Corinne M. Doll

PIK3CA mutational status and overall survival in patients with cervical cancer treated with radical chemoradiotherapy

OBJECTIVE Mutational activation of PIK3CA is associated with poor prognosis in patients with solid tumors, and may predict favorable response to PI3K/AKT/mTOR pathway inhibitors. However, PIK3CA mutational status has not previously been evaluated in patients with cervical carcinoma treated with radical chemoradiotherapy (CRT). The aims of this study were (1) to evaluate the frequency of PIK3CA mutations in patients with cervical cancer treated with radical CRT and (2) to examine the effect of tumor PIK3CA mutational status in pre-treatment biopsies on overall survival (OS) and progression-free survival (PFS). METHODS Patients with cervical cancer, treated at a single institution with radical CRT, from 1999 to 2008, were eligible for this retrospective study. Pre-treatment tumor biopsies (n=157) were retrieved. Genomic DNA was extracted from tumor blocks, and exons 9 and 20 of the PIK3CA gene were sequenced for mutations. RESULTS Eighty-two tumors were sequenced for both exon 9 and exon 20. 19/82 (23%) tumors were PIK3CA mutation positive; of these 84% were squamous cell carcinomas. 79% of mutations were in exon 9. PIK3CA mutation status was strongly associated with overall survival (OS) in FIGO stage IB/II patients, unadjusted HR 6.0 (95% CI 2.1-17.5), p=0.0002, but not stage III/IVA patients, unadjusted HR 1.0 (95% CI 0.32-3.1), p=0.98. CONCLUSIONS In cervical cancer patients treated with CRT, tumor PIK3CA mutation status was associated with overall survival in FIGO stage IB/II cervix cancers. Further evaluation with a larger dataset will be required to validate these findings to inform potential clinical trials designs involving PI3K/AKT/mTOR pathway inhibitors.

Image-guided brachytherapy for cervical cancer: a Canadian Brachytherapy Group survey.

PURPOSE To survey the current use and future plans for image-guided brachytherapy (BT) for cervical cancer by radiation oncologists in Canada. METHODS AND MATERIALS Canadian radiation oncologists treating gynecologic malignancies were identified in January 2009. A 29-item questionnaire (English and French) querying the current practice in the use of imaging in BT planning, and plans for transition to three-dimensional (3D) image guidance for BT for cervical cancer (curative intent, intact cervix), was electronically circulated. Questionnaire responses were tabulated and analyzed by respondent and by center. RESULTS Response rate was 62% (36 of 58 radiation oncologists), representing 71% (22 of 31) of Canadian radiation oncology centers with a gynecologic BT facility. Most of the centers were using high-dose-rate BT (68%), followed by low-dose-rate BT (23%) and pulsed dose-rate BT (10%). Main imaging used for treatment planning by center was plain X-ray (50%), computerized tomography (CT) (45%), and magnetic resonance imaging (MRI) (5%). For respondents using CT or MRI for planning, point A was the most common dose prescription point (50%), followed by gross tumor volume/clinical target volume as per Groupe Européen de Curiethérapie and the European Society for Therapeutic Radiology and Oncology guidelines (44%). For centers using plain X-rays for planning, 73% planned to transition to a 3D image-based approach, with the majority to adopt CT imaging. Eighty percent of respondents agreed that 3D image-guided BT should become standard of care for treatment of cervical cancer in Canada, and additionally support the development of national guidelines. CONCLUSIONS Most of the Canadian radiation oncologists surveyed and Canadian cancer centers are either using 3D imaging and planning or transitioning to a 3D image-based approach within the next year. Point A remained a commonly documented prescription point. Access to MRI was very low. These results may lead to national treatment guidelines.

Low ERCC1 mRNA and protein expression are associated with worse survival in cervical cancer patients treated with radiation alone

PURPOSE To evaluate the association of excision repair cross-complementation group 1 (ERCC1) expression, using both mRNA and protein expression analysis, with clinical outcome in cervical cancer patients treated with radical radiation therapy (RT). EXPERIMENTAL DESIGN Patients (n=186) with locally advanced cervical cancer, treated with radical RT alone from a single institution were evaluated. Pre-treatment FFPE biopsy specimens were retrieved from 112 patients. ERCC1 mRNA level was determined by real-time PCR, and ERCC1 protein expression (FL297, 8F1) was measured using quantitative immunohistochemistry (AQUA®). The association of ERCC1 status with local response, 10-year disease-free (DFS) and overall survival (OS) was analyzed. RESULTS ERCC1 protein expression levels using both FL297 and 8F1 antibodies were determined for 112 patients; mRNA analysis was additionally performed in 32 patients. Clinical and outcome factors were comparable between the training and validation sets. Low ERCC1 mRNA expression status was associated with worse OS (17.9% vs 50.1%, p=0.046). ERCC1 protein expression using the FL297 antibody, but not the 8F1 antibody, was significantly associated with both OS (p=0.002) and DFS (p=0.010). After adjusting for pre-treatment hemoglobin in a multivariate analysis, ERCC1 FL297 expression status remained statistically significant for OS [HR 1.9 (1.1-3.3), p=0.031]. CONCLUSIONS Pre-treatment tumoral ERCC1 mRNA and protein expression, using the FL297 antibody, are predictive factors for survival in cervical cancer patients treated with RT, with ERCC1 FL297 expression independently associated with survival. These results identify a subset of patients who may derive the greatest benefit from the addition of cisplatin chemotherapy.

Brachytherapy for carcinoma of the cervix: A Canadian survey of practice patterns in a changing era

BACKGROUND AND PURPOSE This survey aimed to document practices of Canadian radiation oncologists performing gynecologic brachytherapy for carcinoma of the cervix and to determine what the effect of the phasing-out of LDR after-loading systems from the commercial market is having on practice. MATERIALS AND METHODS A 26-item questionnaire was developed to survey various aspects of brachytherapy practice to include: number of patients treated, prescription points/volume, dose and fractionation, timing, critical structure delineation, expected changes due to the phasing-out of support for low dose rate systems, and support for the development of national guidelines. A link to a web-based survey collection instrument was emailed to each radiation oncologist in Canada practicing gynecologic brachytherapy. RESULTS A 67% response rate was achieved in this web-based survey. Radiation oncologists currently using HDR brachytherapy are most commonly delivering 5 fractions of 6 Gy in addition to an EBRT dose of 45 Gy in 25 fractions. The median total dose equivalents to Point A was 82.9 Gy for both early and advanced disease. In response to the announcement by a major vendor that they would be phasing-out service for a popular LDR after-loader, 49% of Canadian radiation oncologists who practice brachytherapy for cervix cancer are changing to an HDR technique with a further 9% changing to a PDR technique. Eighty-six percent of respondents would support the development of national guidelines for cervix brachytherapy in Canada. CONCLUSIONS Variation in practice exists in Canada in brachytherapy for cervix cancer. Many centers are in the process of phasing-out LDR techniques in response to the withdrawal of commercial support for these systems. Support for the development of Canadian national guidelines is high.

High stromal carbonic anhydrase IX expression is associated with decreased survival in P16-negative head-and-neck tumors.

PURPOSE Head-and-neck squamous cell carcinoma (HNSCC) is the fifth most common malignancy worldwide. Alcohol use and tobacco use are the most established risk factors; however, human papilloma virus (HPV) infection is a major risk factor for a subset of HNSCCs. Although HPV-positive tumors typically present at a more advanced stage at diagnosis, they are associated with a better prognosis. Tumor hypoxia confers poor prognosis and treatment failure, but direct tumor oxygen measurement is challenging. Endogenous markers of hypoxia (EMHs) have been proposed but have not replicated the prognostic utility of direct oxygen measurement. The expression of endogenous markers of hypoxia may be influenced by oxygen-independent factors, such as the HPV status of the tumor. METHODS AND MATERIALS Consecutive cases of locally advanced HNSCC, treated with a uniform regimen of combined radiotherapy and chemotherapy, were identified. Tissue microarrays were assembled from triplicate 0.6-mm cores of archived tumor tissue. HPV status was inferred from semiquantitative p16 immunostaining and directly measured by use of HPV-specific chromogenic in situ hybridization and polymerase chain reaction. Automated quantitative fluorescent immunohistochemistry was conducted to measure epithelial and stromal expression of carbonic anhydrase IX (CAIX) and glucose transporter 1 (GLUT1). RESULTS High stromal CAIX expression was associated with significantly reduced overall survival (p = 0.03) in patients with p16-negative tumors. CONCLUSIONS This is the first study to use quantitative immunohistochemistry to examine endogenous markers of hypoxia stratified by tumor p16/HPV status. Assessment of CAIX expression in p16-negative HNSCC could identify patients with the least favorable prognosis and inform therapeutic strategies.

The prognostic significance of pretreatment leukocytosis in patients with anal cancer treated with radical chemoradiotherapy or radiotherapy.

BACKGROUND: There are emerging data showing the prognostic significance of pretreatment leukocytosis in patients with cervical cancer; it is generally associated with adverse outcome. However, the prognostic impact of leukocytosis in patients with anal cancer has not been previously reported. OBJECTIVE: The purpose of this study was to assess the relationship between pretreatment leukocytosis and clinical outcomes in patients with anal cancer treated with radical chemoradiotherapy or radiotherapy. DESIGN: This is a retrospective cohort study. SETTING AND PATIENTS: One hundred twenty-six patients with invasive anal canal cancer, treated with radical chemoradiotherapy or radiotherapy between 2000 and 2008 at 2 major tertiary cancer centers, were evaluated. MAIN OUTCOME MEASURES: The primary outcomes were disease-free and overall survival. RESULTS: Median follow-up was 24 months. Pretreatment leukocytosis (white blood cell count >10 × 109/L) was identified in 15.9% (20/126) of patients. After adjusting for sex, tumor size, and stage in a multivariate analysis, leukocytosis remained significantly associated with worse disease-free survival (HR, 2.2; 95% CI, 1.1–4.8; p = 0.045) and worse overall survival (HR, 2.9; 95% CI, 1.1–7.9; p = 0.036). Patients with both leukocytosis and anemia (pretreatment hemoglobin <125 g/L) had the worst prognosis: 2-year disease-free survival 42.1% versus 72.9% for patients without these factors (HR, 2.7; 95% CI, 1.1–6.8; p = 0.033); 2-year overall survival 60.9% versus 89.8% (HR, 4.5; 95% CI, 1.5–13.2; p = 0.006). LIMITATIONS: The study was limited by its retrospective nature and lack of patients with multiple hematologic abnormalities (ie, both anemia and leukocytosis). HIV status was unable to be evaluated. CONCLUSIONS: Pretreatment leukocytosis in patients with anal cancer is associated with significantly worse disease-free and overall survival, which appears to be exacerbated with the presence of pretreatment anemia.

Predictors of Pathologic Complete Response After Neoadjuvant Treatment for Rectal Cancer: A Multicenter Study

BACKGROUND Pathologic complete response (pCR) to neoadjuvant chemoradiation (CRT) for rectal cancer is associated with better long-term outcomes, and is used as an early indicator of response to novel agents. To assess the rate and predictors of pCR, we performed a retrospective multicenter study involving 5 Canadian cancer centers. PATIENTS AND METHODS Cancer registries identified consecutive patients with locally advanced rectal adenocarcinoma from the Tom Baker Cancer Centre, Cross Cancer Institute, British Columbia Cancer Agency, Ottawa Hospital Cancer Centre, and the Dr H. Bliss Murphy Cancer Centre who received fluoropyrimidine-based CRT and had curative intent surgery from 2005 to 2012. Patient, tumor, and therapy characteristics were correlated with response. RESULTS Of the 891 patients included, 885 patients had pCR data available. Of the included patients, 161 (18.2%) had a pCR to CRT, and 724 (81.8%) did not. Patients with a pCR had a lower pretreatment carcinoembryonic antigen (CEA) level, and higher hemoglobin level in univariate analysis. In multivariable analysis, statin use at baseline (odds ratio [OR], 1.7; 95% confidence interval [CI], 1.02-2.92; P = .04), lower pretreatment CEA level (OR, 1.03; 95% CI, 1.01-1.06; P = .03), and distance closer to anal verge (OR, 1.07; 95% CI, 1.01-1.15; P = .04) were significant predictors of pCR. The 3-year disease-free survival was 86% in those with a pCR versus 62.5% in those without a pCR (P < .0001) and pCR was associated with improved overall survival (hazard ratio, 0.29; 95% CI, 0.17-0.51; P < .0001). CONCLUSION Lower pretreatment CEA level, proximity to anal verge, and statin use are predictors of pCR in our large retrospective cohort. Clinical trials to investigate statins combined with neoadjuvant CRT might be warranted.

The Prognostic Value of HPV Status and p16 Expression in Patients with Carcinoma of the Anal Canal

Background In anal cancer studies, the detection frequency of high-risk HPV (human papillomavirus) is variable, depending on the method used. There are limited data reporting results of different HPV detection techniques in the same clinical series, and very few correlating results with clinical outcome. Objectives To evaluate tumor expression of p16/HPV16 using three different methods, and to determine their association with clinical outcome in patients with anal canal squamous cell carcinomas (SCC). Design This retrospective study included patients with anal canal SCC treated with definitive radiotherapy or chemoradiotherapy at a single institution between 1992 and 2005. Formalin-fixed paraffin–embedded tumor samples from 53 of the 89 (60%) patient pre-treatment biopsies were adequate for tissue microarray construction. HPV status was determined using: p16 expression by conventional immunohistochemistry (IHC) and quantitative IHC (AQUA), HPV genotype analysis by chromogenic in situ hybridization (CISH) and HPV linear array sub-typing. Expression status was correlated with clinical outcome. Results 80% (28/35) of patient tumors had high p16 expression using conventional IHC. HPV16 CISH was positive in 81% (34/42) of tumors, and 78% (28/36) of tumors were HPV subtype 16. HPV16 CISH correlated with p16 evaluated by conventional IHC (correlation coefficient 0.46; p = 0.01) and by p16 AQUA score (correlation coefficient 0.49; p = 0.001). A subset of cases (15%) had very high p16 quantitative IHC scores (>244) and were associated with a higher incidence of local or distant recurrence (p = 0.04). Conclusions The vast majority (80%) of anal canal SCC in our series were positive for HPV16/p16, regardless of the testing method used. The exploratory analysis of automated quantitative IHC scoring was the only technique to define a subset of patients with a worse prognosis by p16 expression status on univariate analysis. Further exploration of the molecular mechanisms of treatment resistance in association with very high p16 expression is warranted.

The prognostic value of hemoglobin in patients with anal cancer treated with chemoradiotherapy.

PURPOSE: This study aimed to evaluate the impact of hemoglobin level on clinical outcome (local response, progression-free survival, and overall survival) in patients with carcinoma of the anal canal treated with definitive chemoradiotherapy. METHODS: This is a retrospective study of patients with anal cancer treated between 1992 and 2005 with definitive chemoradiotherapy at Tom Baker Cancer Centre. Patient treatment, laboratory, and outcome data were extracted from the chart. RESULTS: Seventy-two patients treated with definitive chemoradiotherapy were identified. The median age was 56 years, the male-to-female ratio was 1:2, and the median tumor size was 3.5 cm. At 6 weeks after the completion of chemoradiotherapy, 62% of patients (38/61) had complete clinical response, and 34% (21/61) had achieved a partial clinical response. At 3 months after treatment, complete clinical response was observed in 78% (49/63) and a partial response in 16% (10/63). The median pretreatment hemoglobin level was 138.5 g/L, and the median on-treatment hemoglobin level was 129 g/L. Distant relapse was associated with hemoglobin levels in the lowest quartiles, pretreatment and on-treatment (P = .007 and P = .008, respectively). Hemoglobin levels were not associated with response at 6 weeks or 3 months. A pretreatment hemoglobin level of <130 g/L was associated with worse progression-free and overall survival (P < .0001, both). A hemoglobin on-treatment level of <121 g/L was associated with progression-free and overall survival (P < .0001 and P = .019, respectively), when stratified by gender. CONCLUSIONS: Hemoglobin status was correlated with progression-free and overall survival, and distant relapse, but not clinical response, in patients with carcinoma of the anal canal treated with chemoradiotherapy. The clinically relevant cut point, and the value of correcting hemoglobin levels before or during treatment, remains to be elucidated.

PIK3CA missense mutation is associated with unfavorable outcome in grade 3 endometrioid carcinoma but not in serous endometrial carcinoma.

OBJECTIVE To evaluate the outcome association of PIK3CA mutational status within histological types of rigorously classified high-grade endometrial carcinomas. METHODS We assessed PIK3CA mutational status in exon 9 and exon 20 hot spots by Sanger sequencing of DNA derived from formalin fixed paraffin embedded tissue of 57 grade 3 endometrioid, 26 serous, 11 clear cell and 5 dedifferentiated carcinomas. We correlated PIK3CA mutation status with clinicopathological and other molecular parameters. Univariate and multivariate disease specific survival analysis was performed using Kaplan-Meier and Cox regression analyses. RESULTS PIK3CA exon 9 or exon 20 missense mutations were identified in 20 of 99 (20%) high-grade endometrial carcinomas without significant difference across histological types (p=0.22). Presence of PIK3CA exon 9 or exon 20 missense mutations was associated with shorter disease specific survival within grade 3 endometrioid (p=0.0029) but not endometrial serous (p=0.57) carcinoma based on univariate analysis. Within grade 3 endometrioid carcinoma, PIK3CA exon 9 or exon 20 missense mutations were more commonly observed in cases that were deficient for mismatch repair protein expression (p=0.0058) and showed loss of ARID1A expression (p=0.037). CONCLUSIONS PIK3CA exon 9 or exon 20 missense mutations are present across all histological types of high-grade endometrial carcinomas but a significant outcome association is only seen in grade 3 endometrioid carcinoma, suggesting a greater biological importance in this tumor type.