RESUMEN

Ovarian cancer patients often suffer from malignant ascites and pleural effusion. Apart from worsening the outcome, this condition frequently impairs the quality of life in patients who are already distressed by ovarian cancer. This study investigated whether single intraperitoneal administration of the anti-VEGF antibody bevacizumab is capable of reducing the ascites-related body surface and prolonging survival. The study was performed in an orthotopic murine model of peritoneal disseminated platin-resistant ovarian cancer. Mice were treated with bevacizumab and/or paclitaxel or buffer (control). Reduction of body surface and increased survival rates were assessed as therapeutic success. Survival of mice in all treatment groups was significantly enhanced when compared to the non-treatment control group. The combination of paclitaxel plus bevacizumab significantly improved body surface as well as overall survival in comparison to a treatment with only one of the drugs. Treatment of malignant effusion with a single dose of bevacizumab as an intraperitoneal application, with or without cytostatic co-medication, may be a powerful alternative to systemic treatment.

RESUMEN

PURPOSE: Targeted oncolytic adenoviruses capable of replication selectively in cancer cells are an appealing approach for the treatment of various cancer types refractory to conventional therapies. The aim of this study was to evaluate the effect of Ad5/3MDR1E1, a multidrug resistance gene 1 (MDR1)-targeted fiber-modified replication-competent adenovirus for the therapy of platinum-pretreated ovarian cancer in combination with cytostatic agents. METHODS: MDR1-specific tumor cell killing of Ad5/3MDR1E1 was systematically evaluated in chemotherapy naïve and pretreated ovarian cancer cells in vitro. Combinations of Ad5/3MDR1E1 and cytostatic agents were studied in vivo and in vitro. An in vivo hepatotoxicity model was used to evaluate liver toxicity. RESULTS: We demonstrate efficient oncolysis of Ad5/3MDR1E1 in chemotherapy-resistant ovarian cancer cells as well as therapeutic efficacy in an orthotopic mouse model. Further, combining Ad5/3MDR1E1 with paclitaxel resulted in greater therapeutic benefit than either agent alone. CONCLUSION: These preclinical data suggest that a fiber-modified adenovirus vector under the control of the MDR1 promoter represents a promising treatment strategy for platinum-pretreated ovarian cancer as a single agent or in combination with conventional anticancer drugs.

Asunto(s)

Viroterapia Oncolítica/métodos , Virus Oncolíticos/fisiología , Neoplasias Ováricas/terapia , Paclitaxel/farmacología , Subfamilia B de Transportador de Casetes de Unión a ATP , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/genética , Adenoviridae/genética , Adenoviridae/fisiología , Animales , Antineoplásicos/farmacología , Proteínas de la Cápside/genética , Línea Celular , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Cisplatino/farmacología , Terapia Combinada , Resistencia a Antineoplásicos , Femenino , Humanos , Ratones , Ratones Endogámicos C57BL , Virus Oncolíticos/genética , Neoplasias Ováricas/patología , Neoplasias Ováricas/virología , Regiones Promotoras Genéticas/genética , Análisis de Supervivencia , Resultado del Tratamiento , Replicación Viral , Ensayos Antitumor por Modelo de Xenoinjerto

RESUMEN

STUDY OBJECTIVE: The aim of this study was to estimate the rate of intrauterine adhesions and subsequent pregnancy outcome in patients with residual trophoblastic tissue treated with hysteroscopic resection versus ultrasound-guided dilation and evacuation (D&E). DESIGN: Cohort study from 2 centers (Canadian Task Force classification II-2). SETTING: Two surgical teams at the University of Duesseldorf Medical Center and the PAN Clinic in Cologne, Germany. PATIENTS: Women with residual trophoblastic tissue after first- or second-trimester miscarriage or term delivery. INTERVENTION: Two techniques were used for the removal of residual trophoblastic tissue: ultrasound-guided evacuation with a curette (D&E) and hysteroscopic resection of trophoblastic tissue (HR). MEASUREMENTS AND MAIN RESULTS: We evaluated 95 patients who underwent secondary intervention for residual trophoblastic disease. A total of 42 patients underwent dilation of the cervix and ultrasound-guided curettage. In a second series of 53 patients, a resectoscope fitted with a 4-mm cutting loop was used for the removal of residual trophoblastic tissue used without application of current. Three months after the intervention, second-look office hysteroscopy was performed. Differences between both treatment groups were statistically significant. After HR, mild intrauterine adhesions were found in 2 patients (4.2%). After D&E, 12 patients (30.8%) presented with intrauterine adhesions (mild intrauterine adhesions: n = 7 [17.9%]; single dense adhesions: n = 3 [7.7%]; and extensive endometrial fibrosis n = 1 [2.6%]). Eighty-two patients wanted to become pregnant. Conception rate of all patients examined was 68.8% (HR) and 59.9% (D&E) (p < .05). In patients younger than 35 years of age who underwent HR, the pregnancy rate was significantly (p < .05) increased compared with patients who underwent D&E (78.1% vs 66.6%). In addition, patients from the HR group demonstrated a significantly (p < .05) shorter time to conception (11.5 month vs 14.5 month). CONCLUSION: The results of this study indicate that selective HR of residual trophoblastic tissue significantly reduces the incidence of intrauterine adhesions and increases pregnancy rates.

Asunto(s)

Legrado/efectos adversos , Endometrio/cirugía , Histeroscopía/efectos adversos , Trofoblastos/patología , Enfermedades Uterinas/cirugía , Aborto Espontáneo/patología , Aborto Espontáneo/cirugía , Adulto , Estudios de Cohortes , Legrado/métodos , Endometrio/patología , Femenino , Humanos , Histeroscopía/métodos , Embarazo , Resultado del Embarazo , Adherencias Tisulares/etiología , Adherencias Tisulares/cirugía , Resultado del Tratamiento , Enfermedades Uterinas/patología

RESUMEN

OBJECTIVE: Multidrug resistance gene 1 (MDR1) mediated resistance to chemotherapeutic agents is a major obstacle for the therapy of various cancer types. The use of conditionally replicating adenoviruses (CRAds) is dependent on molecular differences between tumor cells and non tumor cells. Transcriptional targeting of CRAd replication is an effective way to control replication regulation. The aim of this study was to evaluate the effect of a MDR1 targeted fiber-modified CRAd against chemotherapy resistant ovarian cancer. METHODS: MDR1 expression was evaluated in chemotherapy naïve and pretreated ovarian cancer cells and various control cells. We constructed 2 variants of a fiber-modified CRAd, Ad5/3MDR1E1 and Ad5/3MDR1E1∆24 containing the MDR1 promoter to control viral replication via the E1A gene. The MDR promoter activity and cell killing efficacy were evaluated in vitro. Orthotopic murine models of peritoneally disseminated ovarian cancer were utilized to evaluate the preclinical efficacy of MDR targeted CRAds in vivo. To evaluate the liver toxicity of MDR1 targeted CRAds, we compared Ad5/3MDR1E1 with Ad5/3∆24, a CRAd that replicates in cancer cells inactive in the Rb/p16 pathway by use of an in vivo hepatotoxicity model. RESULTS: We demonstrate efficient oncolysis of Ad5/3MDR1E1 in both chemotherapy resistant ovarian cancer cell lines and in primary tumor cells from pretreated patients as well as therapeutic efficacy in an orthotopic mouse model. Ad5/3MDR1E1 demonstrated significantly decreased liver toxicity compared to other 5/3-fiber modified control vectors examined. CONCLUSIONS: In summary, Ad5/3MDR1E1 is an efficient and safe gene therapy approach for specific targeting of chemotherapy resistant cancer cells.

Asunto(s)

Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/genética , Viroterapia Oncolítica/métodos , Virus Oncolíticos/fisiología , Neoplasias Ováricas/terapia , Subfamilia B de Transportador de Casetes de Unión a ATP , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/biosíntesis , Adenoviridae/genética , Adenoviridae/fisiología , Animales , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/terapia , Neoplasias de la Mama/virología , Línea Celular Tumoral , Resistencia a Antineoplásicos , Femenino , Técnicas de Transferencia de Gen , Humanos , Hígado/virología , Ratones , Ratones Endogámicos C57BL , Viroterapia Oncolítica/efectos adversos , Virus Oncolíticos/genética , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/genética , Neoplasias Ováricas/virología , Regiones Promotoras Genéticas , Replicación Viral

RESUMEN

STUDY OBJECTIVE: The aim of this study was to estimate the benefit of excision of the endocervix during laparoscopic supracervical hysterectomy (LSH) with regard to postoperative cyclical bleeding. DESIGN: Cohort study from 2 centers (Canadian Task Force classification II-2). SETTING: Two surgical teams at the University of Duesseldorf Medical Center and PAN Clinic, Cologne, Germany. PATIENTS: Women with menstrual bleeding disorders resistant to medical treatment, symptomatic leiomyomata, dysmenorrhea. INTERVENTION: Laparoscopic supracervical hysterectomy. The uterus was transsected from the cervix with 2 techniques with and without excision of cervical canal. MEASUREMENTS AND MAIN RESULTS: We evaluated 300 patients who underwent consecutive LSH procedures. In 150 patients the uterus was transsected from the cervix using a monopolar loop. In a second series of 150 patients a unipolar needle electrode was used for the uterine amputation and the excision of cervical canal. The mean duration of the transsection was 65 seconds (monopolar loop) versus 168 seconds (monopolar needle). The excision of the endocervix was performed without any complications in 148 procedures. Histologic examination of the removed tissue revealed endocervical tissue in 83.3% (n = 125), endometrium in 9.4% (n = 14), cervicoisthmic mucosa in 3.3% (n = 5), and myometrium only in 4% (n = 4). All 300 patients were contacted 12 months after surgery to inquire about bleeding status, and 282 (94%) responded. In patients who underwent excision of the endocervix, postoperative cyclical bleeding was significantly reduced compared with the control group (1.4% vs 10.7%). CONCLUSION: The results of this study indicate that the routine excision of the endocervix is a quick safe procedure which allows a significant reduction of postoperative cyclical bleeding in patients who undergo LSH.

Asunto(s)

Histerectomía/efectos adversos , Laparoscopía/métodos , Trastornos de la Menstruación/etiología , Metrorragia/etiología , Adulto , Femenino , Humanos , Histerectomía/métodos , Persona de Mediana Edad , Resultado del Tratamiento

RESUMEN

OBJECTIVE: To evaluate a vascular endothelial growth factor (VEGF)-targeted gene therapy for the treatment of endometriosis. DESIGN: Analysis of the VEGF gene expression and promoter activity in ectopic and eutopic endometrium. Evaluation of the specific replication and cell-killing effect of a VEGF-targeted adenovirus (Ad5VEGFE1) in endometriotic cells. PATIENT(S): Four patients who underwent hysterectomy for benign disease, 30 women with moderate superficial, and 30 women with deep infiltrating endometriosis. INTERVENTION(S): Immunostaining and gene expression of VEGF was examined in eutopic endometrium, endometriotic lesions, and normal peritoneum. The VEGF promoter activity was evaluated in eutopic endometrium and endometriotic lesions. A VEGF-targeted conditionally replicative adenovirus (Ad5VEGFE1) was evaluated regarding specific viral replication in endometriosis cells and induction of apoptosis. The biodistribution of the VEGF-targeted conditionally replicative adenovirus was examined in a mouse model. RESULT(S): The VEGF gene was highly expressed in ectopic endometrium compared with eutopic endometrium and normal peritoneum. The VEGF promoter was active in endometriotic cells. Ad5VEGFE1 showed efficient viral replication and induction of apoptosis in purified primary endometriotic cells and demonstrated a similar lower targeting to the liver and the uterus in a mouse model. CONCLUSION(S): Ad5VEGFE1 is a promising candidate for treating endometriosis and holds potential for clinical testing.

Asunto(s)

Adenoviridae/genética , Endometriosis/terapia , Terapia Genética/métodos , Factor A de Crecimiento Endotelial Vascular/genética , Animales , Apoptosis , Modelos Animales de Enfermedad , Endometriosis/patología , Endometriosis/virología , Femenino , Humanos , Ratones , Ratones Endogámicos C57BL

RESUMEN

For 20 years laparoscopic pelvic and para-aortal lymph node dissection has become increasingly popular as part of minimally invasive surgical treatment concepts for women suffering from gynaecological malignancies. Especially patients suffering from early-stage cervical or endometrial cancers can benefit from the general advantages of a minimally invasive procedure if a comparable degree of radical surgery is achieved. The feasibility and case-control studies published so far suggest comparable indicators of radicality, such as the number of dissected lymph nodes, but also demonstrate potential advantages like a lower intra-operative blood loss, shorter hospital stay and lower postoperative complication rate in comparison with the conventional approach. Regarding long-term survival, reliable data from prospective randomized studies are still lacking but can be expected to be available in the near future.

Asunto(s)

Neoplasias de los Genitales Femeninos/secundario , Neoplasias de los Genitales Femeninos/cirugía , Laparoscopía/tendencias , Escisión del Ganglio Linfático/métodos , Femenino , Neoplasias de los Genitales Femeninos/patología , Humanos , Metástasis Linfática

RESUMEN

UNLABELLED: Endometrial carcinoma is the most common genital cancer in women. While patients usually present with vaginal bleeding, in 10-20% this characteristic symptom is absent. Endometrial thickness (double layer) is measured by transvaginal sonography and thickening indicates an increased risk of malignancy or other pathology (hyperplasia or polyps). OBJECTIVE: We sought to correlate hysteroscopic and pathological findings in asymptomatic postmenopausal women with sonographically thickened endometrium (>6mm). STUDY DESIGN: A prospective observational study in a university hospital of 304 postmenopausal women referred between 1996 and 2006 because of a sonographically thickened endometrium in the absence of abnormal bleeding, who underwent continuous flow hysteroscopy (4.5mm Storz hysteroscope) and fractionated curettage of the uterine cervix and corpus (D & C) in addition to vaginal sonography (5MHz probe). RESULTS: The mean age of the women was 64.8 (range 57.7-71.9) years. Average endometrial thickness measured by ultrasound was 12mm+/-6.7mm. Hysteroscopy suggested the presence of endometrial polyps in 226 women (74.3%), simple endometrial hyperplasia in 34 (11.2%), atrophic endometrium in 18 (5.9%), complex endometrial hyperplasia in 2 (0.7%), atypical hyperplasia in 3 (1%) and leiomyoma in 9 (3.0%). In 12 women (3.9%), the hysteroscopic appearance suggested malignancy and histology revealed endometrial adenocarcinoma. All hysteroscopic results were confirmed by histological examination. CONCLUSION: Hysteroscopy represents an easy, safe and effective method for the investigation of asymptomatic women with a thickened endometrium found with transvaginal ultrasound. The commonest pathology was endometrial polyps.

Asunto(s)

Hiperplasia Endometrial/diagnóstico , Neoplasias Endometriales/diagnóstico , Histeroscopía , Neoplasias Uterinas/diagnóstico , Anciano , Atrofia/diagnóstico , Hiperplasia Endometrial/patología , Endometrio/diagnóstico por imagen , Endometrio/patología , Femenino , Humanos , Histeroscopía/métodos , Leiomioma/diagnóstico , Persona de Mediana Edad , Pólipos/diagnóstico , Posmenopausia , Estudios Prospectivos , Factores de Riesgo , Ultrasonografía , Enfermedades Uterinas/diagnóstico

RESUMEN

PURPOSE: Recently, virotherapy has been proposed as a new therapeutic approach for ovarian cancer. Conditionally replicative adenoviruses (CRAd) may contain tumor-specific promoters that restrict virus replication to cancer cells. Mesothelin, a cell surface glycoprotein, is overexpressed in ovarian cancer but not in normal ovarian tissues. The purpose of this study was to explore the therapeutic utility of a mesothelin promoter-based CRAd in a murine model of ovarian cancer, using noninvasive in vivo imaging. EXPERIMENTAL DESIGN: We constructed a mesothelin promoter-based CRAd with a chimeric Ad5/3 fiber (AdMSLNCRAd5/3) that contains an Ad5 tail, Ad5 shaft, and an Ad3 knob. Previously, a chimeric Ad5/3 fiber has shown improved infectivity in many ovarian cancer cells. Viral replication and oncolysis were assessed in a panel of ovarian cancer cell lines. To test the oncolytic efficacy of AdMSLNCRAd5/3 in a murine model, bioluminescence imaging of tumor luciferase activity and survival analysis were done. RESULTS: AdMSLNCRAd5/3 achieved up to a 10,000-fold higher cell killing effect and up to 120-fold higher levels of viral replication in all human ovarian cancer cells, compared with wild-type Ad5. AdMSLNCRAd5/3 significantly inhibited tumor growth as confirmed by in vivo imaging (P < 0.05). Survival with AdMSLNCRAd5/3 was significantly enhanced when compared with no virus or with a wild-type Ad5-treated group (P < 0.05). CONCLUSIONS: The robust replication, oncolysis, and in vivo therapeutic efficacy of AdMSLNCRAd5/3 showed that this CRAd is a promising candidate for treating ovarian cancer. Importantly, we have applied in vivo imaging that has allowed repeated and longitudinal measurements of tumor growth after CRAd treatment.

Asunto(s)

Adenoviridae/genética , Glicoproteínas de Membrana/genética , Viroterapia Oncolítica/métodos , Neoplasias Ováricas/terapia , Regiones Promotoras Genéticas , Proteínas E1A de Adenovirus/genética , Animales , Femenino , Proteínas Ligadas a GPI , Vectores Genéticos , Humanos , Mesotelina , Ratones , Ratones SCID , Neoplasias Ováricas/virología , Reacción en Cadena de la Polimerasa , Ensayos Antitumor por Modelo de Xenoinjerto

RESUMEN

PURPOSE: Adenovirus serotype 5 (Ad5) has been used for gene therapy with limited success due to insufficient infectivity in cells with low expression of the primary receptor, the coxsackie and adenovirus receptor (CAR). Evidence that adenovirus serotype receptors other than CAR may be of use was presented in previous studies that showed that the Ad3 receptor is expressed at high levels in ovarian cancer cells. We hypothesized that combined use of unique chimeric fibers in the context of novel mosaic adenovirus vectors would enhance infectivity via non-CAR pathways in ovarian cancer cells. EXPERIMENTAL DESIGN: We constructed and characterized Ad5 vectors that use Ad3 knob and reovirus fibers to generate a mosaic fiber virion. Serotype 3 Dearing reovirus uses a fiber-like sigma 1 protein to infect cells expressing sialic acid and junction adhesion molecule 1. We therefore constructed a mosaic fiber Ad5 vector, designated Ad5/3-sigma 1, encoding two fibers: a sigma 1 chimeric fiber and the chimeric Ad5/3 fiber composed of an Ad3 knob. RESULTS: Functionally, Ad5/3-sigma 1 used sialic acid, junction adhesion molecule 1, and Ad3 receptor for cell transduction and achieved maximum infectivity enhancement in ovarian cancer cells with low CAR expression. Furthermore, Ad5/3-sigma 1 achieved infectivity enhancement in primary tissue slices of human ovarian tumor. CONCLUSIONS: We have developed a new type of Ad5 vector with the novel tropism, possessing fibers from Ad3 and reovirus, which exhibits enhanced infectivity via CAR-independent pathway(s). In addition, the flexible genetic platform of vector allows different combination of fiber variants that can be incorporated within the same particle.

Asunto(s)

Adenoviridae/genética , Terapia Genética/métodos , Vectores Genéticos/genética , Vectores Genéticos/metabolismo , Neoplasias Ováricas/terapia , Receptores Virales/metabolismo , Proteínas de la Cápside/genética , Línea Celular Tumoral , Proteína de la Membrana Similar al Receptor de Coxsackie y Adenovirus , Femenino , Humanos , Ácido N-Acetilneuramínico/farmacología , Neoplasias Ováricas/química , Neoplasias Ováricas/metabolismo , Receptores Virales/análisis , Transducción Genética/métodos

RESUMEN

Ovarian cancer is currently the most lethal gynecologic malignancy in Europe and the US. Platin analogues and paclitaxel demonstrate high remission rates, but unfortunately the efficacy of cytostatic agents is limited by the development of multidrug resistance (mdr). Clinical paclitaxel resistance is often associated with mdr1 overexpression. In a recent study, we introduced a highly specific quantitative real-time reverse transcriptase polymerase chain reaction for the quantification of mdr1 transcripts. In the present study, we demonstrate that primary tumor cells from patients with recurrent ovarian cancer overexpress mdr1. To evaluate mdr1 expression, we collected tumor cells from 77 ovarian cancer patients (13 chemotherapy-naive ovarian cancer, 64 recurrent ovarian cancer). Cancer cells were aspirated from 49 solid specimens (63%) and 28 ascitic fluids (37%). Subsequently, cancer cells were exposed in 221 short-term cultures either to blank medium (control) or to a single anticancer drug, cisplatin, doxorubicin or paclitaxel. The drug concentrations applied referred to clinical relevant doses. mdr1 mRNA expression was significantly higher in specimens from recurrent ovarian cancer incubated in paclitaxel than in specimens from chemotherapy-naive ovarian cancer. No significant differences were detectable between the mean value of mdr1 mRNA expression in tumor specimens from recurrent ovarian cancer incubated in cisplatin or doxorubicin. Differences within the untreated patients group were also not statistically significant. The result of this study confirms clinical observations, as well as in-vitro studies based on tumor cell lines, that paclitaxel resistance is correlated with mdr1 overexpression.

Asunto(s)

Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/genética , Antineoplásicos/farmacología , Recurrencia Local de Neoplasia/tratamiento farmacológico , Neoplasias Ováricas/tratamiento farmacológico , Adulto , Anciano , Línea Celular Tumoral , Cisplatino/farmacología , Doxorrubicina/farmacología , Femenino , Humanos , Persona de Mediana Edad , Recurrencia Local de Neoplasia/genética , Neoplasias Ováricas/genética , Paclitaxel/farmacología , ARN Mensajero/análisis

RESUMEN

Gene transfer presents a potentially useful approach for the treatment of diseases refractory to conventional therapies. Various preclinical and clinical strategies have been explored for treatment of gynecological diseases. Given the direst need for novel treatments, much of the work has been performed with gynecological cancers and ovarian cancer in particular. Although the safety of many approaches has been demonstrated in early phase clinical trials, efficacy has been mostly limited so far. Major challenges include improving gene transfer vectors for enhanced and selective delivery and achieving effective penetration and spread within advanced and complex tumor masses. This review will focus on current and developmental gene transfer applications for gynecological diseases.

Asunto(s)

Terapia Genética/métodos , Enfermedades de los Genitales Femeninos/terapia , Neoplasias de los Genitales Femeninos/terapia , Neoplasias Ováricas/terapia , Adenoviridae/genética , Adenoviridae/inmunología , Adenoviridae/metabolismo , Animales , Ensayos Clínicos como Asunto , Proteína de la Membrana Similar al Receptor de Coxsackie y Adenovirus , Femenino , Genes Transgénicos Suicidas , Genes Supresores de Tumor , Terapia Genética/efectos adversos , Vectores Genéticos , Humanos , Neovascularización Patológica/terapia , Viroterapia Oncolítica/métodos , Neoplasias Ováricas/metabolismo , Receptores de Factores de Crecimiento/metabolismo , Receptores Virales/metabolismo

RESUMEN

Adenovirus (Ad)-based cancer gene therapy is a promising, novel approach for treating cancer resistant to established treatment modalities. Unfortunately, the efficacy of nonreplicative first generation Ads was low and data from clinical trials were disappointing. To address this problem, conditionally replicating Ads have been constructed. Infection of tumor cells with conditionally replicating Ads results in tumor-specific replication, subsequent oncolysis and release of the virus progeny. Recently, it has been suggested that the low expression of the coxsackie-Ad receptor is the rate-limiting factor for infectivity with serotype 5 (Ad5). Unfortunately, coxsackie-Ad receptor expression is highly variable and often low on many tumor types. Consequently, molecular strategies have been applied for the development of coxsackie-Ad receptor-independent oncolytic Ads. This review describes recent developments of Ad-based cancer gene therapy, including novel engineering techniques of the Ad capsid for efficient tumor targeting, as well as targeting techniques, to restrict transgene expression to cancer cells.

Asunto(s)

Adenoviridae/genética , Terapia Genética/tendencias , Neoplasias/terapia , Adenoviridae/fisiología , Protocolos de Quimioterapia Combinada Antineoplásica/administración & dosificación , Proteína de la Membrana Similar al Receptor de Coxsackie y Adenovirus , Vectores Genéticos , Humanos , Receptores Virales/genética , Receptores Virales/metabolismo , Replicación Viral

RESUMEN

Gene therapy with adenoviral (Ad) vectors is a promising new approach in the treatment of cancer. Strategies to restrict adenoviral-mediated transgene expression are important to avoid gene transfer into normal cells. Heparanase (HPR) is overexpressed in breast cancer but downregulated in differentiated normal tissue. Expression of the HPR gene was evaluated in breast cancer cells. Biodistribution and liver tropism was evaluated in a mouse model. HPR is highly expressed in breast cancer tissue. The HPR promoter retained its fidelity in an adenovirus context and was activated in breast cancer cells but showed low activity in normal breast cells and the murine liver. We conclude that the HPR pathway is a promising target for the development of breast cancer directed gene therapy strategies.

Asunto(s)

Neoplasias de la Mama/terapia , Terapia Genética , Glucuronidasa/genética , Regiones Promotoras Genéticas/genética , Adenoviridae/genética , Adenoviridae/metabolismo , Animales , Neoplasias de la Mama/enzimología , Neoplasias de la Mama/genética , Línea Celular Tumoral , Femenino , Genes Reporteros , Vectores Genéticos , Glucuronidasa/metabolismo , Humanos , Hígado/metabolismo , Luciferasas , Ratones , ARN Mensajero/metabolismo , Transfección

RESUMEN

PURPOSE: The use of conditionally replicating adenoviruses (CRAD) is dependent on molecular differences between tumor cells and nontumor cells. Transcriptional targeting of CRAD replication via tumor-specific promoters is an effective way to control replication regulation. Genetic fiber pseudotyping is an approach for circumventing low expression of the primary adenovirus serotype 5 (Ad5) receptor by using the distinct adenovirus serotype 3 (Ad3) receptor for entry into and subsequent killing of ovarian cancer cells. EXPERIMENTAL DESIGN: In this study, we constructed a fiber-modified CRAD containing the secretory leukoprotease inhibitor (SLPI) promoter to control viral replication via the E1A gene (Ad5/3SLPI). To evaluate the liver toxicity of chimeric 5/3 fiber-modified CRADs, we compared Ad5/3SLPI with Ad5/3Cox-2L, a CRAD with E1A under control of the Cox-2 promoter, and Ad5/3Delta24, a CRAD that replicates in cancer cells inactive in the retinoblastoma/p16 pathway by use of an in vivo hepatotoxicity model and by a model system that uses slices of human liver. RESULTS: We show efficient viral replication and oncolysis of Ad5/3SLPI in both multiple ovarian cancer cell lines and primary tumor cell spheroids as well as therapeutic efficacy in an orthotopic mouse model of peritoneal carcinomatosis. Ad5/3SLPI showed significantly decreased liver toxicity compared with other 5/3 fiber-modified control vectors examined. CONCLUSIONS: In summary, Ad5/3SLPI is a promising vector candidate for treating metastatic ovarian cancer and showed robust virus replication, oncolysis, and in vivo therapeutic efficacy. Ad5/3SLPI showed comparatively low liver toxicity and therefore holds potential for patient use in the clinic.

Asunto(s)

Adenoviridae/genética , Proteínas de la Cápside/genética , Neoplasias Ováricas/virología , Regiones Promotoras Genéticas/genética , Proteínas/genética , Adenoviridae/crecimiento & desarrollo , Proteínas E1A de Adenovirus/genética , Animales , Línea Celular Tumoral , Proliferación Celular , Femenino , Vectores Genéticos/administración & dosificación , Vectores Genéticos/genética , Humanos , Técnicas In Vitro , Hígado/metabolismo , Hígado/patología , Hígado/virología , Ratones , Ratones Endogámicos C57BL , Neoplasias Ováricas/patología , Neoplasias Ováricas/terapia , Proteínas Inhibidoras de Proteinasas Secretoras , ARN Mensajero/genética , ARN Mensajero/metabolismo , Inhibidor Secretorio de Peptidasas Leucocitarias , Esferoides Celulares/patología , Esferoides Celulares/virología , Análisis de Supervivencia , Replicación Viral , Ensayos Antitumor por Modelo de Xenoinjerto/métodos

RESUMEN

BACKGROUND: Gene therapy is a novel approach for treatment of patients with advanced, recurrent, or metastatic cervical cancer. One effective way to direct transgene expression to specific tissues or tumors is the use of tissue-specific-promoters (TSPs). In the context of adenovirus (Ad)-mediated cancer gene therapy it is rational to choose a TSP which is highly expressed in the tumor but has potentially low activity in non-tumor cells, especially the liver. In this study, we have investigated several promoters which fulfill these criteria. Candidate cervical cancer specific TSPs include promoters of the genes for secretory leukoprotease inhibitor (SLPI), cyclooxygenase-2 (COX-2), Midkine (MK), vascular endothelial growth factor receptor type 1 (flt-1), vascular endothelial growth factor (VEGF), Survivin and the receptor for chemokine SDS-1 (CXCR4). METHODS: To evaluate the specific gene expression of the different promoters in the context of cervical cancer, we constructed a panel of E1-deleted Ads that express luciferase under the control of the promoters of interest. We investigated various established cervical cancer cell lines, as well as purified primary cancer cells and normal control cells from the cervix uteri. RESULTS: In all cell lines tested, promoters for MK, VEGF and CXCR4 showed the highest activity. Both MK and VEGF promoters also resulted in a high activity in primary cervical cancer cells. Interestingly, gene expression profiles correlate with luciferase activity in both cell lines and primary cancer samples. CONCLUSIONS: Our study demonstrates that the promoters for MK and VEGF are active in cervical cancer. We believe that both promoters can be successfully employed as TSPs for gene therapy targeted to cervical cancer.

Asunto(s)

Regiones Promotoras Genéticas , Neoplasias del Cuello Uterino/genética , Adenoviridae/genética , Línea Celular Tumoral , Ciclooxigenasa 2/genética , Citocinas/genética , Proteínas de la Matriz Extracelular/genética , Femenino , Humanos , Proteínas Inhibidoras de la Apoptosis , Proteínas Asociadas a Microtúbulos/genética , Midkina , Cadenas Pesadas de Miosina , Proteínas de Neoplasias/genética , Miosina Tipo IIB no Muscular , Proteínas Inhibidoras de Proteinasas Secretoras , Proteínas/genética , Receptores CXCR4/genética , Inhibidor Secretorio de Peptidasas Leucocitarias , Survivin , Factor A de Crecimiento Endotelial Vascular/genética

RESUMEN

Successful adenoviral (Ad) vector-mediated strategies for cancer gene therapy mandate gene-delivery systems that are capable of achieving efficient gene delivery in vivo. In many cancer types, in vivo gene-transfer efficiency remains limited due to the low or highly variable expression of the primary Ad receptor, the coxsackie Ad receptor (CAR). In this study, we evaluated the expression of CAR on cervical cancer cells as well as CAR-independent targeting strategies to integrins (Ad5.RGD), heparan sulfate proteoglycans (Ad5.pK7) or both (Ad5.RGD.pK7). We used a panel of established cervical cancer cell lines and primary cervical cancer cells isolated from patients to quantify the expression of CAR mRNA and to evaluate the gene-transfer efficiency of fiber-modified Ads. Of the fiber-modified vectors, Ad5.pK7 and Ad5.RGD.pK7 displayed significantly enhanced gene-transfer efficiency in vitro. Gene-delivery efficiency in vivo was evaluated using an s.c. cervical cancer mouse model. Ad5.RGD.pK7 significantly improves tumor targeting in vivo, resulting in a significantly improved tumor/liver ratio in mice. Our results suggest that the double-modified Ad5.RGD.pk7 vector enhances gene transfer to clinically relevant cervical cancer substrates, while the infectivity of nontarget cells in the mouse is not increased and comparable to Ad5. The fiber-modified virus described here can help achieve higher clinical efficacy of cervical cancer gene therapy.

Asunto(s)

Regulación Neoplásica de la Expresión Génica , Técnicas de Transferencia de Gen , Terapia Genética/métodos , Vectores Genéticos , Receptores Virales/biosíntesis , Receptores Virales/genética , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/terapia , Adenoviridae/genética , Animales , Modelos Animales de Enfermedad , Enterovirus/genética , Femenino , Humanos , Integrinas , Ratones , ARN Mensajero/análisis , Células Tumorales Cultivadas

RESUMEN

This study was performed in order to investigate the role of the apoptotic index (AI) as a prediction parameter for the prognosis of patients with primary breast cancer. AI was determined by DNA fragmentation on 298 primary breast cancer samples and compared to clinically established breast cancer parameters. Additionally, we determined the expression of functional parameters including proliferating cell nuclear antigen, p21waf and p27kip by immunohistochemistry. The mean AI was found to be 11.9% (range, 0-90%). 189 tumors (63.4%) were negative for apoptosis, while 109 tissue samples (36.6%) were apoptotic with >5% positive cells. Using univariate analysis (chi2 test), the AI did not show any significant correlation to one of the established prognostic parameters of primary breast cancer (p > 0.05). In contrast, we found a significant positive correlation to the expression of the cell cycle inhibitors p21waf (p = 0.04) and p27kip (p = 0.024). During the clinical follow-up (median observation time for disease-free survival 87 months), several clinically established prognostic parameters including menopausal status, nodal status, tumor size, tumor grade, and hormone receptor expression could be confirmed and were analyzed with respect to the AI in the tumor. Furthermore, AI displayed a significant positive correlation to disease-free survival using Kaplan-Meier survival analysis (log-rank test, p = 0.04). However, AI lost its prognostic significance in multivariate analysis based on the Cox proportional hazard model (relative risk 0.8, confidence interval 0.52-1.33, p = 0.44). Our data indicate that high apoptotic rates in cancer tissues are indicative of a favorable patient outcome. However, the AI was not an independent factor. The study provides indirect evidence that this process may involve cell cycle inhibitors physiologically.

Asunto(s)

Apoptosis , Neoplasias de la Mama/patología , Carcinoma Ductal de Mama/patología , Proteínas de Ciclo Celular/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/mortalidad , Carcinoma Ductal de Mama/metabolismo , Carcinoma Ductal de Mama/mortalidad , Inhibidor p21 de las Quinasas Dependientes de la Ciclina , Inhibidor p27 de las Quinasas Dependientes de la Ciclina , ADN de Neoplasias/análisis , Femenino , Técnica del Anticuerpo Fluorescente Indirecta , Humanos , Pronóstico , Antígeno Nuclear de Célula en Proliferación/metabolismo , Tasa de Supervivencia

RESUMEN

Treatment options for disseminated cervical cancer remain inadequate. Here, we investigated a strategy featuring Ad5-Delta 24 RGD, an oncolytic adenovirus replication-competent selectively in cells defective in the Rb-p16 pathway, such as most cervical cancer cells. The viral fiber contains an alpha(v)beta(3) and alpha(v)beta(5) integrin-binding RGD-4C motif, allowing coxsackie-adenovirus receptor-independent infection. These integrins have been reported to be frequently upregulated in cervical cancer. Oncolysis of cervical cancer cells was similar to a wild-type control in vitro. In an animal model of cervical cancer, the therapeutic efficacy of Ad5-Delta 24 RGD could be demonstrated for both intratumoral and intravenous application routes. Biodistribution was determined following intravenous administration to mice. Further preclinical safety data were obtained by demonstrating lack of replication of the agent in human peripheral blood mononuclear cells. These results suggest that Ad5-Delta 24 RGD could be useful for local or systemic treatment of cervical cancer in patients with disease resistant to currently available modalities.

Asunto(s)

Adenoviridae , Neoplasias del Cuello Uterino/terapia , Animales , Terapia Biológica/métodos , Modelos Animales de Enfermedad , Femenino , Humanos , Infusiones Intravenosas , Leucocitos Mononucleares , Ratones , Distribución Tisular , Células Tumorales Cultivadas , Replicación Viral

RESUMEN

Approaches to alter the native tropism of adenoviruses (Ads) are beneficial to increase their efficacy and safety profile. Liver tropism is important with regard to potential clinical toxicity in humans. Ad5/3 chimeras in which the Ad5 knob is substituted by the Ad3 knob, such as Ad5/3luc1, have been recently shown to increase infectivity of ovarian cancer cell lines and primary tumor cells, which express low levels of the coxsackie-adenovirus receptor (CAR), without increasing infectivity of liver cells. A novel strategy to address the problem of liver uptake and improve the tumor/liver ratio is genetic replacement of the Ad fiber shaft. Ad5.Ad3.SH.luc1 is an Ad5-based vector that contains the fiber shaft from Ad serotype 3 but the fiber knob from Ad serotype 5. To compare tumor/liver of Ad5.Ad3.SH.luc1 and Ad5/3luc1 in vivo, we created three different tumor and treatment models of ovarian cancer in mice, simulating intraperitoneal and intravenous administration of tumors. Ad5.Ad3.SH.luc1 displayed the lowest liver tropism of all viruses in all models tested. Intravenous administration of all viruses resulted in higher tumor transduction rates compared to intraperitoneal administration. Genetic shortening of the Ad5 fiber shaft significantly increases relative tumor/liver gene transfer. This could improve the effective tumor dose and reduce side effects, thereby increasing the bioavailability of therapeutic agents.

Asunto(s)

Adenovirus Humanos/genética , Técnicas de Transferencia de Gen , Hígado/metabolismo , Neoplasias Ováricas/terapia , Animales , Línea Celular Tumoral , Células Epiteliales/metabolismo , Femenino , Terapia Genética , Vectores Genéticos , Humanos , Inyecciones Intraperitoneales , Inyecciones Intravenosas , Inyecciones Subcutáneas , Luciferasas/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Desnudos , Trasplante de Neoplasias , Especificidad de Órganos , Proteínas Recombinantes/metabolismo , Recombinación Genética , Serotipificación , Transducción Genética , Ensayos Antitumor por Modelo de Xenoinjerto