Eag1 Gene and Protein Expression in Human Retinoblastoma Tumors and its Regulation by pRb in HeLa Cells.

Retinoblastoma is the most common pediatric intraocular malignant tumor. Unfortunately, low cure rates and low life expectancy are observed in low-income countries. Thus, alternative therapies are needed for patients who do not respond to current treatments or those with advanced cases of the disease. Ether à-go-go-1 (Eag1) is a voltage-gated potassium channel involved in cancer. Eag1 expression is upregulated by the human papilloma virus (HPV) oncogene E7, suggesting that retinoblastoma protein (pRb) may regulate Eag1. Astemizole is an antihistamine that is suggested to be repurposed for cancer treatment; it targets proteins implicated in cancer, including histamine receptors, ATP binding cassette transporters, and Eag channels. Here, we investigated Eag1 regulation using pRb and Eag1 expression in human retinoblastoma. The effect of astemizole on the cell proliferation of primary human retinoblastoma cultures was also studied. HeLa cervical cancer cells (HPV-positive and expressing Eag1) were transfected with RB1. Eag1 mRNA expression was studied using qPCR, and protein expression was assessed using western blotting and immunochemistry. Cell proliferation was evaluated with an MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. RB1 transfection down-regulated Eag1 mRNA and protein expression. The human retinoblastoma samples displayed heterogeneous Eag1 mRNA and protein expression. Astemizole decreased cell proliferation in primary retinoblastoma cultures. Our results suggest that Eag1 mRNA and protein expression was regulated by pRb in vitro, and that human retinoblastoma tissues had heterogeneous Eag1 mRNA and protein expression. Furthermore, our results propose that the multitarget drug astemizole may have clinical relevance in patients with retinoblastoma, for instance, in those who do not respond to current treatments.

The Diagnosis of Fungal Neglected Tropical Diseases (Fungal NTDs) and the Role of Investigation and Laboratory Tests: An Expert Consensus Report.

The diagnosis of fungal Neglected Tropical Diseases (NTD) is primarily based on initial visual recognition of a suspected case followed by confirmatory laboratory testing, which is often limited to specialized facilities. Although molecular and serodiagnostic tools have advanced, a substantial gap remains between the desirable and the practical in endemic settings. To explore this issue further, we conducted a survey of subject matter experts on the optimal diagnostic methods sufficient to initiate treatment in well-equipped versus basic healthcare settings, as well as optimal sampling methods, for three fungal NTDs: mycetoma, chromoblastomycosis, and sporotrichosis. A survey of 23 centres found consensus on the key role of semi-invasive sampling methods such as biopsy diagnosis as compared with swabs or impression smears, and on the importance of histopathology, direct microscopy, and culture for mycetoma and chromoblastomycosis confirmation in well-equipped laboratories. In basic healthcare settings, direct microscopy combined with clinical signs were reported to be the most useful diagnostic indicators to prompt referral for treatment. The survey identified that the diagnosis of sporotrichosis is the most problematic with poor sensitivity across the most widely available laboratory tests except fungal culture, highlighting the need to improve mycological diagnostic capacity and to develop innovative diagnostic solutions. Fungal microscopy and culture are now recognized as WHO essential diagnostic tests and better training in their application will help improve the situation. For mycetoma and sporotrichosis, in particular, advances in identifying specific marker antigens or genomic sequences may pave the way for new laboratory-based or point-of-care tests, although this is a formidable task given the large number of different organisms that can cause fungal NTDs.

Diagnosis and Management of Fungal Neglected Tropical Diseases In Community Settings-Mycetoma and Sporotrichosis.

BACKGROUND: This is a retrospective, analytic observational study where we describe cases of sporotrichosis and mycetoma from Acapulco General Hospital and Community Dermatology Mexico C.A. over 25 years. Analysis of environmental features that favour the development of such diseases has been made, as well as the limitations in the study and treatment of such diseases in resource poor settings. METHODS: We reviewed the information on 76 sporotrichosis and 113 mycetoma patients out of a total of 14,000 consultations at Acapulco General Hospital and from Community Dermatology clinics. We analysed the epidemiological and mycological characteristics and the investigations used for diagnosis such as direct examination, culture, intradermal test reactions, and biopsy. RESULTS: In total 91 confirmed cases of actinomycetoma, 22 of eumycetoma and 76 of sporotrichosis have been identified including diagnostic studies for both diseases and their treatment. DISCUSSION: The results obtained have been analysed and interpreted in patients with mycetoma and sporotrichosis in the state of Guerrero, México, along with limitations in their management in areas with limited economic and logistical resources. The prevalence of mycetoma in our setting is compared with other centres where patients from all over the country are seen. The possible causes for variations in prevalence in specific areas has been looked for, in one of the poorest states of the Mexican Republic.

[Avances recientes en dermatología comunitaria 2016-2017. Teledermatología, un modelo de enseñanza y asistencia en atención primaria a la salud].

INTRODUCTION: Community dermatology (CD) is integrated by dermatologists, epidemiologists and interested people. It has 27 years of experience at the Guerrero State. Due to unsafety issues, the teaching and medical attention model had to be changed from presential to virtual trough telemedicine. METHOD: Transversal, descriptive, observational and analytic study, in which results from teaching-learning of the health personal that was capacitated throughout the telemedicine network at the Guerrero State were evaluated, during nine courses that reached 35 telemedicine centers. RESULTS: 2465 health workers were capacitated in basic dermatology, which were evaluated in their general knowledge of dermatology before and after every course. The utility of this modality of teaching was statistically evaluated with a random simple in 611 attendees with satisfactory results. DISCUSSION: Teaching through telemedicine network, represents an improvement for CD by reaching a higher number of attendees to several reception centers simultaneously, minimizing transportation expenses and improving the safety of the participants at the activities, with the advantage of allowing continuity of the CD purposes.

INTRODUCCIÓN: La dermatología comunitaria (DC) está integrada por dermatólogos, epidemiólogos y personal afín. Tiene 27 años de existencia en el Estado de Guerrero. Por condiciones de inseguridad se cambió el modelo presencial de enseñanza y atención dermatológica a la forma virtual, a través de teledermatología. OBJETIVO: Determinar la utilidad de la capacitación y enseñanza virtual a través de la telemedicina en DC. MÉTODO: Estudio transversal, descriptivo, observacional y analítico, en el que se evaluaron los resultados de la enseñanza-aprendizaje en el personal de salud capacitado por la red de telemedicina mediante nueve cursos que llegaron a 35 centros de telemedicina en el Estado de Guerrero. RESULTADOS: Se capacitó en dermatología básica a 2465 trabajadores de la salud, evaluando sus conocimientos antes y después de cada curso. Se validó estadísticamente la utilidad de esta modalidad de enseñanza con una muestra al azar de 611 asistentes, con resultados satisfactorios. DISCUSIÓN: La enseñanza a través de la red de telemedicina representa un avance en DC al llegar a un mayor número de asistentes en varios centros de recepción en forma simultánea, minimizando los gastos de transporte y salvaguardando la integridad de los participantes, con la ventaja de dar continuidad a los propósitos de DC.

Resistive Part of Impedance as a Possible Indicator of Hepatocellular Carcinoma.

BACKGROUND AND AIMS: In this work, the multi-frequency impedance both in normal and liver cancer tissues was studied. This was to investigate the feasibility to detect liver cancer by a low cost, easy to use, and a relatively non-invasive electrical impedance measure technique, and thus potentially improving liver cancer diagnosis. METHODS: Hepatocellular carcinoma (HCC) was induced in male Wistar rats by the administration of diethylnitrosamine (DEN) during 16 weeks. The electrical impedances at a frequency sweep of 10-100 KHz in the whole body and 10-60 KHz in the liver were taken at the end of the treatment. RESULTS: The electrical impedance showed that the real component values of the impedance change in HCC. In addition, we found that the imaginary component was not associated with HCC. CONCLUSION: Our results suggest that the electrical impedance may be used as a diagnostic HCC tool.

The combination astemizole-gefitinib as a potential therapy for human lung cancer.

Lung cancer is a major cause of cancer mortality. Thus, novel therapies are urgently needed. Repositioning of old drugs is gaining great interest in cancer treatment. Astemizole is an antihistamine proposed to be repositioned for cancer therapy. This drug targets several molecules involved in cancer including histamine receptors, ABC transporters and the potassium channels Eag1 and HERG. Astemizole inhibits the proliferation of different cancer cells including those from cervix, breast, leukemia and liver. Gefitinib is widely used to treat lung cancer; however, no response or drug resistance occurs in many cases. Here, we studied the combined effect of astemizole and gefitinib on the proliferation, survival, apoptosis and gene and protein expression of Eag1 channels in the human lung cancer cell lines A549 and NCI-H1975. Cell proliferation and survival were studied by the MTT method and the colony formation assay, respectively; apoptosis was investigated by flow cytometry. Gene expression was assessed by real-time polymerase chain reaction (RT-PCR), and protein expression was studied by Western blot analysis and immunocytochemistry. We obtained the inhibitory concentrations 20 and 50 (IC20 and IC50, respectively) values for each drug from the cell proliferation experiments. Drug combination at their IC20 had a superior effect by reducing cell proliferation and survival in up to 80% and 100%, respectively. The drugs alone did not affect apoptosis of H1975 cells, but the drug combination at their IC20 increased apoptosis roughly four times in comparison to the effect of the drugs alone. Eag1 mRNA levels and protein expression were decreased by the drug combination in A549 cells, and astemizole induced subcellular localization changes of the channel protein in these cells. Our in vitro studies strongly suggest that the combination astemizole-gefitinib may be a novel and promising therapy for lung cancer patients.

Panfolliculoma: A Clinical Case Report.

Panfolliculoma is a benign neoplasm of follicular differentiation, and its morphological characteristics are similar to those of trichoblastoma, but it shows greater follicular differentiation. A 55-year-old female patient, visited for a dermatological consultation, presented comedo-like lesions on the eyelids, which were accompanied by moderate pruritus that spread gradually. On examination, the patient had localized dermatosis on the face, which symmetrically affected both upper eyelids and to a lesser extent the lower eyelids. The lesions consisted of papulonodular neoformations, comedones, and redundant skin, and a biopsy was performed. The histopathological report mainly described the presence of atrophic epidermis and dermis that was occupied entirely by a neoplasm of epithelial strain, comprising cystic structures that were surrounded by infundibular epithelium of an internal radicular sheath. Panfolliculoma is a rare benign neoplasm of follicular differentiation, and its morphological characteristics are similar to those of trichoblastoma, the pathologist must perform the clinicopathological correlation to establish the correct diagnosis.

Astemizole inhibits cell proliferation in human prostate tumorigenic cells expressing ether à-go-go-1 potassium channels.

Prostate cancer (PC) is the main cause of cancer mortality in men worldwide. Therefore, novel treatments for PC are needed. Ether à-go-go-1 (Eag1) potassium channels display oncogenic properties, and have been suggested as early tumor markers and therapeutic targets for different cancers. These channels are overexpressed in many human tumors including PC. Astemizole targets several molecules involved in cancer including Eag1 channels, histamine receptors and ABC transporters. Here we studied Eag1 mRNA expression and protein levels in the non-tumorigenic and non-invasive human prostate RWPE-1 cell line, and in the tumorigenic and highly invasive human prostate WPE1-NB26 cell lines. The effect of astemizole on cell proliferation and apoptosis was also studied. The human prostate cell lines RWPE-1 and WPE1-NB26 were cultured following the provider´s instructions. Eag1 mRNA expression and protein levels were studied by real time RT-PCR and immunocytochemistry, respectively. Cell proliferation and apoptosis were studied by a fluorescence AlamarBlue®  assay and flow cytometry, respectively. No difference in Eag1 mRNA expression was observed between the cell lines. However, high Eag1 protein levels were observed in the invasive WPE1-NB26 cells, in contrast to the weak protein expression in RWPE-1 cells. Accordingly, astemizole decreased cell proliferation at nanomolar concentrations only in the invasive WPE1-NB26 cells.  Our results suggest that astemizole may have clinical relevance for prostate cancer treatment in patients with high Eag1 protein levels.

Eag1 channels as potential early-stage biomarkers of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is a major cause of cancer death worldwide. HCC is usually asymptomatic at potential curative stages, and it has very poor prognosis if detected later. Thus, the identification of early biomarkers and novel therapies is essential to improve HCC patient survival. Ion channels have been proposed as potential tumor markers and therapeutic targets for several cancers including HCC. Especially, the ether à-go-go-1 (Eag1) voltage-gated potassium channel has been suggested as an early marker for HCC. Eag1 is overexpressed during HCC development from the cirrhotic and the preneoplastic lesions preceding HCC in a rat model. The channel is also overexpressed in human HCC. Astemizole has gained great interest as a potential anticancer drug because it targets several proteins involved in cancer including Eag1. Actually, in vivo studies have shown that astemizole may have clinical utility for HCC prevention and treatment. Here, we will review first some general aspects of HCC including the current biomarkers and therapies, and then we will focus on Eag1 channels as promising tools in the early diagnosis of HCC.

Differential Expression of Ion Channels and Transporters During Hepatocellular Carcinoma Development.

BACKGROUND: Ion channels and transporters are potential markers and therapeutic targets for several cancers. However, their expression during hepatocellular carcinoma (HCC) development remains unclear. AIM: To investigate the mRNA expression of Na(+), K(+) and Ca(2+) channels and ABC transporters during rat HCC development, as well as Abcc3 protein in human liver biopsies. METHODS: Wistar rats were treated with diethylnitrosamine (DEN) and developed both cirrhosis (12 weeks of treatment) and either pre-neoplastic lesions (16 weeks of treatment) or multinodular HCC (16 weeks of treatment plus 2 weeks DEN-free). The mRNA expression of 12 ion channels and two ABC transporters was studied using real-time RT-PCR. Tumor-containing or tumor-free liver sections were isolated by laser-capture microdissection. Abcc3 protein expression was studied by immunohistochemistry in healthy, cirrhotic and HCC human biopsies. RESULTS: We observed expression changes in seven genes. Kcna3, Kcnn4, Kcnrg and Kcnj11 potassium channel mRNA expression reached peak values at the end of DEN treatment, while Scn2a1 sodium channel, Trpc6 calcium channel and Abcc3 transporter mRNA expression reached their highest levels in the presence of HCC (18 weeks). Whereas Kcnn4 and Scn2a1 channel expression was similar in non-tumor and tumor tissue, the Abcc3 transporter and Kcna3 potassium channels were preferentially overexpressed in the tumor sections. We observed differential Abcc3 protein subcellular localization and expression in human samples. CONCLUSIONS: The ion channel/transporter expression profile observed suggests that these genes are potential early markers or therapeutic targets of HCC. The differential localization of Abcc3 may be useful in the diagnosis of cirrhosis and HCC.