G2/M block in native MDA-MB-231 cells but when PACR cells were treated with the same dose of paclitaxel no G2/M block was observed, suggesting that PACR cells had developed a mechanism for escaping G2/M block. ZR75-1 native lines were also investigated and we established that treatment with paclitaxel also exerted a G2/M block in these lines. In future studies this process will be repeated to investigate the effect of taxane treatment on the ZR75-1 PACR and DOCR lines. CD 1 nude mice were injected with cells from all five cell lines to grow xenografts, unfortunately MDA-MB-231 PACR cells failed to grow so they could not be used for further xenograft experiments. PACR, DOCR and Native ZR75-1 cells did successfully grow as xenografts in mice and confirmed that all 3 groups showed very similar growth patterns. A cross resistance experiment was conducted and it was determined that the DOCR xenografts maintained a taxane resistant phenotype to docetaxel, and not paclitaxel and the PACR xenografts may be perpetuate the paclitaxel resistant phenotype in xenografts and that there may be cross resistance to docetaxel in the paclitaxel resistant xenografts. This is the first time that taxane resistant cell lines grown in this way have been established as xenografts in mice. These cross resistance experiments represent novel findings and merit further investigation. Extensive genomic and transcriptomic analyses were carried out on the cell lines to help identify potential taxane resistance markers. aCGH experiments were carried out to compliment the illumina experiments. The first set of experiments used DNA from pooled whole female blood as ref sample and DNA from each of the native and taxane resistant cell lines as test samples. The second set of experiments used DNA from native cells as a ref sample and DNA from their respective taxane resistant cells as a test, which allowed areas of loss or gain to be tracked in the genome as resistance increased. In the MDA-MB-231 cell lines the following areas of loss extended with increasing resistance: 1p36.13-q44, 6p25.3-q12, 8p, 10p, 19q, X Chr and the following areas of gain 2p25.3-23.3, 3p24.3-q13.3, 4p16.1-q12, 5q14.3-q31.1, 8q21.13-24.3, 11q15.1-q25, centromeric 12, and centromeric 14. In the ZR75-1 PACR and DOCR cell lines the areas of loss extended with increasing resistance in the following regions: 7q, 12p and 16q. For gene expression analysis RNA was extracted from the MDA-MB-231 cell lines, labelled and hybridised them to illumina human ref 8 vs. 2 chips. Data showed a progressive increase in mRNA dysregulation as paclitaxel resistance increased. Eleven genes were dysregulated across all resistance levels in the PACR MDA-MB-231 cells when compared to the relative cell lines; RGS16, CLDN1, IL7R, P&PP1R14C, COBL, TRPV4, TSPAN8, CD33, NLRP2, P13, and PAGE5. The experiment was repeated using MDA-MB-231 PACR, ZR75-1 PACR and DOCR cells and resulting data was analysed to determine genes commonly dysregulated across resistance levels, between MDA-MB-231 PACR and ZR75-1 PACR and between ZR75-1 PACR and DOCR cell lines. An extensive literature search was conducted and established four genes of interest in the context of our genomic and transcriptomic experiments including AURKA, Mdr-1, Stathmin and YY1. The novel biomarkers identified in the illumina experiments were validated with complimentary qPCR gene expression experiments looking at expression levels of the eleven commonly dysregulated genes identified and a panel of 19 other genes with significantly increased or decreased expression as resistance increased including AURKA, Mdr-1, Stathmin and YY1. Western blots were performed with lysates from the cell lines using a standard panel of predictive breast cancer markers and AURKA, Mdr-1, Stathmin and YY1. Combining the data from the genomic study, the gene expression profile, qPCR and Western blotting it was established that Mdr-1 had increased expression in the taxane resistant ZR75-1 lines and YY1 had increased expression in the MDA-MB-231 PACR line. Material from the LAPATAX trial was used to observe any transcriptomic changes occurring in tumours following treatment with docetaxel and to compare them to changes identified in our in vitro and xenograft models, this allowed the final step to be taken into a translational environment. LAPATAX (EORTC 10054) is a phase I-II study of Lapatanib and Docetaxel as neoadjuvant treatment for HER-2 +ve locally advanced/inflammatory or large operable breast cancer. Tumour material from eighteen core biopsies pre and post treatment was obtained, the mRNA was extracted, labelled and hybridised to the illumina array. This allowed the changes in gene expression pre and post docetaxel treatment to be tracked. The gene expression data from the LAPATAX trial was combined with gene expression data from our cell line panel and identified two novel putative markers of taxane resistance DUSP1 and FOS. Although sample size is small this has provided extremely valuable evidence directly from the clinic. These two novel putative biomarkers are extremely intriguing and certainly merit further investigation, ideally using additional taxane treated breast tumour tissue. Ultimately, an isogenic in vitro model of taxane resistance was developed in two different cell lines and with two different taxanes within one cell line. The cell lines were characterised and the effect of the taxanes on the cell cycle was determined in the native and taxane resistant lines. Selected cell lines were grown as xenografts in mice and performed successful cross resistance studies upon them. A large transcriptomic and genomic analysis was conducted and has identified a panel of potential taxane resistance markers and areas of loss and gain in the genome perpetuated by increasing taxane resistance. This analysis was validated using qPCR and Western blotting. This allowed a panel of novel taxane resistance markers to be identified. In future studies it is hoped that these targets will be knocked down with shRNA to observe if the taxane resistant cell lines revert to the parental phenotype. In vitro studies will be conducted to find agents that may be used to reduce expression of these markers and restore sensitivity to taxanes and consequently restore the efficacy of these drugs in a clinical setting. As far as the author is aware this is the first time that isogenic taxane resistant cell lines have been generated and investigated in this way.

No description available.

Taxane (wie Paclitaxel oder Cabazitaxel) sind bewährte Arzneimittel in den systemischen Therapieschemata vieler bösartiger Erkrankungen, einschließlich Brust- und Eierstockkrebs. Sie fördern die Stabilisierung der Mikrotubuli, was zu einem Stillstand des Zellzyklus während der Mitose führt, auf den die Apoptose folgt. Neben dieser antimitotischen Wirkung von Taxanen ist seit einiger Zeit auch eine gefäßverändernde Wirkung von Taxanen bekannt. Kürzlich wurde gezeigt, dass Taxane tatsächlich Störungen in der Gefäßarchitektur verursachen, indem sie den Kalziumeinstrom über TRPC6, einen unselektiven Kationenkanal, auslösen. Der erhöhte intrazelluläre Ca2+-Spiegel bewirkt eine Rundung der Endothelzellen, was zu einer Störung des endothelialen Monolayers, Serumausfluss und Gefäßkollaps führt. In dieser Arbeit konzentrierten wir uns auf die Gefäßbetten von peripheren Organen wie dem Herzen oder der Niere in Abhängigkeit vom Tumorstadium und der Taxol-Behandlung. Die Organe wurden mit immunhistochemischen Techniken angefärbt, um Veränderungen in der Architektur und Morphologie der Blutgefäße zu untersuchen. Wir fanden Veränderungen in der Morphologie der Kapillaren des Herzens und darüber hinaus Veränderungen in der Expression endothelialer Antigene in Abhängigkeit vom Tumorstadium, insbesondere eine zunehmende endotheliale Expression von TRPC6 in Abhängigkeit vom Tumorstadium. Diese Ergebnisse liefern neue Erkenntnisse für das Verständnis der systemischen Auswirkungen maligner Erkrankungen und tragen dazu bei, Folgeerkrankungen bei Patienten mit fortgeschrittenem Krebs zu verhindern. / Taxanes (like Paclitaxel or Cabazitaxel) are well-established drugs in the systemic therapy regimens of many malignancies, including breast and ovarian cancer. They promote the stabilization of microtubules leading to an arrest of the cell cycle during mitosis which is followed by apoptosis. Beside this anti-mitotic action of taxanes, a vascular-directed effect of taxanes has been known for some time. We recently showed that taxanes actually cause vascular disruption by triggering calcium influx via TRPC6. The increased intracellular Ca2+-level causes rounding of endothelial cells, leading to a disruption of the endothelial monolayer, serum efflux, and vascular collapse. In this study, we focused on the vascular beds of peripheral organs like the heart or the kidney in dependency on tumor stage and Taxol-therapy. The organs were stained via immunohistochemical techniques to examine changes in the architecture and morphology of the blood vessels. We found changes in the morphology of the cardiac capillaries and furthermore changes in the expression of endothelial antigens in dependence on the tumor stage, especially an increasing endothelial expression of TRPC6 dependent on the tumor stage. These findings provide new insights for the understanding of the systemic effects of malignant diseases and help to prevent secondary diseases in patients with advanced cancer.

Calophyllum inophyllum is a medicinal plant that is rich in bioactive natural products. Calophinone¡]29¡^¡Bcaloxanthone I¡]30¡^¡Bbrasilixanthone B¡]31¡^¡Bpyranojacareubin¡]32¡^ and osajaxanthone¡]33¡^are five compounds which were isolated from the bark in this experiment. It is the first time to isolate calophinone¡]29¡^ from a matural source. In order to identify calophinone¡]29¡^, 6-acetylcalophinone¡]34¡^ was prepared via acylation. All structures were determined primarily on the basis of 1D, 2D NMR¡BUV¡BIR and Mass spectral analyses. Besides, biological studies don¡¦t reveal that Calophinone¡]29¡^¡Bcaloxanthone I¡]30¡^¡Bbrasilixanthone B¡]31¡^¡Bpyranojacareubin¡]32¡^¡Bosajaxanthone¡]33¡^and 6-acetylcalophinone¡]34¡^, exhibited in vitro cytotoxicity against human liver carcunoma¡BHuman oral epidermoid carcunoma and Human cervical epidermoid carcunoma. In addition, four new Taxoid derivatives that were 13-O-camphanyl-7-O -nicotinoylbaccatin III¡]34¡^¡B13-O-camphanyl-1-deoxybaccatin VI¡]35¡^¡B13-O-¡]4-chlorobenzoyl¡^-7-O-nicotinoylbaccatin III¡]36¡^and 13-O-benzoyl-7-O -nicotinoylbaccatin III¡]37¡^have been prepared via esterification under sonication starting from 13-deacetyl-7-O-nicotinoylbaccatin III¡]32¡^ and 13-deacetyl-1-deoxybaccatin VI¡]31¡^. All the structures were established primarily on the basis of 1D and 2D NMR techniques including DEPT, COSY, HMBC experiments, as well as comparison with known related compounds. It was deemed quite promising to investigate the structure-activity relationship ( SAR ) for the C - 13 side chain analogues of Taxol with some modification of the baccatin III¡]30¡^ moiety in order to discover more effective anticancer agents with better pharmacological properties. Compounds 34 and 35 showed significant cytotoxicity against prostate cancer cell line¡]PC-3¡^. Under concentration of 10£gM, the cell survival percent was 76% and 65% in case of compounds 34 and 35 compared to 60 % in case of Taxol. According to the structure-activity relationship, nicotinoyl and camphanic acyl group should be the source of activity in compounds 30 and 31. Consequently, it is necessary to introduce nicotinoyl chloride and camphanic acyl chloride groups via chemical reaction to improve the bioactivity.

Indiana University-Purdue University Indianapolis (IUPUI) / The main focus of this study is to determine the impact of Set-Binding Factor 2 (SBF2) on human-derived neurons in the context of taxane-induced peripheral neuropathy. Taxane-induced peripheral neuropathy (TIPN) is a devastating survivorship issue for many cancer patients; SBF2 has been previously identified as a potential germline predictor that has been found to be significantly associated with severe TIPN in African American (AA) patients. The work described here provides ex vivo support for the use of SBF2 as a genotypic biomarker to identify a priori which patients are at a higher risk of manifesting severe TIPN. This study demonstrates that diminished expression of SBF2 exacerbated the effect of paclitaxel on viability and morphology and altered the functional response of a neuronal model exposed to paclitaxel treatment. Furthermore, transcriptomic work showed that reduced expression of SBF2 in a neuronal model treated with paclitaxel impacted the expression of genes that modulate stress-induced cell death and pain threshold. Altogether, these findings suggest that SBF2 plays a role in the development of TIPN. This work sheds light on the pathways potentially involving SBF2 that can be studied to further evaluate the function of this gene in neurons and its contribution to severe TIPN. Further functional approaches investigating these pathways will be pivotal in elucidating the underlying biological mechanism for this toxicity and identifying novel targeted therapeutic strategies to prevent or treat TIPN. / 2021-05-17

Ο σκοπός της παρούσας μελέτης ήταν να εξετάσει τον ρόλο της ταξόλης (paclitaxel) και της καρβοπλατίνης σε σχέση με τις παραμέτρους του ανοσοποιητικού συστήματος σε ασθενείς πάσχοντες από μη μικροκυτταρικό καρκίνο του πνεύμονα και από καρκίνο των ωοθηκών· πριν, κατά τη διάρκεια και μετά από χημειοθεραπεία και η επίδραση που είχε ο ανωτέρω συνδυασμός φαρμάκων στην συνολική επιβίωση των ασθενών. Υλικό και Μέθοδος: Εξετάσθηκαν 24 ασθενείς με μη-μικροκυτταρικό καρκίνο του πνεύμονα και 20 με καρκίνο των ωοθηκών (όλοι μεταστατικοί), όπου χωρίστηκαν σε δύο ομάδες με κριτήριο την επιβίωση και που εν συνεχεία τους χορηγήθηκε συνδυασμός καρβοπλατίνης και ταξόλης για έξι θεραπευτικούς κύκλους. Ομάδα Α. Ασθενείς με καλή επιβίωση (>12 μήνες για μη-μικροκυτταρικό καρκίνο του πνεύμονα, >30 μήνες για καρκίνο των ωοθηκών) Ομάδα Β. Ασθενείς με «φτωχή» επιβίωση (<12 μήνες για μη-μικροκυτταρικό καρκίνο του πνεύμονα, <18 μήνες για καρκίνο των ωοθηκών) Την ίδια χρονική περίοδο εξετάσθηκαν οι λεμφοκυτταρικοί υποπληθυσμοί (CD3, CD4, CD8, CD56, CD34) καθώς και οι κυτταροκίνες ιντερλευκίνη-3 (IL-3) και ιντερφερόνη-γ (IFN-γ), σε σχέση με την ποιότητα ζωής και το προσδόκιμο επιβίωσης κατά την διάρκεια της χημειοθεραπευτικής αγωγής. Η στατιστική ανάλυση των αποτελεσμάτων έγινε με την μέθοδο ANOVA. Αποτελέσματα: Από την εξαγωγή των αποτελεσμάτων παρατηρήθηκε μία στατιστικώς σημαντική διαφορά ανάμεσα στις τιμές των λεμφοκυτταρικών 207 υποπληθυσμών CD4 και CD4/CD8 μετά από χημειοθεραπεία μεταξύ των δύο ομάδων ασθενών Α και Β (p=0,001 και p=0,006). Αυτό σημαίνει ότι η περαιτέρω αύξηση του αριθμού των βοηθητικών Τ-λεμφοκυττάρων (T-helper) μετά από χημειοθεραπεία συμβάλλει θετικά στο προσδόκιμο επιβίωσης. Επιπροσθέτως, στατιστικώς ενδιαφέρων σε σημείο που να μπορούμε να μιλήσουμε και για προγνωστικό παράγοντα, ήταν η διαφορά ανάμεσα στις τιμές της ιντερφερόνης-γ μεταξύ των ομάδων Α και Β πριν και μετά τη χημειοθεραπεία (p=0,039 και p=0,027, αντιστοίχως). Οι ασθενείς με υψηλές τιμές ιντερλευκίνης-3 παρουσίαζαν επίσης χαμηλή τοξικότητα. Συμπεράσματα: Στην παρούσα μελέτη η προσπάθεια μας επικεντρώθηκε στο να καταδείξουμε την επίδραση που ασκείται, από την χρήση του συνδυασμού καρβοπλατίνης-ταξόλης, στους λεμφοκυτταρικούς υποπληθυσμούς και στις κυτταροκίνες καθώς και την επιρροή που ασκούν και τα δύο αυτά στοιχεία του ανοσοποιητικού συστήματος στο προσδόκιμο επιβίωσης και στην εν γένει ποιότητα ζωής. / The aim of the present study was to exam the role of Paclitaxel (Taxane) and Carboplatin in the parameters of the immune system in patients with non-smallcell lung cancer and in patients with ovarian cancer before, during and after chemotherapy treatment, and the effect of this combination in the overall survival of the patients. Methods: 24 patients with non-small-cell lung cancer (NSCLC) and 20 patients with ovarian cancer (all in stage IIIb-IV) were treated with a combination of paclitaxel and carboplatin for six treatment cicles and they were separated into two groups in terms of survival. GROUP (A). Long survival (>12 months for NSCLC, >30 months for ovarian Ca) GROUP (B). Long survival (<12 months for NSCLC, <18 months for ovarian Ca) At the same time we combined the relevant parameters (CD3, CD4, CD8, CD56, CD34, IL-3, IFN-γ) with the quality of life during treatment with chemotherapy. The results were analyzed using ANOVA system. Results: We observed a significant statistical difference between the values of CD4 and CD4/CD8 after chemotherapy between group A and group B (p=0,001 and p=0,006). This means that the further increase of T-helper cells after chemotherapy has a better prognosis concerning survival. In addiction, statistically interesting, which we may call a prognostic factor, was the difference in values of IFN-γ between individuals of groups and B before and after chemotherapy (p=0,039 and p=0,027, respectively). Patients with high IL-3 had little chance of toxicity. Conclusions: In the current study we tried to demonstrate the effects from the use of the combination of carboplatin-paclitaxel in the whole population of Tcells/ cytokines and the reaction of them in the quality of life.

No description available.