Kristina Schee

Clinical relevance of microRNA miR-21, miR-31, miR-92a, miR-101, miR-106a and miR-145 in colorectal cancer

BackgroundMicroRNAs (miRNAs) regulate gene expression by binding to mRNA, and can function as oncogenes or tumor suppressors depending on the target. In this study, using qRT-PCR, we examined the expression of six miRNAs (miR-21, miR-31, miR-92a, miR-101, miR-106a and miR-145) in tumors from 193 prospectively recruited patients with colorectal cancer, and associations with clinicopathological parameters and patient outcome were analyzed. The miRNAs were chosen based on previous studies for their biomarker potential and suggested biological relevance in colorectal cancer.MethodsThe miRNA expression was examined by qRT-PCR. Associations between miRNA expression and clinicopathological variables were explored using Mann–Whitney U and Kruskal-Wallis test while survival was estimated using the Kaplan-Meier method and compared using the log-rank test.ResultsMiR-101 was hardly expressed in the tumor samples, while for the other miRNAs, variable expression levels and expression ranges were observed, with miR-21 being most abundantly expressed relative to the reference (RNU44). In our study cohort, major clinical significance was demonstrated only for miR-31, as high expression was associated with advanced tumor stage and poor differentiation. No significant associations were found between expression of the investigated miRNAs and metastasis-free or overall survival.ConclusionsInvestigating the expression of six miRNAs previously identified as candidate biomarkers in colorectal cancer, few clinically relevant associations were detected in our patient cohort. Our results emphasize the importance of validating potential tumor markers in independent patient cohorts, and indicate that the role of miRNAs as colorectal cancer biomarkers is still undetermined.

Deep Sequencing the MicroRNA Transcriptome in Colorectal Cancer

Colorectal cancer (CRC) is one of the leading causes of cancer related deaths and the search for prognostic biomarkers that might improve treatment decisions is warranted. MicroRNAs (miRNAs) are short non-coding RNA molecules involved in regulating gene expression and have been proposed as possible biomarkers in CRC. In order to characterize the miRNA transcriptome, a large cohort including 88 CRC tumors with long-term follow-up was deep sequenced. 523 mature miRNAs were expressed in our cohort, and they exhibited largely uniform expression patterns across tumor samples. Few associations were found between clinical parameters and miRNA expression, among them, low expression of miR-592 and high expression of miR-10b-5p and miR-615-3p were associated with tumors located in the right colon relative to the left colon and rectum. High expression of miR-615-3p was also associated with poorly differentiated tumors. No prognostic biomarker candidates for overall and metastasis-free survival were identified by applying the LASSO method in a Cox proportional hazards model or univariate Cox. Examination of the five most abundantly expressed miRNAs in the cohort (miR-10a-5p, miR-21-5p, miR-22-3p, miR-143-3p and miR-192-5p) revealed that their collective expression represented 54% of the detected miRNA sequences. Pathway analysis of the target genes regulated by the five most highly expressed miRNAs uncovered a significant number of genes involved in the CRC pathway, including APC, TGFβ and PI3K, thus suggesting that these miRNAs are relevant in CRC.

MicroRNAs as biomarkers in colorectal cancer.

Colorectal cancer is a leading cause of cancer-related morbidity and mortality in the Western world. While improved diagnostic surveillance and treatment strategies involving surgery, chemo-, and radiotherapy have all contributed to earlier detection and improved survival, treatment decisions are still made almost exclusively based on the cancers clinicopathological stage at diagnosis. Therefore, the search for new biomarkers to facilitate early diagnosis and individualized treatment is particularly warranted. MicroRNAs (miRNAs) are short, noncoding RNAs that regulate gene expression through posttranscriptional interactions with mRNA, thereby potentially leading to a vast range of downstream effects that depend on the target proteins affected. The discovery that miRNAs may act as either oncogenes or tumor suppressors has initiated extensive research in the cancer field, leading to the identification of numerous miRNAs implicated in carcinogenesis and tumor progression. MiRNAs are chemically stable and can thus be detected in a broad range of clinical samples, making these molecules particularly attractive as potential biomarkers in cancer. While the knowledge of miRNA involvement in colorectal cancer biology is less extensive than for other cancer types and several targets with potential biological and clinical relevance have been identified, a significant amount of research is still needed. In this review, we explore the literature regarding the relevance of miRNAs in colorectal cancer, focusing in particular on miRNAs as potential diagnostic, prognostic, and predictive biomarkers.

Clinical Significance of Long Intergenic Noncoding RNA-p21 in Colorectal Cancer

BACKGROUND Long intergenic noncoding RNAs (lincRNAs) have been shown to be novel regulators for both transcription and posttranscriptional/translation. One of them, lincRNA-p21, was regulated by p53 and contributed to apoptosis in mouse embryonic fibroblasts. However, the impact of such regulation on colorectal cancer (CRC) remains to be determined. METHODS Total RNA was extracted from CRC cell lines and snap fresh frozen CRC samples from 2 CRC patient cohorts. The expression of lincRNA-p21 was quantified by quantitative real-time polymerase chain reaction analysis. RESULTS We discovered that the expression level of lincRNA-p21 was increased by elevated wild-type p53 induced by nutlin-3 in HCT-116 colon cancer cells. The expression level of lincRNA-p21 was significantly (P = .0208) lower in CRC tumor tissue when compared with the paired normal tissue from the same patient. There was no significant correlation of lincRNA-p21 with p53 status (wild-type vs. mutant). Tumors in the rectum showed a higher level of lincRNA-p21 than tumors in the colon (P = .00005). In addition, lincRNA-p21 in patients with stage III tumors was significantly higher than in those with stage I tumors (P = .007). Elevated levels of lincRNA-p21 were significantly associated with higher pT (P = .037 between pT 2 and 3) and vascular invasion (P = .017). CONCLUSIONS These results indicate that lincRNA-p21 may contribute to CRC disease progression.

Self-Sampling for Human Papillomavirus Testing among Non-Attenders Increases Attendance to the Norwegian Cervical Cancer Screening Programme

Increasing attendance to screening offers the best potential for improving the effectiveness of well-established cervical cancer screening programs. Self-sampling at home for human papillomavirus (HPV) testing as an alternative to a clinical sampling can be a useful policy to increase attendance. To determine whether self-sampling improves screening attendance for women who do not regularly attend the Norwegian Cervical Cancer Screening Programme (NCCSP), 800 women aged 25–69 years in the Oslo area who were due to receive a 2nd reminder to attend regular screening were randomly selected and invited to be part of the intervention group. Women in this group received one of two self-sampling devices, Evalyn Brush or Delphi Screener. To attend screening, women in the intervention group had the option of using the self-sampling device (self-sampling subgroup) or visiting their physician for a cervical smear. Self-sampled specimens were split and analyzed for the presence of high-risk (hr) HPV by the CLART® HPV2 test and the digene® Hybrid Capture (HC)2 test. The control group consisted of 2593 women who received a 2nd reminder letter according to the current guidelines of the NCCSP. The attendance rates were 33.4% in the intervention group and 23.2% in the control group, with similar attendance rates for both self-sampling devices. Women in the self-sampling subgroup responded favorably to both self-sampling devices and cited not remembering receiving a call for screening as the most dominant reason for previous non-attendance. Thirty-two of 34 (94.1%) hrHPV-positive women in the self-sampling subgroup attended follow-up. In conclusion, self-sampling increased attendance rates and was feasible and well received. This study lends further support to the proposal that self-sampling may be a valuable alternative for increasing cervical cancer screening coverage in Norway.

Cervical cancer prevented by screening: Long-term incidence trends by morphology in Norway.

Both major morphologic types of cervical cancer, squamous cell carcinoma (SCC) and adenocarcinoma (AC), are causally related to persistent infection with high‐risk human papillomavirus (hrHPV), but screening has primarily been effective at preventing SCC. We analysed incidence trends of cervical cancer in Norway stratified by morphologies over 55 years, and projected SCC incidence in the absence of screening by assessing the changes in the incidence rate of AC. The Cancer Registry of Norway was used to identify all 19,530 malignancies in the cervix diagnosed in the period 1956–2010. The majority of these (82.9%) were classified as SCCs, 10.5% as ACs and the remaining 6.6% were of other or undefined morphology. By joint‐point analyses of a period of more than five decades, the average annual percentage change in the age‐standardised incidence was −1.0 (95%CI: −2.1–0.1) for cervical SCC, 1.5 (95%CI:1.1–1.9) for cervical AC and −0.9 (95%CI: −1.4 to −0.3) for cervical cancers of other or undefined morphology. The projected age‐standardised incidence rate of cervical SCC in Norway, assuming no screening, was 28.6 per 100,000 woman‐years in 2010, which compared with the observed SCC rate of 7.3 corresponds to an estimated 74% reduction in SCC or a 68% reduction due to screening in the total cervical cancer burden. Cytology screening has impacted cervical cancer burden more than suggested by the overall observed cervical cancer incidence reduction since its peak in the mid‐1970s. The simultaneous substantial increase in cervical adenocarcinoma in Norway is presumably indicative of an increase in exposure to HPV over time.

Cervical cancer prevented by screening

Both major morphologic types of cervical cancer, squamous cell carcinoma (SCC) and adenocarcinoma (AC), are causally related to persistent infection with high‐risk human papillomavirus (hrHPV), but screening has primarily been effective at preventing SCC. We analysed incidence trends of cervical cancer in Norway stratified by morphologies over 55 years, and projected SCC incidence in the absence of screening by assessing the changes in the incidence rate of AC. The Cancer Registry of Norway was used to identify all 19,530 malignancies in the cervix diagnosed in the period 1956–2010. The majority of these (82.9%) were classified as SCCs, 10.5% as ACs and the remaining 6.6% were of other or undefined morphology. By joint‐point analyses of a period of more than five decades, the average annual percentage change in the age‐standardised incidence was −1.0 (95%CI: −2.1–0.1) for cervical SCC, 1.5 (95%CI:1.1–1.9) for cervical AC and −0.9 (95%CI: −1.4 to −0.3) for cervical cancers of other or undefined morphology. The projected age‐standardised incidence rate of cervical SCC in Norway, assuming no screening, was 28.6 per 100,000 woman‐years in 2010, which compared with the observed SCC rate of 7.3 corresponds to an estimated 74% reduction in SCC or a 68% reduction due to screening in the total cervical cancer burden. Cytology screening has impacted cervical cancer burden more than suggested by the overall observed cervical cancer incidence reduction since its peak in the mid‐1970s. The simultaneous substantial increase in cervical adenocarcinoma in Norway is presumably indicative of an increase in exposure to HPV over time.

Safety and acceptability of human papillomavirus testing of self-collected specimens: A methodologic study of the impact of collection devices and HPV assays on sensitivity for cervical cancer and high-grade lesions

BACKGROUND Comparative data on different self-collection methods is limited. OBJECTIVES To assess the impact of hrHPV testing of two self-collection devices for detection of cervical carcinoma and high-grade lesions. STUDY DESIGN Three hundred ten patients collected two cervicovaginal specimens using a brush (Evalyn®Brush) and a swab (FLOQSwabs™), and filled a questionnaire at home. Then, a physician at the clinic took a cervical specimen into PreservCyt® buffer for hrHPV testing and cytology. All specimens were tested using Anyplex™ II HPV28, Cobas® 4800 HPV Test and Xpert®HPV. RESULTS Performance comparison included 45 cervical carcinomas and 187 patients with premalignant lesions. Compared to the physician-specimen, hrHPV testing of Evalyn®Brush showed non-inferior sensitivity for CIN3+ (relative sensitivity of Anyplex™ 0.99; Cobas® 0.96; Xpert®HPV 0.97) while hrHPV testing of FLOQSwabs™ showed inferior sensitivity (relative sensitivity of Anyplex™ 0.91; Cobas® 0.92; Xpert®HPV 0.93). Similar results were observed for invasive carcinomas albeit that FLOQSwabs™ was statistically non-inferior to the physician-specimen. Self-collection by either Evalyn®Brush or FLOQSwabs™ was more sensitive for CIN3+ than LSIL or worse cytology. Significant decrease in sensitivity for CIN3+ were observed for FLOQSwabs™ when specimens were preprocessed for hrHPV testing after 28 days. Both devices were well accepted, but patients considered Evalyn®Brush easier and more comfortable than FLOQSwabs™. CONCLUSIONS Self-collection is comparable to current screening practice for detecting cervical carcinoma and CIN3+ but device and specimen processing effects exist. Only validated procedure including collection device, hrHPV assay and specimen preparation should be used.

Investigation of nonspecific cross-reacting antigen 2 as a prognostic biomarker in bone marrow plasma from colorectal cancer patients

Carcinoembryonic antigen (CEA) is still the only routinely used biomarker in colorectal cancer (CRC), but its utility is hampered by poor specificity and sensitivity, and the search for novel biomarkers is highly warranted. The nonspecific cross-reacting antigen 2 (NCA-2), a truncated CEA species molecule which is transcribed from the same gene, has been suggested as an alternative biomarker to CEA. In the present work, specific immunofluorometric assays were used for detection of NCA-2 and full-length CEA in bone marrow plasma from 277 CRC patients to assess their value as prognostic biomarkers, and detection was also performed in tumor tissue and a CRC cell line. Elevated plasma CEA was associated with advanced tumor stage at diagnosis and adverse patient outcome, while for NCA-2, although the same trends were observed, no additional prognostic information was gained. While specific detection of NCA-2 was clearly achieved in plasma samples, cross-reactivity with full-length CEA was observed when the antigen was exposed to common fixation chemicals. The results from this study indicate that NCA-2 is probably not a prognostic biomarker in CRC and, furthermore, underline the issue of antibody specificity when investigating CEA species molecules.

Abstract 4135: Improving attendance to the cervical cancer screening program: Does self-sampling at home improve cervical cancer prevention

With over half a million new cases and 275,000 deaths each year, cervical cancer constitutes a major public health problem world-wide, ranking as the third most common cancer in women. Screening with Pap-smear has reduced cervical cancer related morbidity and mortality considerably. However, the challenge lies in making screening and treatment available to all women at risk. Main objective of this study was to evaluate performance of high-risk human papillomavirus (hrHPV) testing on self-collected smears by women who did not attend to screening regularly. Specifically we were interested to observe if: i) use of different self-sampling devices for hrHPV was feasible in Norway and ii) will self-sampling enhance attendance among women with suboptimal screening? iii) which HPV test should be used to ensure optimal follow-up? Following approval from the Regional Ethical Committee, we identified in Oslo, Norway, during April-May 2013, 3,393 women with sub-optimal screening history. To motivate screening attendance, two different interventions were randomly allocated: 1) 300 women in age 26-34 and 300 women in age of 35-49 years, and 200 women in age of 50-69 years, received either lavage based self-screener (Delphi ScreenerTM, Delphi Bioscience BV) or dry brush self-screener (Evalyn Brush®, Rovers, The Netherlands); 2) 2,593 received a routine reminder letter to make an appointment for a Pap-smear. Those, who returned self-sampling devices or who had screening smear taken in 6 months period were defined as attendees. The population-based screening registry was used to identify those who had a screening smear. Returned self-sampling devices were tested for hrHPV by Hybrid Capture2 (HC2) High-Risk HPV DNA Test® (QIAGEN, Gaithersburg, MD, USA) and HPV genotyped (CLART, Genomica, Madrid, Spain). Out of 800, 80 women declined to participate in the self-sampling study and 720 devices was mailed out; 168 (23.3%) self-sampling devices were returned, 88 dry brush and 80 lavage-based self-sampling devices; in addition 59 had a Pap-smear (8.2%). 388 (14.7%) out of 2,593 who received a letter had a Pap-smear. Overall hrHPV positivity rate, defined as positive either by CLART or HC2, was over 20% being highest in 26-34 years old (>30% ) and lowest in 50-69 years old ( The preliminary results show 31.5% participation for self-sampling group compared to 14.7% participation for the routine, reminder letter group. Preliminary results indicate that i) it is feasible to use self-sampling device in routine screening; ii) use of self-sampling improves overall attendance of the screening program; iii) comparison of two different hrHPV detection methods will be presented in the conference. Citation Format: Kristina Schee, Lonnberg Stefan, Helle Pedersen, Jesper Bonde, Mari Nygard. Improving attendance to the cervical cancer screening program: Does self-sampling at home improve cervical cancer prevention. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4135. doi:10.1158/1538-7445.AM2014-4135