{"paper_id":"31f9b4fe-d3d6-46d6-a78d-8fd530aac281","body_text":"Science Letters 2025; 14(2):1326230sl \n1 \n \n \n \n \n \n \n \n \nAssociation of Serum Cancer Antigen \n125 and Carcinoembryonic Antigen with \nMenstrual Irregularities in Young Non-\nHospitalized Females  \nMuhammad Salman Khalid 1,2*, Areej Fatima 2, Tuba Shabbir 3, Ali \nZaman1, Ahmed Abbas 1, Muzdalfa Liaquat 1, Marium Tufail Rao 2, \nAbeera Amir4 \n1Department of Allied Health Sciences, IQRA University Chak Shehzad Campus, Islamabad, \nPakistan  \n2Department of Biomedical Engineering and Sciences, School of Mechanical and \nManufacturing Engineering, National University of Sciences and Technology, Islamabad, \nPakistan  \n3Atta ur Rehman School of Applied Biosciences, National  University of Science s and \nTechnology, Islamabad, Pakistan \n4Department of Clinical Services, School of Health Sciences, University of Management \nand Technology, Lahore, Pakistan  \nAbstract \nMenstrual irregularities affect a substantial proportion of young females in \nSouth Asia and are often accompanied by mesothelial endometriosis, pelvic \ninflammatory disorders, and polycystic ovary syndrome (PCOS). This study \naimed to document serum carcinoembryonic antigen (CEA) and serum cancer \nantigen 125 (CA 125) levels in young, non-hospitalized female students with \nmenstrual irregularities across higher education institutions in Islamabad and \nRawalpindi. A cross-sectional study was conducted on 200 female students \n(aged 18 to 25) with pre-screened menstrual irregularities. A structured form \ndocumented demographics and clinical parameters. Venous blood samples \nwere collected, and the serum concentrations of both markers (CA 125 and \nCEA) were quantified using enzyme -linked immunosorbent assay (ELISA), \nwith rigorous statistical validation employing Chi -square categorical \nassociation (p < 0.05). In the population, 2.5% (n = 5) had elevated CEA levels \n(>5.0 ng/mL), and 7.0% (n = 14) had abnormal CA 125 antigen levels (>35.0 \nU/mL). Abnormally high CA 125 levels were strongly associated with severe \ndysmenorrhea (χ2 = 6.31, p = 0.012) and increased body mass index (25.0 \nkg/m2). Elevated CA 125 in young females was associated with dysmenorrhea \nseverity and increased body mass index, a pattern consistent with benign \npelvic pathology such as endometriosis or pelvic inflammatory disease rather \nthan malignancy. However, this cross-sectional sample lacked clinical and \nimaging correlation, and these findings should be confirmed in longitudinal \nstudies. \n \nResearch article \n2026 | Volume 14 | Issue 2 | Article ID 1326230sl \n                            \nL \nS \nDOI: https://doi.org/10.47262/SL/14.2.132026230 \n \nScience Letters ISSN 2345-5463 \nA R T I C L E  I N F O  \n \n \nReceived  \nJuly 18, 2026 \nRevised \nSeptember 23, 2026           \n Accepted   \nSeptember 27, 2026             \nPublished  \nOctober 29, 2026 \n \n \n*Corresponding author  \nMuhammad Salman Khalid  \nE-mail   \nsalman.khalid@iqrauni.edu.pk \nPhone \n+92-3040153816 \n \n \nKeywords \nCancer antigen 125 \nCarcinoembryonic antigen   \nDysmenorrhea  \nMenstrual irregularities  \nTumor markers  \nYoung females \n \n \nHow to Cite \nKhalid MS, Fatima A, Shabbir T, \nZaman A, Abbas A, Liaquat M, \nRao MT, Amir A. Association of \nserum cancer antigen 125 and \ncarcinoembryonic antigen with \nmenstrual irregularities in young \nnon-hospitalized females. \nScience Letters 2026; \n14(2):1326230sl \n \n \nOpen Access \nThis work is licensed under the Creative Commons Attribution -Non-\nCommercial 4.0 International License.  \n \nhttp://thesciencepublishers.com/images/paste66.jpg\nSCAN ME \n\n\n \nScience Letters 2026; 14(2):1326230sl \n2 \n \nIntroduction  \nMenstrual irregularities are among the most neglected \nhealth concerns among young females worldwide, \naffecting a substantial number (20 -30%) of females \nduring the reproductive years. In South Asian \ncountries, especially Pakistan, polymenorrhea , \noligomenorrhea, and severe dysmenorrhea are the \nmost widely reported menstrual abnormalities among \nuniversity-enrolled females across higher education \ninstitutions. These abnormalities are indirectly related \nto structural or functional pelvic disorders, including, \nbut not limited to, early -stage endometriosis, benign \novarian cysts, polycystic ovarian syndrome (PCOS), \nor pelvic inflammatory disorders (PID). Despite this \nhigh burden, young females in resource -restricted \nsettings in Pakistan rarely undergo p reliminary \nbiomarker screening or any diagnostic evaluations \nuntil symptoms compromise their fertility.  \nCancer antigen (CA 125) is a dense, high -\nmolecular-weight glycoprotein used as a preliminary \nbiomarker for evaluating epithelial cells related to \novarian cancer. However, this diagnostic marker is \nnot limited to ovarian -related anomalies; its \nexpression is upregulated  during mechanical \nstretching, mesothelial regulation of the peritoneum, \ncytokine dysregulation, and interleukin -induced \ninflammation. Consequently, elevated serum CA 125 \nlevels (>35 U/mL) are documented in various non -\nmalignant conditions, including uteri ne fibroid \nformation, active endometriosis, phases of the \nmenstrual cycle, and pelvic infections. This non -\nspecific mesothelial release of CA 125 is a better \npredictor of inflammation than an exact indicator of \novarian-related abnormalities. Apart from CA 125, \ncarcinoembryonic antigen (CEA) is another \nglycoprotein analyzed during the management of \ncolorectal, mucinous, and gastrointestinal disorders, \nwith mild increases in baseline CEA (>5.0  ng/mL) \nobserved during systemic inflammation, exposure to \npassive smoking, and metabolic dysfunctions [7-9].  \nWhile the diagnostic utility of CEA and CA 125 \nin late-stage oncology is well established in tertiary \ncare facilities across various districts of Pakistan, \nscreening among young and non-hospitalized groups \nremains underreported [10].  Evaluating these \nantigenic markers among university students with \nmenstrual irregularities provides a valuable baseline \nfor assessing epidemiological data. It also helps \nidentify benign pelvic pathologies, warranting early, \nnon-invasive clinical follow -up across diagnostic \nlaboratories [11]. This study aims to quantify serum \nCEA and CA 125 among female university students \nwith menstrual irregularities in Islamabad and \nRawalpindi and assess their relationship with body \nmass index (BMI) and symptom severity.  \nMaterials and Methods \nStudy design and setting  \nWe conducted a cross -sectional observational \nanalysis at four major higher education institutions in \nPakistan's twin cities: IQRA University, Pir Mehr Ali \nShah Arid Agriculture University (PMAS -AAUR), \nNational University of Sciences and Technology \n(NUST), and Fatimah Jinnah Women University \n(FJWU). The laboratory procedures followed \nstandard medical laboratory technology protocols \nissued by the National Institute of Health (NIH) and \nthe National Bioethics Committee (NBC).  \nPopulation and eligibility criteria  \nThe observed population comprises female university \nstudents aged 18 to 25 years who have  been \ndiagnosed with persistent menstrual irregularities. All \n200 analyzed participants had menstrual \nirregularities, and no regular-cycle comparison group \nwas enrolled, so all analyses are within -cohort. For \nthe evaluation of serum CEA and CA 125 levels, \nvenous blood (3 ml) was collected, along with a well-\nstructured questionnaire to assess lifestyle factors and \ngynecological history.  \nInclusion and exclusion criteria  \nParticipants were unmarried females aged 18 to 25 \nyears who reported menstrual irregularities persisting \nfor at least three consecutive months, categorized as \neither Polymenorrhea (cycle duration <21 days) or \noligomenorrhea (cycle duration >35 days). All \nparticipants voluntarily provided written informed \nconsent before enrollment and sample collection. \nFemales with a prior diagnosis of systemic, cancer, or \npelvic disorders were excluded. We also excluded \nstudents with active or recent pregnancy . We also \nexcluded students with autoimmune, inflammatory, \nor chronic hepato-renal conditions. \nSample size  \nThe sample size for this cross -sectional study was \ncalculated using the World Health Organization \n(WHO) [12]. The confidence interval, prevalence of \nmenstrual irregularities, and the error margin were set \nto 95%, 15%, and 5%, respectively. The required \nsample was 196; to account for potential hemolysis, \n200 students were enrolled.  \n\n \nScience Letters 2025; 14(2):1326230sl \n3 \n \nClinical scoring: data collection  \nBefore blood collection, students completed a well -\nstructured, pretested questionnaire to collect data on \nage, weight, height, dysmenorrhea severity (mild, \nmoderate, or severe based on analgesic use and \nrestrictions in daily activity), menstrual irregularity, \nand first -degree family history of anomalies. Body \nmass index was calculated as kg/m 2 and further \nstratified as per the criteria of WHO [13, 14]: \nunderweight: (<18.5 kg/m²) ; n ormal: (18.5 –24.9 \nkg/m²) and overweight: (≥25.0 kg/m²)  \nLaboratory procedure  \nBlood samples (3mL) were extracted from each \nparticipant using sterile equipment (aseptic \nconditions) and transferred into gel -barrier serum \nseparator tubes (SST). Samples were allowed to clot \nat room temperature (27 0C) for 30 minutes and \ncentrifuged at 3000 rpm for 10 minutes to separate \nserum from plasma. The serum aliquots were \nimmediately stored at 20 0C till further analysis. \nSerum CA-125 (Cat No. ELH-CA125-1, RayBiotech, \nInc., Peachtree Corners, GA, USA) and CEA (Cat No. \nKE00281, Proteintech Group, Inc., Rosemont, IL, \nUSA) levels were quantified via sandwich ELISA \nfollowing standard manufacturer incuba tion, \nwashing, and TMB substrate protocols. Absorbance \nwas recorded at 450 nm using a microplate reader \n(Model xMark, Bio-Rad Laboratories, Inc., Hercules, \nCA, USA), with concentrations calculated in \nduplicate against standard curves in accordance with \nstandard biosafety and clinical protocols [15-17].  \nEthical considerations  \nThe departmental institutional review board (IRB) at \nIQRA University Chak Shehzad approved the \nproposed protocol and questionnaire  (approval no \nIRB/IUCS/AHS/2026/011, dated: May 04, 2026) , \nand strict measures were taken to obtain written \ninformed consent from participants. Confidentiality \nof data was maintained through de -identification; \nunique numbers were assigned to each specimen, and \naccess was given only  to authorized investigators \n[18].  \nStatistical analysis \nData were computed using SPSS Version 26.0 (IBM \nCorporation. Categorical variables (types of \nirregularities, BMI groups, biomarker elevations ) \nwere presented as percentages, means, and \nfrequencies. Pearson Chi-Square (x2) test was used to \ndetermine categorical associations between elevated \nserum levels and observed clinical parameters.  \nExpected cell counts were verified before applying \nthe Pearson chi-square test; where any expected count \nwas below 5 (family history of cancer and the BMI \ncomparison), Fisher's exact test was used , and exact \np-values are reported.  The p-value was set to <0.05 \nfor the data to be statistically significant.  No \ncorrection for multiple comparisons was applied \nbecause the analyses were exploratory and \nhypothesis-generating; p-values should therefore be \ninterpreted with caution. \nResults \nPopulation baseline profile and clinical \nparameters  \nThe mean age of all subjects was 22.4 ± 1.9 years, and \nmost subjects (n = 116, 58.0%) were between 19 and \n22 years. Regarding BMI, 114 participants (57.0%) \nwere within the optimal range, while 58 participants \n(29% of the population) were in the overweight \ncategory. About the menstrual patterns, most of the \nsubjects were dealing with oligomenorrhea (n = 92, \n46.0%), followed by painful cycles of dysmenorrhea \n(n = 72, 36.0%) and mild Polymenorrhea  (n = 36, \n18.0%) (Fig. 1). Moreover, a positive family history \nof subsequent anomalies was also documented in n = \n22 subjects (11.0%) of the cohort under observation \n(Table 1).  \n \n \nFig. 1 Distribution of menstrual irregularity types in the study \ncohort (n = 200). Oligomenorrhea was the most common \nirregularity (46.0%, n = 92), followed by severe dysmenorrhea \n(36.0%, n = 72) and polymenorrhea  (18.0%, n = 36). \nPercentages represent the proportion of the total cohort (n = \n200). Numbers in parentheses indicate the number of \nparticipants in each category. \n\n\n \nScience Letters 2026; 14(2):1326230sl \n4 \n \nTable 1 Anthropometric measures, baseline characteristics and irregularity profile of population under observation. \nVariable Category Frequency (n) Percentage (%) \nAge Group 18–21 years 116 58.0 \n22–25 years 84 42.0 \nBMI Stratification \nUnderweight (<18.5 kg/m²) 28 14.0 \nNormal (18.5–24.9 kg/m²) 114 57.0 \nOverweight (≥ 25.0 kg/m²) 58 29.0 \nDominant Menstrual Irregularity \nOligomenorrhea (>35-day cycles) 92 46.0 \nPolymenorrhea (<21-day cycles) 36 18.0 \nSevere Dysmenorrhea  72 36.0 \nFamily History of Cancer Present (First-degree relative) 22 11.0 \nAbsent  178 89.0 \n \nTable 2 Dysregulation of serum CA-125 and CEA levels among participants (n = 200). \nBiomarker Mean ± SD Reference Cutoff Normal (≤ Cutoff) n (%) Elevated (> Cutoff) n (%) \nSerum CA-125 18.4 ± 11.2 U/mL 35.0 U/mL 186 (93.0) 14 (7.0) \nSerum CEA 1.8 ± 1.1 ng/mL 5.0 ng/mL 195 (97.5) 5 (2.5) \n \nPrevalence and tumor marker quantification \nThe mean serum CEA concentration was 1.8 ± 1.1  \nng/mL, and the mean serum CA -125 concentration \nwas 18.4 ± 11.2 U/mL. Both markers were right -\nskewed; the median (IQR) was 15.6 (10.2 –24.8) \nU/mL for CA-125 and 1.6 (1.0–2.4) ng/mL for CEA, \nwith observed ranges of 4.1 –58.7 U/mL and 0.3–7.9 \nng/mL, respectively. Serum CA -125 was elevated \n(>35.0 U/mL) in 14 participants (7.0%), and serum \nCEA was elevated (>5.0 ng/mL) in 5 participants \n(2.5%) (Table 2).  \nBivariate analysis: categorical association  \nCross-tabulation analysis revealed a statistically \nsignificant association between dysmenorrhea \nseverity and abnormal CA 125 levels (χ² = 6.31, p = \n0.012). Furthermore, elevated CA 125 levels were \npresent in 9 of 72 participants with severe \ndysmenorrhea (12.5%) compared with 5 of 128 \nparticipants with mild/moderate dysmenorrhea \n(3.9%) (χ² = 6.31, p = 0.012). Moreover, serum CA \n125 levels were elevated in participants with higher \nBMI (≥ 25.0 kg/m²), with 7 (12.1%) overweight \nparticipants showing elevated titers compared with 7 \n(4.9%) normal ones ( χ² = 4.31, p = 0.038) . No \nstatistically significant association was observed \nbetween family history of malignancies and  CA 125 \n(p = 0.345) (Table 3).  \nDiscussion \nThe current study documented the serum CEA and \nCA 125 profiles in a cohort of non -hospitalized \nfemale university students aged 18 to 25 years, \npresenting with menstrual irregularities in public and \nprivate higher education sectors in Islamabad and \nRawalpindi. The main goal was to establish screening \nprofiles to determine elevations of subclinical \nbiomarkers with menstrual parameters and \nsociodemographic factors. Because this was a cross -\nsectional study with no imaging or surgical \nconfirmation, the associations reported here are \nunadjusted and cannot establish causality or a specific \nunderlying diagnosis.  \nOur finding s highlighted elevations of serum \nCEA (n= 5) and CA 125 (n = 14) among the cohort. \nIn asymptomatic participants, higher levels of CA 125 \nare primarily driven by non -malignant pelvic \ndisorders rather than cancer -related disorders  [19, \n20]. The elevated levels of CA 125 are secreted by the \nmesothelial linings of the fallopian tubes and \nperitoneum walls in response to abnormal levels of \ncytokines, pelvic remodeling, and microhemorrhage \n(tiny, localized bleeding)  [21]. A critical finding of \nthis investigation is the significant link between \nseverity of dysmenorrhea and higher levels of CA 125 \n(p = 0.012). Females  with extreme, disabling pain \nduring the late stage of the menstrual cycle showed a \n12.5% rate of serum CA 125 elevation  [22-24]. This \nresult aligns w ith reported pathophysiological \nprofiles, highlighting CA 125 as a secondary \nbiomarker for initial screening of adenomyosis, \npelvic inflammatory disorders (PID) , and \nmesothelial-related endometriosis  [25, 26]. \nFurthermore, during early stages of endometriosis, \nendometrial implants are documented to produce a \nlocalized immune response (upregulated cytokines \nand interleukins that support inflammation) during \nlate-stage menstruation, leading to retroperitoneal \nshredding with subsequent vascular permeability , \nallowing CA 125 to move into systemic circulatio n\n\n \nScience Letters 2026; 14(2):1326230sl \n5 \n \nTable 3 Bivariate cross-tabulation analysis of factors associated with serum CA-125 elevation (n = 200). \nClinical Parameter CA-125 Normal (≤ \n35 U/mL) n (%) \nCA-125 Elevated (> 35 \nU/mL) n (%) \nTotal n \n(%) χ² p-value \nDysmenorrhea Severity \n   \n6.31 0.012 \nMild/Moderate (n = 128) 123 (96.1) 5 (3.9) 128 (100) \n  \nSevere (n = 72) 63 (87.5) 9 (12.5) 72 (100) \n  \nBMI Category \n   \n4.31 0.038 \nUnderweight/Normal (n = 142) 135 (95.1) 7 (4.9) 142 (100) \n  \nOverweight (n = 58) 51 (87.9) 7 (12.1) 58 (100) \n  \nFamily History of Cancer \n   \n0.89 0.345 \nPresent (n = 22) 20 (90.9) 2 (9.1) 22 (100) \n  \nAbsent (n = 178) 166 (93.3) 12 (6.7) 178 (100) \n  \nPearson chi-square was used for dysmenorrhea severity; Fisher's exact test was used for BMI category and family history of cancer because  of expected cell counts \nbelow 5 \nallowing CA 125 to move into systemic circulation \n[27]. In unadjusted analysis, elevated CA 125 levels \nwere associated with BMI (p = 0.038); excess adipose \ntissue is metabolically active and secretes adipokines \nand TNF -α, producing chronic low -grade \ninflammation [28, 29], which may explain this \nassociation. Because no multivariable adjustment was \nperformed, this relationship may be confounded by \nage or dysmenorrhea severity [28, 29]. This metabolic \ncondition is closely related to polycystic ovarian \nsyndrome (PCOS), where low-grade inflammation in \nthe pelvis, along with follicular arrest, is associated \nwith low -to-high biomarker fluctuations [30]. In \ncontrast, CEA levels did not surpass the minimum \nthreshold (97.5% reference limit) in the studied \npopulation. This comparatively low prevalence \n(2.5%) of elevated serum CEA levels reflects the \nspecific nature of this antigenic marker for mucinous \ncarcinomas and mucosal lesions in the \ngastrointestinal tract, which are very rare in young \nfemales [10, 31]. Minor variation in CEA levels may \nreflect limited environmental exposure or non -\nspecific dietary factors among the enrolled \nparticipants.  \nFrom the perspective of a laboratory \ntechnologist, these findings reveal the diagnostic \nvalue of integrating enzyme -linked immunosorbent \nassay (ELISA) screening in combination with clinical \nsymptom scoring for better gynecological outcomes. \nDetecting CA 125 elevations on subclinical grounds \nin young non -hospitalized females  with \ndysmenorrhea provides a non -invasive screening \nprotocol for better identification of candidates \nrequiring either gynecological evaluation or pelvic \nultrasonography.   \nLimitations \nThe study has many limitations, including failure to \nestablish a causal relationship over varying time \nframes (cross-sectional study design) and the inability \nto perform ultrasonographic  procedures to better \nunderscore expected conditions like polycystic \novarian syndrome or endometriosis. Prospective \nstudies should focus on multicenter longitudinal \ndesigns that integrate imaging modalities with \nbiomarker-based diagnostics.     \nAcknowledgments \nThe authors thank Dr. Aneela Shaukat, Assistant \nProfessor, and Mr. Dawood Khan, Lecturer, \nDepartment of Allied Health Sciences, IQRA \nUniversity, Chak Shehzad Campus, Islamabad, for \ntheir continuous support in documentation, refining \nthe methodology, and data analysis. The authors are \nalso thankful to the supporting staff and laboratory \ntechnologists of the biochemistry laboratory at Nayab \nLab and City Lab for assisting in the conduction and \ninterpretation of ELISA.  \nConflicts of Interests  \nThe authors share no competing interests in the \npreparation of this manuscript.  \nReferences   \n[1] Taşpınar A, Söyler AK, Günaydın S. A descriptive study \nof the prevalence of dysmenorrhea in university students \nand its relationship with quality of life. 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Dig Dis Sci 2022; 67(9):4541-4548.","source_license":"CC0","license_restricted":false}