The morphological evaluation of tumor-infiltrating lymphocytes (TILs) in breast cancer (BC) is gaining momentum as evidence strengthens for the clinical relevance of this immunological biomarker. Accumulating evidence suggests that the extent of lymphocytic infiltration in tumor tissue can be assessed as a major parameter by evaluation of hematoxylin and eosin (H&E)-stained tumor sections. TILs have been shown to provide prognostic and potentially predictive value, particularly in triple-negative and human epidermal growth factor receptor 2-overexpressing BC. A standardized methodology for evaluating TILs is now needed as a prerequisite for integrating this parameter in standard histopathological practice, in a research setting as well as in clinical trials. This article reviews current data on the clinical validity and utility of TILs in BC in an effort to foster better knowledge and insight in this rapidly evolving field, and to develop a standardized methodology for visual assessment on H&E sections, acknowledging the future potential of molecular/multiplexed approaches. The methodology provided is sufficiently detailed to offer a uniformly applied, pragmatic starting point and improve consistency and reproducibility in the measurement of TILs for future studies.

Purpose: The exact definition of pathologic complete response (pCR) and its prognostic impact on survival in intrinsic breast cancer subtypes is uncertain. Methods: Tumor response at surgery and its association with long-term outcome of 6,377 patients with primary breast cancer receiving neoadjuvant anthracycline-taxane–based chemotherapy in seven randomized trials were analyzed. Results: Disease-free survival (DFS) was significantly superior in patients with no invasive and no in situ residuals in breast or nodes (n = 955) compared with patients with residual ductal carcinoma in situ only (n = 309), no invasive residuals in breast but involved nodes (n = 186), only focal-invasive disease in the breast (n = 478), and gross invasive residual disease (n = 4,449; P < .001). Hazard ratios for DFS comparing patients with or without pCR were lowest when defined as no invasive and no in situ residuals (0.446) and increased monotonously when in situ residuals (0.523), no invasive breast residuals but involved nodes (0.623), and focal-invasive disease (0.727) were included in the definition. pCR was associated with improved DFS in luminal B/human epidermal growth factor receptor 2 (HER2) –negative (P = .005), HER2-positive/nonluminal (P < .001), and triple-negative (P < .001) tumors but not in luminal A (P = .39) or luminal B/HER2-positive (P = .45) breast cancer. pCR in HER2-positive (nonluminal) and triple-negative tumors was associated with excellent prognosis. Conclusion: pCR defined as no invasive and no in situ residuals in breast and nodes can best discriminate between patients with favorable and unfavorable outcomes. Patients with noninvasive or focal-invasive residues or involved lymph nodes should not be considered as having achieved pCR. pCR is a suitable surrogate end point for patients with luminal B/HER2-negative, HER2-positive (nonluminal), and triple-negative disease but not for those with luminal B/HER2-positive or luminal A tumors.

Purpose: Preclinical data suggest a contribution of the immune system to chemotherapy response. In this study, we investigated the prespecified hypothesis that the presence of a lymphocytic infiltrate in cancer tissue predicts the response to neoadjuvant chemotherapy. Methods: We investigated intratumoral and stromal lymphocytes in a total of 1,058 pretherapeutic breast cancer core biopsies from two neoadjuvant anthracycline/taxane-based studies (GeparDuo, n = 218, training cohort; and GeparTrio, n = 840, validation cohort). Molecular parameters of lymphocyte recruitment and activation were evaluated by kinetic polymerase chain reaction in 134 formalin-fixed, paraffin-embedded tumor samples. Results: In a multivariate regression analysis including all known predictive clinicopathologic factors, the percentage of intratumoral lymphocytes was a significant independent parameter for pathologic complete response (pCR) in both cohorts (training cohort: P = .012; validation cohort: P = .001). Lymphocyte-predominant breast cancer responded, with pCR rates of 42% (training cohort) and 40% (validation cohort). In contrast, those tumors without any infiltrating lymphocytes had pCR rates of 3% (training cohort) and 7% (validation cohort). The expression of inflammatory marker genes and proteins was linked to the histopathologic infiltrate, and logistic regression showed a significant association of the T-cell–related markers CD3D and CXCL9 with pCR. Conclusion: The presence of tumor-associated lymphocytes in breast cancer is a new independent predictor of response to anthracycline/taxane neoadjuvant chemotherapy and provides useful information for oncologists to identify a subgroup of patients with a high benefit from this type of chemotherapy.

We have previously shown the prognostic importance of tumor-infiltrating lymphocytes (TILs) in newly diagnosed triple-negative breast cancer (TNBC) using tumor samples from a large clinical trial cohort. In this study, we aimed to validate these findings and also investigate associations with trastuzumab benefit in HER2-overexpressing disease (HER2+). A prospective–retrospective study was conducted using samples from the FinHER adjuvant, phase III trial that enrolled 1010 early-stage BC patients, 778 of whom were HER2-nonamplified. Those with HER2+ disease (n = 232) were randomized to 9 weeks of trastuzumab or no trastuzumab in addition to chemotherapy. Two pathologists independently quantified stromal TILs in 935 (92.6%) available slides. The primary end point of distant disease-free survival (DDFS) and interactions with trastuzumab were studied in Cox regression models. Confirming our previous findings, in TNBC (n = 134) each 10% increase in TILs was significantly associated with decreased distant recurrence in TNBC; for DDFS the hazard ratio adjusted for clinicopathological factors: 0.77; 95% confidence interval (CI) 0.61–0.98, P = 0.02. In HER2+ BC (n = 209), each 10% increase in lymphocytic infiltration was significantly associated with decreased distant recurrence in patients randomized to the trastuzumab arm (DDFS P_interaction = 0.025). Higher levels of TILs present at diagnosis were significantly associated with decreased distant recurrence rates in primary TNBC. These results confirm our previous data and further support that TILs should be considered as a robust prognostic factor in this BC subtype. We also report for the first time an association between higher levels of TILs and increased trastuzumab benefit in HER2+ disease. Further research into why some TN and HER2+ BCs can or cannot generate a host antitumor immune response and how trastuzumab can favorably alter the immune microenvironment is warranted.