Elevated Serum βhcg in a 34-Year-Old Woman with Breast Cancer Zhen Ni Zhou, 1 * Joshua Hayden, 2 Timothy M. D'Alfonso, 2 and Kevin Holcomb 3 CASE DESCRIPTION A 34-year-old G5P4014 (5 pregnancies, 4 children born at term, 0 preterm, 1 abortion, and 4 living children) with stage IV triple-positive (ER+/ PR+/Her2+) breast cancer was admitted to the medical oncology service for recurrent shortness of breath and right-sided pleuritic chest pain secondary to disease progression. The patient had widespread metastases to the bone, liver, and lungs and was being treated with docetaxel, trastuzumab, pertuzumab, and zoledronic acid. A gynecology consult was requested after the patient was incidentally noted to have a positive urine human chorionic gonadotropin (hcg) 4 (Beckman Coulter Icon 25) and serum βhcg of 102 miu/ml (Beckman UniCel DxI). There was no evidence of impaired renal function (estimated glomerular filtration rate levels all within normal limits), and no medications that can affect hcg concentration were noted in the patient's chart. The patient's last menstrual period was approximately 1 year ago. She experienced premature ovarian failure because of chemotoxic effects on the ovary. The patient last had unprotected sexual intercourse about 6 months before her presentation. She complained of chronic lower back pain and nausea, which were attributed to chemotherapy, but denied having any vaginal bleeding, spotting, lower abdominal pain, or cramping. There was no evidence of an intrauterine or ectopic pregnancy seen on transvaginal ultrasound. The βhcg remained elevated 2 days after the initial result and increased 4 days later (Fig. 1). At this time, the patient developed vaginal spotting without any accompanying lower abdominal pain or cramps. In light of this new symptom and the inappropriately increasing serum βhcg levels, the patient was counseled regarding the concern for an abnormal pregnancy, ectopic or intrauterine. The risks and benefits of further expectant management (serum βhcg level < 200 miu/ml is associated with an 88% chance of spontaneous resolution) (1) and immediate treatment with methotrexate (MTX) were discussed with the patient; she opted for immediate treatment. The serum βhcg initially decreased after MTX administration but subsequently increased 2 weeks later (Fig. 1). Given the continued concern for an abnormal pregnancy, the patient was retreated with MTX and a similar βhcg trend was observed (Fig. 1). A repeat transvaginal ultrasound examination remained unremarkable. Given the broad differential diagnosis, i.e., early pregnancy (intrauterine and ectopic), false-positive serum βhcg, pituitary secretion of hcg, and malignancy, the patient was given the option to 1 Department of Obstetrics and Gynecology, Weill Cornell Medical College New York Presbyterian Hospital, New York, NY; 2 Departments of Pathology and Laboratory Medicine, Weill Cornell Medical College New York Presbyterian Hospital, New York, NY; 3 Department of Obstetrics and Gynecology, Division of Gynecologic Oncology, Weill Cornell Medical College New York Presbyterian Hospital, New York, NY. *Address correspondence to this author at: 525 East 68th Street, Box 122, New York, NY 10065. Fax 212-746-6656; e-mail zhz9006@nyp.org. DOI: 10.1373/jalm.2017.025288 2018 American Association for Clinical Chemistry 4 Nonstandard abbreviations: hcg, human chorionic gonadotropin; MTX, methotrexate; IHC, immunohistochemistry; ACOG, American College of Obstetricians and Gynecologists. July 2018 03:01 000 JALM 1 Copyright 2018 by American Association for Clinical Chemistry.
Fig. 1. Levels of βhcg (Beckman UniCel DxI) during hospitalization of patient. The arrows indicate administration of MTX (50 mg/m 2 intramuscular). The stars indicate when urine pregnancy tests (Beckman Coulter Icon 25) were performed; these tests were positive on day 1 and negative on day 33. undergo a dilation and curettage or expectant management with serial βhcg measurements. The patient opted for the latter, and the decision was made to hold any further treatment and to follow serum βhcg levels expectantly. Serial measurements of βhcg remained increased with continued follow-up evaluation (Fig. 1). A negative urine pregnancy test on day 33 raised concern for a false-positive serum result. Therefore, the serum sample from day 33 (104 miu/ml) was evaluated further. The βhcg level remained increased (112 miu/ml) following linear dilutions (1:2, 1:5, and 1:10) and treatment with heterophile antibody blocking tubes (Scantibodies ). In addition, increased hcg levels were confirmed with alternative platforms (βhcg: 88 miu/ml, TOSOH Bioscience, AIA-2000; total hcg: 91 miu/ml, Elecsys Cobas hcg+b). In light of the irregular βhcg trend, alternate explanations for the increased and rising βhcg were considered. Case discussion Pregnancies of unknown location (positive pregnancy test with no evidence of ectopic or intrauterine pregnancy) are common diagnostic dilemmas in the field of obstetrics and gynecology; a wide range of potential diagnoses for a persistently positive low-level βhcg exists. False-positive βhcg test results should always be considered when there is a discordance between the patient's clinical findings and laboratory results. Pituitary secretion of hcg is another possibility, especially in perimenopausal and postmenopausal women (2). Less commonly, increased serum βhcg levels are caused by malignancies such as gestational trophoblastic neoplasias, choriocarcinomas, and, in rare cases, nontrophoblastic malignancies. Although these tumors typically produce intact hcg, many nontrophoblastic tumors, such as breast carcinomas, produce only βhcg (3 5). Analytical artifacts, i.e., phantom hcg, should always be considered when a laboratory value is inconsistent with the clinical picture (6). Depending on the source or condition, hcg can be present in the serum in a variety of different forms, such as the intact dimer, free β subunit, or a modified form of the β subunit (7). Commercial assays used to determine the presence and concentrations of serum and urine hcg differ with regard to the specificity of the reagents used to determine the molecular form of hcg present (7). Because these tests are typically designed as sandwich immunoassays, they are susceptible to interferences by heterophile antibodies, resulting in a false-positive test result and unwarranted interventions. Falsely increased serum βhcg levels in particular can occur and lead to unwarranted treatment and harmful consequences for the patient. For this reason, clear guidance has been published on how to rule out possible artifacts, including urine tests, linear dilutions, removal of heterophilic antibodies, and repeat testing on alternative platforms (8). Three of these 4 approaches confirmed that this patient had a true elevation in serum βhcg levels. It is unclear why the urine sample was negative on day 33. False-negative urine hcg tests have been reported that are caused by the variant hook effect (excess of the β core fragment suppresses signal from the 2 JALM 000 03:01 July 2018
βhcg in the urine, leading to a false-negative result) (9). A second possibility is that the urine immunoassay used antibodies with poor crossreactivity to the βhcg variant produced and/or excreted in the urine of this patient such that whatever level was present was below the limit of detection of the assay. In support of this, we analyzed the urine sample on our serum platform by diluting the patient's urine 10-fold with male serum. This returned a positive signal (9 miu/ml), suggesting that the urine did contain βhcg that was not, or was poorly, recognized by the antibodies in the urine immunoassay. Overly dilute urine can also result in false-negative urine hcg tests; specific gravity was not measured on this urine sample and cannot be definitively ruled out. However, the presence of detectable βhcg by a serum assay when the urine was spiked into male serum strongly suggests the negative urine test was not simply because of overdilution. The laboratory evaluation confirmed a true-positive serum βhcg and false-negative urine hcg result on this day. Pituitary secretion of hcg is not an infrequent cause of increased βhcg in perimenopausal and postmenopausal women (2). This explanation was deemed unlikely because the levels of βhcg (>50 miu/ml) far exceeded those generally observed for pituitary βhcg (5 14 miu/ml) (2). Clinical confirmation of a pituitary source of hcg can be made empirically by treating the patient with a 3-week course of oral contraceptives; this intervention suppresses release of gonadotropin-releasing hormone and, in turn, pituitary hcg production. Unfortunately, the patient died from complications of metastatic breast cancer, and empiric oral contraceptive therapy was not performed. Age and levels of follicle-stimulating hormone can also be helpful in determining pituitary hcg secretion (2); however, the applicability of this algorithm to younger female patients who have undergone premature ovarian failure is unclear. Fig. 2. Needle core biopsy of primary tumor (A) and IHC showing negative staining for βhcg (B). Core biopsy showed invasive ductal carcinoma, poorly differentiated (A). IHC staining for βhcg was negative (B). The βhcg was from Leica Biosystems; clone PA0014 polyclonal and placenta was used as a positive control. The final diagnostic consideration, which we believe is the most likely explanation, is malignancy. Previous studies have shown that serum from patients with nontrophoblastic tumors displays immunoreactivity to hcg (3, 4). In particular, metastatic breast carcinomas have been shown to express βhcg in up to 60% of patients (5). Immunohistochemical (IHC) staining for βhcg expression was performed on a core needle biopsy of this patient's breast cancer obtained at the time of initial diagnosis and was negative (Fig. 2). Unfortunately, whether the metastatic lesions or the pituitary could be the source of ectopic βhcg production could not be confirmed secondary to the patient's death. Although the IHC for hcg expression performed on the patient's breast cancer was negative, the metastatic lesions could have been responsible for the ectopic production of βhcg. This would explain the high βhcg serum level, the temporary decrease following MTX administration, the negative IHC of the primary lesion, and the discordance in the serum and urine immunoassays (βhcg produced by malignancy has shown greater molecular heterogeneity and, thus, would be expected to show greater discrepancies between antibody-based immunoassays) (10). In summary, there are many potential explanations for increased hcg, including both benign and July 2018 03:01 000 JALM 3
malignant diagnoses as well as analytic artifacts. When there is a discrepancy between the patient's clinical presentation and laboratory results, the laboratory plays a crucial role in assisting the physician by investigating the unexpected test results. Although current American College of Obstetricians and Gynecologists (ACOG) Committee Opinion acknowledges the presence and subsequent treatment of patients with false-positive hcg test results, our case report illustrates specific circumstances whereby patients can have positive hcg tests that are discordant with clinical findings and are not analytic artifacts (8). Although we were unable to definitively identify the source of hcg, the fact that the patient's hcg was increased on 3 platforms (Beckman, TOSOH, and Roche), after Scantibodies treatment, and when diluted linearly all support the conclusion that the positive hcg was not an analytic artifact. Contrary to the ACOG Committee Opinion, a negative urine hcg result in this patient did not definitively identify an analytic artifact. Laboratorians should be aware that not all unexpected βhcg tests are analytic TAKEAWAYS The most common causes of persistent low levels of βhcg include early pregnancy (intrauterine or ectopic), physiologic artifact ( phantom hcg ), pituitary hcg, and active or quiescent gestational trophoblastic disease. A negative urine test finding is not definitive evidence of a false-positive serum result; true-positive serum βhcg and negative urine hcg results can be observed. Malignancy, although an uncommon cause of persistent low elevations of βhcg, needs to remain high in the differential diagnosis, especially for patients with known carcinoma. artifacts. True elevations owing to nontrophoblastic malignancy are possible and should be kept in mind. Author Contributions: All authors confirmed they have contributed to the intellectual content of this paper and have met the following 4 requirements: (a) significant contributions to the conception and design, acquisition of data, or analysis and interpretation of data; (b) drafting or revising the article for intellectual content; (c) final approval of the published article; and (d) agreement to be accountable for all aspects of the article thus ensuring that questions related to the accuracy or integrity of any part of the article are appropriately investigated and resolved. Authors Disclosures or Potential Conflicts of Interest: Upon manuscript submission, all authors completed the author disclosure form. Employment or Leadership: Z.N. Zhou, T.M. D Alfonso, K. Holcomb, and J. Hayden, New York-Presbyterian/Weill Cornell Medical College. Consultant or Advisory Role: None declared. Stock Ownership: None declared. Honoraria: None declared. Research Funding: None declared. Expert Testimony: None declared. Patents: None declared. Other Remuneration: New York Presbyterian Hospital, Weill Cornell Medical. Role of Sponsor: The funding organizations, New York Presbyterian Hospital and Weill Cornell Medical, had a role in the review and interpretation of data, and preparation and final approval of the manuscript. REFERENCES 1. American College of Obstetricians and Gynecologists. ACOG practice bulletin no. 94: medical management of ectopic pregnancy. Obstet Gynecol 2008;111:1479 85. 2. Snyder J, Haymond S, Parvin C, Gronowski A, Grenache D. Diagnostic considerations in the measurement of human chorionic gonadotropin in aging women. Clin Chem 2005;51:1830 5. 3. Stenman UH, Alfthan H, Hotakainen K. Human chorionic gonadotropin in cancer. Clin Biochem 2004;37:549 61. 4. Agnantis NJ, Patra F, Khaldi L, Filis S. Immunohistochemical expression of subunit beta HCG in breast cancer. Eur J Gynaecol Oncol 1992; 13:461 6. 5. Tormey DC, Waalkes TP, Simon RM. Biological markers in 4 JALM 000 03:01 July 2018
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