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ASPM and microcephalin expression in epithelial ovarian cancer correlates with tumour grade and survival.

Brüning-Richardson A, Bond J, Alsiary R, Richardson J, Cairns DA, McCormack L, Hutson R, Burns P, Wilkinson N, Hall GD, Morrison EE, Bell SM - Br. J. Cancer (2011)

Bottom Line: Two microcephaly-associated proteins, abnormal spindle-like microcephaly-associated protein (ASPM) and microcephalin, are involved in mitosis and DNA damage repair.Their expression is deregulated at the RNA level in EOC.Furthermore, low levels of nuclear microcephalin correlated with reduced patient survival.

View Article: PubMed Central - PubMed

Affiliation: Section of Ophthalmology and Neurosciences, Leeds Institute of Molecular Medicine, Welcome Trust Brenner Building, St James's University Hospital, Leeds LS9 7TF, UK.

ABSTRACT

Background: The clinico-pathological and molecular heterogeneity of epithelial ovarian cancer (EOC) complicates its early diagnosis and successful treatment. Highly aneuploid tumours and the presence of ascitic fluids are hallmarks of EOC. Two microcephaly-associated proteins, abnormal spindle-like microcephaly-associated protein (ASPM) and microcephalin, are involved in mitosis and DNA damage repair. Their expression is deregulated at the RNA level in EOC. Here, ASPM and microcephalin protein expression in primary cultures established from the ascites of patients with EOC was determined and correlated with clinical data to assess their suitability as biomarkers.

Methods: Five established ovarian cancer cell lines, cells derived from two benign ovarian ascites samples and 40 primary cultures of EOC derived from ovarian ascites samples were analysed by protein slot blotting and/or immunofluorescence to determine ASPM and microcephalin protein levels and their cellular localisation. Results were correlated with clinico-pathological data.

Results: A statistically significant correlation was identified for ASPM localisation and tumour grade, with high levels of cytoplasmic ASPM correlating with grade 1 tumours. Conversely, cytoplasmic microcephalin was only identified in high-grade tumours. Furthermore, low levels of nuclear microcephalin correlated with reduced patient survival.

Conclusion: Our results suggest that ASPM and microcephalin have the potential to be biomarkers in ovarian cancer.

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Related in: MedlinePlus

Nuclear and/or cytoplasmic microcephalin levels are associated with tumour grade and survival. (A) Weak microcephalin levels in the nucleus are associated with higher-grade tumours. (B) Cytoplasmic microcephalin is only detected in higher-grade tumours (P=0.047). (C) Kaplan–Meier survival plot showing survival for groups with weak nuclear microcephalin levels and medium/strong nuclear microcephalin levels (P=0.081).
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fig6: Nuclear and/or cytoplasmic microcephalin levels are associated with tumour grade and survival. (A) Weak microcephalin levels in the nucleus are associated with higher-grade tumours. (B) Cytoplasmic microcephalin is only detected in higher-grade tumours (P=0.047). (C) Kaplan–Meier survival plot showing survival for groups with weak nuclear microcephalin levels and medium/strong nuclear microcephalin levels (P=0.081).

Mentions: Immunofluorescence staining with the microcephalin antibody on 36 samples revealed three distinct staining patterns: nuclear, cytoplasmic and nuclear foci. After establishing cutoff points following calculation of staining intensities, microcephalin levels in the nucleus were classified as weak, medium or strong for each sample. Overall, nuclear microcephalin expression in the majority of the samples fell into the medium or strong category. Cytoplasmic staining was seen in only 25% of the samples with all exhibiting a weak staining pattern. Twenty-five percent of the samples displayed strong multiple nuclear foci staining with the remaining samples displaying only one or two strong foci per nucleus. Figure 4 shows examples of each staining pattern (Figure 4A) and a summary of the microcephalin localisation pattern (Figure 4B). The two control samples, 1078-A1 and 1153-A1, displayed strong nuclear staining and strong foci staining was also detected in 1153-A1 (Supplementary Figure 5B). These are therefore likely to represent the ‘normal' localisation and expression level of microcephalin in cultured ovarian epithelial cells. The most common staining pattern observed in the tumour-derived primary cultures was nuclear staining (Figure 5A); therefore, the fluorescence data from these observations were plotted for each individual primary culture sample in comparison to the established cell lines (Figure 5B). The majority of the samples had nuclear staining intensities similar to the cell lines with the exception of 1108-A1 (markedly lower) and 1143-A1 (higher). Clinical data were available for 32 primary cultures. Interestingly, weak nuclear staining was found only in the higher-grade tumours (moderately differentiated, grade 2, to poorly differentiated, high grade 3). In primary cultures of all patients with well-differentiated tumours (grade 1), medium or strong nuclear staining was observed (Figure 6A). An absence of cytoplasmic microcephalin immunostaining was observed in low/well differentiated (grade 1) tumours, whereas cytoplasmic staining correlated with grade 2 and 3 tumours exclusively (P=0.047; Figure 6B). This finding suggests that there is a positive correlation between the aberrant cytoplasmic localisation of microcephalin and tumour development. No association was found between nuclear foci staining and the clinical data.


ASPM and microcephalin expression in epithelial ovarian cancer correlates with tumour grade and survival.

Brüning-Richardson A, Bond J, Alsiary R, Richardson J, Cairns DA, McCormack L, Hutson R, Burns P, Wilkinson N, Hall GD, Morrison EE, Bell SM - Br. J. Cancer (2011)

Nuclear and/or cytoplasmic microcephalin levels are associated with tumour grade and survival. (A) Weak microcephalin levels in the nucleus are associated with higher-grade tumours. (B) Cytoplasmic microcephalin is only detected in higher-grade tumours (P=0.047). (C) Kaplan–Meier survival plot showing survival for groups with weak nuclear microcephalin levels and medium/strong nuclear microcephalin levels (P=0.081).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3101901&req=5

fig6: Nuclear and/or cytoplasmic microcephalin levels are associated with tumour grade and survival. (A) Weak microcephalin levels in the nucleus are associated with higher-grade tumours. (B) Cytoplasmic microcephalin is only detected in higher-grade tumours (P=0.047). (C) Kaplan–Meier survival plot showing survival for groups with weak nuclear microcephalin levels and medium/strong nuclear microcephalin levels (P=0.081).
Mentions: Immunofluorescence staining with the microcephalin antibody on 36 samples revealed three distinct staining patterns: nuclear, cytoplasmic and nuclear foci. After establishing cutoff points following calculation of staining intensities, microcephalin levels in the nucleus were classified as weak, medium or strong for each sample. Overall, nuclear microcephalin expression in the majority of the samples fell into the medium or strong category. Cytoplasmic staining was seen in only 25% of the samples with all exhibiting a weak staining pattern. Twenty-five percent of the samples displayed strong multiple nuclear foci staining with the remaining samples displaying only one or two strong foci per nucleus. Figure 4 shows examples of each staining pattern (Figure 4A) and a summary of the microcephalin localisation pattern (Figure 4B). The two control samples, 1078-A1 and 1153-A1, displayed strong nuclear staining and strong foci staining was also detected in 1153-A1 (Supplementary Figure 5B). These are therefore likely to represent the ‘normal' localisation and expression level of microcephalin in cultured ovarian epithelial cells. The most common staining pattern observed in the tumour-derived primary cultures was nuclear staining (Figure 5A); therefore, the fluorescence data from these observations were plotted for each individual primary culture sample in comparison to the established cell lines (Figure 5B). The majority of the samples had nuclear staining intensities similar to the cell lines with the exception of 1108-A1 (markedly lower) and 1143-A1 (higher). Clinical data were available for 32 primary cultures. Interestingly, weak nuclear staining was found only in the higher-grade tumours (moderately differentiated, grade 2, to poorly differentiated, high grade 3). In primary cultures of all patients with well-differentiated tumours (grade 1), medium or strong nuclear staining was observed (Figure 6A). An absence of cytoplasmic microcephalin immunostaining was observed in low/well differentiated (grade 1) tumours, whereas cytoplasmic staining correlated with grade 2 and 3 tumours exclusively (P=0.047; Figure 6B). This finding suggests that there is a positive correlation between the aberrant cytoplasmic localisation of microcephalin and tumour development. No association was found between nuclear foci staining and the clinical data.

Bottom Line: Two microcephaly-associated proteins, abnormal spindle-like microcephaly-associated protein (ASPM) and microcephalin, are involved in mitosis and DNA damage repair.Their expression is deregulated at the RNA level in EOC.Furthermore, low levels of nuclear microcephalin correlated with reduced patient survival.

View Article: PubMed Central - PubMed

Affiliation: Section of Ophthalmology and Neurosciences, Leeds Institute of Molecular Medicine, Welcome Trust Brenner Building, St James's University Hospital, Leeds LS9 7TF, UK.

ABSTRACT

Background: The clinico-pathological and molecular heterogeneity of epithelial ovarian cancer (EOC) complicates its early diagnosis and successful treatment. Highly aneuploid tumours and the presence of ascitic fluids are hallmarks of EOC. Two microcephaly-associated proteins, abnormal spindle-like microcephaly-associated protein (ASPM) and microcephalin, are involved in mitosis and DNA damage repair. Their expression is deregulated at the RNA level in EOC. Here, ASPM and microcephalin protein expression in primary cultures established from the ascites of patients with EOC was determined and correlated with clinical data to assess their suitability as biomarkers.

Methods: Five established ovarian cancer cell lines, cells derived from two benign ovarian ascites samples and 40 primary cultures of EOC derived from ovarian ascites samples were analysed by protein slot blotting and/or immunofluorescence to determine ASPM and microcephalin protein levels and their cellular localisation. Results were correlated with clinico-pathological data.

Results: A statistically significant correlation was identified for ASPM localisation and tumour grade, with high levels of cytoplasmic ASPM correlating with grade 1 tumours. Conversely, cytoplasmic microcephalin was only identified in high-grade tumours. Furthermore, low levels of nuclear microcephalin correlated with reduced patient survival.

Conclusion: Our results suggest that ASPM and microcephalin have the potential to be biomarkers in ovarian cancer.

Show MeSH
Related in: MedlinePlus