Prognostic significance of FOXP3+ tumor-infiltrating lymphocytes in breast cancer depends on estrogen receptor and human epidermal growth factor receptor-2 expression status and concurrent cytotoxic T-cell infiltration.

INTRODUCTION: The infiltration of FOXP3+ regulatory T cells into invasive tumors has been reported to be associated with survival in a variety of cancers. The prognostic significance of FOXP3+ tumor-infiltrating lymphocytes (TILs) in breast cancer, however, remains controversial. METHODS: FOXP3+ TILs were assessed by immunohistochemistry on tissue microarrays constructed from a well-defined cohort of 3,992 breast cancer patients linked to detailed demographic, biomarker, treatment and outcome data. Survival analyses were performed using the Kaplan-Meier function and Cox proportional hazards regression models to evaluate the association of FOXP3+ TILs with breast cancer-specific survival, stratified by intrinsic subtype and cytotoxic T-cell infiltration status (as defined by CD8 immunohistochemistry). RESULTS: The presence of high numbers of FOXP3+ TILs was significantly associated with young age, high grade, estrogen receptor (ER) negativity, concurrent CD8+ cytotoxic T-cell infiltration, and human epidermal growth factor receptor-2 positive (HER2+)/ER- and core basal subtypes. On multivariate survival analysis, a high level of FOXP3+ TILs was significantly associated with poor survival in ER+ breast cancers that lacked CD8+ T-cell infiltrates (hazard ratio (HR) = 1.30, 95% confidence interval (CI) = 1.02 to 1.66). However, in ER- breast cancers, FOXP3+ TILs were strongly associated with improved survival in the HER2+/ER- subgroup, particularly in those with co-existent CD8+ T-cell infiltrates (HR = 0.48, 95% CI = 0.23 to 0.98), for which the presence of high levels of FOXP3+ TILs was independent of standard clinical prognostic factors. CONCLUSIONS: FOXP3+ regulatory TILs are a poor prognostic indicator in ER+ breast cancer, but a favorable prognostic factor in the HER2+/ER- subtype. The prognostic value of FOXP3+ TILs in breast cancer differs depending on ER and HER2 expression status and CD8+ T-cell infiltration.

Immunohistochemical characterization of prototypical endometrial clear cell carcinoma--diagnostic utility of HNF-1ß and oestrogen receptor.

AIMS: The great majority of ovarian clear cell carcinomas have a hepatocyte nuclear factor 1 homeobox B (HNF-1ß)-positive and oestrogen receptor (ER)-negative immunoprofile. However, the pattern of HNF-1ß and ER immunostaining in clear cell carcinomas of the endometrium and the usefulness of this panel in distinguishing clear cell carcinoma from other histological types of endometrial carcinoma have yet to be well defined. METHODS AND RESULTS: We examined the immunostaining patterns of HNF-1ß, ER and p53 in 15 morphologically classic pure endometrial clear cell carcinomas, and compared these patterns with 15 endometrioid and 15 serous carcinomas of the endometrium. We observed the presence of diffuse (>70%) moderate to strong nuclear HNF-1ß staining and negative ER staining in 14 of 15 clear cell carcinomas, with the remaining case showing both diffuse strong nuclear HNF-1ß staining and focal ER staining. In comparison, only one of 15 serous carcinomas and none of 15 endometrioid carcinomas showed a combination of diffuse moderate to strong HNF-1ß nuclear staining and negative ER staining. Aberrant p53 immunostaining was observed in five of 15 (33%) clear cell carcinomas. CONCLUSIONS: Overall, our findings demonstrate that, similarly to the situation for the ovary, a diagnostic panel of HNF-1ß and ER may be considered for separating clear cell carcinoma from endometrioid and serous carcinoma of the endometrium.

Immunohistochemical survey of mismatch repair protein expression in uterine sarcomas and carcinosarcomas.

Uterine sarcomas and carcinosarcomas are an aggressive group of uterine malignancies. The frequency of mismatch repair (MMR) protein loss by immunohistochemical evaluation has not been comprehensively characterized in this group of tumors; hence, the appropriateness of applying an immunohistochemical panel to screen for Lynch syndrome in these tumors remains unclear. We examined for the immunohistochemical loss of 4 MMR proteins (MLH1, MSH2, MSH6, and PMS2) in a series of 67 uterine carcinosarcomas and 51 uterine sarcomas (20 leiomyosarcomas, 11 adenosarcomas, 9 low-grade endometrial stromal sarcomas, 8 high-grade endometrial stromal sarcomas/undifferentiated endometrial sarcomas, and 3 rhabdomyosarcomas) at our institution. Four of the 67 (6.0%) carcinosarcomas demonstrated abnormal MMR protein expression. Two tumors showed concurrent loss of MLH1 and PMS2 in both the carcinomatous and sarcomatous components. One tumor showed the loss of only PMS2 in both components. The remaining tumor showed an isolated loss of MLH1 and PMS2 in only the small cell carcinoma component, whereas the non-small-cell carcinoma and sarcoma components demonstrated normal staining patterns for MMR proteins. Two of 20 leiomyosarcomas (10%) showed the loss of MMR proteins: one with loss of PMS2 and the other with loss of MSH2 and MSH6. All other uterine sarcoma types examined showed intact MMR protein expression. These observations provide a basis for MMR protein screening in uterine carcinosarcomas and leiomyosarcomas but not in other types of uterine mesenchymal or mixed epithelial/mesenchymal malignancies.

A survey of immunohistochemical biomarkers for basal-like breast cancer against a gene expression profile gold standard.

Gene expression profiling of breast cancer delineates a particularly aggressive subtype referred to as 'basal-like', which comprises ∼15% of all breast cancers, afflicts younger women and is refractory to endocrine and anti-HER2 therapies. Immunohistochemical surrogate definitions for basal-like breast cancer, such as the clinical ER/PR/HER2 triple-negative phenotype and models incorporating positive expression for CK5 (CK5/6) and/or EGFR are heavily cited. However, many additional biomarkers for basal-like breast cancer have been described in the literature. A parallel comparison of 46 proposed immunohistochemical biomarkers of basal-like breast cancer was performed against a gene expression profile gold standard on a tissue microarray containing 42 basal-like and 80 non-basal-like breast cancer cases. Ki67 and PPH3 were the most sensitive biomarkers (both 92%) positively expressed in the basal-like subtype, whereas CK14, IMP3 and NGFR were the most specific (100%). Among biomarkers surveyed, loss of INPP4B (a negative regulator of phosphatidylinositol signaling) was 61% sensitive and 99% specific with the highest odds ratio (OR) at 108, indicating the strongest association with basal-like breast cancer. Expression of nestin, a common marker of neural progenitor cells that is also associated with the triple-negative/basal-like phenotype and poor breast cancer prognosis, possessed the second highest OR at 29 among the 46 biomarkers surveyed, as well as 54% sensitivity and 96% specificity. As a positively expressed biomarker, nestin possesses technical advantages over INPP4B that make it a more ideal biomarker for identification of basal-like breast cancer. The comprehensive immunohistochemical biomarker survey presented in this study is a necessary step for determining an optimized surrogate immunopanel that best defines basal-like breast cancer in a practical and clinically accessible way.

Morphologic and Molecular Characteristics of Mixed Epithelial Ovarian Cancers.

Epithelial ovarian cancer (EOC) consists of 5 major histotypes: high-grade serous carcinoma (HGSC), endometrioid carcinoma (EC), clear cell carcinoma (CCC), mucinous carcinoma (MC), and low-grade serous carcinoma (LGSC). Each can have a broad spectrum of morphologic appearances, and 1 histotype can closely mimic histopathologic features more typical of another. Historically, there has been a relatively high frequency of mixed, defined by 2 or more distinct histotypes present on the basis of routine histopathologic assessment, histotype carcinoma diagnoses (3% to 11%); however, recent immunohistochemical (IHC) studies identifying histotype-specific markers and allowing more refined histotype diagnoses suggest a much lower incidence. We reviewed hematoxylin and eosin-stained slides from 871 cases of EOC and found the frequency of mixed carcinomas to be 1.7% when modern diagnostic criteria are applied. Through international collaboration, we established a cohort totaling 22 mixed EOCs, consisting of 9 EC/CCC, 4 EC/LGSC, 3 HGSC/CCC, 2 CCC/MC, and 4 other combinations. We interrogated the molecular differences between the different components of each case using IHC, gene expression, and hotspot sequencing analyses. IHC data alone suggested that 9 of the 22 cases were not mixed tumors, as they presented a uniform immuno-phenotype throughout, and these cases most probably represent morphologic mimicry and variation within tumors of a single histotype. Synthesis of molecular data further reduces the incidence of mixed carcinomas. On the basis of these results, true mixed carcinomas with both morphologic and molecular support for the presence of >1 histotype within a given tumor represent <1% of EOCs.