Expression of KATP channels in human cervical cancer: Potential tools for diagnosis and therapy.

Various ion channels, including ATP-sensitive potassium (KATP) channels, are expressed in cancer and have been suggested as potential tumor markers and therapeutic targets. KATP channels are composed of at least two types of subunit, an inwardly rectifying K+ channel (Kir6.x) and a sulfonylurea receptor (SUR). However, the association between KATP channels and cervical cancer remains elusive. The present study determined that the Kir6.2, SUR1 and SUR2 subunits are expressed in cervical cancer cell lines and/or human biopsies. The potential association of subunit expression with tumor differentiation and invasion was analyzed. The effect of the KATP channel blocker glibenclamide on the proliferation of cervical cancer cell lines was also studied. Five cervical cancer cell lines, two primary cultures of cervical cancer cells, one normal keratinocyte cell line and 74 human biopsies were used in the experiments. The mRNA and protein levels of the Kir6.2 subunit were assessed by reverse transcription-polymerase chain reaction and immunochemistry, respectively. Cell proliferation was evaluated by MTT assay. Kir6.2 subunit overexpression compared with control, was observed in some cervical cancer cell lines and cervical tumor tissues. Additionally, increased KATP channel expression was observed in high-grade, poorly differentiated and invasive human cervical cancer biopsies. Kir6.2 subunit expression was not observed in the majority of the non-cancerous cervical tissues. The effect of the KATP channel blocker glibenclamide on the proliferation of five different cervical cancer cell lines was studied, revealing that as Kir6.2 mRNA expression increased, the inhibitory effect of glibenclamide also increased. The results of the present study suggest, for the first time to the best of our knowledge, that the KATP channel subunits, Kir6.2 and SUR2, could potentially represent tools for diagnosing and treating cervical cancer.

Eag1 potassium channels as markers of cervical dysplasia.

Human ether à-go-go 1 (Eag1) potassium channels are potential tumor markers and therapeutic targets for several types of malignancies, including cervical cancer. Estrogens and human papilloma virus oncogenes regulate Eag1 gene expression, suggesting that Eag1 may already be present in pre-malignant lesions. Therefore, Eag1 could be used as an early marker and/or a potential risk indicator for cervical cancer. Consequently, we studied Eag1 protein expression by immunochemistry in cervical cancer cell lines, normal keratinocytes, cervical cytologies from intraepithelial lesions, biopsies from cervical intraepithelial neoplasias (CIN 1, 2 and 3) and in normal smears from patients taking or not taking estrogens. Two hundred and eighty-six samples obtained by liquid-based cytology and fifteen CIN biopsies were studied. We observed Eag1 protein expression in the cervical cancer cell lines, as opposed to normal keratinocytes. Eag1 was found in 67% of the cervical cytologies from low-grade intra-epithelial lesions and in 92% of the samples from high-grade intraepithelial lesions, but only in 27% of the normal samples. Noteworthy, morphologically normal cells obtained from dysplastic samples also exhibited Eag1 expression. In CIN biopsies we found that the higher the grade of the lesion, the broader the Eag1 protein distribution. Almost 50% of the normal patients taking estrogens displayed Eag1 expression. We suggest Eag1 as a potential marker of cervical dysplasia and a risk indicator for developing cervical lesions in patients taking estrogens. Eag1 detection in cervical cancer screening programs should help to improve early diagnosis and decrease mortality rates from this disease.

Functional expression of voltage-gated sodium channels in primary cultures of human cervical cancer.

Cervical cancer (CaC) is the third most frequent cause of death from cancer among women in the world and the first in females of developing countries. Several ion channels are upregulated in cancer, actually potassium channels have been suggested as tumor markers and therapeutic targets for CaC. Voltage-gated sodium channels (VGSC) activity is involved in proliferation, motility, and invasion of prostate and breast cancer cells; however, the participation of this type of channels in CaC has not been explored. In the present study, we identified both at the molecular and electrophysiological level VGSC in primary cultures from human cervical carcinoma biopsies. With the whole cell patch clamp technique, we isolated and identified a voltage-gated Na(+) current as the main component of the inward current in all investigated cells. Sodium current was characterized by its kinetics, voltage dependence, sensitivity to tetrodotoxin (TTX) block and dependence to [Na(+)](o). By analyzing the expression of mRNAs encoding TTX-sensitive Na(+) channel alpha subunits with standard RT-PCR and specific primers, we detected Na(v)1.2, Na(v)1.4, Na(v)1.6, and Na(v)1.7 transcripts in total RNA obtained from primary cultures and biopsies of CaC. Restriction enzyme analysis of PCR products was consistent with the molecular nature of the corresponding genes. Notably, only transcripts for Na(v)1.4 sodium channels were detected in biopsies from normal cervix. The results show for the first time the functional expression of VGSC in primary cultures from human CaC, and suggest that these channels might be considered as potential molecular markers for this type of cancer.

Ether a go-go potassium channels as human cervical cancer markers.

Ether a go-go (EAG) potassium channels display oncogenic properties. In normal tissues, EAG mRNA is almost exclusively expressed in brain, but it is expressed in several somatic cancer cell lines, including HeLa, from cervix. Antisense experiments against eag reduce cell proliferation in some cancer cell lines, and inhibition of EAG-mediated currents has been suggested to decrease cell proliferation in a melanoma cell line. Because of the potential clinical relevance of EAG, we investigated EAG mRNA expression in the following fresh samples from human uterine cervix: 5 primary cultures obtained from cancerous biopsies, 1 cancerous fresh tissue, and 12 biopsies of control normal tissue. All of the control cervical samples came from patients with negative pap smears. Reverse transcription-PCR and Southern-blot experiments revealed eag expression in 100% of the cancerous samples and in 33% of the normal biopsies. Immunochemistry experiments showed the presence of EAG channel protein in cells from the primary cultures and in cervical cancer biopsies sections from the same patients. In addition, we looked for EAG-mediated currents in the cultures from cervical cancer cells. Here we show for the first time EAG channel activity in human tumors. Patch-clamp recordings showed typical EAG-mediated currents modulated by magnesium and displaying a pronounced Cole-Moore shift. Because EAG expression and channel activity have been suggested to be important in cell proliferation, our findings strongly support the idea of considering EAG as a tumor marker as well as a potential membrane therapeutic target for cervical cancer.

Quimiterapia neoadyuvante en pacientes con carcinoma epidermoide de cérvix estadio IB2 y IIA: resultados preliminares de un estudio aleatorizado / Neoadyuvant chemotherapy in patients with cervical carcinoma stages IB2 and IIA: preliminary results in aletorized study

Antecedentes. El cáncer de cérvix es el tumor maligno más frecuente en México. Estudios fase II de quimioterapia neoadyuvante seguida de cirugía en pacientes con estadios clínicos IB2 y IIA sugieren un beneficio en cuanto a control local y supervivencia en comparación con radioterapia sola. Objetivo. Determinar si la quimioterapia neoadyuvante seguida de cirugía mejora la supervivencia en comparación a radioterapia en pacientes con cáncer epidermoide de cérvix estadios IB2 y IIA. Pacientes y métodos. Pacientes en estadios IB2 y IIA de acuerdo a la clasificación de la Federación Internacional de Ginecología y Obstetricia (FIGO) fueron asignadas de manera aleatoria para recibir quimioterapia neoadyuvante con el siguiente esquema: cisplatino 50 mg/m2 d1, vincristina 1.5 mg/m2 d1 y bleomicina 20 mg/m2 d1, d2 y d3 (PVB) en infusión intravenosa continua. Los ciclos se administraron cada 10 días por tres veces. Después de la quimioterapia, las pacientes fueron sometidas a histerectomía radical y linfadenectomía pélvica bilateral. Se administró radioterapia adyuvante en caso de ganglios pélvicos positivos; afección parametrial; margen quirúrgico positivo e invasión estromal mayor de dos tercios del espesor cervical. Las pacientes del brazo de radioterapia recibieron una combinación de teleterapia y braquiterapia a una dosis de 8,500 y 5,500 cGy a los puntos A y B, respectivamente. Resultados. El estudio fue planeado para incluir 80 pacientes por brazo, pero se terminó de manera prematura por lo que sólo se incluyeron 20 enfermas (10 por brazo). La respuesta global a la quimioterapia fue del 90 por ciento y nueve de ellas fueron sometidas a cirugía. Nueve de las diez pacientes asignadas a radiación que completaron el tratamiento obtuvieron respuesta completa. El tratamiento fue bien tolerado en ambos grupos de mujeres. A un seguimiento máximo de 14 meses, una paciente en cada brazo ha recaído. Conclusión. Los resultados preliminares de este estudio sugieren que la quimioterapia neoadyuvante con PVB es factible, produce alto índice de respuestas y parece disminuir la presencia de factores patológicos de alto riesgo para recurrencia.

ADN del virus del papiloma humano en plasma de pacientes con carcinoma cervicouterino: un potencial marcador de enfermedad residual mínima / DNA-human paillomavirus in serum of patients with cervical carcinoma: a potential marker in minimum residual disease

Antecedentes. El carcinoma cervicouterino es la primera causa de muerte por cáncer en mujeres mexicanas. Estudios moleculares han demostrado que el virus del papiloma humano (VHP) es el principal agente etiológico de esta enfermedad. En este tumor no existe un marcador sensible y específico que pudiera utilizarse como marcador de enfermedad residual mínima y para prevenir recaídas tempranamente. Dado que la amplificación en plasma de secuencias de ADN específicas a tumores se ha utilizado con este fin, en este trabajo se determinó la factibilidad de amplificar, mediante reacción en cadena de la polimerasa en el ADN del plasma, secuencias de VPH en pacientes con cáncer cervicouterino. Material y métodos. Se analizaron 34 pacientes con carcinoma cervicouterino invasor con diferentes estadios clínicos sin tratamiento previo y 24 mujeres con infección cervical por VPH pero sin lesiones invasoras. La extracción del ADN se realizó mediante técnicas estándar: digestión con proteinasa K, extracción con fenol-cloroformo y precipitación con etanol. Para verificar la calidad del ADN, se amplificó un fragmento del gen de la ß-globina. Las secuencias del VPH se amplificaron con los ligonucleótidos generales LICI-LIC2. Las muestras positivas también se amplificaron con oligonucleótidos específicos para VPH tipos 16 y 18. Resultados. ADN del VPH se encontró en el 70 por ciento de los casos de pacientes con cáncer y no se amplificó en los 24 controles. No se encontró correlación entre el resultado de la reacción en caderna de la polimerasa con el estadio clínico ni con el tipo viral, peso se observó menor frecuencia de positividad en los tumores adenoescamosos. Conclusión. Los resultados sugieren que la detección del ADN del VPH en el plasma de las pacientes es específica de aquéllas con cáncer invasor y que esto podría constituir un marcador de enfermedad residual mínima y predictor temprano de recaídas