The current goal of prognostic testing in early-stage, estrogen receptor-expressing breast cancer is to provide information to aid in the choice between using hormonal therapy alone or hormonal therapy in conjunction with cytotoxic chemotherapy. Prior clinical studies with unstratified patient populations have not established whether there is a clinical benefit by adding adjuvant chemotherapy to tamoxifen in estrogen receptor-expressing patients >60 years old (1
). In general, there is a bias in younger patients, who have overall worse prognosis relative to older patents, to treat with cytotoxic chemotherapy. Cytotoxic chemotherapy is used more cautiously in elderly or frail patients with otherwise limited expected life expectancy.
This work shows that a prognostic model that uses immunohistochemistry staining using five monoclonal antibodies combined in a multivariate index assay is able to identify a group of estrogen receptor-expressing, tamoxifen-treated, node-negative patients who have a relatively poor prognosis and a greater absolute benefit from chemotherapy compared with unstratified patient populations. In the study using samples from B-20 adjuvant cytotoxic chemotherapy trial, high-risk patients and low-risk patients both showed an absolute benefit from chemotherapy; however, the magnitude of benefit in high-risk patients was four times greater than in low-risk patients. The intermediate-risk group was not well separated from the low-risk group in these studies and therefore, with current data, is uninformative for managing patients classified as intermediate risk. These studies were blinded, prospectively designed retrospective trials in which no specimens from the test cohorts were used in the development of the test.
The strong contribution of age to multivariate models containing the immunohistochemistry prognosticator prompted us to explore in more detail the strength of the immunohistochemistry model in different age strata. The training cohort for the five-antibody test was composed of 83% of patients >50 years old, whereas both the B-14 and B-20 clinical trial cohorts were significantly younger (69% and 55%, respectively). Younger patients in the low-risk group identified by this immunohistochemistry test had a 20% risk of disease progression that warrants consideration of aggressive treatment strategies, consistent with current clinical practice. In contrast, in patients of ages ≥60 years with whom cytotoxic chemotherapy is currently used much more cautiously, the test identified high-risk patients with a 22% risk of breast cancer-specific death compared with 6% in low-risk patients. This, combined with the absolute 21% decrease in recurrence rate associated with the administration of adjuvant chemotherapy identified in the high-risk patient strata in the B-20 study, suggests that elderly high-risk patients may gain more clinical benefit relative to low-risk patients from accepting the risk and morbidity of cytotoxic chemotherapy. It should be noted that stratification into age groups was not a prespecified analysis in the trial design and therefore should be confirmed in additional studies.
The B-14 and B-20 paraffin-block trial samples were also used in the development and validation of the 21-gene predictor, OncoTypeDX
). This test uses preparation of RNA from macrodissected paraffin-block sections and RT-PCR of prepared RNA to detect levels of gene expression. Because 20% and 30%, respectively, of blocks were depleted by the RT-PCR analysis before this immunohistochemistry study, a direct comparison of results is not possible. However, a number of interesting trends are worth noting. The two overlapping studied cohorts had comparable 10-year recurrence rates overall, but the RT-PCR test identified a low-risk group with modestly better outcomes compared with the immunohistochemistry test (7% for the molecular test compared with 11% for the immunohistochemistry test). The difference in outcomes seems to be biased toward differences in performance in younger patients. For example, in patients >60 years of age, the RT-PCR and immunohistochemistry cohorts had very similar recurrence rates before risk stratification, and the tests done comparably in assigning patients to low-risk strata (low-risk distant recurrence-free interval of 7.2% for the immunohistochemistry test compared with 7.5% for the 21-gene test), whereas in younger patients, the RT-PCR cohort had lower rates of tumor progression in its low-risk group. Thus, a difference in performance in younger patients seems to account for much of the overall difference in performance of the two tests. Because epidemiologic work suggests that breast tumors that occur in premenopausal as opposed to postmenopausal patients are biologically distinct and the immunohistochemistry test was trained predominantly on postmenopausal patients, it is possible that the immunohistochemistry test is best suited to postmenopausal patients (15
). Whether the difference in performance in younger patients is due to such intrinsic biological differences between the two tests or to biased depletion of cohort samples between the molecular work and tissue array construction will require additional trials wherein more comparable cohorts are studied.
Few previous studies have evaluated biomarkers in randomized clinical trial populations relative to their utility in prediction of chemotherapy benefit. Only patient age, tumor size, and tumor grade have gained widespread use as markers of prognosis. There is relatively strong evidence to support the use of immunohistochemistry markers of proliferation (e.g., Ki67/MIB-1) and measurements of hormone receptor status (e.g., quantitative estrogen receptor and progesterone receptor) as prognosticators for the natural history of hormonally treated breast cancer (2
). The utility of additional diagnostic testing hinges on both the clinical effect of the result and the technical reliability of the test. Every technology applied to diagnostic testing, whether it is morphologic, immunologic, or molecular based, requires demonstration of the analytic reproducibility and clinical effect of testing. Morphology and immunohistochemistry have mixed records as diagnostic tests, but both have shown remarkably good reproducibility, when done by skilled specialists and in experienced high-volume laboratories. Immunohistochemistry-based assays have the advantage over molecular assays in that detection of biomarker expression is done and evaluated in situ
, which gives confidence that measurement of expression is not confounded by expression in a variable admixture of normal or benign breast tissue and/or contaminating inflammatory cells and stroma. The ~85% to 95% reproducibility of the replicate stains and stain interpretation in this study using the very limited tissue available in 600-μm tissue array cores suggest that these immunohistochemistry biomarkers are technically robust markers. Assay performance should improve further under clinical practice conditions wherein whole sections are available for interpretation (16
). In addition, standardization of laboratory practice, automated staining, and quantitative image analysis approaches promise to improve laboratory consistency and performance of immunohistochemistry (17
). Information from biomarker expression in situ
such as offered by immunohistochemistry assays may be prudent in the setting of limited tissue such as that available through the use of needle or core biopsies, which are becoming more common as diagnostic specimens with the emergence of neoadjuvant approaches to treating breast cancer.
The demonstration that five immunohistochemistry antibodies distinguish breast cancer patients at high risk of recurrence and who seem to have a robust response to chemotherapy suggests that this test may be useful for helping manage early-stage, estrogen receptor-expressing breast cancer. Patients identified as high risk for recurrence may be more inclined to elect aggressive treatment approaches, whereas patients predicted to have low risk of tumor progression may elect to forgo the morbidity associated with cytotoxic therapy because their baseline prognosis is very good although they may still derive some benefit from chemotherapy. Further testing in postmenopausal patients as well as exploration of additional markers selectively on premenopausal patients should be done to continue to tailor the test to accommodate the recognized biological and clinical diversity of breast cancer patients. Because immunohistochemistry technology is widely distributed and relatively inexpensive, this five-antibody test may be a cost-effective alternative to molecular-based tests. Given the cost advantage, the ease of testing, and the confidence engendered by in situ visualization of marker expression, immunohistochemistry approaches will likely continue to have a significant role in managing decision making for early-stage breast cancer.