Archives of Academic Emergency Medicine. 2019; 7 (1): e24 OR I G I N A L RE S E A RC H Value of CA-125 Glycoprotein in Predicting Acute Appen- dicitis; a Diagnostic Accuracy Study Mahboub Pouraghaei1, Kavous Shahsavarinia1, Farzad Kakaei2, Sevda Gholipour-Khalili3∗, Babak Mohammadpour1, Payman Moharamzadeh1, Moloud Balafar1 1. Emergency Medicine Research Center, Tabriz University of Medical Sciences, Tabriz, Iran. 2. Department of Surgery and Transplantation, Tabriz University of Medical Sciences, Tabriz, Iran. 3. Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran. Received: February 2019; Accepted: March 2019; Published online: 6 April 2019 Abstract: Introduction: Carcinogen antigen 125 (CA-125) is a glycoprotein antigen, which has shown potentials in pre- dicting peritoneal inflammation. The aim of this study is to determine the value of CA-125 in predicting acute appendicitis (AA). Methods: This prospective diagnostic accuracy study was conducted on 15 – 70 year-old pa- tients with acute abdominal pain, suspected to AA, referred to emergency department. The serum level of CA- 125 was measured for all patients before appendectomy and its screening characteristics in detection of AA case (confirmed by histology findings) were calculated and reported with 95% confidence interval (CI). Results: 95 patients with the mean age of 31.65 ± 12.9 (15-75) years were studied (54.3% male). Based on the histologic findings, 72 (75.8%) cases were categorized as AA (23 cases as severe). AA and non-AA (NAA) groups were similar regarding the mean age (p = 0.59), mean duration of symptoms (p = 0.08), mean white blood cell (WBC) count (p = 0.37), and mean PMN percentage (p = 0.55). Mean CA-125 level was 16.5 ± 20.0 U/mL in the AA group and 30.5 ± 6.1 U/mL in the NAA group (p = 0.001). Adjustment of analysis based on gender revealed a significant correlation between CA-125 level and diagnosis of AA only in females (34.23 ± 39 U/mL in NAA versus 20.7 ± 26.7 U/mL in AA, p = 0.012). The area under the ROC curve of CA-125 was 0.62 (95%CI: 0.51 to 0.72). Sensitivity, specificity, NPV, PPV, NLR, and PLR of CA-125 in 16.4 U/mL cut off (best point) were 77.8% (95%CI: 66.4 - 86.7), 50.0% (95%CI: 28.2 - 71.8), 83.6% (95%CI: 76.7 - 88.7), and 40.7% (95%CI: 27.4 - 55.6), 0.44 (95%CI 0.2 - 0.8), and 1.56 (95%CI: 1.0 - 2.4), respectively. Conclusion: Considering the lower levels of CA-125 in patients with AA compared with NAA cases and also weak screening performance characteristics, it seems that it could not be considered as an accurate screening tool in this regard. Keywords: Appendix; appendicitis; CA-125 antigen; biomarkers; abdominal pain Cite this article as: Pouraghaei M, Shahsavarinia K, Kakaei F, Gholipour-Khalili S, Mohammadpour B, Moharamzadeh P, Balafar M. Value of CA-125 Glycoprotein in Predicting Acute Appendicitis; a Diagnostic Accuracy Study. Arch Acad Emerg Med. 2019; 7(1): e24. 1. Introduction Acute appendicitis (AA) is one of the common causes of ab- dominal emergency surgeries. The possibility of one facing this condition over their lifetime varies between 6.7% to 8.6% (1, 2). The accurate and timely diagnosis of acute appendici- tis plays an essential role in preventing life-threatening com- plications such as perforation associated with other morbidi- ties or mortality (3). The diagnosis of appendicitis is made ∗Corresponding Author: Sevda Gholipour-Khalili; Imam Reza Hospi- tal, Golgasht Street, Tabriz, Iran. Postal Code: 51666-14756 Email: sevymed@yahoo.com Phone: 00989144089490 based on clinical examination followed by laboratory and ra- diographic studies. These patients may undergo unneces- sary hospital admissions and operations as the result of false- positive diagnosis, while the false-negative findings can lead to extreme consequences (4). Studies have shown that 40-83% of cases are detected based on the classic clinical symptoms (5-7). Currently, laboratory parameters such as white blood cell count, neutrophil per- centage, and C-reactive protein concentration are used in various combinations to improve sensitivity and specificity of assessments for determining risk of appendicitis (8, 9). Carcinogen antigen 125 (CA-125) has shown potentials to be considered as a diagnostic test in this regard (10). CA-125 is a glycoprotein antigen, which is well known as a marker This open-access article distributed under the terms of the Creative Commons Attribution NonCommercial 3.0 License (CC BY-NC 3.0). Downloaded from: http://journals.sbmu.ac.ir/aaem M. Pouraghaei et al. 2 Figure 1: The area under the receiver operating characteristics (ROC) curve of CA-125 in predicting acute appendicitis. for epithelial ovarian cancer. Increased levels of CA-125 have been identified in benign and malignant conditions, includ- ing uterine leiomyoma, endometriosis, pelvic inflammatory disease, cirrhosis, and pleural or peritoneal effusion. Dif- ferent types of coelomic epithelial cells such as peritoneal mesothelial cells have a role in producing CA-125, which may increase in conditions of peritoneal inflammation (11). Some studies have applied CA-125 as a diagnostic tool in AA, which found the serum level of CA-125 to be higher in patients suf- fering from appendicitis (10, 12). However, some researchers opposed this notion with results indicating absence of a pre- dictive role for this tumor marker (13, 14). According to the divergent findings, the association between CA-125 and AA is not well understood. The aim of this study is to determine the value of CA-125 in predicting AA. 2. Methods 2.1. Study design and setting This prospective diagnostic accuracy study was conducted on patients with acute abdominal pain, suspected to AA, re- ferred to emergency departments of Imam Reza and Sina University Hospitals, Tabriz, Iran, from 1st of January 2016 to the end of December 2017. The research protocol was con- firmed by the Ethics committee of Tabriz University of Med- ical Sciences (TBZMED.REC.94/3-7/23) and all the partici- pants gave informed written consent prior to the study. 2.2. Participants Using non-probability consecutive sampling method, the participants were chosen from patients between the ages of 15 to 70 years with acute abdominal pain suspected to acute appendicitis who were candidates for surgical appen- dectomy. Patients with history of smoking, diabetes, hyper- tension or any medical conditions that associated with in- creased level of serum CA-125, such as cirrhosis, congestive heart failure, inflammatory bowel disease, malignancy, en- dometriosis, pregnancy and recent abdominal surgery were excluded. 2.3. Procedure The serum level of CA-125 was checked for all patients (us- ing 5 cc blood sampling from left/right brachial vein) in the emergency department before the operation. An Electro- chemiluminescence (ECL) assessment was done using E411 Cobas machine and kits produced by Germany Roche Com- pany (based on Company reference) for CA-125 measure- ment. In addition, routine laboratory testing including white blood cell (WBC) level and polymorph nuclear (PMN) cell percentage were also done for each patient. Ultrasonogra- phy was also performed on each patient as part of the diag- nostic approach. The gold standard for diagnosis of AA was positive histological findings described in pathology reports. Patients with an Alvarado score of 7 or above were considered for surgery. 2.4. Data gathering A checklist consisting of patients’ demographic, clinical, and histopathological variables (after the surgery) was filled out for all patients by a senior emergency medicine resident un- der super vision of an emergency medicine specialist and su- pervisor of the project. The pathology reports were classified in three categories including negative for appendicitis, sim- ple appendicitis or severe ones (phlegmonous, abscess, per- forated or gangrenous appendix). 2.5. Statistical Analysis Sample size was calculated as 93 patients, considering 60% sensitivity, 100% specificity and a confidence interval of 95%, and 80% power. Statistical analysis was performed using SPSS software version 19.0 (IBM Corp., Armonk, N.Y., USA). Categorical variables were reported as percentages and con- tinuous variables as mean ± standard deviation (SD). Con- tinuous variables were compared using independent t-test and categorical variables were compared via chi-square test. Sensitivity, specificity, positive predictive value (PPV ), neg- ative predictive value (NPV ), positive likelihood ratio (PLR), and negative likelihood ratio (NLR) as well as area under the receiver operating characteristics (ROC) curve of CA-125 in This open-access article distributed under the terms of the Creative Commons Attribution NonCommercial 3.0 License (CC BY-NC 3.0). Downloaded from: http://journals.sbmu.ac.ir/aaem 3 Archives of Academic Emergency Medicine. 2019; 7 (1): e24 Table 1: Comparison of the baseline characteristics between patients with acute appendicitis and those with normal appendix on histologic findings Variable Acute appendicitis P value Yes (n=72) No (n=23) Gender Male 38 (52.7) 5 (21.7) 0.03 Female 34 (42.3) 18 (78.3) Age (year) Mean ± SD 37.25 ± 44.8 29.73 ± 14.9 0.59 Duration of symptoms (hour) Mean ± SD 12.53 ± 16.9 11.32 ± 18.2 0.08 PMN (%) Mean ± SD 75.1 ± 11.5 76.2 ± 12.2 0.55 WBC count (/ mm3) Mean ± SD 12022.3 ± 3871.6 10209.5 ± 4387.4 0.37 Data are reported as mean ± standard deviation (SD) or frequency (%). PMN: polymorph nuclear; WBC: white blood cell. predicting AA were calculated using Medcalc software (ver- sion 18.2) and reported with 95% confidence interval (CI). P- values less than 0.05 were considered as significant. 3. Results 95 patients with the mean age of 31.65 ± 12.9 (15-75) years were studied (54.3% male). Based on the histologic findings, 72 (75.8%) cases were categorized as AA (23 cases as severe) and the remaining 23 (24.2%) cases were normal (NAA). Table 1 compares the baseline characteristics of AA patients with others. The two groups were similar regarding mean age (p = 0.59), mean duration of symptoms (p = 0.08), mean white blood cell (WBC) count (p = 0.37), and mean PMN percentage (p = 0.55). 3.1. CA-125 level Mean CA-125 level was 16.5 ± 20.0 U/mL in the AA group and 30.5 ± 6.1 U/mL in the NAA group (p = 0.001). It was signif- icantly higher in female patients (25.4 ± 31.8 vs 12.9 ± 10.7 U/mL; p = 0.03). Adjustment of analysis based on gender re- vealed a significant correlation between CA-125 level and di- agnosis of AA only in females (34.23 ± 39 in NAA versus 20.7 ± 26.7 U/mL in AA, p = 0.012). Adjustment of analysis based on severity of AA revealed an insignificant correlation between severity of AA and CA-125 level (p = 0.058). 3.2. Screening characteristics of CA-125 Figure 1 shows the area under the ROC curve of CA-125 in identifying cases with AA. The area under the ROC curve of CA-125 was 0.62 (95%CI: 0.51 to 0.72). Based on the ROC curve analysis, the best cut off point of CA-125 in predict- ing AA was estimated to be 16.4 U/mL. Sensitivity, speci- ficity, NPV, PPV, NLR, and PLR of CA-125 in 16.4 U/mL cut off were 77.8% (95%CI: 66.4 - 86.7), 50.0% (95%CI: 28.2 - 71.8), 83.6% (95%CI: 76.7 - 88.7), and 40.7% (95%CI: 27.4 - 55.6), 0.44 (95%CI 0.2 - 0.8), and 1.56 (95%CI: 1.0 - 2.4), respectively. 4. Discussion In this study, we assessed the possible diagnostic value of CA- 125 in detecting AA. We hypothesized that patients with AA have higher CA-125 levels compared with NAA patients. Sur- prisingly, NAA patients had higher CA-125 levels compared with AA patients. In the cut off value of 16.4 U/mL, CA-125 had a sensitivity of 77.8% and specificity of 50% in differenti- ating NAA patients from AA cases. Literature supported that some tumor markers such as CA- 125 may have additional value in not only the diagnosis of AA, but also the differentiation between severe complicated cases and simple cases (11). CA-125 is commonly used as a marker in gynecological cancers. Considering the possi- ble role of peritoneal cells in secreting CA-125 during the phase of inflammation, this biomarker may increase over the course of peritonitis resulting from AA. Basaran et al. proposed that secretion of CA-125 starts six hours prior to the onset of inflammation (15). Zeimet et al. compared the release of CA-125 from peritoneal cells and malignant ovarian cells. They reported higher CA-125 synthesis in peritoneal cells due to inflammation (16). It is also pro- posed that CA-125 is secreted by apical surface of mesothelial monolayers as a response to inflammatory cytokines such as interleukin-1 beta and tumor necrotizing factor-alpha, and E coli lipopolysaccharide (17). Berger et al. reported a significantly higher CA-125 level in males with severe appendicitis compared to simple cases, whereas, CA-125 in males with AA didn’t differ significantly from NAA patients. However, Sevinc et al. found a significant positive correlation between CA-125 levels and AA diagnosis, which also decreased after the surgical appendectomy (11). In contrast, in the present study, the serum levels of CA-125 in patients with severe AA did not significantly differ from sim- This open-access article distributed under the terms of the Creative Commons Attribution NonCommercial 3.0 License (CC BY-NC 3.0). Downloaded from: http://journals.sbmu.ac.ir/aaem M. Pouraghaei et al. 4 ple types of disease, even after the categorization of the study population based on gender. Cetinkaya et al. also examined the correlation between CA- 125 and severe AA. They found high serum levels of CA-125 in severe AA cases with the cut off value of 35 U/mL. The cal- culated sensitivity, specificity, and PPVs were 60%, 100%, and 100%, respectively (18). In our study, the female group had significantly higher levels of CA-125. After analyzing CA-125 levels in gender specific subgroups, we found CA-125 levels to be higher in NAA cases compared to AA cases in the female subgroup. However, there was not a significant correlation between CA-125 levels and AA diagnosis in the male group. Secretion of CA-125 from ovarian cells in females may have affected the final results of the study. Some of the previous researchers also evaluated CA-125 levels only in male cases. 5. Limitation The study population was limited to 95 patients and in stud- ies with larger sample groups the results may differ from the present report. In addition, we did not evaluate CA-125 levels after the surgery to compare cases with AA and NAA. Hence, more studies with larger study populations and with higher cut off values are required to assess the possible role of CA- 125 in AA diagnosis. 6. Conclusion The results of the present study showed a low sensitivity and specificity for serum levels of CA-125 in differentiating pa- tients with AA from those with NAA. Additionally, it was not an effective marker for diagnosis of the complicated form of AA. However, more research on a greater sample of patients is required to evaluate its predictive value. 7. Appendix 7.1. Acknowledgements We would like to thank Yousef Asgharzadeh and Zahra Seifar for their contribution to English editing of the paper. This study was retrieved from a medical dissertation conducted in Tabriz University of Medical Sciences. 7.2. Author contribution M.P. study design, K.S.N. and B.M study conduct, F.K. and M.P advisory, and supervision, B.M. and M.B data gathering and writing, P.M. and S.Gh.K. analysis. Authors ORCIDs Mahboub Pouraghaei: 0000-0002-8421-6110 Kavous Shahsavarinia: 0000-0001-7359-4880 Farzad Kakaei: 0000-0003-1632-7393 7.3. Funding/Support This study was fully supported by Tabriz University of Medi- cal Sciences. 7.4. Conflict of interest All the authors declared that there is no conflict of interest. References 1. Chamisa I. A clinicopathological review of 324 appen- dices removed for acute appendicitis in Durban, South Africa: a retrospective analysis. Ann R Coll Surg Engl. 2009 Nov;91(8):688–92. 2. Ricci MA, Trevisani MF, Beck WC. Acute appendicitis. A 5-year review. Am Surg. 1991 May;57(5):301–5. 3. Stringer MD. Acute appendicitis. J Paediatr Child Health. 2017 Nov;53(11):1071–6. 4. 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High serum cancer antigen 125 level indicates perforation in acute appendicitis. The American journal of emergency medicine. 2015;33(10):1465-7. This open-access article distributed under the terms of the Creative Commons Attribution NonCommercial 3.0 License (CC BY-NC 3.0). Downloaded from: http://journals.sbmu.ac.ir/aaem Introduction Methods Results Discussion Limitation Conclusion Appendix References