Title: Symptoms, CA125 and HE4 for the preoperative prediction of ovarian malignancy in Brazilian women with ovarian masses

Transcription

1 Author's response to reviews Title: Symptoms, CA125 and HE4 for the preoperative prediction of ovarian malignancy in Brazilian women with ovarian masses Authors: Denise R Pitta (deni@fcm.unicamp.br) Luis O Sarian (luis.sarian@gmail.com) Amilcar Barreta (abarreta@yahoo.com.br) Elisabete A Campos (betecamp@unicamp.br) Liliana L A Andrade (lucci@unicamp.br) Ana M Fachini (oncologia@caism.unicamp.br) Leonardo M Campbell (leo.m.campbell@gmail.com) Sophie Derchain (derchain@fcm.unicamp.br) Version: 2 Date: 11 March 2013 Author's response to reviews: see over

2 Reviewer's report 1# Title: Symptoms, CA125 and HE4 for the preoperative prediction of ovarian malignancy in Brazilian women with ovarian masses Version: 1 Date: 7 January 2013 Reviewer: Toon Van Gorp Reviewer's report: Dear editor, This is a nicely written paper on the importance of symptoms in the diagnosis of ovarian cancer. The authors compare a healthy cohort (n=178), with patients with a benign mass (n =117), and with patients with a malignant mass (n = 61). This is a prospective study with an adequate power analysis based on previous studies. To my opinion it certainly deserves publication. Response: Thank you very much for the time and effort you put reviewing our manuscript. We are also grateful for the encouraging words. We performed a thorough revision of the text, carefully addressing each of the referees concerns. However, in light of the conceptual nature of the reviewer s comments, the text had to be completely reformulated. We put a special effort in making it clear to the reader that our main objective was to assess the usefulness of symptoms and tumor markers in the preoperative discrimination of women with malignant from those with benign ovarian tumors when these women were already diagnosed with an adnexal mass. However, there are also a couple of revisions that need to be addressed before publication: Major Compulsory Revisions 1) In the introduction, the authors are a little bit overoptimistic about the usefulness of HE4 in the diagnosis of ovarian cancer. The authors write: HE4 (human epididymis protein 4) has been proven to be more sensitive for the detection of ovarian cancer compared to CA125 alone. First, in most of the studies HE4, on an individual basis, did not excel CA125. As an example the meta-analysis of Li et al. which was published recently in BMC Cancer: Fake Li, Ruxiu Tie, Kai Chang, Feng Wang, Shaoli Deng, Weiping Lu, Lili Yu, Ming Chen Does risk for ovarian malignancy algorithm excel human epididymis protein 4 and ca125 in predicting epithelial ovarian cancer: A meta-analysis. BMC Cancer 2012, 12:258. Second, even for the combination of CA125 and HE4 not all studies were positive. Some examples of studies that were not able to confirm these results: -Jacob F, Meier M, Caduff R, Goldstein D, Pochechueva T, Hacker N, Fink D, Heinzelmann- Schwarz V. No benefit from combining HE4 and CA125 as ovarian tumor markers in a clinical setting. Gynecol Oncol Jun 1;121(3): Van Gorp T, Cadron I, Despierre E, Daemen A, Leunen K, Amant F, Timmerman D, De Moor B, Vergote I. HE4 and CA125 as a diagnostic test in ovarian cancer: prospective validation of the Risk of Ovarian Malignancy Algorithm. Br J Cancer Mar 1;104(5):

3 I don t mind that the authors are generally positive about HE4 and the above mentioned references do not have to be cited, but I do feel that this statement should be more balanced and less overoptimistic. Response: The introduction has now been modified, in order to tone down the optimistic view regarding the performance of HE4. The pioneer studies (on which we based our introduction) were essentially based on Dr. Moores s results, which are overly optimistic about the performance of HE4. In fact, based on newer studies, which we neglected in the original version of the manuscript, we are prone to agree with you in that the performance of HE4 may not be so shinning, and maybe performs worse than CA 125. We incorporated Li s and Van Gorp s works to our reference list, because these studies are necessary readings to someone who seeks for a balanced view of the performance of these markers. Because Dr. Timmerman, who has also reviewed this manuscript, made several questionings (items 7, 8 and 9 of his review) about the performance of the markers, we modified our analytical approach. The manuscript is now much more focused on the performance indicators of the symptom clusters and the biomarkers, which led us to re-focus the introduction as well. 2) A more important remark is the fact that there is a major source of bias in this study: Attention bias and recall bias. Cases were asked to participate in this study when the diagnosis was known. Patients with ovarian cancer are more likely to mention symptoms then patients with a diagnosis of a benign mass or more so than patients without any ovarian mass. The symptoms may be exaggerated because of the recent diagnosis. This source of bias should be discussed in the discussion section as an important drawback. Response: The questionnaires were applied before surgery, i.e. before any potential final pathological diagnoses had been ascertained. Patients, as their doctors, were only aware that they had an adnexal mass. This has now been clarified in the revised text; the modifications spanned from the objectives to the discussion, and a good notion of these can be derived from the methods section: Women with adnexal masses were interviewed before surgery to avoid recall bias, since the main purpose of the study was to investigate whether symptoms could help discriminate preoperatively women with malignant ovarian tumors.. As mentioned before, this information is reiterated at several points in the text. 3) Something else which should be clarified: The authors used the Ward agglomerative method to cluster of symptoms. Clusters of symptoms were identified when symptoms agglomerated at the lowest level of the cluster dendogram. Symptoms that did not agglomerate at the lowest level of the cluster dendogram were considered as isolated symptoms. When looking at the dendogram it can be noticed that, indeed, leg swelling and able to feel abdominal mass remained isolated symptoms. However, it is not clear to me why back pain is not considered as an isolated symptom but clustered together with pelvic an abdominal pain. Back pain is at the same level as leg swelling and able to feel abdominal mass. This should be corrected an if needed the statistical calculations should be redone. Response: You are correct. The definitions of symptom clusters were loosely based on the product of the Ward agglomerative method. We have now redefined the clusters, adhering strictly to the lowest level of the dendogram. All calculations were thus remade. In addition, we modified the presentation of the results with redesigned tables, this time trying to address the modifications proposed by you and referee #2. Minor Essential Revisions

4 1) Page 4 (introduction): Serum analysis of tumor markers and ultrasound were the traditional approach to women with adnexal masses and CA125 measurement is an important component in the workup of a woman with an adnexal mass [4, 5]. The references cited in this sentence do not refer to content of the sentence since neither of the references refer to ultrasound or CA125 measurements. More appropriate references should be stated here. Response: We agree. Pertinent works are now referenced in the text, which has been completely reformulated. 2) The last paragraph on page 12 and beginning of page 13: The proportion of women with the abdomen cluster of symptoms. The bladder cluster of symptoms was significantly more prevalent in women with benign tumors compared with healthy women (p=0.03). can be shortened or omitted since all this information can be found in Table 5. Response: We agree. Because the text has now been completely revised, this and several other text inconsistencies and redundancies have been addressed. 3) reference should be cited in ascending order: -bottom of page 5: [14, 15, 11, 16]. -bottom of page 14: [23, 17, 18, 15] I hope that these remarks can improve the manuscript. Response: We apologize for these and many other edition problems.

5 Reviewer's report 2# Title: Symptoms, CA125 and HE4 for the preoperative prediction of ovarian malignancy in Brazilian women with ovarian masses Version: 1 Date: 14 January 2013 Reviewer: Dirk Timmerman Reviewer's report: This manuscript covers an interesting and important topic. However it needs several Major Compulsory Revisions. Response: Thank you very much for your diligent review of our manuscript. In light of the conceptual nature of your comments, the text had to be rewritten in a different manner. Our main objective was to assess the usefulness of symptoms and tumor markers in the preoperative discrimination of women with malignant from those with benign ovarian tumors when these women had already been diagnosed with an adnexal mass. We took special care addressing your comments 7 to 9: a performance comparison of the isolated symptoms, symptom clusters and tumors marker has now been made. We calculated sensitivity, specificity, PPV and NPV for the detection of malignancy. For that reason, although we respond to each of the points raised by you, it is not possible to pinpoint each modification made to the text. 1. The authors do not mention and do not make any use of the information obtained from imaging (e.g. ultrasound findings). Response: The objective of the study was to we investigate preoperatively the presence of specific symptoms and tumor markers in women with suspected adnexal masses in order to assess the role of each symptoms, a symptoms index and tumor marker expression in the discrimination of malignant and benign ovarian tumors. We also evaluate the presence of these symptoms among a group of healthy women to assess the likelihood that controls have a similar experience of symptoms as cases. In this new version we tried to make it clear to reader why we focused on this discrimination, in the present study. We are aware that ultrasound (US) and other clinical data are pivotal for the discrimination of adnexal masses. In this current version of our manuscript, we added entire paragraphs regarding the importance of US, in introduction, discussion and conclusion (Kaijser et al. 2013). Also recently, we published our results using ultrasound (Hartman CA, Juliato CR, Sarian LO, Toledo MC, Jales RM, Morais SS, Pitta DR, Marussi EF, Derchain S. Ultrasound criteria and CA-125 as predictive variables of ovarian cancer in women with adnexal tumors. Ultrasound Obstet Gynecol 2012,40(3):360-6 ). In that study, we based US methodology on IOTA paradigms, testing the IOTA simple ultrasound rules (as proposed by Timmerman et al., 2000, and using other references like Timmerman et al. 2005; Timmerman et al., 2007, Timmerman et al., 2008, Van Holsbeke et al., 2009, Timmerman et al., 2010, Van Holsbeke et al., 2012) to identify malignant tumors among women with adnexal masses, resulting in a net sensitivity of 90%, specificity of 87%, positive predictive value (PPV) of 69% and negative predictive value (NPV) of 97%. 2. The authors do not prove that including symptoms and tumor markers will improve distinction between malignant and benign tumors compared to ultrasound or other diagnostic tests.

6 Response: We agree. In the present study, we are interested in assessing whether using very simple, affordable tools such as a brief questionnaire about symptoms and blood tests can guide the general practitioner in the referral of a patient to a specialized center. In the current version of the manuscript, we clearly acknowledge that US is the major tool for the discrimination of adnexal masses. We hope the text can now clearly convey our opinion about this subject.. 3. The authors do not discuss state-of-the-art tests or algorithms to distinguish preoperatively between benign and malignant tumors (see IOTA studies). Response: We agree in full. Indeed, our first version missed several important aspects of the current knowledge on the discrimination of adnexal masses. These omissions have now been tackled. 4. The manuscript and especially the discussion seem to mix screening with preoperative diagnosis. Response: We agree; we are now focusing only on the preoperative discrimination of malignant from benign ovarian tumors in women with US detected adnexal masses. I do not agree with the authors stating in their introduction: ROMA index has performed better than RMI. Since other studies for instance Van Gorp et al. Eur J Cancer 2012 found different results. Response. You are right, the text was modified accordingly. 5. I do not fully understand how symptoms contribute to a preoperative diagnosis of ovarian cancer in clinical practice? From a screening perspective they may be valuable to identify women at high risk of having ovarian cancer that need further diagnostic evaluation. However, in a diagnostic setting, or preoperative setting symptoms may increase your awareness for cancer but you then still need to use a diagnostic tool to decide upon the nature of an ovarian mass. Response: We are not interested or, say, not capable at this point, to address ovarian câncer screening in the general population. It may seem useless to investigate symptoms and CA125 levels in women already diagnosed with an adnexal mass, since the decision as to whether operate or will be most likely made on the basis of the sonographic features of the lesion. However, it seems clear to us that our results suggest that symptom investigation in association with CA125 measurements can substantially augment the likelihood of a woman being correctly referred to a specialized cancer center. 6. The authors state that selection of symptomatic patients seems a promising way of detecting low volume disease and increase resectability. However, in the DOvE trial although they could identify cases presenting with less tumor burden than those found in women referred to the same hospital through the regular channels, seven of nine HGSC (type II disease) in DOvE patients were still stage III IV because the disease originated outside the ovaries. Therefore it remains to be seen whether symptom based screening will eventually reduce overall mortality. Response: We agree, and we are not going to address screening issues in the present study, as mentioned before; we modified the discussion regarding stage I disease.

7 Our data demonstrated that symptoms may be used even to differentiate early stage ovarian disease from women with benign ovarian tumors. In the present study, 53% of the patients had stage I disease, regardless of the histological type of the tumor, and 78% of these had positive symptom index. Rossing et al. [N] demonstrated that the symptom index was positive in 62.3% of women with early stage disease and Goff et al (2004) obtained a 57% sensitivity. However, in both studies, most of the women were surveyed after diagnosis, whereas in our study we surveyed the women before surgery and the diagnosis of cancer. Because we aimed at identifying women who would benefit from a referral to a specialized center, we grouped together women with epithelial, germ cell and sex cord malignant tumors. On the other hand, we allocated to the same group women with borderline epithelial tumors and those with lowor high-grade invasive epithelial carcinomas. It is well known that these different histological types display varying clinical behaviors. Based on a dualistic model of carcinogenesis epithelial ovarian carcinoma can be classified as type I and type II. Type I included low-grade epitelial carcinomas, generally indolent and easily detected in stage I. Type II ovarian carcinoma, comprise high-grade and undifferentiated carcinomas (Kurman and Shi 2011). It has been well demonstrated that high-grade serous tumors are rarely diagnosed before it has spread, and for this type of tumors, diagnostic approaches should be aimed at diagnosing low-volume tumors, not only at early stage (Brown and Palmer 2009, Gilbert 2012). 7. What are the cut off levels used to indicate malignant disease for CA125, HE4 and ROMA? Response: We used standard receiver operator characteristics (ROC) analysis to determine the best CA125, HE4 and ROMA index cutoff points in discriminating benign from malignant ovarian tumors. The optimal cutoff point for CA125 was 36.4U/L; HE4 was 41.6pmol/L for premenopausal women and 96.6pmol/L for postmenopausal women. we based the risk estimation on the optimal cutoff points of the ROMA predictive index (PI) in pre (5.01%) and postmenopausal (18.2%) women. ROC AUC comparisons were performed with the DeLong method. These informations are now cited in the text. 8. Why do the authors not provide test performance of ROMA, HE4, or CA125 for preoperative prediction of ovarian cancer? And compare this to use of a symptom index combined with biomarkers as a diagnostic tool? Response: Thank you very much for the nice suggestion. We have now done that. We performed pairwise comparisons of the prevalence of symptom clusters, symptom index, and the positivity rate of CA125, HE4 and ROMA index according to tumor malignancy and stage strata, using unadjusted odds ratios with 95%CI. We also calculated the performance indicators (sensitivity, specificity, with 95% confidence intervals, positive and negative predictive values) of the tumor markers in strata of women positive or negative symptom index, for the discrimination of all malignant and stage I malignant tumor from benign ovarian tumors. 9. From a clinical perspective it is still unclear to me to what extent symptoms contribute to the diagnosis of ovarian cancer? You provide sufficient data that specific clusters of symptoms are more prevalent among malignant tumors. But since this study focuses on the preoperative prediction of an ovarian mass I would expect to see diagnostic test results (sensitivity/specificity/lr+, LR-, AUCs) Response: Thank you again for the insightful suggestion. Please see tables 4 to 6.

8 10. I would recommend shortening of the first paragraph of the discussion. From: in this sample..benign tumors and the control group. Response: This and many other modifications have now been made. Minor essential revisions: The abstract does not describe whether the study was prospective or retrospective. Please revise the English text by a native English speaker. E.g. P5: correct grammar in the following sentence: and what would be the best strategy does diminish delays in diagnosis. P14: filling full quickly, should read feeling full In the tables commas should be replaced by dots (e.g. p<0.01 instead of p<0,01) Response: We apologize for these and many other edition problems. Because the text has now been completely revised, this and several other text inconsistencies and redundancies have been addressed.