Elevation of Serum Riboflavin Carrier Protein in Breast Cancer 1 Prakash N. Rao, 2 Edward Levine, Michael O. Myers, Veena Prakash, James Watson, Alan Stolier, Jeffrey J. Kopicko, Patricia Kissinger, Shailaja G. Raj, and Madhwa H. G. Raj Departments of Medicine [P. N. R., V. P., A. S., J. J. K., P. K.], Surgery [E. L., M. O. M., J. W.], Obstetrics and Gynecology [S. G. R., M. H. G. R.], and Biochemistry [M. H. G. R.], Louisiana State University Medical Center, New Orleans, Louisiana 70112 Abstract Presently available tumor markers have had a limited clinical impact. Riboflavin carrier protein (RCP) is an estrogen inducible protein that occupies a key position in riboflavin metabolism. Because other vitamin carrier proteins (VCP) have been shown to be overexpressed in patients with malignant disease, we evaluated serum RCP levels in patients with adenocarcinoma of the breast. In this prospective blinded study, patients with breast cancer, benign breast disease, and healthy controls were analyzed for RCP levels. Using a highly sensitive RIA, we observed that serum RCP levels were significantly elevated in women with breast cancer (n  52) as compared with control subjects [n  50; 6.06  7.27 ng/ ml versus 0.70  0.19 ng/ml (mean  SD), respectively; P < 0.0001]. A serum RCP level of >1.0 ng/ml was highly predictive of the presence of breast cancer, detecting 88% of tumors in stages I-II and 100% of tumors in stages III-IV. Overall, this RCP assay has a sensitivity of 92.3%, a specificity of 88%, a positive predictive value of 88.9%, and a negative predictive value of 91.7%. These results show increased serum levels of RCP in breast adenocarcinoma patients and suggest that RCP levels may be useful as a new marker for breast cancer. The positive predictive value in early-stage breast cancer suggests that the RCP assay may be a useful adjunct to present screening technology. Introduction Although lung cancer is the leading cause of cancer death in women in the United States, breast cancer remains the leading cause of cancer-related death in women between the ages of 40 and 55 (1). Because early detection, diagnosis, and therapy is associated with greater than 90% survival rate, it is of critical importance that reliable screening techniques be optimized. Riboflavin and related compounds are essential require- ments for cell growth and development. The transport and subsequent deposition of riboflavin in the developing embryo is facilitated by a phosphoglycoprotein called RCP, 3 which binds the protein in a 1:1 molar ratio (2). RCP plays a key role in growth and development. This is underscored by gestational studies in rodents and primates, in which immunological inter- ference of RCP causes an inhibition of riboflavin transport to the developing embryo that results in acute fetal wastage and pregnancy termination (3). In the chicken, the inability to produce RCP is associated with embryonic death in the eggs (4). This vitamin carrier protein has been shown to be an estrogen inducible protein (3). Studies on the role of RCP in the mammary gland have shown that it is synthesized and secreted into the milk by mammary epithelial cells under estrogen and progesterone con- trol (3). Because breast cancer is also estrogen-related, and be- cause other VBPs like FBP (5) and RBP 4 have been shown to be overexpressed in malignant breast tissue, it was of interest to evaluate RCP dynamics in women with breast cancer. We hypothesized that we would observe an increased synthesis and secretion of RCP by the breast cancer cell, and that this up- regulated synthesis would be reflected by elevated levels of the protein in the blood of women with breast cancer. We further hypothesized that the increased synthesis would be reflected in increased immunohistochemical RCP expression in breast tis- sue. Materials and Methods The protocol and patient consent forms for the study were reviewed and approved by the Institutional Review Board of Louisiana State University, New Orleans, LA. A total of 102 women, including 52 with intact breast tumors and 50 control subjects, were studied. The breast cancer patients were consec- utively recruited from the Surgical Oncology clinics at the Louisiana State University Medical Center and included 27 Caucasians and 25 African Americans. The average age of the Caucasian women was 56.5  14.3 years and was similar to that of the African American women at 50.5  10.73 (range for the whole population, 27–79 years). Forty-one of the 52 tumors presented in stages I and II (stage I, n = 11; IIa, n = 20; IIb, n = 10). Of the remaining 11, 6 were in stage IIIa, 4 were in stage IIIb, and 1 was in stage IV. In this study, a similar distribution of tumor stages was observed in Caucasian and African American women. Data on tumor stage and hormone receptor profile form part of the clinical protocol and were obtained from the clinical faculty. Control subjects included 8 women with fibrocystic breast Received 12/10/98; revised 9/1/99; accepted 9/1/99. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 Supported by Grant CA 71743-01 from The National Cancer Institute. 2 To whom requests for reprints should be addressed, at Department of Medicine, Section of Hematology & Oncology, Louisiana State University Medical Center, 1542 Tulane Avenue, New Orleans, LA 70112. Phone: (504) 568-4636; Fax: (504) 568-2127. 3 The abbreviations used are: RCP, riboflavin carrier protein; FBP, folate-binding protein; RBP, retinol-binding protein; ROC, receiver operating curve; VBP, vitamin-binding protein. 4 Unpublished results. 985 Vol. 8, 985–990, November 1999 Cancer Epidemiology, Biomarkers & Prevention on February 2, 2016. © 1999 American Association for Cancer Research. cebp.aacrjournals.org Downloaded from