Assessing dynamic change in muscle during treatment of patients with cancer: Precision testing standards.

BACKGROUND: Computed tomography images acquired during routine cancer care provide an opportunity to determine body composition with accuracy and precision. Quantification of skeletal muscle is of interest owing to its association with clinical outcomes. However, the standards of precision testing considered mandatory in other areas of radiology are lacking from the literature in this area. We aim to describe the change in skeletal muscle over time at different anatomical levels using the precision error. METHODS: Thirty-eight male patients with squamous cell carcinoma of the head and neck were evaluated at two time points encompassing their treatment plan. Precision testing consisted of analyzing the cross-sectional area (CSA) of the skeletal muscle and total adipose tissue of 76 CT studies (38 images at baseline repeated twice and 38 follow-up images repeated twice) measured by a skilled observer. The % coefficient of variation (%CV), the root-mean-square standard deviation (RMS SD) and the corresponding 95% least significant change (LSC) were calculated for four anatomical levels: upper arm, thigh, chest and abdomen. RESULTS: The median time between scans was 223.6 (SD 31.2) days. Precision error (% CV) for total skeletal muscle cross sectional area was 0.86% for upper arm, 0.26% for thigh, 0.39% for chest and 0.63% for abdomen. The corresponding LSC values in upper arm, thigh, chest and abdomen were 2.4%, 0.7%, 1.1% and 1.8%, respectively. Based on the LSC for RMS SD, patients were classified in two categories according to muscle cross-sectional area: stable (i.e within LSC value) or gained and loss. To compare the four anatomical levels, the proportion of patients with muscle loss exceeding the LSC value was 74.3% for arm, 86.2% for thigh, 82.9% for chest and 76.3% for abdomen. For these same anatomic regions, the mean muscle loss for those patients classified below the LSC was 14.6% (SD 9.3), 13.4% (SD 7.8), 11.9% (SD 6.5) and 11.6% (SD 5.5), respectively. Only the loss of muscle area was significantly higher in thigh (p = 0.023), using L3 as the reference level. CONCLUSIONS: We recommend the uniform use of a standard precision test when reporting muscle change over time. LSC values vary from 0.7 to 2.4% depending on anatomic site; with the lowest precision error to detect change in the thigh. Based on this analysis, muscle wasting appears to be systemic and while present in limbs and trunk is significantly higher in the thigh than in the chest, abdomen or upper arm.

Biologic treatments for psoriasis: the cost of alternative methods of dispensation of medication in Spain.

OBJECTIVE: Hospital pharmacy services have adapted to the COVID­19 pandemic. The aim of the study is to determine the economic  consequences of replacing hospital pharmacy dispensation with other  dispensing methods in the context of biological treatments for psoriasis in  Spain. METHOD: Multiple dispensation scenarios were evaluated, combining different  dispensation frequencies and sites, and telepharmacy followup intervals. Self- injectable biological medicines for psoriasis (interleukin and tumour necrosis  factor alpha inhibitors) were included. All costs (in 2020 euros) were  considered from the perspective of the National Health System. RESULTS: The annual cost of hospital pharmacy-based dispensations every 4  weeks combined with telepharmacy monitoring at each administration ranged  from €194.9 to €2,088.0 per patient. Across the different simulated scenarios,  biological medicines associated with the lowest cost were those administered  less frequently (every 12 weeks). CONCLUSIONS: In the post-COVID-19 era, new models of hospital  pharmaceutical care that include changes in drug dispensation and  telepharmacy strategies will have economic consequences for the National  Health System that merit consideration.

Objetivo: Los servicios de farmacia hospitalaria se han adaptado a la pandemia de COVID-19. El objetivo del estudio es determinar las  consecuencias económicas de sustituir la dispensación de medicamentos en el servicio de farmacia hospitalaria por otros métodos de dispensación en el  contexto de los tratamientos biológicos para la psoriasis en España.Método: Se evaluaron múltiples escenarios de dispensación, combinando diferentes frecuencias y lugares de dispensación, y frecuencias del seguimiento de telefarmacia. Se incluyeron los medicamentos biológicos autoinyectables para la psoriasis (inhibidores de interleucinas y del  factor de necrosis tumoral alfa). Todos los costes (euros de 2020) se  consideraron desde la perspectiva del Sistema Nacional de Salud.Resultados: Considerando la dispensación en la farmacia hospitalaria, la  frecuencia de dispensación cada 4 semanas y la telefarmacia en cada  administración, el coste anual de dispensación por paciente osciló entre 194,9  € y 2.088,0 €. En los diferentes escenarios simulados, los fármacos biológicos  que se asociaron a un coste inferior fueron los que se administran de forma  más espaciada en el tiempo (cada 12 semanas).Conclusiones: En la era post-COVID-19, los nuevos modelos de atención farmacéutica hospitalaria que consideran cambios en la dispensación  farmacológica y la telefarmacia tendrán consecuencias económicas para el Sistema Nacional de Salud que merecen consideración.

Clinical outcomes after more conservative management of patent ductus arteriosus in preterm infants / Desfechos clínicos após tratamento mais conservador da persistência do canal arterial em prematuros

Abstract Objective: Management of patent ductus arteriosus is still controversial. This study aimed to describe the impact of a more conservative approach on treatment rates and on main outcomes of prematurity, especially in preterm infants with <26 weeks of gestation. Method: Clinical charts review of infants ≤30 weeks with patent ductus arteriosus between 2009 and 2016 at two centers. In 2011, the authors changed patent ductus arteriosus management: in first period (2009-2011), patients who failed medical treatment underwent surgical closure; in second period (2012-2016), only those with cardiopulmonary compromise underwent surgical ligation. Medical treatment, surgical closure, mortality, and survival-without-morbidity were compared. Results: This study included 188 patients (27 ± 2 weeks, 973 ± 272 grams); 63 in P1 and 125 in P2. In P2, significantly lower rates of medical treatment (85.7% P1 versus 56% P2, p < 0.001) and surgical closure (34.5% P1 versus 16.1% P2, p < 0.001) were observed. No differences were found in chronic lung disease (28.8% versus 13.9%, p = 0.056), severe retinopathy of prematurity (7.5% versus 11.8%, p = 0.403), necrotizing enterocolitis (15.5% versus 6.9%, p = 0.071), severe intraventricular hemorrhage (25.4% versus 18.4%, p = 0.264), mortality (17.5% versus 15.2%, p = 0.690) or survival-without-morbidity adjusted OR = 1.10 (95% CI: 0.55-2.22); p = 0.783. In P2, 24.5% patients were discharged with patent ductus arteriosus. The subgroup born between 23 and 26 weeks (n = 82) showed significant differences: lower incidence of chronic lung disease (50% versus 19.6%, p = 0.019) and more survival-without-morbidity (20% versus 45.6%, p = 0.028) were found. Conclusion: A conservative approach in preterm infants with patent ductus arteriosus can avoid medical and surgical treatments, without a significant impact in survival-without-morbidity. However, two-thirds of preterm infants under 26 weeks are still treated.

Resumo Objetivo O tratamento da persistência do canal arterial ainda é controverso. Nosso objetivo foi descrever o impacto de uma abordagem mais conservadora em nossas taxas de tratamento e nos principais desfechos da prematuridade, especialmente em prematuros com < 26 semanas de gestação. Método Revisão de prontuários de lactentes com ≤ 30 semanas e persistência do canal arterial entre 2009-2016 em dois centros. Em 2011, mudamos o manejo da persistência do canal arterial: no primeiro período (2009-2011), os pacientes que não apresentaram sucesso com o tratamento clínico foram submetidos a fechamento cirúrgico; no segundo período (2012-2016), apenas aqueles com comprometimento cardiopulmonar foram submetidos ao fechamento cirúrgico. Comparamos o tratamento clínico, fechamento cirúrgico, mortalidade e sobrevida sem morbidade. Resultados Foram incluídos 188 pacientes (27 ± 2 semanas, 973 ± 272 gramas); 63 em P1 e 125 em P2. Em P2, foram observadas taxas significativamente mais baixas de tratamento clínico (85,7% no P1 versus 56% no P2, p < 0,001) e fechamento cirúrgico (34,5% no P1 versus 16,1% no P2, p < 0,001). Não foram encontradas diferenças em relação à doença pulmonar crônica (28,8% versus 13,9%, p = 0,056), retinopatia grave da prematuridade (7,5% versus 11,8%, p = 0,403), enterocolite necrosante (15,5% versus 6,9%, p = 0,071), hemorragia intraventricular grave (25,4% versus 18,4%, p = 0,264), mortalidade (17,5% versus 15,2%, p = 0,690) ou OR ajustado pela sobrevida sem morbidade = 1,10 (IC95%: 0,55-2,22); p = 0,783. Em P2, 24,5% dos pacientes receberam alta com persistência do canal arterial. O subgrupo nascido entre 23 a 26 semanas (n = 82) apresentou diferenças significativas, foram encontradas menor incidência de doença pulmonar crônica (50% versus 19,6%, p = 0,019) e maior sobrevida sem morbidade (20% versus 45,6%, p = 0,028). Conclusão Uma abordagem conservadora em prematuros com persistência do canal arterial pode evitar tratamentos clínicos e cirúrgicos, sem um impacto significativo na sobrevida sem morbidade. No entanto, dois terços dos prematuros com menos de 26 semanas ainda são tratados.

Tumor suppressive function of E2F-1 on PTEN-induced serrated colorectal carcinogenesis.

Many human cancers present Phosphatase and tensin homolog (PTEN) deficiency and between 20 and 30% of colorectal tumors show PTEN loss. The transcription factor, E2 promoter binding factor 1 (E2F-1), exhibits tumor promoter or suppressive functions depending on cellular type and tissue context, but its role in the progression and development of colorectal carcinogenesis was largely unknown. Here, using a tamoxifen-inducible PTEN knockout mouse model, we have demonstrated that loss of PTEN leads to the development of colorectal tumorigenesis through the serrated pathway. Next, we studied PTEN loss-driven colorectal lesions in the context of E2F-1 deficiency in vivo. Our results revealed that monoallelic and biallelic absence of E2F-1 led to an increased incidence and progression of serrated tumorigenesis induced by PTEN loss. Finally, we investigated the mechanisms by which double PTEN/E2F-1 deficiency leads to enhanced tumorigenesis. We found that colorectal tumors from PTEN/E2F-1 double knockout mice and the human colorectal carcinoma cell line HT29 with shRNA-mediated downregulation of PTEN and E2F-1 exhibit hyperactivation of the RAS-MAPK pathway, accumulation of DNA damage and resistance to apoptosis. To date, this is the first preclinical study evaluating the effect of genetic deletion of E2F-1 in colorectal malignancies driven by PTEN deficiency. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Diagnostic Accuracy of 18F-FDG PET/CT in Patients With Biochemical Evidence of Recurrent, Residual, or Metastatic Medullary Thyroid Carcinoma.

OBJECTIVE: Medullary thyroid carcinoma (MTC) is a rare malignancy. Location of residual, recurrent, or metastatic disease is crucial to treatment management and outcome. We aimed to evaluate the use of F-FDG PET/CT in localizing MTC foci in patients with biochemical relapse. METHODS: This is a retrospective cohort study. Review of 51 FDG PET/CT studies of 45 patients referred to restage MTC due to increased calcitonin (Ctn) and carcinoembryonic antigen (CEA) values at follow-up. FDG PET/CT diagnostic accuracy was determined through a patient-based analysis, using histology as criterion standard when available, or other imaging studies and clinical follow-up otherwise (mean, 4 years). RESULTS: There were 25 positive scans. Sensitivity, specificity, positive and negative predictive values, diagnostic accuracy, and positive likelihood ratio were 66.7%, 83.3%, 88.0%, 57.7%, 72.5%, and 4.0, respectively. Using a Ctn cutoff of 1000 pg/mL, sensitivity increased to 76.9%. There were significant differences of Ctn and CEA values between positive and negative FDG PET/CT (P < 0.05). Regarding true-positive studies, average SUVmax comparing locoregional and metastatic disease was at the limit of significance (P = 0.046). CONCLUSIONS: PET/CT can be useful to restage patients with biochemical relapse of MTC, with a better performance in higher Ctn levels. Its high positive predictive value (88%) may impact in the therapeutic management, although its low negative predictive value (57.7%) makes strict follow-up mandatory in examinations without pathologic findings.

Invasive Sinonasal Lesions: From the Nasal Fossa and Paranasal Sinuses to the Endocranium.

The aim of this work is to review the spectrum of sinonasal lesions that extend to the endocranium and to present key points that may narrow the differential diagnosis. The most frequent sinonasal lesions that extend into the endocranium are malignant; however, benign entities are not unusual. Imaging diagnosis is difficult because malignant lesions and benign entities share similar clinical, epidemiologic, and imaging features. Tumor features in relation to bone, intratumor homogeneity and structure, magnetic resonance imaging signal, along with clinical and epidemiologic aspects may allow an appropriate diagnostic focus with important management implications.

2-phenylethynesulphonamide (PFT-µ) enhances the anticancer effect of the novel hsp90 inhibitor NVP-AUY922 in melanoma, by reducing GSH levels.

Heat shock proteins (HSPs), are molecular chaperones that assist the proper folding of nascent proteins. This study aims to evaluate the antitumour effects of the hsp90 inhibitor NVP-AUY922 in melanoma, both in vitro and in vivo. Our results show that NVP-AUY922 inhibits melanoma cell growth in vitro, with down regulation of multiple signalling pathways involved in melanoma progression such as NF-ĸB and MAPK/ERK. However, NVP-AUY922 was unable to limit tumour growth in vivo. Cotreatment of A375M xenografts with NVP-AUY922 and PFT-µ, a dual inhibitor of both hsp70 and autophagy, induced a synergistic increase of cell death in vitro, and delayed tumour formation in A375M xenografts. PFT-µ depleted cells from the reduced form of glutathione (GSH) and increased oxidative stress. The oxidative stress induced by PFT-µ further enhanced NVP-AUY922-induced cytotoxic effects. These data suggest a potential therapeutic role for NVP-AUY922 used in combination with PFT-µ, in melanoma.

Bioluminescence Imaging to Monitor the Effects of the Hsp90 Inhibitor NVP-AUY922 on NF-κB Pathway in Endometrial Cancer.

PURPOSE: In this study, we first aimed to evaluate the effects in vitro and in vivo, of the Hsp90 inhibitor NVP-AUY922, in endometrial cancer (EC). We also aimed to track nuclear factor kappa B (NF-κB) signalling, a key pathway involved in endometrial carcinogenesis and to check whether NVP-AUY922 treatment modulates it both in vitro and in vivo. PROCEDURES: I n vitro effects of NVP-AUY922 on EC cell growth and the signalling pathways were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), clonogenic assays, Western Blot and luciferase assay. NVP-AUY922 effect on Ishikawa (IK) xenograft growth was evaluated in vivo, and NF-κB activity was monitored using bioluminescence imaging. RESULTS: NVP-AUY922 inhibited the growth of three endometrial cell lines tested in vitro. In vivo, NVP-AUY922 reduced tumour growth of 47 % (p = 0.042) compared to control condition. Moreover, the bioluminescence signal of the tumours harbouring IK NF-κB-LUC cells was significantly reduced in NVP-AUY922-treated animals compared to untreated ones. CONCLUSIONS: NVP-AUY922 reduced EC tumour growth and NF-κB signalling both in vitro and in vivo. As therapeutic resistance of EC remains a challenge for oncologists nowadays, we think that NVP-AUY922 represents a valid alternative to conventional chemotherapy, and we believe that this approach for assessing and tracking the activation of NF-κB pathway may be of therapeutic benefit.

Combinatorial therapy using dovitinib and ICI182.780 (fulvestrant) blocks tumoral activity of endometrial cancer cells.

Mutations in fibroblast growth factor receptor 2 (FGFR2) have been recently described as a molecular-specific feature in endometrial carcinomas and the presence of activated FGFR2 mutations is associated with poor prognosis. For that reason, inhibition of FGFR2 could be a therapeutic target in the treatment of endometriod carcinomas. In this work, we investigated the antitumoral activity of dovitinib (a multiple kinase inhibitor) in human endometrial cancer cell (ECC) lines. We found that dovitinib caused cell growth arrest, loss of clonogenic growth, and cell-cycle arrest in FGFR2-mutated ECCs in in vitro and in vivo experiments. Next, we investigated the mechanistic basis of dovitinib effects. We could determine that dovitinib modified expression levels of well-known key cell-cycle regulatory proteins that induce cellular senescence. To further investigate the role of dovitinib, we analyzed its effect on estrogen receptor α (ER-α) expression. Surprisingly, we discovered that dovitinib enhances ER-α expression in FGFR2-mutant ECCs. Because blocking one signaling pathway is often not sufficient to cause total tumor regression and the effectiveness of individual inhibitors is often short-lived, we examined the impact of targeting FGFR2 with dovitinib in combination with a selective ER antagonist, fulvestrant (ICI182.780). Combination of dovitinib plus ICI182.780 resulted in a significantly higher inhibition of cell growth than dovitinib treatment alone. These findings suggest that combinatory therapies using dovitinib plus ICI182.780 treatment can be truly effective in patients with endometrial carcinomas carrying FGFR2 mutations.

Long-term estradiol exposure is a direct mitogen for insulin/EGF-primed endometrial cells and drives PTEN loss-induced hyperplasic growth.

Loss of tumor-suppressor PTEN is the most common alteration in endometrial carcinoma. However, the relationship between loss of PTEN, growth factors [eg, insulin/insulin-like growth factor (IGF)-1], epidermal growth factor (EGF), and hyperestrogenism in the development of endometrial carcinoma is still controversial. By using three-dimensional (3D) cultures of PTEN(+/+) and PTEN(+/-) endometrial epithelial cells, we investigated the effects of EGF, insulin/IGF, and estradiol in endometrial cell proliferation. We have previously demonstrated that 3D cultures of endometrial cells require EGF and insulin/IGF to proliferate. Herein, we demonstrate that, in the presence of EGF and insulin/IGF, long-term estradiol treatment directly induces proliferation of 3D cultures. Moreover, we show that the mitogenic effects of estradiol require the presence of insulin/IGF and EGF, because withdrawal of such factors completely abolishes estradiol-induced proliferation. In the presence of EGF and insulin/IGF, PTEN(+/-) and PTEN(+/+) spheroids display a similar rate of proliferation. However, the addition of estradiol causes an exaggerated proliferation of PTEN(+/-) cultures, leading to formation of complex structures, such as those observed in endometrial hyperplasia or carcinoma. In summary, we demonstrate that EGF and insulin/IGF prime endometrial epithelial cells to direct the mitogenic effects of estradiol. Furthermore, PTEN deficiency results in enhanced responsiveness to this combination, leading to the development of hyperplasia of endometrial cells in culture.

Nuclear factor-κB2/p100 promotes endometrial carcinoma cell survival under hypoxia in a HIF-1α independent manner.

Endometrial carcinoma (EC) is a common female cancer, treated mainly by surgery and adjuvant radiotherapy. Relapse following treatment is associated with increased risk of metastases. Hypoxia, a common microenvironment in solid tumors, correlates with malignant progression, rendering tumors resistant to ionizing therapy. Hence, we assessed here the immunohistochemical expression of hypoxia-inducible factor-1α (HIF-1α) and members of the NF-κB family in 82 primary EC and 10 post-radiation recurrences of EC. Post-radiation recurrences were highly hypoxic, with a higher expression of HIF-1α and also RelA (p65) and p52 when compared with primary EC. We next investigated the effects of hypoxia on EC cell lines. We found that EC cell lines are highly resistant to hypoxia-induced apoptosis. We thus focused on the molecular mechanisms involved in conferring hypoxic cell death resistance. We show that in addition to the classical NF-κB, hypoxia activates the alternative NF-κB pathway. To characterize the upstream kinases involved in the activation of these pathways, we used lentiviral-mediated knockdown and mouse embryonic fibroblasts lacking IKKα and IKKß kinases. Both IKKα and IKKß kinases are required for RelA (p65) and p100 accumulation, whereas p52 processing under hypoxia is IKKα dependent. Furthermore, Ishikawa endometrial cell line harboring either RelA (p65) or p52 short-hairpin RNA was sensitive to hypoxia-induced cell death, indicating that, in addition to the known prosurvival role of RelA (p65) under hypoxia, alternative NF-κB pathway also enhances hypoxic survival of EC cells. Interestingly, although HIF-1α controlled classical NF-κB activation pathway and survival under hypoxia through RelA (p65) nuclear accumulation, the alternative pathway was HIF-1α independent. These findings have important clinical implications for the improvement of EC prognosis before radiotherapy.

KSR1 is overexpressed in endometrial carcinoma and regulates proliferation and TRAIL-induced apoptosis by modulating FLIP levels.

The Raf/MEK/extracellular signal-regulated kinase (ERK) pathway participates in many processes altered in development and progression of cancer in human beings such as proliferation, transformation, differentiation, and apoptosis. Kinase suppressor of Ras 1 (KSR1) can interact with various kinases of the Raf/MEK/extracellular signal-regulated kinase pathway to enhance its activation. The role of KSR1 in endometrial carcinogenesis was investigated. cDNA and tissue microarrays demonstrated that expression of KSR1 was up-regulated in endometrial carcinoma. Furthermore, inhibition of KSR1 expression by specific small hairpin RNA resulted in reduction of both proliferation and anchorage-independent cell growth properties of endometrial cancer cells. Because inhibition of apoptosis has a pivotal role in endometrial carcinogenesis, the effects of KSR1 in regulation of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis were investigated. KSR1 knock-down sensitized resistant endometrial cell lines to both TRAIL- and Fas-induced apoptosis. Sensitization to TRAIL and agonistic anti-Fas antibody was caused by down-regulation of FLIP (FLICE-inhibitory protein). Also investigated was the molecular mechanism by which KSR1 regulates FLIP protein levels. It was demonstrated that KSR1 small hairpin RNA did not affect FLIP transcription or degradation. Rather, FLIP down-regulation was caused by Fas-associated death domain protein-dependent inhibition of FLIP translation triggered after TRAIL stimulation in KSR1-silenced cells. Re-expression of heterologous KSR1 in cells with down-regulated endogenous KSR1 restored FLIP protein levels and TRAIL resistance. In conclusion, KSR1 regulates endometrial sensitivity to TRAIL by regulating FLIP levels.

Promoter hypermethylation and expression of sprouty 2 in endometrial carcinoma.

Sprouty 2 is a key antagonist regulator of receptor tyrosine kinases, and downstream signaling pathways, like fibroblastic growth factor (FGF) and Ras-mitogen-activated protein kinase (RAS-MAPK). By controlling these pathways, sprouty 2 is involved in regulation of cell proliferation, differentiation, and angiogenesis. Alterations in fibroblastic growth factor receptor (FGFR) and members of the RAS-MAPK pathway are frequent in endometrial carcinoma. The expression of sprouty 2 has been found to be decreased in several types of human cancer, by mechanisms of promoter methylation. In the present study, we have assessed the expression of sprouty 2 in endometrial carcinoma, in correlation with sprouty 2 promoter methylation. Sprouty 2 immunohistochemical expression was assessed using 3 different tissue microarrays: one constructed from paraffin blocks of 80 samples of normal endometrium and 2 tissue microarrays containing samples of 157 endometrial carcinoma (1 tissue microarray constructed with 95 endometrial carcinomas previously studied for microsatellite instability and alterations in phosphatase and tensin homolog (PTEN), k-ras, and b-catenin, and 1 tissue microarray containing 62 endometrial carcinoma, which were also subjected to sprouty 2 promoter methylation analysis). The immunohistochemical expression of sprouty 2 was correlated with cellular proliferation (Ki67) and clinicopathologic data. Sprouty 2 promoter methylation was assessed by methylation-specific polymerase chain reaction, with DNA obtained from fresh-frozen samples of endometrial carcinoma and corresponding normal tissues, and correlated with promoter methylation of RAS association domain family-1A (RASSF1A). A highly significant decrease in sprouty 2 immunoexpression was seen in the proliferative phase of normal endometrium (P < .001). Differences were detected between types I and II endometrial carcinoma, but they were not statistically significant. Reduced immunoexpression of sprouty 2 was seen in 19.85% of endometrial carcinoma and was strongly and inversely associated with increased cell proliferation (Ki67; r = -0.367; P = .001). Sprouty 2 promoter methylation was detected in 31 (53.4%) of 58 endometrial carcinomas. Results from our study show that alterations in sprouty 2 may be involved in endometrial carcinogenesis by controlling cell proliferation.

A novel three-dimensional culture system of polarized epithelial cells to study endometrial carcinogenesis.

Development of three-dimensional (3D) cultures that mimic in vivo tissue organization has a pivotal role in the investigation of the involvement of cell adhesion and polarity genes in the pathogenesis of epithelial cancers. Here we describe a novel 3D culture model with primary mouse endometrial epithelial cells. In this model, isolated endometrial epithelial cells develop single-lumened, polarized glandular structures resembling those observed in endometrial tissue. Our in vitro 3D culture model of endometrial glands requires the use of serum-free defined medium with only epidermal growth factor and insulin as growth supplements and 3% Matrigel as reconstituted extracellular matrix. Under these culture conditions, glands of epithelial cells displaying typical apicobasal polarity and proper positioning of tight and adherent junctions are formed by hollowing as early as 7 to 8 days in culture. Addition of the phosphatidylinositol 3-kinase inhibitor LY294002 completely inhibits bromodeoxyuridine incorporation and cyclinD1 expression, confirming that in vitro growth of endometrial glands depends on phosphatidylinositol 3-kinase/Akt signaling. To prove that our culture method is a good model to study endometrial carcinogenesis, we knocked down E-cadherin or phosphatase and tensin homolog expression by lentivirus-delivered short hairpin RNAs. Down-regulation of E-cadherin resulted in complete loss of epithelial cell polarity and glandular formation, whereas phosphatase and tensin homolog down-regulation resulted in increased proliferation of glandular epithelial cells. These properties indicate that our 3D culture model is suitable to study the effect of growth factors, drugs, and gene alterations in endometrial carcinogenesis and to study normal endometrial biology/physiology.