Carbonic anhydrase IX-based tumor imaging and therapy: a review / 生物工程学报

Carbonic anhydrase IX (CAIX) is a transmembrane protein that is specifically overexpressed on the surface of hypoxic tumor cells. With the function of regulating the acidity of tumor cells both inside and outside, CAIX is closely related to tumor proliferation, invasion and metastasis. Therefore, CAIX is a promising target for tumor imaging and therapy. Herein, we summarized recent advances in CAIX-based tumor imaging, therapy and theranostics, and prospected future applications of using CAIX as an anti-tumor target.

Quantitative Characterization of the Chemical Space Governed by Human Carbonic Anhydrases and selenium-containing derivatives of solfonamides

Abstract Due to the fact that different isoforms of carbonic anhydrase play distinct physiological roles, their diseases/disorders involvement are different as well. Involvement in major disorders such as glaucoma, epilepsy, Alzheimer's disease, obesity and cancers, have turned carbonic anhydrase into a popular case study in the field of rational drug design. Since carbonic anhydrases are highly similar with regard to their structures, selective inhibition of different isoforms has been a significant challenge. By applying a proteochemometrics approach, herein the chemical interaction space governed by acyl selenoureido benzensulfonamides and human carbonic anhydrases is explored. To assess the validity, robustness and predictivity power of the proteochemometrics model, a diverse set of validation methods was used. The final model is shown to provide valuable structural information that can be considered for new selective inhibitors design. Using the supplied information and to show the applicability of the constructed model, new compounds were designed. Monitoring of selectivity ratios of new designs shows very promising results with regard to their selectivity for a specific isoform of carbonic anhydrase.

Enzyme inhibition and antibacterial potential of 4-Hydroxycoumarin derivatives

The 4-Hydroxycoumarin derivatives are known to show a broad spectrum of pharmacological applications. In this paper we are reporting the synthesis of a new series of 4-Hydroxycoumarin derivatives synthesized through Knovenegal condensation; they were characterized by using UV-Vis, FT-IR, NMR spectroscopies. The synthesized compounds were evaluated for antibacterial activity against Staphylococcus aureus and Salmonella typhimurium strains. The compounds (2), (3) and (8) showed favorable antibacterial activity with zone of inhibitions 26.5± 0.84, 26.0 ± 0.56 and 26.0 ± 0.26 against Staphylococcus aureus (Gram-positive) respectively. However, the compounds (5) and (9) were found more active with 19.5 ± 0.59 and 19.5 ± 0.32 zone of inhibitions against Salmonella typhimurium (Gram-negative). Whereas, in urease inhibition assay, none of the synthesized derivatives showed significant anti-urease activity; although, in carbonic anhydrase-II inhibition assay, the compound (2) and (6) showed enzyme inhibition activity with IC50 values 263±0.3 and 456±0.1, respectively.

A Validation Study of a Multiple Reaction Monitoring-Based Proteomic Assay to Diagnose Breast Cancer / 한국유방암학회지

Increased Expression of the Matrix-Modifying Enzyme Lysyl Oxidase-Like 2 in Aggressive Hepatocellular Carcinoma with Poor Prognosis

BACKGROUND/AIMS: Lysyl oxidase-like 2 (LOXL2), a collagen-modifying enzyme, has been implicated in cancer invasiveness and metastasis. METHODS: We evaluated the expression of LOXL2 protein, in addition to carbonic anhydrase IX (CAIX), keratin 19, epithelial cell adhesion molecule, and interleukin 6, in 105 resected hepatocellular carcinomas (HCCs) by immunohistochemistry. RESULTS: LOXL2 positivity was found in 14.3% (15/105) of HCCs, and it was significantly associated with high serum α-fetoprotein levels, poor differentiation, fibrous stroma, portal vein invasion, and advanced TNM stage (p < 0.05 for all). Additionally, LOXL2 positivity was significantly associated with CAIX (p=0.005) and stromal interleukin 6 expression (p=0.001). Survival analysis of 99 HCC patients revealed LOXL2 positivity to be a poor prognostic factor; its prognostic impact appeared in progressed HCCs. Furthermore, LOXL2 positivity was shown to be an independent predictor of overall survival and disease-specific survival (p < 0.05 for all). Interestingly, co-expression of LOXL2 and CAIX was also an independent predictor for overall survival, disease-specific survival, disease-free survival, and extrahepatic recurrence-free survival (p < 0.05 for all). CONCLUSIONS: LOXL2 expression represents a subgroup of HCCs with more aggressive behavior and is suggested to be a poor prognostic marker in HCC patients.

Mining and engineering of microbial carbonic anhydrases for biomimetic carbon dioxide sequestration / 生物工程学报

The increasing atmospheric carbon dioxide levels have been correlated with global warming. Carbonic anhydrases (CA) are the fastest among the known enzymes to improve carbon capture. The capture of carbon dioxide needs high temperature and alkaline condition, which is necessary for CaCO₃ precipitation in the mineralization process. In order to use CAs for biomimetic carbon sequestration, thermo-alkali-stable CAs are, therefore, essential, and polyextremophilic microbes are one of the important sources of these enzymes. The current review focuses on both those isolated by thermophilic organisms from the extreme environments and those obtained by protein engineering techniques, and the industrial application of the immobilized CAs is also briefly addressed. To reduce the greenhouse effect and delay global warming, we think further research efforts should be devoted to broadening the scope of searching for carbonic anhydrase, modifying the technology of protein engineering and developing highly efficient immobilization strategies.

An Intrarenal Adrenocortical Carcinoma Arising in an Adrenal Rest

We describe a case of a 61-year-old Korean man who was diagnosed with renal cell carcinoma that was discovered on abdominopelvic computed tomography obtained after the patient complained of back pain. A radical nephrectomy was performed, and the surgical specimen showed a relatively well-circumscribed and yellowish lobulated hard mass. Microscopically, the tumor showed sheets and nests of hypercellular pleomorphic cells with thick fibrous septation, frequent mitoses, and areas of adrenal cortical-like tissue. Immunohistochemical staining revealed that the tumor cells were positive for inhibin-α, vimentin, synaptophysin, and melan A. It also revealed that the tumor cells were negative for pan-cytokeratin, epithelial membrane antigen, paired box 8, α-methylacyl-coenzyme A racemase, CD10, cytokeratin 7, carbonic anhydrase 9, c-Kit, renal cell carcinoma, transcription factor E3, human melanoma black 45, desmin, smooth muscle actin, S-100, chromogranin A, CD34, anaplastic lymphoma kinase, and integrase interactor 1. Based on these histopathological and immunohistochemical findings, we diagnosed the tumor as intrarenal adrenocortical carcinoma arising in an adrenal rest. Several cases of intrarenal adrenocortical carcinoma have been reported, although they are very rare. Due to its poor prognosis and common recurrence or metastasis, clinicians and pathologists must be aware of this entity.

Role of carbonic anhydrases in skin wound healing

Skin wound closure occurs when keratinocytes migrate from the edge of the wound and re-epithelialize the epidermis. Their migration takes place primarily before any vascularization is established, that is, under hypoxia, but relatively little is known regarding the factors that stimulate this migration. Hypoxia and an acidic environment are well-established stimuli for cancer cell migration. The carbonic anhydrases (CAs) contribute to tumor cell migration by generating an acidic environment through the conversion of carbon dioxide to bicarbonate and a proton. On this basis, we explored the possible role of CAs in tissue regeneration using mouse skin wound models. We show that the expression of mRNAs encoding CA isoforms IV and IX are increased (~25 × and 4 ×, respectively) during the wound hypoxic period (days 2–5) and that cells expressing CAs form a band-like structure beneath the migrating epidermis. RNA-Seq analysis suggested that the CA IV-specific signal in the wound is mainly derived from neutrophils. Due to the high level of induction of CA IV in the wound, we treated skin wounds locally with recombinant human CA IV enzyme. Recombinant CA IV significantly accelerated wound re-epithelialization. Thus, CA IV could contribute to wound healing by providing an acidic environment in which the migrating epidermis and neutrophils can survive and may offer novel opportunities to accelerate wound healing under compromised conditions.

Antihypertensive effect of Ganjang (traditional Korean soy sauce) on Sprague-Dawley Rats

BACKGROUND/OBJECTIVES: Although Korean fermented foods contain large amounts of salt, which is known to exacerbate health problems, these foods still have beneficial effects such as anti-hypertension, anti-cancer, and anti-colitis properties. We hypothesized that ganjang may have different effects on blood pressure compared to same concentrations of salt. MATERIALS/METHODS: Sprague-Dawley rats were divided into control (CT), NaCl (NC), and ganjang (GJ) groups and orally administered with 8% NaCl concentration for 9 weeks. The systolic blood pressure (SBP), serum chemistry, Na⁺ and K⁺ concentrations and renal gene expressions were measured. RESULTS: The SBP was significantly increased in the NC group compared to the GJ and CT groups. In addition, the Na+ concentration in urine was higher in the GJ and NC groups than the CT group, but the urine volume was increased in the GJ group compared to the other groups. The serum renin levels were decreased in the GJ group compared to the CT group, while the serum aldosterone level was decreased in the GJ group relative to the NC group. The mRNA expression of the renin, angiotensin II type I receptor, and mineralocorticoid receptor were significantly lower in the GJ group compared to other groups. Furthermore, GJ group showed the lowest levels of genes for Na⁺ transporter in kidney cortex such as Na⁺/K⁺ ATPaseα1 (NKAα1), Na⁺/H⁺ exchanger 3 (NHE3), Na⁺/HCO₃⁻ co-exchanger (NBC), and carbonic anhydrases II (CAII). CONCLUSIONS: The decreased SBP in the GJ could be due to decreased renin and aldosterone levels in serum and increased urinary volume and excretion of Na⁺ with its transporter gene alteration. Therefore, ganjang may have antihypertensive effect despite its high contents of salt.

Expression and clinical pathological implications of carbonic anhydrase 9 and P glycoprotein in laryngeal squamous cell carcinoma / 临床耳鼻咽喉头颈外科杂志

OBJECTIVE@#To identify the difference of CA IX and P-gp expression level between laryngeal squamous cell carcinoma (LSCC) and benign tissues, evaluate the relationship of these two proteins in LSCC, and their correlation with clinical and pathological features.@*METHOD@#Immunohistochemical detection of CA IX and P-gp were performed in 47 cases of LSCC and 20 cases of vocal cord polyps.@*RESULT@#Overexpression of CA IX and P-gp both in LSCC and in vocal cord polyp (P < 0.05) were confirmed, with a correlation between the two proteins in LSCC (r = 0.324, P < 0.05). The expression of CA IX was related to clinical staging and lymph node metastasis in LSCC (P < 0.05). While P-gp was related to clinical staging and histological grading in LSCC (P < 0.05).@*CONCLUSION@#The overexpression of CA IX and P-gp may play a role in LSCC progression.

The Expression of Carbonic Anhydrase (CA) IX/XII and Lymph Node Metastasis in Early Breast Cancer / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: The aim of study was to test by immunohistochemical (IHC) staining whether carbonic anhydrase (CA) 9 and 12 have an effect on sentinel lymph node (SLN) metastasis in early breast cancer and to find clinicopathologic factors associated with SLN metastasis. MATERIALS AND METHODS: Between June 2003 and June 2011, medical records of 470 patients diagnosedwith breast cancer with pT1-2, pN0-2, and M0 were reviewed. Of these 470, 314 patients who underwent SLN biopsy+/-axillary dissection were subjects of this study. Using tissue microarray, IHC staining for CA9 and CA12 was performed. Clinicopathologic factors such as patient age, tumour size, lymphatic invasion, hormone receptor status, and the Ki-67 labeling index were analysed together. RESULTS: The mean age of all patients was 51.7 years. The mean number of harvested SLN was 3.62, and 212 patients (67.5%) had negative SLN. Lymphatic invasion, the Ki-67 labelling index of primary tumours, and CA9 staining of stromal cells, were independent risk factors for SLN metastasis in the multivariate analysis. In 33 patients (10.5%) without the three risk factors, no patient had SLN metastasis. In 80 patients without lymphatic invasion of primary tumours or CA9 staining of stromal cells, only four patients (5%) had positive SLN. CONCLUSION: CA9 staining of stromal cells is an independent risk factor for SLN metastasis as well as lymphatic invasion and a low Ki-67 labelling index of primary tumours in patients with early breast cancer. IHC staining of primary tumours for CA12was not associatedwith SLN metastasis.

Knowledge and practice of Brazilian pediatricians on gastroesophageal reflux disease in infants / Conhecimento e prática de pediatras brasileiros sobre a doença do refluxo gastroesofágico em lactentes

OBJECTIVE: To assess the knowledge and practice of pediatricians about infants with physiological reflux and gastroesophageal reflux disease. METHODS: 140 pediatricians were interviewed during two scientific events in 2009 and 2010. The questions referred to two clinical cases of infants. One with symptoms of infant regurgitation (physiological reflux) and another with gastroesophageal reflux disease. RESULTS: Among 140 pediatricians, 11.4% (n=16) and 62.1% (n=87) would require investigation tests, respectively for infant regurgitation (physiological reflux) and gastroesophageal reflux disease. A series of upper gastrointestinal exams would be the first requested with a higher frequency. Medication would be prescribed by 18.6% (n=6) in the case of physiological reflux and 87.1% (n=122) in the case of gastroesophageal reflux disease. Prokinetic drugs would be prescribed more frequently than gastric acid secretion inhibitors. Sleeping position would be recommended by 94.2% (n=132) and 92.9% (n=130) of the respondents, respectively for the case of physiological reflux and gastroesophageal reflux disease; however, about half of the respondents would recommend the prone position. Only 10 (7.1%) of the pediatricians would exclude the cow's milk protein from the infants' diet. CONCLUSIONS: Approaches different from the international guidelines are often considered appropriate, especially when recommending a different position other than the supine and prescription of medication. In turn, the interviews enable us to infer the right capacity of the pediatricians to distinguish physiologic reflux and gastroesophageal reflux disease correctly. .

OBJETIVO: Avaliar o conhecimento e a prática de pediatras brasileiros na assistência ao lactente com refluxo fisiológico e doença do refluxo gastroesofágico. MÉTODOS: Foram entrevistados 140 médicos pediatras em dois eventos científicos em 2009 e 2010. As perguntas referiam-se a dois casos clínicos de lactentes, um com quadro compatível com regurgitação do lactente (refluxo fisiológico) e outro com doença do refluxo gastroesofágico. RESULTADOS: Dos 140 participantes, 11,4% (n=16) e 62,1% (n=87) solicitariam exame para lactentes, respectivamente, com refluxo fisiológico e doença do refluxo gastroesofágico. O primeiro exame solicitado com maior frequência seria a radiografia contrastada de esôfago, estômago e duodeno. Medicação seria prescrita por 18,6% (n=26) para o caso de refluxo fisiológico e 87,1% (n=122) para o caso de doença do refluxo gastroesofágico. Procinéticos seriam prescritos com maior frequência do que os redutores da secreção ácida gástrica. Prescrição de posição para dormir fez parte das recomendações de 94,2% (n=132) e 92,9% (n=130) dos entrevistados, respectivamente, para os casos de refluxo fisiológico e doença do refluxo gastroesofágico. Entretanto, cerca da metade dos entrevistados não recomendaria o decúbito dorsal. Prescrição de dieta de exclusão do leite de vaca para um lactente com quadro de doença do refluxo gastroesofágico seria feita por apenas 10 (7,1%) dos participantes. CONCLUSÕES: Condutas diferentes das diretrizes internacionais são frequentemente consideradas adequadas, especialmente quanto à recomendação de posição diferente do decúbito dorsal e prescrição de medicamentos. As respostas permitem inferir a capacidade de correta diferenciação entre refluxo fisiológico e doença do refluxo gastroesofágico. .

Significance of carbonic anhydrase IX protein in fibrotic foci of mammary invasive ductal carcinomas / 中华病理学杂志

<p><b>OBJECTIVE</b>To study the relationship between fibrotic focus (FF) and carbonic anhydrase (CA) IX in invasive ductal carcinomas (IDC) of the breast.</p><p><b>METHODS</b>In 167 cases of IDC, the FF was assessed morphologically, and expression of ER, PR and CA IX was evaluated using MaxVision immunohistochemistry.</p><p><b>RESULTS</b>The expression of CA IX in IDC with and without FF was 56.3% (45/80) and 28.7% (25/87) respectively, with significant difference (P=0.001). In IDC with FF, the CA IX expression of tumor cells in tumors with CA IX-positive fibroblasts (35/40, 87.5%) was significantly (P<0.001) higher than that in tumors with CA IX-negative fibroblasts (10/40, 25.0%). In IDC with FF, the CA IX expression of fibroblasts of FF in grade 3 IDC (23/33, 69.7%) was significantly (P=0.006) higher than that in grade 1+2 tumors (17/47, 36.2%). The ER and PR expression of tumor cells in tumors containing CA IX-negative fibroblasts was 72.5% (29/40) and 65.0% (26/40) respectively, whereas the ER and PR expression of tumor cells in tumors containing CA IX-positive fibroblasts was 50.0% (20/40) and 42.5% (17/40) respectively; the difference was statistically significant (for both ER and PR, P=0.04). The age of patients with tumors containing CA IX-negative fibroblasts was significantly (P=0.002) older than those containing CA IX-positive fibroblasts. The FF diameter/tumor diameter in tumors containing CA IX-positive fibroblasts was significantly larger than those containing CA IX-negative fibroblasts. (3) For the groups of tumor size≤2 cm and tumor size between 2 cm to 5 cm, the diameter of the fibrotic focus was significantly (P<0.01) smaller than the fibrotic focus size of tumors>5 cm in size.</p><p><b>CONCLUSIONS</b>CA IX expression is correlated with FF, and that in fibroblasts of FF correlated with patients' age, tumor grade, hormone receptors and FF diameter/tumor diameter. CA IX expression in FF might be a marker for poor prognosis in patients with breast cancer.</p>

Low-Dose Acetazolamide in the Treatment of Premenstrual Dysphoric Disorder: A Case Series

The treatment of premenstrual dysphoric disorder (PMDD) is far from satisfactory, as there is a high proportion of patients who do not respond to conventional treatment. The antidiuretic sulfonamide, acetazolamide, inhibits carbonic anhydrase and potentiates GABAergic transmission; the latter is putatively involved in PMDD. We therefore tried acetazolamide in a series of women with intractable PMDD. Here, we describe a series of eight women diagnosed with DSM-IV-TR PMDD, five of whom had comorbidity with a mood disorder and one with an anxiety disorder, who were resistant to treatment and responded with symptom disappearance after being added-on 125 mg/day acetazolamide for 7-10 days prior to menses each month. Patients were free from premenstrual symptoms at the 12-month follow-up. We suggest that acetazolamide may be used to improve symptoms of PMDD in cases not responding to other treatments. GABAergic mechanisms may be involved in counteracting PMDD symptoms.

DNA Hypomethylation-Mediated Overexpression of Carbonic Anhydrase 9 Induces an Aggressive Phenotype in Ovarian Cancer Cells

PURPOSE: Both genetic and epigenetic alterations can lead to abnormal expression of metastasis-regulating genes in tumor cells. Recent studies suggest that aberrant epigenetic alterations, followed by differential gene expression, leads to an aggressive cancer cell phenotype. We examined epigenetically regulated genes that are involved in ovarian cancer metastasis. MATERIALS AND METHODS: We developed SK-OV-3 human ovarian carcinoma cell xenografts in mice. We compared the mRNA expression and DNA methylation profiles of metastatic tissues to those of the original SK-OV-3 cell line. RESULTS: Metastatic implants showed increased mRNA expression of the carbonic anhydrase 9 (CA9) gene and hypomethylation at CpG sites in the CA9 promoter. Treatment of wild-type SK-OV-3 cells with the DNA methyltransferase inhibitor 5-aza-2'-deoxycytidine reduced methylation of the CA9 promoter and increased CA9 mRNA expression. Eight CpGs, which were located at positions -197, -74, -19, -6, +4, +13, +40, and +86, relative to the transcription start site, were hypomethylated in metastatic tumor implants, compared to that of wild-type SK-OV-3. Overexpression of CA9 induced an aggressive phenotype, including increased invasiveness and migration, in SK-OV-3 cells. CONCLUSION: Alterations in the DNA methylation profile of the CA9 promoter were correlated with a more aggressive phenotype in ovarian cancer cells.

Association of Carbonic Anhydrase IX Expression on the Prognosis and Therapeutic Response in Clear Cell Renal Cell Carcinoma / 계명의대학술지

Recent studies of Carbonic anhydrase IX (CAIX) expression and clinical significance in renal cell carcinoma (RCC) have given rise to disagreements in the usefulness of CAIX as a prognostic factor. The purpose of this study was to evaluate the association between CAIX expression and clinical factors in RCC. The medical record of 172 RCC patients in hospital (from January 1999 and December 2007) were reviewed retrospectively. Patients were divided into a high expression group (109 cases) and low expression group (63 cases) according to their degree of CAIX expression. We evaluated the association between CAIX expression and age, body mass index (BMI), type of renal neoplasm, tumor stage, nuclear grade, metastasis after surgery and tumor-specific survival rate. The mean age of the high expression group and the low expression group were 56 years and 54 years respectively. The mean BMI of the high expression group and the low expression group were 24.2 kg/m2 and 24.5 kg/m2 respectively. Comparing the difference between clear cell RCC and non clear cell RCC, CAIX was significantly more expressed in clear cell RCC. There was no significant differences between high expression clear cell RCC and low expression clear cell RCC according to age, BMI, nuclear grade, metastasis after surgery and tumor-specific survival rate (p=0.237, p=0.802, p=0.382, p=0.551). However, in clear cell RCC, CAIX expression was significantly more expressed in patients with higher T or N stages (p=0.015, p=0.033). CAIX was significantly higher expressed in clear cell RCC and was significantly lower expressed in patients with higher T stage or N stage.

The Effects of Topical Carbonic Anhydrase Inhibitors on Nitric Oxide Production in Trabecular Meshwork Cells

PURPOSE: To investigate and compare the effects of topical carbonic anhydrase inhibitors on the production and expression of nitric oxide in cultured human trabecular meshwork cells (HTMC). METHODS: Primarily cultured HTMC were exposed to 0, 10, and 100 microM dorzolamide and brinzolamide using serum-deprived media for 3 days. Production of nitric oxide was assessed with Griess assay. Expressions of eNOS mRNA were assessed with RT-PCR. RESULTS: Both dorzolamide and brinzolamide increased the production of nitric oxide eNOS activity (p < 0.05). Dorzolamide had a more potent effect than brinzolamide on the production of nitric oxide and the expression of eNOS mRNA. CONCLUSIONS: Topical carbonic anhydrase inhibitors increased the production of nitric oxide, which was accompanied by increased eNOS activity. Dorzolamide had a more potent effect than brinzolamide on the production of nitric oxide and expression of eNOS mRNA in HTMC. The increased production of nitric oxide by topical carbonic anhydrase inhibitors involves mechanisms other than carbonic anhydrase inhibition.

Relationship between dental caries and salivary proteome by electrospray ionization ion-trap tandem mass spectrometry in children aged 6 to 8 years / 华西口腔医学杂志

<p><b>OBJECTIVE</b>To explore the relationship between salivary proteome and dental caries and to promote the biomarker studies of dental caries susceptibility by comparing the salivary proteome of caries-active children and caries-free children with electrospray ionization ion-trap tandem mass spectrometry (ESI-MS/MS).</p><p><b>METHODS</b>Ten caries-active children and ten caries-free children were sampled. The salivary proteome of the two groups was studied, and the differential protein between the two groups was analyzed by ESI-MS/MS after sodium dodecyl sulfate polyacrylamide gel electrophoresis, filter-aided sample preparation, and liquid chromatography.</p><p><b>RESULTS</b>The concentration of salivary protein was higher in the caries-active group than in the caries-free group. The polypeptide counts of thecaries-active and caries-free groups were 602 and 481, which belonged to 286 and 227 proteins, respectively. The differential polypeptide count of the two groups was 361, and the differential protein count was 118. The detected proteins included matrix metalloproteinase-9 (MMP9), mucin-7 (MUC7), lactotransferrin (LTF), carbonic anhydrase 6 (CA6), azurocidin (AZU), and cold agglutinin.</p><p><b>CONCLUSION</b>The total salivary protein was higher in the caries-active group than in the caries-free group. The preliminary detection of differential proteins (MMP9, MUC7, LTF, CA6, AZU, and cold agglutinin) may lay some foundation for biomarker research of dental caries susceptibility.</p>

Carbonic Anhydrase IX is Not a Predictor of Outcomes in Non-Metastatic Clear Cell Renal Cell Carcinoma - A Digital Analysis of Tissue Microarray

Introduction The knowledge about the molecular biology of clear cell renal cell carcinoma (ccRCC) is evolving, and Carbonic Anhydrase type IX (CA-IX) has emerged as a potential prognostic marker in this challenging disease. However, most of the literature about CA-IX on ccRCC comes from series on metastatic cancer, with a lack of series on non-metastatic cancer. The objective is to evaluate the expression of CA-IX in a cohort of non-metastatic ccRCC, correlating with 1) overall survival, and 2) with established prognostic parameters (T stage, tumor size, Fuhrman nuclear grade, microvascular invasion and peri-renal fat invasion). Materials and Methods This is a retrospective cohort study. We evaluated 95 patients with non-metastatic clear cell renal cell carcinoma, as to the expression of CA-IX. The analyzed parameters where: overall survival (OS), TNM stage, tumor size (TS), Fuhrman nuclear grade (FNG), microvascular invasion (MVI), peri-renal fat invasion (PFI). We utilized a custom built tissue microarray, and the immunoexpression was digitally quantified using the Photoshop® software. Results: Th e mean follow-up time was 7.9 years (range 1.9 to 19.5 years). The analysis of CA-IX expression against the selected prognostic parameters showed no correlation. The results are as follows: Overall survival (p = 0.790); T stage (p = 0.179); tumor size (p = 0.143); grouped Fuhrman nuclear grade (p = 0.598); microvascular invasion (p = 0.685), and peri-renal fat invasion (p = 0.104). Conclusion Carbonic anhydrase type IX expression does not correlate with overall survival and conventional prognostic parameters in non-metastatic clear cell renal cell carcinoma. .