Microsatellite Instability Analysis in Uterine Cavity Washings to Detect Endometrial Cancer in Lynch Syndrome

Discussion

Herein we report a proof-of-concept study of MSI analysis in uterine cavity washings in LS patients with various indications of hysterectomy. While the procedure appeared feasible and the results were interpretable, both cases of unstable endometrial cancers yielded MSI in uterine washings and there was no false-negative case. This pilot study suggests that molecular tests could be of interest in the screening of endometrial cancer in LS patients.

Although the natural history of LS-related endometrial carcinoma has not been yet clearly elucidated, it seems that hyperplasia precedes development of cancer for a short period (13-15). Loss of expression of the MMR genes is followed by MSI which has been reported in both hyperplasia with or without atypia and invasive cancer (13, 14). Our study confirms these data: 3 out of 5 patients with loss of expression of MMR genes had benign tissue. Nieminen et al. analyzed precursor lesions of endometrioid endometrial cancer in DNA MMR gene mutation carriers (16). They showed on 110 samples that decreased MMR protein expression was present in 7% of normal endometrium, 40% in simple hyperplasia, 100% in complex hyperplasia without atypia, 92% in complex hyperplasia with atypia, and 100% in endometrial carcinoma. MSI frequencies were lower (6%, 17%, 67%, 38%, and 64%, respectively). Notably, molecular changes in endometrial tissue were detectable up to 12 years before endometrial carcinoma. This study suggests that, contrary to the traditional concept, both complex hyperplasia and atypical hyperplasia are precursor lesions of endometrial carcinoma (16). To identify occult hereditary non-polyposis colorectal cancer individuals among endometrial carcinoma patients, Sutter et al. examined complex atypical hyperplasia and endometrial carcinomas of 60 women of less than 50 years using MSI, immunohistochemistry, and DNA sequence analysis. They reported that all complex atypical hyperplasias with high-level MSI progressed to endometrial carcinoma; only one third of the complex atypical hyperplasias with MSI progressed to high-level MSI endometrial cancer. MSI analysis of complex atypical hyperplasia in young patients may therefore be a useful prognostic marker for predicting possible progression to high-level MSI endometrial carcinomas (17).

The MSI concept has already been assessed and validated in urinary tract carcinomas. Steiner et al. reported 10 cases of recurrent transitional-cell carcinoma (TCC) diagnosed by MSI analysis in urine in the routine follow-up of 11 patients (18). Similarly, Amira et al. found that MSI analysis allowed the detection of 92% of recurrence in TCC follow-up (19). Mourah et al. showed that MSI analysis for the detection of bladder TCC was a non-invasive reproducible tool, associated with a good diagnostic value (sensitivity of 83% and specificity of 100%) (20). Furthermore, this technique offered an early diagnosis of recurrence, prior to symptomatic evidence of disease (18, 19).

These findings have paved the way of our hypothesis that MSI identification in uterine cavity washings cells could be a good marker for detecting pre-malignant or malignant transformation of endometrial mucosa at a pre-cancerous state or an early stage of the disease.

We previously reported that microsatellite analysis in uterine cavity washings revealed instability in 2 cases of unstable endometrial cancer (12). As far as we are aware of, this was the first report in endometrial cancer, and led to the current study, highlighting the potential benefit of molecular tools for screening in LS-related endometrial cancer. Here we describe a simple procedure to retrieve endometrial cells, which reflect the surrounding pathological environment. This screening tool would not replace endometrial biopsy, which is currently the gold standard diagnostic tool. Although we reported a case of Lynch syndrome related-endometrial cancer diagnosed by MSI revealed on routine endometrial biopsy (21), the technique we describe in the present study also offers the advantage to explore the whole uterine cavity, and thus would not constitute a blinded sampling as endometrial biopsy does.

Nevertheless, a few limits should be considered. Firstly, this is a proof-of-concept study, with a small cohort, although our Institution if a referral Center for LS. These encouraging results warrant further investigations in a wider population, which are currently performed. Secondly, whereas our pilot study included only previously discovered malignant lesions, there was no case of hyperplasia, and therefore we were unable to assess the diagnostic value of the test to detect pre-cancerous lesions or asymptomatic hyperplasia, which could be of paramount interest. Indeed, screening and early diagnosis of asymptomatic lesion would be the ultimate goal of MSI analysis. Once again, the on-going study will hopefully respond to this issue. At last, uterine washings were carried-out under general anaesthesia in this pilot study, but we have no data whether such a technique would be tolerated in the ou-patient setting in awake patients. Respective invasiveness of endometrial biopsy and uterine washings should, thus, be compared. This is namely an essential pre-requisite for a screening tool. This point is currently addressed in our institution for further progression of this concept. Moreover, if general anaesthesia allowed us infusing a significant amount of saline, we speculate that lower volume could be as informative and better-tolerated in the absence of analgesia. Therefore, this innovative procedure could be performed during out-patient hysteroscopy or during the routine follow-up at the office. Indeed, to be of use, uterine washings should ideally be done in primary care by nurse practioners or general practioners, without requiring hysteroscopy.