Centrosome aberrations and tumor development

Sammanfattning: The transformation of a normal cell to a tumor cell is a result of a failure somewhere in a complicated system of coordination, checkpoints and control mechanisms of inhibitors and activators working as triggers and transactivators of genes of importance for cell cycle progression. In the present thesis, the focus is on an important function that controls the segregation of the chromosomes during cell division to the next generation of two daughter cells. The conductor of this nice orchestration is the centrosome, which picks up the exact number chromosomes assembled in metaphase plate and brings them to each of the two creating nuclei. Since transformation to tumor cells includes rearrangements of the genetic material, the centrosome plays a crucial role and is therefore of interest to investigate. We have used both in vivo and in vitro systems to elucidate the role of centrosomes in tumor development. Studying the progression of cancer in the human cervical epithelia, we analyzed condyloma, intraepithelial dysplasia and neoplasia (CIN I-III) and cervical cancer, and found an increased number of aberrant centrosomes first detected in the mild CIN I lesions. Thereafter in moderate CIN II and severe CIN III to invasive cancer a steadily increasing number of centrosomes above the normal number of 2 were registered in immunohistochemical staining with ã-tubulin. The results stress attention to the question whether the disturbances in centrosome function precedes malignification or not. We also found a transient appearance of tetraploid cells in the intraepithelial lesions. Tetraploidization results in an increased number of centrosomes for the cell to deal with when the tetraploid cell enters mitosis enhancing the risk for missegregation of chromosomes. All invasive cervical cancers investigated were aneuploid. The literature reveals a low frequency of p53 mutation in these cancers. But a high frequency of high-risk HPV revealed p53 function disturbance. In adrenocortical tumors, both adenomas and carcinomas were analyzed with regard to aberrant centrosome numbers. Among fibroblasts, either one or two signals in ã-tubulin immunohistochemical staining was found in 99.7%. However in the benign adenomas, a mean of 4% of the cells showed more than two signals indicating amplification. In adrenocortical carcinomas, a mean of 10% of the cells had more than two signals. Again the question of centrosome aberrations to occur early in carcinogenesis is raised. The literature reveals a low frequency of p53 mutation in these cancer but most tumors are aneuploid. The in vitro system used was human breast cancer cell lines. In four selected lines, one was diploid-tetraploid (Hs578T), one tetraploid (MCF7) and one aneuploid (MDA-MB-231). Hs578T and MDAMB-231 were p53 mutated. We also added a non-transformed fibroblast line (HDF). The two p53 mutated lines revealed the highest frequency of aberrant centrosomes. By perturbing the mitosis with griseofulvin, we pushed in various frequencies all four lines to a higher ploidy level and found the p53 mutated lines to be more vulnerable according to centrosome function. By adding 13-cis-retinoic acid (13cRA) to MDA-MB-231, a reduction in aberrant centrosomes was obtained. However, no effect on cyclinA, D and E could be seen and thereby no restoration of G1 checkpoint function occurred related to the cyclins investigated. We conclude 13cRA to exert its effect upon structures as cytoskeleton and mitotic spindle formation and also push cells into apoptosis. Centrosomes have come into focus in the cancer literature the last decade. The present thesis shows that they must be studied further and maybe also be used in cancer diagnosis.