Biomedical subjects

M I Koukourakis

Publications and source records attributed to M I Koukourakis.

At least 19 recordsLinked to original sources

PubMed2006

Hypoxia inducible factors 1alpha and 2alpha are associated with VEGF expression and angiogenesis in gallbladder carcinomas.

AIMS: To investigate the significance of the hypoxia inducible factors HIF-1alpha and HIF-2alpha in gallbladder adenocarcinomas and their relation to angiogenesis and to the expression of VEGF, an angiogenic factor transcriptionally regulated by HIFalphas. METHODS: HIF-1alpha and 2alpha expression was assessed immunohistochemically in 60 patients with early gallbladder adenocarcinomas, treated with surgery alone. In addition, the vascular density (VD) and the expression of the angiogenic factors VEGF and thymidine phosphorylase (TP) were examined. The results were correlated with clinico-pathological features and prognosis. RESULTS: Overexpression of HIF-1alpha and 2alpha was significantly associated with increased tumor angiogenesis and VEGF expression, while HIF-2alpha was linked with upregulation of TP. None of these factors were associated with T-stage and tumor grade. Although HIFs did not relate significantly with prognosis, patients with HIF-1/2 expression who failed to switch-on VEGF or intratumoral angiogenesis had a favorable outcome. CONCLUSION: Hypoxia inducible factors are upregulated in a large proportion of gallbladder adenocarcinomas, a feature strongly related to increased expression of VEGF and intensified angiogenesis.

Adenocarcinoma↗

PubMed2005

LYVE-1 immunohistochemical assessment of lymphangiogenesis in endometrial and lung cancer.

AIMS/METHODS: Normal and malignant pulmonary and endometrial tissues were analysed for lymphatic vessels to assess the process of lymphangiogenesis and its role at these sites, using specific immunostaining for LYVE-1 and the panendothelial marker CD31. RESULTS: Lymphatics were clearly demonstrated in some normal tissues (myometrium, bronchial submucosa, and intestinal submucosa), but not in others (endometrium and alveolar tissue). LYVE-1 positive lymphatic vessels were detected at the tumour periphery of endometrial and lung carcinomas, but not within the main tumour mass. Double staining for LYVE-1 and the MIB1 proliferation marker revealed a higher proliferation index in lymphatic endothelial cells at the invading front of endometrial carcinomas, compared with myometrial areas distal to the tumour. Lung and endometrial carcinomas did not have an intratumorous lymphatic network. CONCLUSIONS: Although lymphangiogenesis may occur at the invading tumour front, incorporated lymphatics do not survive. Therefore, the dissemination of cancer cells through the lymphatics may occur by invasion of peripheral cancer cells into the adjacent normal lymphatics, or through shunts eventually produced at the invading tumour front as a consequence of active angiogenesis and lymphangiogenesis.

Adenocarcinoma↗

PubMed2005

Proliferating fibroblasts at the invading tumour edge of colorectal adenocarcinomas are associated with endogenous markers of hypoxia, acidity, and oxidative stress.

BACKGROUND: Stroma frequently forms at sites of active tumour invasion, and may be important for tumour growth and progression. The term "stromatogenesis" is used to describe this unique process that involves host peritumorous fibroblasts and is very different to reactive fibrosis. AIMS/METHODS: To investigate the activation status of host fibroblasts at the invading tumour edge, assessed as MIB1 proliferation index and thymidine phosphorylase (TP) expression. Results were related to vascular density and certain properties of invading cancer cells-MIB1 proliferation activity, TP expression, expression of endogenous markers of hypoxia (hypoxia inducible factor-1alpha; HIF1alpha) and acidity (lactate dehydrogenase-5; LDH5). Standard immunohistochemical techniques were applied to 150 colorectal adenocarcinomas. RESULTS: Normal fibroblasts at the tumour edge had a median MIB1 index of 2%-significantly higher than normal submucosal fibroblasts (0.3%) and significantly lower than cancer cells (40%). Normal peritumorous fibroblasts with a proliferation rate above the median strongly expressed TP and were supported by an increased vascular network. Cancer cells close to these fibroblasts had a high MIB1 proliferative index, high HIF1alpha and LDH5 reactivity, and a clear trend to extramural extension. All associations were significant. CONCLUSIONS: These results suggest that activated fibroblastic status at the invading tumour front sets the stage for stromatogenesis and new blood vessel formation, facilitating deep transmural invasion in colorectal adenocarcinomas. This complicity of peritumorous fibroblasts in the overall aggressiveness/invasive and metastatic ability of colorectal tumours, occurring within the framework of cancer-stromal cell interactions, is probably favoured by the altered microenvironmental conditions of hypoxia and acidity.

Adenocarcinoma↗

PubMed2004

Angiogenesis and vascular survival ability in ovarian adenocarcinomas.

This study was designed to investigate the prognostic relevance of ovarian tumour angiogenesis in terms of tumour angiogenic activity (TAA) and vascular survival ability (VSA), i.e. the ability of newly formed vessels to survive once incorporated into the main tumour mass. TAA was assessed at the edge of the invading tumour, while VSA was evaluated in inner tumour areas, always in comparison with the invading edge. A series of 46 ovarian grade-1 adenocarcinomas of the endometrioid and the serous cell type was assessed. Endothelial cells were revealed after using a standard immunohistochemical technique and the CD31 antibody. Vascular density was, in general, higher at the periphery of the tumour than in the inner tumour areas, although in both cases, a rich vascular supply was associated with a poor survival. By combining counts at the tumour edge versus inner tumour areas (edvin), four groups of tumour vascularity emerged: edvin type 1 (low TAA/low VSA), edvin type 2 (low TAA/high VSA), edvin type 3 (high TAA/low VSA) and edvin type 4 (high TAA/high VSA). Edvin type-4 tumours were related to the most unfavourable prognosis. It is concluded that VSA and TAA are complementary procedures in assessing ovarian tumour vasculature and, therefore, prognosis, and by combining the two parameters, a more precise impression of the state of vascularisation in the ovary is obtained, which may prove useful in designing anti-angiogenic therapies.

Adenocarcinoma↗

PubMed2004

Tumour angiogenic activity and vascular survival ability in bladder carcinoma.

BACKGROUND: Tumour angiogenic activity (TAA) is an important prognostic factor in many human tumours, including transitional cell carcinomas of the urinary bladder. The new tumour vessels are formed in the invading tumour front. This peripheral tumour area is internalised as soon as the growing tumour forms a new front. AIMS: To investigate and compare TAA with the ability of the tumour vasculature to survive (VSA) in inner tumour areas. METHODS: Fifty one cystectomy specimens with transitional cell carcinoma of the urinary bladder were studied. Sections were stained immunohistochemically for endothelial cells and proliferation activity, using the monoclonal antibodies CD31 and MIB-1, respectively. TAA was studied at the invading tumour edge-designated as the mean number of blood vessels in three "hot spots" at this site. VSA was assessed by comparing the vascular density in peripheral and inner tumour areas. RESULTS: High TAA at the invading tumour edge significantly correlated with lymph node involvement, but not with patient survival. Extensive lymphocytic infiltration was more frequent in tumours with high TAA. VSA was significantly higher in tumours of high proliferation index, high histological grade, advanced T stage, and poor prognosis. However, there was no association with metastasis to regional lymph nodes. CONCLUSION: VSA and TAA provide a more complete profile of the tumour vasculature and are associated with aggressive tumour behaviour in transitional cell carcinomas of the urinary bladder. The qualitative information provided by VSA may be important for the identification of angiogenic tumours with differential responses to various antiangiogenic treatments.

Adult↗

PubMed2003

Lactate dehydrogenase-5 (LDH-5) overexpression in non-small-cell lung cancer tissues is linked to tumour hypoxia, angiogenic factor production and poor prognosis.

Lactate dehydrogenase-5 (LDH-5) catalyses the reversible transformation of pyruvate to lactate, having a principal position in the anaerobic cellular metabolism. Induction of LDH-5 occurs during hypoxia and LDH-5 transcription is directly regulated by the hypoxia-inducible factor 1 (HIF1). Serum LDH levels have been correlated with poor prognosis and resistance to chemotherapy and radiotherapy in various neoplastic diseases. The expression, however, of LDH in tumours has never been investigated in the past. In the present study, we established an immunohistochemical method to evaluate the LDH-5 overexpression in tumours, using two novel antibodies raised against the rat muscle LDH-5 and the human LDH-5 (Abcam, UK). The subcellular patterns of expression in cancer cells were mixed nuclear and cytoplasmic. In direct contrast to cancer cells, stromal fibroblasts were reactive for LDH-5 only in a minority of cases. Serum LDH, although positively correlated with, does not reliably reflect the intratumoral LDH-5 status. Lactate dehydrogenase-5 overexpression was directly related to HIF1alpha and 2alpha, but not with the carbonic anhydrase 9 expression. Patients with tumours bearing high LDH-5 expression had a poor prognosis. Tumours with simultaneous LDH-5 and HIF1alpha (or HIF2alpha) overexpression, indicative of a functional HIF pathway, had a particularly aggressive behaviour. It is concluded that overexpression of LDH-5 is a common event in non-small-cell lung cancer, can be easily assessed in paraffin-embedded material and provides important prognostic information, particularly when combined with other endogenous markers of hypoxia and acidity.

Adult↗

PubMed2003

Molecular analysis of local relapse in high-risk breast cancer patients: can radiotherapy fractionation and time factors make a difference?

Large primary breast tumours and extensive lymph node involvement are linked to a high rate of local recurrence after surgery. In 10-20% of such high-risk breast cancer patients, local relapse will occur despite postoperative radiotherapy. In the present study, we investigated whether molecular features, such as angiogenesis, cancer cell proliferation, steroid receptor expression, c-erbB-2 oncoprotein overexpression, p53 protein nuclear accumulation or bcl-2 antiapoptotic protein expression, can predict failure of local therapy. We further examined as to which subgroups of patients could benefit from altered fractionation schemes of radiotherapy. In univariate analysis, high intratumoural angiogenesis, c-erbB overexpression and mutant-p53 nuclear accumulation were significantly associated with increased relapse rate (P=0.0002, 0.009 and 0.05, respectively). In multivariate analysis, the microvessel density and the c-erbB-2 status were independent and significant factors related to local relapse (P=0.001, t-ratio 3.36 and P=0.02, t-ratio 2.26, respectively). Hypofractionated and accelerated radiotherapy supported with amifostine (HypoARC regimen) was significantly more effective than standard radiotherapy in cases with high cancer cell proliferation index, c-erbB-2 and p53 overexpression. High angiogenesis, however, was linked with local relapse regardless of the radiotherapy regimen.

Breast Neoplasms↗

PubMed2003

Vascular endothelial growth factor (VEGF) expression in operable gallbladder carcinomas.

AIM: To investigate the angiogenic and prognostic role of vascular endothelial growth factor (VEGF) in operable gallbladder carcinomas. METHODS: Sixty patients with early gallbladder carcinomas, treated with surgery alone, were investigated immunohistochemically for the expression of VEGF, thymidine phosphorylase (TP) and new blood vessel formation. The results were correlated with clinico-pathological features and prognosis. RESULTS: An increased VEGF secretion in gallbladder carcinomas was significantly associated with increased angiogenesis but not with patients survival, although high angiogenesis did relate with poor prognosis. TP was also associated with angiogenesis, but only the combined VEGF/TP expression was associated with unfavourable survival. Histological grade was another independent factor of prognosis. CONCLUSION: Both VEGF and TP expression are associated with high rate of angiogenesis, a factor directly associated with prognosis. The combined expression of these angiogenic factors confer a particularly poor post-operative outcome, speculature.

Adult↗

PubMed2003

Hypoxia inducible factor 1alpha and 2alpha overexpression in inflammatory bowel disease.

AIMS: Hypoxia inducible factors 1alpha and 2alpha (HIF1alpha and HIF2alpha) are hypoxia regulated transcriptional factors, which control the expression of a variety of genes responsible for angiogenesis, glycolysis, and the inhibition of apoptosis. Because angiogenesis and tissue regeneration are integral components of the inflammatory process, this study was designed to investigate the role of HIFalpha molecules in inflammatory bowel disease. METHODS: Surgical specimens from patients with active ulcerative colitis (UC) and Crohn's disease (CD) were assessed immunohistochemically for HIF1alpha and HIF2alpha reactivity, and the expression of these molecules was compared with the expression of the angiogenic factors thymidine phosphorylase (TP), vascular endothelial growth factor (VEGF), and VEGF-KDR activated vasculature. The vascular density of the lesions was also assessed using anti-CD31 immunostaining. RESULTS: HIF1alpha was expressed focally (epithelial cells, stromal fibroblasts, and myocytes) in both UC and CD, whereas HIF2alpha was expressed focally in UC and diffusely in CD. TP expression was uniformly positive in both diseases. VEGF expression was absent in CD, and weakly positive in UC. The VEGF-KDR reactivity of the submucosal vasculature was only slightly increased in UC and CD compared with normal tissue. The inflammatory cells stained with HIF2alpha and TP in all cases, but the reactivity was generalised in CD and focal in UC. In both diseases, vascular density was significantly higher than that seen in normal tissue. CONCLUSIONS: The discordant expression of HIF2alpha and VEGF in CD suggests an inherent deficiency of the intestine to respond to various stresses by the induction of VEGF. This finding should be investigated further.

Adult↗

PubMed2002

Concurrent administration of Docetaxel and Stealth liposomal doxorubicin with radiotherapy in non-small cell lung cancer : excellent tolerance using subcutaneous amifostine for cytoprotection.

The substantial augmentation of the radiation sequelae during chemo-radiotherapy with novel drugs masks the real potential of such regimens. In this study we examined whether subcutaneous administration of amifostine can reduce the toxicity of a highly aggressive chemo-radiotherapy scheme with Stealth liposomal doxorubicin (Caelyx and Docetaxel (Taxotere in non-small cell lung cancer. Twenty-five patients with stage IIIb non-small cell lung cancer were recruited in a phase I/II dose escalation trial. The starting dose of Taxotere was 20 mg m(-2) week and of Caelyx was 15 mg m(-2) every two weeks, during conventionally fractionated radiotherapy (total dose of 64 Gy). The dose of Taxotere/Caelyx was, thereafter, increased to 20/25 (five patients) and 30/25 mg m(-2) (15 patients). Amifostine 500 mg was given subcutaneously before each radiotherapy fraction, while an i.v. amifostine dose of 1000 mg preceded the infusion of docetaxel. The 'in-field' radiation toxicity was low. Grade 3 esophagitis occurred in 9 out of 25 (36%) patients. Apart from a marked reduction of the lymphocyte counts, the regimen was deprived from any haematological toxicity higher than grade 1. No other systemic toxicity was noted. The CR and CR/PR rates in 15 patients treated at the highest dose level was 40% (6 out of 15) and 87% (13 out of 15) respectively. It is concluded that the subcutaneous administration of amifostine during high dose Taxotere/Caelyx chemo-radiotherapy is a simple and effective way to render this aggressive regimen perfectly well tolerated, by reducing the systemic and the 'in-field' toxicity to the levels expected from simple conventional radiotherapy. The impressive tolerance and the high CR rate obtained encourages the conduct of a relevant randomized trial to assess an eventual survival benefit in patients with non-small cell lung cancer.

Amifostine↗

PubMed2002

Thymidine phosphorylase expression in normal, hyperplastic and neoplastic prostates: correlation with tumour associated macrophages, infiltrating lymphocytes, and angiogenesis.

Thymidine phosphorylase is an angiogenic factor primarily expressed by cancer cells, stromal cells and tumour-associated macrophages in many human malignancies. These different types of thymidine phosphorylase-expressing cells, however, may have a distinct place in the angiogenic process, and this question was addressed in the present study. A series of 20 normal/hyperplastic prostate glands and 60 prostate carcinomas was investigated by immunohistochemistry, using specific antibodies for thymidine phosphorylase (P-GF.44C), tumour-associated macrophages (CD68), endothelium (CD31) and prostate specific antigen (ER-PR8). Thymidine phosphorylase expression by normal and hyperplastic epithelial or stromal cells occurred almost exclusively in the context of an intense lymphocytic infiltrate. High thymidine phosphorylase cancer cells and thymidine phosphorylase stromal cells expression was associated with high angiogenesis in prostate carcinomas, and this significant association was extended to include both tumour-associated macrophages and tumour-infiltrating lymphocytes. Thymidine phosphorylase expression and tumour-infiltrating lymphocytes were related inversely with prostate specific antigen reactivity. In conclusion, thymidine phosphorylase is a major angiogenic factor in prostate carcinomas and its up-regulation is likely to occur in the context of a host immune response.

Cell Transformation, Neoplastic↗

PubMed2002

Diminutive polyps of large bowel should be an early target for endoscopic treatment.

BACKGROUND AND AIMS: Aim of the present study is to ascertain the importance of diminutive colorectal polyps and define the need for removal according to their characteristics and malignant potential. PATIENTS AND METHODS: A total of 4,723 patients who underwent colonoscopy were evaluated and 624 patients with 826 polyps were recorded. There were 352 patients with 443 diminutive polyps, studied according to their distribution. Of these, 371 were removed, histologically examined and correlated to patient characteristics and occurrence of synchronous neoplasms. RESULTS: Of the right colon polyps, 81/115 were diminutive, versus 362/711 of the left colon (p<0.0001). Adenomas were more common in patients over 50 years of age, (p<0.0001). In all colonic segments, diminutive adenomas prevailed over hyperplastic polyps, whereas the proportion of diminutive adenomas predominated in the right colon (p=0.0015). Adenomas were classified as tubular 39%, tubulovillous 55.7% and villous 5.3%. The degree of dysplasia was mild in 45.5%, moderate in 51% and severe in 3.5%. The prevalence of synchronous neoplasms was 37.4%. They were more frequently found in males over 50 years of age and in patients with diminutive adenomas compared to those with diminutive hyperplastic polyps (p=0.0078). CONCLUSIONS: The majority of right colon polyps are diminutive. The proportion of diminutive adenomas is higher in patients over 50 years and in the right vs left colon. Diminutive polyps should be removed taking into account the high prevalence of adenomas with a villous component and their significant degree of dysplasia.

Adenomatous Polyposis Coli↗

PubMed2002

Patterns of episialin/MUC1 expression in endometrial carcinomas and prognostic relevance.

AIMS: To investigate episialin/MUC1 expression in the normal, hyperplastic and neoplastic endometrium, and relate patterns of tumour MUC1 reactivity with histopathological characteristics, oestrogen and progesterone receptor (ER and PR) status, bcl-2 and p53 oncoproteins and with clinical behaviour. METHODS AND RESULTS: We studied 42 normally cycling endometria, 45 endometrial hyperplasias of various forms, and 111 endometrial carcinomas of endometrioid and non-endometrioid cell types with specific monoclonal antibodies employing standard immunohistochemical techniques. The follow-up period ranged from 34 to 182 months with a median of 86 months. Epithelial mucin episialin/MUC1 was consistently expressed in the normal endometrium, following a cyclical pattern: "apical membrane staining" in early and mid-proliferative endometrium; "purely cytoplasmic staining" in late proliferative endometrium; and "cytoplasmic staining with intraluminal secretions" in secretory endometrium. Immunostaining patterns in simple and complex hyperplasia were similar to late proliferative endometrium, while atypical hyperplasias and endometrial carcinomas either simulated patterns of proliferative endometrium or lacked MUC1 reactivity. Membranous MUC1 positivity was statistically more frequent in endometrioid carcinomas compared with carcinomas of non-endometrioid type (P = 0.006). Cytoplasmic MUC1 positivity was significantly associated with poor prognosis, while MUC1-negative carcinomas were associated with PR expression and an improved survival (P=0.04). There was no association of MUC1 patterns with bcl-2 and p53 immunoreactivity or with other histopathological variables. CONCLUSIONS: Episialin/MUC1 is an integral component of the normal premenopausal endometrium and is probably hormonally regulated. It is frequently expressed in endometrial hyperplasias and carcinomas. The loss of MUC1 expression from endometrial carcinomas is associated with a favourable prognosis.

Adenocarcinoma↗

PubMed2002

Prognostic role of angiogenesis in operable carcinoma of the gallbladder.

The prognostic significance of intratumoral angiogenesis was investigated in 62 patients with stage I-III carcinomas of the gallbladder treated with simple cholecystectomy. Microvessel density (MVD) was assessed immunohistochemically, using the alkaline phosphatase/anti-alkaline phosphatase method and the monoclonal antibody CD31. The mean MVD was 30.5 vessels per x 200 optical field. Using the thirty-third and the sixty-sixth percentile, the patients were grouped into three MVD categories: low (MVD 9-18; 20 patients), medium (MVD 19-31; 20 patients), and high (MVD 32-86; 22 patients). A high MVD was more frequent in well-differentiated adenocarcinomas compared with moderately and poorly differentiated tumors (p = 0.04), but there was no statistically significant association between MVD and T stage, or patients' age or sex. Multivariate analysis, including MVD, T stage, and histologic grade, showed that MVD was a significant independent prognostic factor in carcinomas of the gallbladder (p = 0.001, t ratio 3.3). It is believed that the assessment of intratumoral angiogenesis in patients with operable gallbladder carcinomas may be useful in predicting prognosis and, perhaps, in decision making for postoperative adjuvant treatment.

Adult↗

PubMed2002

Amifostine in clinical oncology: current use and future applications.

Amifostine (Ethyol), an inorganic thiophosphate, is a selective broad-spectrum cytoprotector of normal tissues that provides cytoprotection against ionizing radiation and chemotherapeutic agents, thus preserving the efficacy of radiotherapy and chemotherapy. This review summarizes the preclinical data and clinical experience with amifostine, and provides insight into future clinical directions. Amifostine, an inactive pro-drug, is transformed to an active thiol after dephosphorylation by alkaline phosphatase found in the normal endothelium. The absence of alkaline phosphatase in the tumoral endothelium and stromal components, and the hypovascularity and acidity of the tumor environment, may explain its cytoprotective selectivity. The cytoprotective mechanism of amifostine is complicated, involving free radical scavenging, DNA protection and repair acceleration, and induction of cellular hypoxia. Intravenous administration of amifostine 740-900 mg/m(2) before chemotherapy and 250-350 mg/m(2) before each radiotherapy fraction are widely used regimens. The US Food and Drug Administration has approved the use of amifostine as a cytoprotector for cisplatin chemotherapy and for radiation-induced xerostomia. Ongoing trials are being conducted to determine the efficacy of amifostine in reducing radiation-induced mucositis and other toxicities. Novel schedules and routes of administration are under investigation, and may further simplify the use of amifostine and considerably broaden its applications.

Amifostine↗

PubMed2001

The angiogenic "vascular endothelial growth factor/flk-1(KDR) receptor" pathway in patients with endometrial carcinoma: prognostic and therapeutic implications.

BACKGROUND: Vascular endothelial growth factor (VEGF) is an important endothelial cell mitogen associated with increased angiogenesis and aggressive tumor behavior. Its stimulating effect on endothelial cells basically is dependent on the presence of specific VEGF receptors, such as the flk-1(KDR) receptor. This study investigates the roles of VEGF and of a functionally intact angiogenic pathway, "VEGF/flk-1(KDR)," in patients with endometrial carcinoma and their significance in prognosis and therapy. METHODS: A series of 121 endometrial carcinomas were studied. The expression of VEGF by endometrial tumor cells was assessed using the monoclonal antibody (MoAb) VG1. VEGF/KDR complexes on tumor endothelium or activated microvessel density (aMVD) were identified using the MoAb 11B5. In addition, the standard microvessel density (sMVD) was assessed with anti-CD31. In all tumors, the alkaline phosphatase/antialkaline phosphatase technique was employed. A Fisher exact test or an unpaired, two-tailed t test was used for testing correlations between categoric tumor variables, whereas a log-rank test was used to determine statistical differences between life tables. A Cox proportional hazards model was used to assess the effect of tumor variables on overall survival. RESULTS: Cytoplasmic VEGF expression in > 50% of tumor cells was associated significantly with aMVD (P < 0.0001) and with sMVD (P < 0.003). In univariate survival analysis, VEGF (P = 0.0002), aMVD (P = 0.001), and sMVD (P = 0.0009) were significant prognostic variables. Equally important were the histologic parameters tumor type (P = 0.03), tumor grade (P = 0.003), and disease stage (P < 0.0001). In multivariate analysis, disease stage was the most important independent prognostic factor (P < 0.0001), followed by VEGF/KDR (P < 0.01), and VEGF (P < 0.04). Furthermore, VEGF and VEGF/KDR were the only independent prognostic variables for patients with Stage I endometrioid adenocarcinoma. CONCLUSIONS: sMVD and the angiogenic factor VEGF are important indicators of a poor prognosis in patients with endometrial carcinoma. VEGF/KDR complexes define a subgroup of patients with endometrial carcinoma with an even worse prognosis.

Adult↗

PubMed2001

Expression of hypoxia-inducible carbonic anhydrase-9 relates to angiogenic pathways and independently to poor outcome in non-small cell lung cancer.

Carbonic anhydrase-9 (CA9), a transmembrane enzyme with an extracellular active site, is involved in the reversible metabolism of the carbon dioxide to carbonic acid. Up-regulation of CA by hypoxia and the hypoxia-inducible factor (HIF) pathway has been recently postulated (Wykoff et al. Cancer Res., 60: 7075-7083, 2000). In the present study we examined the expression of this enzyme in non-small cell lung cancer. Of 107 cases analyzed, 39 (36.4%) had strong membrane/cytoplasmic expression of CA9 and were grouped as positive. The staining was confined around areas of necrosis, and a significant association of CA9 expression with the extent of necrosis was noted (P = 0.004). Nevertheless, 38 of 74 cases with focal or extensive necrosis did not express CA9. CA9 expression was more frequent in the squamous cell histology (P = 0.001) and with advanced T stage (P = 0.009). A significant coexpression of CA9 with platelet-derived endothelial cell growth factor and basic fibroblast growth factor receptor expression was noted. Double staining of CA9 with anti-CD31 monoclonal antibody revealed an overall higher microvessel density in the areas expressing CA9 than in negative areas (P = 0.0005). Thirty-one of 38 CA9-positive cases were positive for HIF1a/HIF2a, but HIF positivity was a more common event (68 of 107) and their patterns of expression were diffuse (not confined in the necrotic areas). A direct association of CA9 expression with epidermal growth factor receptor, c-erbB-2, and MUC1 expression was also noted (P < 0.04). Survival analysis showed that CA9 expression is related to poor prognosis. CA9 expression in tumors with low vascularization defined a prognosis similar to the one of patients with highly angiogenic tumors. Multivariate analysis revealed that CA9 expression is a significant prognostic factor independent of angiogenesis. We conclude that CA9 is an important molecule in non-small cell lung cancer, the up-regulation of which occurs in highly hypoxic/necrotic regions of the tumors. The expression of CA9 is linked to the expression of a constellation of proteins involved in angiogenesis, apoptosis inhibition, and cell-cell adhesion disruption, which explains the strong association of CA9 with poor outcome.

Adenocarcinoma↗

PubMed2001

Relation of hypoxia inducible factor 1 alpha and 2 alpha in operable non-small cell lung cancer to angiogenic/molecular profile of tumours and survival.

Hypoxia inducible factors HIF1alpha and HIF2alpha are important proteins involved in the regulation of the transcription of a variety of genes related to erythropoiesis, glycolysis and angiogenesis. Hypoxic stimulation results in rapid increase of the HIF1alpha and 2alpha protein levels, as a consequence of a redox-sensitive stabilization. The HIFalphas enter the nucleus, heterodimerize with the HIF1beta protein, and bind to DNA at the hypoxia response elements (HREs) of target genes. In this study we evaluated the immunohistochemical expression of these proteins in 108 tissue samples from non-small-cell lung cancer (NSCLC) and in normal lung tissues. Both proteins showed a mixed cytoplasmic/nuclear pattern of expression in cancer cells, tumoural vessels and tumour-infiltrating macrophages, as well as in areas of metaplasia, while normal lung components showed negative or very weak cytoplasmic staining. Positive HIF1alpha and HIF2alpha expression was noted in 68/108 (62%) and in 54/108 (50%) of cases respectively. Correlation analysis of HIF2alpha expression with HIF1alpha expression showed a significant association (P< 0.0001, r = 0.44). A strong association of the expression of both proteins with the angiogenic factors VEGF (P< 0.004), PD-ECGF (P< 0.003) and bFGF (P< 0.04) was noted. HIF1alpha correlated with the expression of bek-bFGF receptor expression (P = 0.01), while HIF2alpha was associated with intense VEGF/KDR-activated vascularization (P = 0.002). HIF2alpha protein was less frequently expressed in cases with a medium microvessel density (MVD); a high rate of expression was noted in cases with both low and high MVD (P = 0.006). Analysis of overall survival showed that HIF2alpha expression was related to poor outcome (P = 0.008), even in the group of patients with low MVD (P = 0.009). HIF1alpha expression was marginally associated with poor prognosis (P = 0.08). In multivariate analysis HIF2alpha expression was an independent prognostic indicator (P = 0.006, t-ratio 2.7). We conclude that HIF1alpha and HIF2alpha overexpression is a common event in NSCLC, which is related to the up-regulation of various angiogenic factors and with poor prognosis. Targeting the HIF pathway may prove of importance in the treatment of NSCLC.

Adenocarcinoma↗