Working together to shape the endometrial cancer research agenda: The top ten unanswered research questions.

BACKGROUND: Endometrial cancer (EC) is the most common gynaecological cancer in developed nations and its incidence is rising. As a direct consequence, more women are dying from EC despite advances in care and improved survivorship. There is a lack of research activity and funding, as well as public awareness about EC. We sought to engage patients, carers and healthcare professionals to identify the most important unanswered research questions in EC. METHODOLOGY: The priority setting methodology was developed by the James Lind Alliance and involved four key stages: gathering research questions; checking these against existing evidence; interim prioritisation; and a final consensus meeting during which the top ten unanswered research questions were agreed using modified nominal group methodology. RESULTS: Our first online survey yielded 786 individual submissions from 413 respondents, of whom 211 were EC survivors or carers, and from which 202 unique unanswered research questions were generated. 253 individuals, including 108 EC survivors and carers, completed an online interim prioritisation survey. The resulting top 30 questions were ranked in a final consensus meeting. Our top ten spanned the breadth of patient experience of this disease and included developing personalised risk scoring, refining criteria for specialist referral, understanding the underlying biology of different types of EC, developing novel personalised treatment and prevention strategies, prognostic and predictive biomarkers, increasing public awareness and interventions for psychological issues. CONCLUSION: Having established the top ten unanswered research questions in EC, we hope this galvanises researchers, healthcare professionals and the public to collaborate, coordinate and invest in research to improve the lives of women affected by EC.

Clinical and immunologic results of a phase II trial of sequential imiquimod and photodynamic therapy for vulval intraepithelial neoplasia.

PURPOSE: High-risk human papillomavirus (HPV)-associated vulval intraepithelial neoplasia (VIN) is difficult to treat by excision or ablation because of high recurrence rates. Small studies of photodynamic therapy (PDT) and imiquimod treatments have shown some success and function at least in part through stimulation of local immune responses. Indeed, there is evidence that immunosuppressed individuals have higher rates of VIN, suggesting immune control is relevant. EXPERIMENTAL DESIGN: In the study, 20 women with high-grade VIN were treated with topical imiquimod and the PDT sequentially. Vulval biopsy and blood were taken pretreatment and, after imiquimod and PDT, with follow up for 1 year. Clinical response was assessed by measuring lesion size. Biopsies were analyzed for HPV DNA and tumor-infiltrating lymphocytes including T regulatory cells. RESULTS: The treatment was well-tolerated. There was an overall response rate of 55% by intention treat and 64% per protocol. The 52-week symptom response was 65% asymptomatic, compared with 5% at baseline. The nonresponders showed a significantly higher level of T regulatory cells in the lesions after imiquimod treatment. CONCLUSIONS: The response rates are clinically relevant, and the treatment regimen was feasible for the majority. Initial nonresponders to imiquimod seem to be relatively refractory, and this may derive from their unfavorable local immune environment, in particular, the increased proportions of T regulatory cells, possibly the limiting action and/or development of any HPV T-cell immunity. The potential benefit of this treatment is its ability to treat multifocal disease.

Immunological and clinical responses in women with vulval intraepithelial neoplasia vaccinated with a vaccinia virus encoding human papillomavirus 16/18 oncoproteins.

This study assessed the immunological and clinical responses of women with human papillomavirus (HPV) 16-associated high-grade vulval intraepithelial neoplasia (VIN) vaccinated with TA-HPV, a recombinant vaccinia virus encoding modified HPV 16 and 18 E6 and E7. Eighteen women with HPV 16-positive high-grade VIN were vaccinated with TA-HPV. The extent of their baseline disease was compared after 24 weeks by lesion measurements and histological analysis. Viral load was assessed pre- and postvaccination by real time PCR. Cell-mediated immunity to HPV 16 E6 and/or E7 peptides (HLA-A2 epitopes) or vaccinia-infected cell lysates was determined by IFN-gamma enzyme-linked immunospot (ELISPOT) and T cell proliferation using an HPV 16 L2E6E7 fusion protein. Antibodies were measured by ELISA using vaccinia-infected cell lysates or HPV 16 and 18 E6 and E7 glutathione S-transferase-fusion proteins. Lesion-infiltrating CD4(+), CD8(+), CD1a(+), and CD68(+) immune cells were assessed by immunohistochemistry. The single vaccination with TA-HPV was well tolerated, and all patients showed an increased ELISPOT and/or antibody response to vaccinia. There were significant differences in HPV-16 E7-specific ELISPOT and L2E6E7 proliferative responses in the patients at one or more time points postvaccination as compared with the prevaccination status; two patients showed transient increased antibody responses. Overall, 13 women showed an increased HPV 16-specific immune response by one or more methodologies after immunization. Eight patients demonstrated a reduction in lesion diameter of at least 50% and a further four patients showed significant symptom relief. Viral load was reduced or cleared in six of eight lesion responders but also in six of ten nonresponders. Before vaccination, clinical responders had significantly higher levels of lesion-associated CD4(+), CD8(+), and CD1a(+)-immune cells than nonresponders. There were no differences in CD68 (macrophages) between responders and nonresponders before or after vaccination. Nonresponders did show a significant increase in CD4(+)- and CD8(+)- but not CD1a(+)-immune cells postvaccination but at lower levels overall than responder patients. Local immune infiltration may be a critical factor in potential responsiveness to vaccine therapy in HPV-associated neoplasia and should be carefully monitored in future placebo-controlled trials of immunotherapy for VIN.

p38 mitogen-activated protein kinase is required for TGFbeta-mediated fibroblastic transdifferentiation and cell migration.

Transforming growth factor beta (TGFbeta) contributes to tumor progression by inducing an epithelial to mesenchymal transdifferentiation (EMT) and cell migration. We found that TGFbeta-induced EMT was blocked by inhibiting activation of p38 mitogen-activated protein kinase (MAPK) with H-7, a protein kinase C inhibitor, and with SB202190, a direct inhibitor of p38MAPK. Inhibition of the p38MAPK pathway affected TGFbeta-mediated phosphorylation of ATF2, but did not inhibit phosphorylation of Smad2. SB202190 impaired TGFbeta-mediated changes in cell shape and reorganization of the actin cytoskeleton. Forced expression of dominant-negative (DN) MAPK kinase 3 (MKK3) inhibited TGFbeta-mediated activation of p38MAPK and EMT. Expression of DN-p38alpha impaired TGFbeta-induced EMT. Inhibition of p38MAPK blocked TGFbeta-induced migration of non-tumor and tumor mammary epithelial cells. TGFbeta induced activation of the p38MAPK pathway within 15 minutes. Expression of TGFbeta type II (TbetaRII) and type I (TbetaRI/Alk5) kinase-inactive receptors blocked EMT and activation of p38MAPK, whereas expression of constitutively active Alk5-T204D resulted in EMT and phosphorylation of MKK3/6 and p38MAPK. Finally, dominant-negative Rac1N17 blocked TGFbeta-induced activation of the p38MAPK pathway and EMT, suggesting that Rac1 mediates activation of the p38MAPK pathway. These studies suggest that the p38MAPK pathway is required for TGFbeta-mediated EMT and cell migration.