Serum markers in breast cancer: clinical usefulness and possible improvements.

Introduction

In contrast to tissue-based markers [1,2], serum-based markers in breast cancer have received relatively little attention. Nevertheless, a number of serum-based markers are in use as indicators for breast cancer. These include carcinoembryonic antigen (CEA), tissue polypeptide antigen (TPA), tissue polypeptide-specific antigen (TPS) and human epidermal growth factor receptor 2 (HER2; the extracellular domain). Of these, the most widely used are CA 15-3 and CEA (for review see [3,4]).

In this article, we will review the current status of serum markers in breast cancer and speculate on likely developments in this area.

Use of serum markers in screening and aiding early diagnosis of breast cancer

Lack of sensitivity and specificity preclude the use of all existing serum markers for the early detection of breast cancer. Women with apparently localised breast cancer who present with high pre-operative marker levels (e.g. > 50 kU/l) are likely to have advanced disease [5] and should undergo appropriate investigations to diagnose or exclude this possibility. Low marker levels, however, do not exclude advanced disease. Whereas existing serum markers have no use in detecting early breast cancer, some emerging findings suggest that the use of proteomics [6-8] or auto-antibodies to breast-associated proteins [9] may be superior to existing markers in women with early breast cancer. It is important to stress, however, that these findings are preliminary and have not been validated in external settings.

Use of serum markers for determining prognosis in breast cancer

Most research on prognostic markers in breast cancer has been carried out on tumour tissue. However, serum markers have a number of potential advantages compared to tissue prognostic factors. Firstly, unlike tumour tissue which requires either biopsy or surgery, blood can be obtained with minimal inconvenience to patients. Secondly, automated, relatively cheap and standardised assays are available for a number of serum markers. Thirdly, serum-based markers can be determined in patients with small tumours including those with in situ cancers, whereas for tissue-based markers, especially if cell-free extracts of freshly-frozen tissue are necessary, patients with very small tumours cannot be assessed. Finally, serum marker determinations can be carried out at several stages during the management of patients. Thus, markers can be determined pre-operatively, postoperatively, after adjuvant therapy, at the time of recurrence and during treatment for recurrent disease.

Several studies have shown that elevated preoperative levels of either CA 15-3 or CEA are associated with poor outcome in patients with breast cancer [1]. Although, different assays and cut-off points for CA 15-3 have been used (25-40 kU/l), the conclusion from almost all studies was that high levels at initial presentation were associated with poor patient outcome. Indeed, in some studies, the prognostic impact of CA 15-3 was found to be independent of tumour size and lymph node status 10-12]. Of significance, CA 15-3 levels were found to be prognostically useful in two reports [11,12] in lymph node-negative breast cancer patients, one of the subgroups of patients for whom new prognostic factors are most urgently required. Based on these findings, a pre-operative CA 15-3 level might be combined with existing prognostic factors in planning the optimum management of patients with newly diagnosed breast cancer.

Use of serum markers for surveillance following surgery for breast cancer

Following surgery for breast cancer, it is now common practice to follow up patients on a regular basis with clinical and radiological examinations, and tumour marker determinations. This practice is based on the belief that the early detection of recurrent or metastatic disease, followed by treatment, improves the chance of cure, or results in an improved survival. From a biological point of view, it might be expected that the early detection of recurrent disease, followed by the iniation of therapy, would improve outcome compared with starting therapy when recurrence/metastasis is clinically evident. There is, however, little evidence available to support this hypothesis. Indeed, two large multicentre randomised prospective trials (each with more than 1000 patients) compared outcome in patients followed-up with clinical visits and mammography versus follow-up with an intensive regimen that included radiology and traditional laboratory testing [13,14]. The conclusion from both of these studies was that use of an intensive follow-up programme failed to improve patient outcome.

It is important to point out that this work was published almost 15 years ago. Thus, its relevance to current clinical practice is unclear [3]. In the meantime, more sensitive imaging procedures such as computed (CT) and positron emission tomography (PET), and magnetic resonance imaging (MRI) have become available. Furthermore, currently used markers such as CA 15-3 and CEA are more sensitive than standard biochemical tests for detecting early metastasis. These markers were not used in the two randomised trials mentioned above. Of more importance, in the last 15 years we have seen major advances in treatment for breast cancer such as the use of the third-generation aromatase inhibitors, taxanes and trastuzumab. However, one needs to question whether research carried out 15 years ago should dictate policy on breast cancer surveillance today. In an ideal situation, the type of randomised controlled trial carried out in the early 1990s should be repeated, this time with the inclusion of markers such as CA 15-3 and CEA. Ethical issues, however, would now make it difficult to conduct such a study.

In the absence of evidence-based information, guidelines vary regarding the use of markers in postoperative surveillance of asymptomatic women following a diagnosis of breast cancer. For example, the American Society of Clinical Oncology (ASCO), the European Society of Medical Oncology and the European Society of Mastology (EUSOMA) recommend against the use of markers in the routine surveillance of patients following primary treatment for breast cancer [5,15-17]. By contrast, the European Group on Tumour Markers (EGTM) recommends the use of markers during follow-up [18]. The National Academy of Clinical Biochemistry (NACB) (USA) recommends against routine CA 15-3 (or BR 27.29) testing in asymptomatic patients following diagnosis of operable breast cancer:

'The Panel, however, would like to note that there are a number of small studies suggesting that the early initiation of therapy based on increasing serum markers levels can lead to an enhanced outcome. Although these studies do not provide high-level evidence that early treatment based on rising tumor marker levels positively impacts on patient outcome, some doctors as well as some patients may wish to have serial levels of CA 15-3 (or BR 27.29) determined following primary surgery. The ultimate decision about whether or not to use CA 15-3 (BR 27.29) in this situation must be taken by the doctor in consultation with the patient' (from [19]).

Monitoring therapy in advanced disease

Following the commencement of therapy for advanced disease, it is important to know as early as possible whether the patient is responding. If the patient is benefiting, then clearly treatment should be continued. If, on the other hand, treatment is not effective, an alternative therapy might be given. If an alternative therapy is unavailable, these patients might be willing to participate in clinical trials of new therapies or they could decide to avoid further therapy.

A convenient and relatively inexpensive approach for helping to establish response is to measure serum markers such as CA 15-3 or CEA. In general, decreasing marker levels correlate with tumour regression, whereas increasing marker levels correlate with tumour progression [20-23]. According to the EGTM guidelines [18], markers should be measured prior to every chemotherapy course and at least at 3-monthly intervals for patients receiving hormone therapy. The EGTM defines an increase in marker concentration of at least 25% to be significant [18]. It is recommended that such an increase be confirmed with a second test within 1 month. If the increase is confirmed, this provides evidence of progressive disease. Similarly, it has been suggested that a confirmed decrease in serum levels of more than 50% is consistent with tumour regression [18].

In contrast to the EGTM recommendations, the ASCO guidelines state that neither CA 15-3 nor CEA should be routinely used for monitoring therapy in patients with advanced breast cancer [5,15]. However, this panel also stated:

'... that in exceptional circumstances such as the presence of osseous metastasis, which are difficult to evaluate clinically, the marker level may be able to support the clinical estimate of disease status. However, the marker cannot in any situation stand alone to define response to treatment' (from [5]).

EUSOMA also recommends against the general use of serum markers for monitoring therapy in advanced breast cancer [17]. However, similar to the ASCO guidelines, the EUSOMA guidelines stated that:

'... in the absence of evaluable disease, increase in tumor marker accompanied by an increase in symptoms (e.g. bone pain) should be taken as indicating disease progression'. '... an increase in serum markers without symptoms of progression should prompt a complete work-up to investigate for progression of known disease sites or appearance of new sites' (from [17]).

Although most changes in serum markers are due to increasing or decreasing tumour bulk, it is important to bear in mind that transient increases in marker concentrations may occur, especially following the start of therapy. The spurious increases or spikes are probably due to therapy-mediated apoptosis or necrosis of tumour cells and not due to tumour progression [3]. Thus, Hayes et al. [24] found a spike for either CA 15-3 or CEA in seven out of 16 patients undergoing chemotherapy. For CA 15-3, the peak of the spike above the initial value was 125% (range, 30-230%) and its duration was 67 days (range, 31-101 days). All patients in whom a spike was observed ultimately either showed disease regression or had stable disease. In another study, spikes in CA 15-3 and CEA returned to pre-treatment levels by 60 days of chemotherapy [25]. As with cytotoxic chemotherapy, treatment with granulocyte colony-stimulating factor (G-CSF) can also cause transient elevations in CA 15-3 concentrations [26]. This increase appears to be due to enhanced expression of the mucin-like protein MUC-1 (i.e. the molecule detected in the CA 15-3 test) in peripheral blood neutrophils caused by G-CSF.

Potential new markers for breast cancer

As well as sensitivity and specificity, a desirable property of a cancer serum marker is organ specificity. Currently, only two organ-specific tumour markers are available [i.e. prostate-specific antigen (PSA) for prostate cancer and thyroglobulin for differentiated thyroid cancer]. All the serum markers currently used for breast cancer lack breast specificity and can be elevated in serum from patients with most types of adenocarcinoma, especially in patients with advanced disease [3].

In recent years, however, a number of proteins have been described that are expressed almost exclusively in breast tissue including breast cancer. These include mammaglobin A, lipophilin B, NY-BR-1, B726P and small breast epithelial mucin (SBEM) [27-33]. The challenge now is to devise sensitive and specific assays for measuring these proteins in serum followed by determination of their potential value in breast cancer.

Will serum markers for breast cancer improve?

As mentioned above, the main problem with all existing serum markers for breast cancer is lack of sensitivity for early disease and lack of specificity for breast cancer. Clearly, new markers must offer improved sensitivity and specificity. One of the most promising approaches in this respect is the use of proteomics. In recent years, there have been a number of claims of detection of breast cancer with relatively high sensitivity and specificity using this technology [6-8].

In 2002, Li et al. applied the SELDI-TOF MS approach to proteomics [6] to serum from 103 patients with breast cancer, 25 patients with benign breast disease and 41 healthy women. Three peaks were selected based on their collective contribution to the optimum separation between patients with early-stage malignancy (i.e. stages 0 and 1) and healthy controls. These three peaks were designated as BC1, BC2 and BC3. Later, BC2 and BC3 were identified as complement [C3a.sub.des] Arg and truncated complement [C3a.sub.des] Arg, respectively [6]. Multivariate logistic regression was then used to combine the three different peaks in order to form a single composite index. Cross-validation using bootstrap analysis resulted in a sensitivity of 97% and a specificity of 91% for breast cancer. Other investigators have also reported that proteomics can differentiate between patients with early breast cancer and controls, with considerably higher sensitivities and specificities than existing markers [7,8]. In contrast to these promising reports, using proteomics Karsan et al. [34] were unable to differentiate between patients with early breast cancer and controls. Clearly 'the jury is still out' as to the value of proteomics for the early detection of breast cancer. Finally, if the relatively breast-specific proteins mentioned above such as mammaglobin A, lipophilin B, NY-BR-1, B726P and SBEM could be detected in serum, they also have the potential to provide a new generation of serum markers for breast cancer.

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Michael J Duffy (1,2), Denis Evoy (2) and Enda McDermott (2)

(1) Department of Pathology and Laboratory Medicine, St Vincent's University Hospital, Dublin, Ireland and (2) School of Medicine and Medical Science, Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Ireland

Correspondence to: Michael J Duffy, Nuclear Medicine Laboratory, St Vincent's University Hospital, Elm Park, Dublin 4, Ireland. (email: [e-mail])