Metastasis is the spread of cancer cells from the primary tumor to distant sites. It is the most dangerous attribute of cancer, and also the principle cause of cancerrelated morbidity and mortality. Metastasis suppressor genes are a group of genes that suppress tumor metastasis without significant effect on tumorigenicity. NM23 was the first identified metastasis suppressor gene, and loss of its expression is a frequent hallmark of metastatic growth in multiple cancers (e.g. melanoma, carcinomas of breast, stomach and liver). NM23-H1 possesses at least three enzymatic activities, including nucleoside diphosphate kinase (NDPK), histidine kinase (hisK), and a more recently described 3f-5f exonuclease (EXO). While the hisK has been shown to be linked to the suppression of cell motility, the NDPK has been reported to be unrelated to the suppression of metastatic potential indirectly. Relevance of EXO has not been addressed. Other known 3f-5f exonuclease are closely associated with DNA repair functions, suggesting NM23-H1 may suppress mutations required for metastasis. As a transcription factor, NM23 has been shown to modestly downregulate the transcription on PDGF-A chain, a growth factor oncogene, either alone or in association with another transcriptional factor, Pur@. At the same time, identification of NM23-H1 as a 3f-5fexonuclease suggests the role of NM23-H1 in DNA repair. Etoposide and cisplatin elicited nuclear translocation of H1 within 4 h in HeLa and HepG2 cells, seen as accumulation of H1 in small intranuclear foci, strongly suggesting the DNA repair function of H1. To investigate the enzymatic function contributing to metastasis suppressor activity of H1, complementation system was used by transfecting NM23-H1 with individually disrupted enzymatic function into 2 melanoma cell lines, 1205LU and WM793. Overexpression of H1 in 1205LU suppressed lung metastasis in vivo without effect on indices of transformation (e.g. proliferation, soft agar colonization). EXO- deficient H1 and NDPK-deficient H1 lost suppression of lung metastasis, while hisK-deficient H1 maintained suppressor activity. Consistent with the results in 1205LU cells, EXO-deficient H1 and NDPKdeficient H1 lost suppression of the progression of WM793 cells in protein-free medium, while WT and hisK-deficient H1 prevented the progression. Taken together, these data suggest that the NDPK and/or 3f-5fEXO activity of H1 inhibits the progression of premetastatic cells to the metastatic phenotype, possibly via a DNA repair function or other structural transactions with DNA.

NM23-H1 represents the first identified metastasis suppressor, exhibiting reduced expression in breast carcinoma and melanoma, and an ability to inhibit metastatic growth without significant impact on the transformed phenotype. Although its molecular mechanism of action is not fully understood, NM23-H1 possesses at least three enzymatic activities that may mediate metastasis suppressor function. It catalyzes nucleoside diphosphate kinase (NDPK) activity, as well as protein histidine kinase and 3’-5’ exonuclease activities. As 3’-5’ exonucleases are generally required for maintenance of genomic integrity, this activity represents a plausible mediator to underlie the metastasis suppressor function of NM23-H1 protein. To investigate the relevant activity of NM23-H1 in metastasis suppression, we constructed a panel of NM23-H1 mutant variants with selective enzymatic lesions. Previous studies have identified some key amino acid residues important for the enzymatic characteristics of NM23-H1. However, none of them are selective for disrupting the 3’-5’ exonuclease activity. In this study, we show that a substitution of Glu5 to alanine results in a dramatic, selective loss in 3’-5’ exonuclease property without significant affecting other enzymatic activities. To measure the extent to which the exonuclease function opposes mutation and metastasis, NM23-deficient and metastatic cell lines with forced expression of NM23-H1 variants are analyzed in nude mice. In spontaneous metastasis models, NM23-H1 mutants deficient in 3‘-5’ exonuclease activity significantly disrupt the capacity of metastasis suppression of wild-type protein, indicating that the 3’-5’ exonuclease activity of NM23-H1 is necessary for the spontaneous metastasis-suppressing effects. As 3'-5' exonucleases are generally associated with DNA repair process, we have also studied the contributions of yeast NM23 homologue YNK1 to genomic integrity in Saccharomyces cerevisiae. Consistent with an antimutator function, ablation of YNK1 significantly results in increased mutation rates following exposure to UV irradiation and the alkylating agent methyl methanesulfonate (MMS). The impaired DNA-damage response of ynk1Δ cells suggests a role of human homologue NM23 in DNA repair. More evidence is being collected in our laboratory to demonstrate a role for NM23-H1 in maintaining genomic integrity. Collectively, our findings of DNA repair activity of NM23-H1 will contribute to the understandings of the mechanisms in metastasis suppression and new drug discoveries.

Metastasis is one of the main causes of death in cancer patients. Metastatic melanoma is a death sentence, as chemotherapeutic agents have a 5% success rate or do not extend survival beyond 10 months. The lack of effective chemotherapeutic agents for treating metastatic melanoma indicates a dire need to identify new drug targets and develop new therapies. Our lab has previously shown that the kinase activity of Abelson family of non-receptor tyrosine kinases (c-Abl and Arg) is elevated in invasive breast cancer cell lines as compared to non-invasive cell lines. Previous studies from our lab have shown that Abl kinases are convergent point of ErbB2 and Src Kinases in melanoma cells and Abl kinases promote invasion by an undefined mechanism. Although Abl kinases promote invasion, it is not known whether they are important for metastastic potential. For the first time, we report that Abl kinases promote melanoma cell proliferation, survival, matrigel-invasion and single-cell 3D invasion. To investigate the mechanism by which Abl kinases promote invasion, we found out that active c-Abl transcriptionally upregulates MMP-1, and using rescue approaches we show that c-Abl promotes invasion via a STAT3àMMP-1 pathway. In contrast, active Arg drives invasion in a STAT3-independent manner, and upregulates the expression of MMP-3 and MT1-MMP, in addition to MMP-1. We also found that Abl kinases promote invasion via lysosomal degradation of a metastasis suppressor, NM23-H1 by activating lysosomal cathepsins B and L, which directly cleave and degrade NM23-H1. Furthermore, c-Abl and Arg are activated in primary melanomas and cAbl/Arg activity is inversely correlated with NM23-H1 expression both in primary melanoma and human melanoma cells. We also demonstrate, for the first time that active Abl kinases promote metastasis in vivo, as inhibition of c-Abl/Arg with nilotinib, dramatically inhibits lung colonization/metastasis in a mouse model using two different melanoma cell lines. In summary, we identify Abl kinases as critical, novel, drug targets in metastatic melanoma, and our data indicate that nilotinib may be useful in preventing metastasis in a select group of patients, harboring active Abl kinases.

Colorectal cancer (CRC) is the fourth most common malignancy in the developed Western countries and represents the second leading cause of death from cancer after lung cancer. Despite better diagnostic tools and improvement of surgical standards, the prognosis of this disease is still unsatisfactory with a mean 5-year cancer-specific survival of approximately 50%, mainly due to our inability to predict liver failure secondary to hepatic involvement and still poorly effective palliative treatments. The metastatic cascade is a complex, multistep process driven by progressive accumulation of genetic alterations which result in proto-oncogene activation and inactivation of tumour suppressor genes. Among the additional pathways involved in this process, secretion of angiogenesis factors, proteolysis of the extra-cellular matrix (ECM) molecules, and probably inhibition of apoptosis are also known to facilitate tumour progression. We undertook a retrospective study in a series of paraffin-embedded human colonic tissues to investigate the prognostic significance of new tumour markers as predictors of liver metastasis and survival in sporadic CRC. This research was conducted in three parts: first, immunohistochemical studies of protein markers and development of a new quantitative method of measurement in a subset of the immunostained sections using a colour video image analysis (VIA) procedure; second, concomitant determination of the expression of the bcl-2 gene at the messenger RNA (mRNA) level, using a more specific methodology of in situ hybridisation (ISH) and investigation of the relationship between bcl-2 mRNA and its encoded protein expression; and third, investigation of microsatellite alterations at four loci using a fluorescent microsatelllite polymerase chain reaction (PCR) assay coupled with an automated DNA sequencer. In our initial immunohistochemical experiment, we found a good correlation between colour VIA and semiquantitative evaluation of nm23 immunoreactivity (IR) confirming the validity of such quantitative analysis (Pearson's correlation coefficient r=0.88; P<0.001). Furthermore, overexpression of nm23 was associated with an increased risk of developing liver metastases (logrank test for trend, P<0.001) and cancer-related death (P=0.002). We used the same quantitative method to determine the expression of urokinase-type plasminogen activator (u-PA) and c-erbB-2 (HER2/neu) proteins and found that neither predicted patient outcome. However, CRC showing overexpression of u-PA (above 85 pixels) had an increased risk of liver metastasis (P=0.013). Since this was a post hoc analysis we can not be confident that this represents a real effect. There were significant positive correlations among expression of all three markers, u-PA, c-erbB-2 and nm23 proteins (u-PA vs c-erbB-2, Spearman rank correlation coefficient, P=0.003; u-PA vs nm23, P<0.001; c-erbB-2 vs nm23, P=0.001) suggesting that, in vivo, all proteins interact or are similarly regulated. Semiquantitative analysis of the vascular endothelial growth factor (VEGF) protein showed that expression of VEGF was significantly reduced in the metastatic liver tumours compared to their matched primary ones (Wilcoxon test, P=0.002), suggesting VEGF activity to be secondarily down-regulated once the tumour cells reach the hepatic parenchyma. There was no strong evidence from our data that the level of VEGF in the primary tumour could predict risk of liver metastasis or survival duration. Finally, when semiquantitatively assessing five protein markers (nm23, p53, c-erbB- 2, u-PA, and VEGF) individually or in combination, we found that only nm23 protein expression was positively related to the risk of liver metastasis (logrank test, P<0.001); p53 protein expression was only marginally associated (P=0.091). Furthermore, patients with Dukes' stage B tumours showing positive expression of nm23 protein also demonstrated an increased risk of liver metastasis (P=0.001). Although the risk of developing liver secondaries was statistically correlated with the number of positive markers (NPM) and the cumulative intensity score (CIS) (logrank test for trend, P=0.004 and P=0.001 respectively), these two parameters did not improve the predictive value over and above that of nm23 protein alone. These results suggest that the only marker of the five we studied that provides prognostic information about the risk of developing liver metastasis in CRC is nm23. Its evaluation may be particularly useful in selecting high risk Dukes' stage B patients who should be considered for adjuvant chemotherapy. In the second part of this study, immunohistochemical analysis using a monoclonal mouse antibody to the bcl-2 protein and in situ hybridisation using digoxigenin-labelled bcl-2 cRNA probes were carried out on the same paraffin-embedded specimens and semiquantitatively assessed. These specimens also included adenomas with various degrees of dysplasia. The expression of bcl-2 protein was gradually and significantly lost during the progression from moderately dysplastic adenoma to primary CRC [moderate/severe dysplasia: Mann-Whitney U-test, P=0.0001; severe dysplasia/primary CRC: P=0.027], whereas the cellular expression of bcl-2 mRNA was progressively increased during the dysplasia/adenoma-carcinoma neoplastic sequence. Our observation suggests that in a proportion of CRCs the bcl-2 proto-oncogene expression may be down-regulated at a post-transcriptional level. In the final section of this study, we investigated the possible relationship between loss of heterozygosity (LOH) and microsatellite instability (MIN) at four microsatellite loci spanning the 17q21-23 region that includes the nm23-H1 and nm23- H2 genes, to the risk of liver metastasis and nm23 protein expression. Genomic DNA was extracted from the same series of paraffin-embedded colorectal specimens and a fluorescent PCR coupled with DNA fragment analysis in an automated DNA sequencer was applied. In 45% and 48% of the primary and secondary lesions, LOH was present in at least one locus. We found a positive correlation when looking for a trend comparing the fraction of sites with LOH at these loci to risk of liver recurrence (logrank test for trend, P=0.005). This remained an independent predictor after adjusting to T-stage (Multivariate Cox regression, P=0.022), N-stage (P=0.007), or Dukes' stage (P=0.012). MIN was present in 2 loci in 41% and 30% of the primary and secondary tumours respectively. Considering only the Dukes' B tumours, we found that an increasing number of sites showing MIN was associated with a reduced risk of liver recurrence (logrank test for trend, P=0.032). When comparing LOH and MIN status of the primary and matched liver secondary tumours with their corresponding normal tissue samples, we found concordant genomic alterations in 72% (NME1 locus) to 43% (D17S579). Finally, we observed a trend in association between the proportion of loci with LOH and nm23 positivity ( ?? 2 test for trend, P=0.024). These findings suggest that, genomic alterations in the 17q21-23 region may affect prognosis of CRC as well as regulation of the nm23 protein expression via a stillmechanism.

Colorectal cancer (CRC) is the fourth most common malignancy in the developed Western countries and represents the second leading cause of death from cancer after lung cancer. Despite better diagnostic tools and improvement of surgical standards, the prognosis of this disease is still unsatisfactory with a mean 5-year cancer-specific survival of approximately 50%, mainly due to our inability to predict liver failure secondary to hepatic involvement and still poorly effective palliative treatments. The metastatic cascade is a complex, multistep process driven by progressive accumulation of genetic alterations which result in proto-oncogene activation and inactivation of tumour suppressor genes. Among the additional pathways involved in this process, secretion of angiogenesis factors, proteolysis of the extra-cellular matrix (ECM) molecules, and probably inhibition of apoptosis are also known to facilitate tumour progression. We undertook a retrospective study in a series of paraffin-embedded human colonic tissues to investigate the prognostic significance of new tumour markers as predictors of liver metastasis and survival in sporadic CRC. This research was conducted in three parts: first, immunohistochemical studies of protein markers and development of a new quantitative method of measurement in a subset of the immunostained sections using a colour video image analysis (VIA) procedure; second, concomitant determination of the expression of the bcl-2 gene at the messenger RNA (mRNA) level, using a more specific methodology of in situ hybridisation (ISH) and investigation of the relationship between bcl-2 mRNA and its encoded protein expression; and third, investigation of microsatellite alterations at four loci using a fluorescent microsatelllite polymerase chain reaction (PCR) assay coupled with an automated DNA sequencer. In our initial immunohistochemical experiment, we found a good correlation between colour VIA and semiquantitative evaluation of nm23 immunoreactivity (IR) confirming the validity of such quantitative analysis (Pearson's correlation coefficient r=0.88; P<0.001). Furthermore, overexpression of nm23 was associated with an increased risk of developing liver metastases (logrank test for trend, P<0.001) and cancer-related death (P=0.002). We used the same quantitative method to determine the expression of urokinase-type plasminogen activator (u-PA) and c-erbB-2 (HER2/neu) proteins and found that neither predicted patient outcome. However, CRC showing overexpression of u-PA (above 85 pixels) had an increased risk of liver metastasis (P=0.013). Since this was a post hoc analysis we can not be confident that this represents a real effect. There were significant positive correlations among expression of all three markers, u-PA, c-erbB-2 and nm23 proteins (u-PA vs c-erbB-2, Spearman rank correlation coefficient, P=0.003; u-PA vs nm23, P<0.001; c-erbB-2 vs nm23, P=0.001) suggesting that, in vivo, all proteins interact or are similarly regulated. Semiquantitative analysis of the vascular endothelial growth factor (VEGF) protein showed that expression of VEGF was significantly reduced in the metastatic liver tumours compared to their matched primary ones (Wilcoxon test, P=0.002), suggesting VEGF activity to be secondarily down-regulated once the tumour cells reach the hepatic parenchyma. There was no strong evidence from our data that the level of VEGF in the primary tumour could predict risk of liver metastasis or survival duration. Finally, when semiquantitatively assessing five protein markers (nm23, p53, c-erbB- 2, u-PA, and VEGF) individually or in combination, we found that only nm23 protein expression was positively related to the risk of liver metastasis (logrank test, P<0.001); p53 protein expression was only marginally associated (P=0.091). Furthermore, patients with Dukes' stage B tumours showing positive expression of nm23 protein also demonstrated an increased risk of liver metastasis (P=0.001). Although the risk of developing liver secondaries was statistically correlated with the number of positive markers (NPM) and the cumulative intensity score (CIS) (logrank test for trend, P=0.004 and P=0.001 respectively), these two parameters did not improve the predictive value over and above that of nm23 protein alone. These results suggest that the only marker of the five we studied that provides prognostic information about the risk of developing liver metastasis in CRC is nm23. Its evaluation may be particularly useful in selecting high risk Dukes' stage B patients who should be considered for adjuvant chemotherapy. In the second part of this study, immunohistochemical analysis using a monoclonal mouse antibody to the bcl-2 protein and in situ hybridisation using digoxigenin-labelled bcl-2 cRNA probes were carried out on the same paraffin-embedded specimens and semiquantitatively assessed. These specimens also included adenomas with various degrees of dysplasia. The expression of bcl-2 protein was gradually and significantly lost during the progression from moderately dysplastic adenoma to primary CRC [moderate/severe dysplasia: Mann-Whitney U-test, P=0.0001; severe dysplasia/primary CRC: P=0.027], whereas the cellular expression of bcl-2 mRNA was progressively increased during the dysplasia/adenoma-carcinoma neoplastic sequence. Our observation suggests that in a proportion of CRCs the bcl-2 proto-oncogene expression may be down-regulated at a post-transcriptional level. In the final section of this study, we investigated the possible relationship between loss of heterozygosity (LOH) and microsatellite instability (MIN) at four microsatellite loci spanning the 17q21-23 region that includes the nm23-H1 and nm23- H2 genes, to the risk of liver metastasis and nm23 protein expression. Genomic DNA was extracted from the same series of paraffin-embedded colorectal specimens and a fluorescent PCR coupled with DNA fragment analysis in an automated DNA sequencer was applied. In 45% and 48% of the primary and secondary lesions, LOH was present in at least one locus. We found a positive correlation when looking for a trend comparing the fraction of sites with LOH at these loci to risk of liver recurrence (logrank test for trend, P=0.005). This remained an independent predictor after adjusting to T-stage (Multivariate Cox regression, P=0.022), N-stage (P=0.007), or Dukes' stage (P=0.012). MIN was present in 2 loci in 41% and 30% of the primary and secondary tumours respectively. Considering only the Dukes' B tumours, we found that an increasing number of sites showing MIN was associated with a reduced risk of liver recurrence (logrank test for trend, P=0.032). When comparing LOH and MIN status of the primary and matched liver secondary tumours with their corresponding normal tissue samples, we found concordant genomic alterations in 72% (NME1 locus) to 43% (D17S579). Finally, we observed a trend in association between the proportion of loci with LOH and nm23 positivity ( ?? 2 test for trend, P=0.024). These findings suggest that, genomic alterations in the 17q21-23 region may affect prognosis of CRC as well as regulation of the nm23 protein expression via a stillmechanism.

Uveal Melanoma (UM) is the most common malignant intra-ocular tumor in adults. Despite the high accuracy of clinical diagnosis and advances in local treatment, more than 50% of UM patients develop metastasis within ten years of initial diagnosis. NM23 is a human metastasis suppressor gene. Reduced Nm23-H1 expression is correlated with high metastatic potential in a variety of different cancers including melanoma. C-Met is a receptor tyrosine kinase (RTK) that has been known to stimulate the invasive growth and increase the metastatic potential of cancer cells. Expression of c-Met is correlated with high mortality rate in UM patients. Treatment with CQX-2 inhibitors showed promise as an adjuvant therapy in adenocarcinoma of the colon. A previous report from our laboratory showed that topical treatment with Nepafenac (a CQX-2 inhibitor) delayed the progression of the primary tumor and the formation of metastasis in the experimental rabbit model of UM. / The purpose of this thesis is to investigate the expression levels ofNm23-H1 in UM cell lines with different metastatic potentials, in paraffin embedded tissues from primary tumors of UM patients, and in an experimental rabbit model. In addition, the aim of this thesis is to determine whether treating human uveal melanoma cell lines with Nepafenac would increase the expression levels of Nm23-H1 and decrease the expression levels of c-Met in vitro (UM cell lines) and in vivo (experimental rabbit model). / To achieve our goal, we used several types of assays in our UM cell lines and paraffin embedded tissues from patient samples and experimental rabbit model, including quantitative immunostaining, quantitative Real-time PCR, and small interference RNA (siRNA). / The Real-time PCR results of five human uveal melanoma cell lines showed that expression of Nm23-HI is higher in cell lines with low metastatic potential compared to those with high metastatic potential. The invasive ability of the uveal melanoma cell lines increased after silencing Nm23-H1 expression with siRNA. The increased immunostaining intensity of Nm23-H1 in patient samples is associated with better survival rate. Moreover, treatment with Nepafenac resulted in increase of Nm23-H1 levels and decrease of c-Met levels in both the UM cell lines and the experimental rabbit model. / In conclusion, Nm23-H1 is a potent prognostic marker to predict the survival rate of UM patients and it has the potential to identify high-risk patients. To the best of our knowledge, this is the first study to show that treatment with COX-2 inhibitor causes an upregulation of Nm23-H1 and downregulation of c-Met in UM. Therefore, treatment with COX-2 inhibitors may be a useful strategy as an adjuvant therapy for UM patients.

Cancer patient mortality is caused by the ability of tumor cells to invade the extracellular matrix and metastasize. Our lab was the first to identify the role of Abl family of non-receptor tyrosine kinases (c-Abl and Arg) in the progression of solid tumor cancers. In our previous studies, we showed that high c-Abl/Arg activity promotes proliferation, invasion, and metastasis in melanoma and breast cancer cells lines. Here, we demonstrate that our previous findings are clinically relevant by showing increased c-Abl/Arg kinase activity in primary melanoma tumor tissue in comparison to low activity as compared to benign nevi. Additionally, in breast cancer tissue, we found aggressive tumor subtypes (triple-negative and high-grade breast cancer) had increased c-Abl/Arg activity as compared to less aggressive subtypes. To define the mechanism by which c-Abl and Arg promote melanoma and breast cancer metastasis, we searched for novel pathways by which c-Abl and Arg promote invasion, a key step in metastasis. Significantly, we found that c-Abl and Arg decrease the expression of non-metastatic protein, NM23-H1, a metastasis suppressor that is lost during metastatic progression. We demonstrate that NM23-H1 is localized and degraded within the lysosome via proteases, cathepsins L and B. Moreover, we show that c-Abl and Arg upregulate cathepsin mRNA levels and activate the cathepsins, which in-turn degrade NM23-H1. We demonstrate that this pathway is functionally significant as c-Abl and Arg require the downregulation of NM23-H1 to promote invasion in melanoma and breast cancer cell lines. We show that the pathway is clinically significant as c-Abl/Arg activity is inversely correlated with NM23-H1 expression in mouse lung metastases, as well as in human primary melanoma and primary breast cancer tissue. In summary, we are the first to demonstrate novel crosstalk between oncogenic and metastasis suppressor signaling pathways, and provide evidence that pharmacological inhibition of Abl family kinases in melanoma and breast cancer patients may prevent metastatic progression by stabilizing a metastasis suppressor.

No description available.

Made available in DSpace on 2015-07-22T20:50:43Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-05-27. Added 1 bitstream(s) on 2015-08-11T03:25:25Z : No. of bitstreams: 1 Publico-00212.pdf: 1245886 bytes, checksum: 4848d0e58992c5a9ff4b387a14c2534a (MD5) / Objetivos: Analisar a expressão das proteínas CD10 e NM23 por estudo imunohistoquímico do tecido do carcinoma colorretal e da mucosa adjacente. Avaliar a relação da expressão dessas proteínas com os aspectos anatomopatológicos da neoplasia, estadiamento clínico, ocorrência de metástases hepáticas e prognóstico dos doentes. Método: Cento e trinta doentes operados por carcinoma colorretal foram analisados. Bloco de tissue microarray foi confeccionado com tecido neoplásico e com a mucosa não neoplásica adjacente. Estudo imuno-histoquímico foi realizado com anticorpos monoclonais NM23 e CD10 no tecido neoplásico e no tecido não neoplásico da mucosa adjacente. A leitura foi realizada por aparelho de escaneamento de lâminas. A imunoexpressão foi avaliada pelo percentual de células coradas e foram obtidos escores de intensidade. Foram considerados como positivos para CD 10 os tumores que expressavam o marcador em mais de 10% das células neoplásicas. Para NM 23 considerou-se dois grupos divididos em fortes expressores (mais de 50%) e expressores fracos (menos de 50%) das células coradas. Os resultados foram relacionados com as características morfológicas e histopatológicas do carcinoma colorretal, estadiamento clínico, presença de metástases hepáticas e com o prognóstico. No estudo estatístico foram utilizados os testes de Mann-Whitney, Kruskal- Wallis e exato de Fisher. A sobrevivência foi avaliada utilizando a curva de Kaplan- Meier, e o desfecho de comparação entre as curvas foi calculado pelo teste de Long rank. Resultados: Ambos os marcadores CD10 e NM23 apresentaram expressão maior no tecido do carcinoma do que na mucosa não neoplásica adjacente (p<0,0001 para ambos). A imunoexpressão tecidual das proteínas NM23 e CD10 não apresentou relação com o grau de diferenciação celular (p=0,57 e p=0,48, respectivamente), invasão vascular (p=0,85 e p=0,67, respectivamente), invasão linfática (p=0,41 e 0,73, respectivamente), infiltração perineural (p=0,46 e p=0,24, respectivamente) e com o estadiamento pela classificação TNM (p=0,19 para ambos). A imunoexpressão de CD10 no tecido do carcinoma colorretal foi maior (p=0,05) nas neoplasias exofíticas do que nos tumores não exofíticos. A expressão das proteínas NM23 e CD10 não apresentou relação com a incidência de metástases linfonodais (p=0,08 e 0,30, respectivamente). A expressão tecidual dos marcadores NM23 e CD10 não se relacionou com a ocorrência de metástases hepáticas (p=0,59 e p=0,31, respectivamente). A sobrevivência livre de doença mostrou relação significante (p=0,01) com a maior intensidade de imunoexpressão da proteína NM23 no tecido do carcinoma colorretal, o mesmo não ocorrendo com a imunoexpressão da proteína CD10 (p=0,18). A sobrevivência global não mostrou relação com as expressões das proteínas NM23 e CD10 (p=0,13 e p=0,24, respectivamente). Conclusões: O tecido neoplásico do carcinoma colorretal expressou mais intensamente as proteínas NM23 e CD10 do que a mucosa não neoplásica adjacente. A imunoexpressão de CD10 no tecido do carcinoma colorretal foi maior (p=0,05) nas neoplasias exofíticas do que nos tumores não exofíticos. A expressão das proteínas NM23 e CD10 não se relacionou com os demais aspectos anatomopatológicos da neoplasia, com a presença de metástase hepática e com o estadiamento do carcinoma colorretal. Os doentes com imunoexpressão aumentada da proteína NM23 apresentaram sobrevivência livre de doença significativamente maior. A intensidade da imunoexpressão tecidual da proteína CD10 não influenciou a sobrevivência livre de doença e a sobrevivência global não se relacionou com a imunoexpressão das proteínas NM23 e CD10. / Aims: To analyze the tissue expression of the proteins CD 10 and NM 23 through the immunohistochemichal study of the colorectal carcinoma and evaluate the expression relation of these proteins with the anatomopathological aspects of the neoplasia, clinical staging, occurrence of hepatic metastasis and patients’ prognostic. Method: One hundred and thirty operated patients of colorectal carcinoma have been analyzed. A block of tissue microarray was produced with the neoplastic mucosa and with the adjacent non-neoplastic mucosa. An immunohistochemichal study was performed with monoclonal antibodies NM23 and CD10 on the neoplastic tissue and non-neoplastic tissue of the adjacent mucosa. The interpretation of the slides was made by a scanner device. The immunoexpression was evaluated by the percentage of colored cells and the obtained intensity scores. The results were related to the morphological and histopathological characteristics of the carcinoma, clinical staging, presence of hepatic metastasis and to the prognostic of the patients. In the statistic study were used the Mann-Whitney test, the Kruskal-Wallis test and Fisher’s exact test. The analysis of survival was conducted with the use of the Kaplan-Meier curve and the comparison conclusion between the curves was calculated through the Longrank test. Results: Both markers CD10 and NM23 presented a higher expression on the carcinoma tissue rather than on the non-neoplastic adjacent mucosa (p<0,0001 for both). The expression of the proteins NM23 and CD10 did not present any relation to the degree of cellular differentiation (p=0,57 and p=0,48, respectively) , vascular invasion (p=0,85 and p=0,67, respectively), lymphatic invasion (p=0,41and 0,73, respectively), perineural infiltration (p=0,46 and p=0,24, respectively) and with the staging by the TNM classification (p=0,19). The immunoexpression of CD10 on the colorectal carcinoma tissue was higher (p=0,15) on the exophytic neoplasias than on the non-exophytic tumors. The expression of the proteins NM23 and CD10 did not present any relation with the incidence of lymphonodal metastasis (p=0,08 and 0,30, respectively). The tissue expression of the markers NM23 and CD10 did not relate to the occurrence of hepatic metastasis (p=0,59 and 0,31 respectively). The disease-free survival disclosed a significant relation (p=0,01) with a higher intensity of immunoexpression of the protein NM23 on the colorectal carcinoma’s tissue. However, the same did not occur with the immunoexpression of the protein CD10 (p=0,18). The global survival did not show any relation with the expression of the proteins NM23 and CD10 (p=0,13 and p=0,24, respectively). Conclusions: The neoplastic tissue of the colorectal carcinoma expresses more intensely the proteins NM23 and CD10 than the adjacent nonneoplastic mucosa. The expression of the proteins NM23 and CD10 does not relate to the presence of lymphonodal metastasis, hepatic metastasis, degree of cellular differentiation, colonic or rectal localization of the neoplasia, presence of vascular and/or lymphatic invasion, presence of neural infiltration and the staging of the colorectal carcinoma. The patients with increased immunoexpression of the protein NM23 presented a disease-free survival significantly higher. The intensity of the tissue immunoexpression of the protein CD10 did not influence the disease-free survival. The global survival does not relate to the immunoexpression of the proteins NM23 and CD10. / TEDE / BV UNIFESP: Teses e dissertações