254 Training set ER EGFR Her2 CK5/6 Bcl2 Table 3 Correlations between mast cells and other biomarkers Kendall’s tau-b Signiﬁcance (2-tailed) N Kendall’s tau-b Signiﬁcance (2-tailed) N Kendall’s tau-b Signiﬁcance (2-tailed) N Kendall’s tau-b Signiﬁcance (2-tailed) N Kendall’s tau-b Signiﬁcance (2-tailed) N Mast cells 0.034 0.148 1,788 –0.029 0.228 1,646 0.049 0.052 1,746 –0.003 0.906 1,624 0.077 0.002 1,616 N—scorable for both markers All scores binarized as detailed in the text was a favourable prognostic marker in the validation set (P = 0.006). Multivariate analysis Cox proportional hazard model was used to carry out the multivariate analysis and included age, tumor grade, tumor size, nodal status, ER and Her2 as independent predictors of BCSS. All the above variables achieved statistical signiﬁcance as shown in Table 4(b). Presence of MCs did not achieve statistical signiﬁcance (P = 0.128) but retained a similar HR = 0.846, 95% CI 0.683–1.049 as in the training set analysis. X-tile analysis results We assessed the association between patient outcome and number of positively stained MCs in the stroma of tumors using X-tile software. This software allowed us to deﬁne an optimal cut-off point that deﬁned the number of MCs needed to predict good prognosis in the cancer patients. The program divided the population into low and high-risk groups based on the number of MCs detected. It converted continuous data into ordinal classes for statistical analysis. The X-tile plot showed that breast cancer patients with any number of stromal MCs had better prognosis compared to those who have no MC inﬁltration in their stroma. MCs ranging between 1 and 22 in the tumor stroma were present in 507 (27.61%) patients and were grouped together in the low population group of the training set. The high popu- lation group comprised of 1,329 (72.39%) patients in the 123 Breast Cancer Res Treat (2008) 107:249–257 training set. It reiterated the ﬁndings obtained by KM survival analysis that the presence of any number of stro- mal MCs predicts good prognosis in invasive breast cancer patients. Discussion The importance of the reciprocal relationship between tu- mor and stroma is being increasingly recognized [21], and is the role of stromal inﬂammatory cells like MCs, macrophages, ﬁbroblasts and T cell subtypes in cancer initiation and progression. cDNA microarray analysis has shown that genes expressed by stromal cells correlate with differences in the biology of the tumors and are prognostic predictors in breast cancer [22]. There is an ongoing debate about possible detrimental or beneﬁcial effects of MC accumulation in the stroma of solid tumors. In-vitro studies have shown that MC inhibits tumor growth [23]. This is in agreement with other studies showing increase in MC count in early stage of non-small cell lung cancer and thereby supporting their anti-tumor role. [3]. Increased islet/stromal MC-ratio and presence of tumor islet MCs were shown to be independent good prognostic indicators in non-small-cell lung cancer [24]. Recent studies in ovarian cancer [13] and colorectal cancer [15] have also conﬁrmed their correlation with prognosis. MCs secrete factors like heparin, interleukin-8 (IL-8) and vascular endothelial growth factor that promote neo- vascularization; histamine which suppresses immune re- sponse; platelet-derived growth factor, nerve growth factor and stem-cell factors which are mitogenic; and proteases that promote metastasis [9]. On the other hand, MCs can inhibit tumor growth by releasing endogenous peroxidase that is cytotoxic to mammalian tumor cells [25], by natural cytotoxicity [21] and by recruiting neutrophils, eosinophils, lymphocytes and macrophages [26]. MCs can secrete protective substances without degranulation in the presence of tumor-derived blockers like oxidized polyamines and thus be detrimental to the tumor [27]. They can destroy tumor cell surface structures directly and indirectly in a fashion similar to the effect of arginase [28]. MCs can also inhibit tumor growth by secreting beneﬁcial cytokines like IL-1, IL-4, IL-6, and tumor necrosis factor-a that induce apoptosis of endothelial cells [29]; and chondroitin sulphate which inhibits metastases [30]. Cathepsin G secreted by MC activates platelets, lymphocytes and macrophages, and is known to be cytotoxic to some mammalian cells [31]. Its inhibition by a tumor cell product was implicated in the progression of advanced squamous cell tumors [32] suggesting its cytotoxic capa- bility against target cells. MCs secrete chymase, which stimulates apoptosis in different target cells [33] and