We have previously identified two stromal signatures that are present in a subset of breast cancers that correspond to clinical-pathologic features: the DTF fibroblast response signature, associated with lower tumor grade and increased survival(6
), and the CSF1 macrophage response signature, associated with decreased survival among low-grade tumors(5
). We have found that these signatures are also present in pre-invasive cancer, i.e. ductal carcinoma in situ(16
). A number of recent studies have identified stromal signatures that predict response to chemotherapy (15
). We attempted to further understand the biology of stromal response by asking the following question: is the stromal response unique to the tumor, or is the stromal response an inherent reaction of the host to tissue alteration?
We approached this question by studying the variability in the stromal responses in clinically distinct tumors from the same patient. If the stromal response is dependent primarily on the tumor, we would not expect significant stromal response concordance between two separate tumors from the same patient. Therefore, in the first part of our study, we compared stromal responses to synchronous breast primaries in the same patient to determine whether stromal response to cancer is patient-dependent or tumor-provoked. If the stromal response is primarily dependent on the host, we would see a significant concordance in stromal responses between the two cancers from the same patient. In contrast, if the stromal response is largely dependent on the inciting tumor, we would not expect to see significant concordance in stromal responses from cancers derived from the same patient. In the second part of our study, we proceeded to ask whether the stromal response is a generalized response to tissue injury, or a specific response generated in reaction to the inciting tumor. We addressed this question by examining stromal signatures in normal breast, biopsy site changes, and carcinoma. If the stromal response is a tumor-specific reaction, we would anticipate significant differences in stromal signature response between biopsy site changes and carcinoma. If the stromal response is a more generalized response to tissue injury, we would anticipate significant differences in stromal signature response between normal tissue and biopsy site changes, but negligible differences between biopsy site changes and carcinoma. We then studied stromal response to biopsy site changes and carcinoma to delineate tumor-specific response from a more generalized tissue injury response. In doing so, we found that the two stromal signatures have divergent associations.
Our studies indicate that the DTF fibroblast response signature is significantly conserved in patients with paired breast primaries. In examining stroma taken from normal breast and biopsy site changes, we find that the response to biopsy site changes is similar to the response to carcinoma, and conclude that DTF fibroblast response is a more generalized response to tissue disruption and injury that appears to be specific to individual patients. This suggests that the DTF fibroblast response is a host-specific response and that the ability to generate this response varies between patients.
In contrast to the DTF fibroblast response, which showed significant concordance within a patient, the CSF1 macrophage response showed no significant concordance within patients with paired breast primaries. However, we cannot exclude that in a larger patient cohort, the CSF1 macrophage response signature may or may not show significant correlation as a couple of the markers within the signature show correlation within a patient. Because the response is increased in association with higher tumor grade, it may be that the CSF1 macrophage response is at least in part invoked by tumor.
These findings are in keeping with a separate study in our group on colon and breast cancer metastases, which demonstrated that DTF stromal expression is dependent on the tissue type (lymph node versus colon), while CSF1 stromal expression is conserved within the same tumor regardless of tissue background(17
). This finding complements our conclusions from studying breast cancer, in that DTF appears to be not only patient specific, but tissue specific, while CSF1 response may be dependent on the individual tumor characteristics.
We have demonstrated two divergent stromal responses to breast cancer by studying marker expression in patients with synchronous breast primaries. A potential confounding factor would be if a patient developed two biologically similar tumors that would then induce the same stromal response. To evaluate whether epithelial markers were more conserved in paired primaries than would be expected by chance, we analyzed the paired breast primaries for conventional tumor biomarkers (ER, PR, Her2neu, and Ki-67) and tumor characteristics (histologic subtype, tumor grade, size, laterality). Of these features, the only one that attained statistical significance for concordance in paired primaries was Ki67 (p= 0.014), suggesting that proliferative index may be conserved in dual primaries. Due to the relatively small sample size and small amount of total variation in ER, PR and Her2neu across all the samples, our study was underpowered for identifying statistically significant conservation of these markers among paired primaries. We did not find any evidence of significant conservation of these makers, but we note that this hypothesis needs to be evaluated in a larger study with higher proportions of hormone receptor negative and Her2 positive patients. Taken together, our findings are consistent with evidence in the literature suggesting that dual breast primaries show distinct patterns of molecular alterations, based on molecular studies (18
), histologic subtype and hormonal receptor status (20
), and by cytogenetics (23
The interaction of cancer and stroma is reminiscent of the seed and soil theory proposed by Stephen Paget in 1889 to explain the preferential metastases of breast cancer to sites such as bone(24
). In the case of cancer treatment, therapy has predominantly relied on attacking the “seed,” i.e. the epithelial component. Study of the “soil,” or the stroma, allows for understanding of the tumor microenvironment and opens up possibilities for a different direction of therapy. The study of the milieu in which breast cancer is situated is challenging, as the stromal response may be a product of tumor provocation as well as the patient's innate healing mechanism. In this paper, we have attempted a better understanding of this “soil,” in hopes that knowledge of the stromal response can in the future lead to novel treatments which create less fertile grounds for tumor growth.
Statement of Translational Relevance
This study examines the coincidence of specific stromal response signatures synchronous breast cancers. The findings provide insight towards understanding how these stromal responses arise in breast cancer and may lead to targeted therapies in the future.