NK-cell and T-cell functions in patients with breast cancer: effects of surgery and adjuvant chemo- and radiotherapy

Breast cancer is globally the most common malignancy in women. Her2-targeted monoclonal antibodies are established treatment modalities, and vaccines are in late-stage clinical testing in patients with breast cancer and known to promote tumour-killing through mechanisms like antibody-dependent cellular cytotoxicity. It is therefore increasingly important to study immunological consequences of conventional treatment strategies. In this study, functional tests and four-colour flow cytometry were used to detect natural killer (NK)-cell functions and receptors as well as T-cell signal transduction molecules and intracellular cytokines in preoperative breast cancer patients, and patients who had received adjuvant radiotherapy or adjuvant combined chemo-radiotherapy as well as in age-matched healthy controls. The absolute number of NK cells, the density of NK receptors as well as in vitro quantitation of functional NK cytotoxicity were significantly higher in preoperative patients than the post-treatments group and controls. A similar pattern was seen with regard to T-cell signalling molecules, and preoperative patients produced significantly higher amounts of cytokines in NK and T cells compared to other groups. The results indicate that functions of NK and T cells are well preserved before surgery but decrease following adjuvant therapy, which may speak in favour of early rather than late use of immunotherapeutic agents such as trastuzumab that may depend on intact immune effector functions

Differential pattern and prognostic significance of CD4+, FOXP3+ and IL-17+ tumor infiltrating lymphocytes in ductal and lobular breast cancers

<p>Abstract</p> <p>Background</p> <p>Clinical relevance of tumor infiltrating lymphocytes (TILs) in breast cancer is controversial. Here, we used a tumor microarray including a large series of ductal and lobular breast cancers with long term follow up data, to analyze clinical impact of TIL expressing specific phenotypes and distribution of TILs within different tumor compartments and in different histological subtypes.</p> <p>Methods</p> <p>A tissue microarray (TMA) including 894 ductal and 164 lobular breast cancers was stained with antibodies recognizing CD4, FOXP3, and IL-17 by standard immunohistochemical techniques. Lymphocyte counts were correlated with clinico-pathological parameters and survival.</p> <p>Results</p> <p>CD4⁺lymphocytes were more prevalent than FOXP3⁺TILs whereas IL-17⁺TILs were rare. Increased numbers of total CD4⁺and FOXP3⁺TIL were observed in ductal, as compared with lobular carcinomas. High grade (G3) and estrogen receptor (ER) negative ductal carcinomas displayed significantly (<it>p </it> < 0.001) higher CD4⁺and FOXP3⁺lymphocyte infiltration while her2/neu over-expression in ductal carcinomas was significantly (<it>p </it> < 0.001) associated with higher FOXP3⁺TIL counts. In contrast, lymphocyte infiltration was not linked to any clinico-pathological parameters in lobular cancers. In univariate but not in multivariate analysis CD4⁺infiltration was associated with significantly shorter survival in patients bearing ductal, but not lobular cancers. However, a FOXP3⁺/CD4⁺ratio > 1 was associated with improved overall survival even in multivariate analysis (<it>p </it>= 0.033).</p> <p>Conclusions</p> <p>Ductal and lobular breast cancers appear to be infiltrated by different lymphocyte subpopulations. In ductal cancers increased CD4⁺and FOXP3⁺TIL numbers are associated with more aggressive tumor features. In survival analysis, absolute numbers of TILs do not represent major prognostic indicators in ductal and lobular breast cancer. Remarkably however, a ratio > 1 of total FOXP3⁺/CD4⁺TILs in ductal carcinoma appears to represent an independent favorable prognostic factor.</p

A different immunologic profile characterizes patients with HER-2-overexpressing and HER-2-negative locally advanced breast cancer: implications for immune-based therapies

INTRODUCTION: The clinical efficacy of trastuzumab and taxanes is at least partly related to their ability to mediate or promote antitumor immune responses. On these grounds, a careful analysis of basal immune profile may be capital to dissect the heterogeneity of clinical responses to these drugs in patients with locally advanced breast cancer undergoing neoadjuvant chemotherapy. METHODS: Blood samples were collected from 61 locally advanced breast cancers (36 HER2- and 25 HER2+) at diagnosis and from 23 healthy women. Immunophenotypic profiling of circulating and intratumor immune cells, including regulatory T (Treg) cells, was assessed by flow cytometry and immunohistochemistry, respectively. Serum levels of 10 different cytokines were assessed by multiplex immunoassays. CD8+ T cell responses to multiple tumor-associated antigens (TAA) were evaluated by IFN-γ-enzyme-linked immunosorbent spot (ELISPOT). The Student's t test for two tailed distributions and the Wilcoxon two-sample test were used for the statistical analysis of the data. RESULTS: The proportion of circulating immune effectors was similar in HER2+ patients and healthy donors, whereas higher percentages of natural killer and Treg cells and a lower CD4+/CD8+ T cell ratio (with a prevalence of naïve and central memory CD8+ T cells) were observed in HER2- cases. Higher numbers of circulating CD8+ T cells specific for several HLA-A*0201-restricted TAA-derived peptides were observed in HER2+ cases, together with a higher prevalence of intratumor CD8+ T cells. Serum cytokine profile of HER2+ patients was similar to that of controls, whereas HER2- cases showed significantly lower cytokine amounts compared to healthy women (IL-2, IL-8, IL-6) and HER2+ cases (IL-2, IL-1β, IL-8, IL-6, IL-10). CONCLUSIONS: Compared to HER2- cases, patients with HER2-overexpressing locally advanced breast cancer show a more limited tumor-related immune suppression. This may account for the clinical benefit achieved in this subset of patients with the use of drugs acting through, but also promoting, immune-mediated effects

Dynamics of notch pathway expression during mouse testis post-natal development and along the spermatogenic cycle

Articles in International JournalsThe transcription and expression patterns of Notch pathway components (Notch 1–3, Delta1 and 4, Jagged1) and effectors (Hes1, Hes2, Hes5 and Nrarp) were evaluated (through RT-PCR and IHC) in the mouse testis at key moments of post-natal development, and along the adult spermatogenic cycle. Notch pathway components and effectors are transcribed in the testis and expressed in germ, Sertoli and Leydig cells, and each Notch component shows a specific cell-type and timewindow expression pattern. This expression at key testis developmental events prompt for a role of Notch signaling in prepubertal spermatogonia quiescence, onset of spermatogenesis, and regulation of the spermatogenic cycle

Lymphocyte depletion and repopulation after chemotherapy for primary breast cancer

Background: Approximately 30 % of breast cancer patients receive chemotherapy, yet little is known about influences of current regimens on circulating lymphocyte levels and phenotypes. Similarly, clinico-pathological factors that modify these influences, and implications for future immune health remain mainly unexplored. Methods: We used flow-cytometry to assess circulating lymphocyte levels and phenotypes in 88 primary breast cancer patients before chemotherapy and at time-points from 2 weeks to 9 months after chemotherapy completion. We examined circulating titres of antibodies against pneumococcal and tetanus antigens using ELISAs. Results: Levels of B, T and NK cells were significantly reduced 2 weeks after chemotherapy (p < 0.001). B cells demonstrated particularly dramatic depletion, falling to 5.4 % of pre-chemotherapy levels. Levels of all cells recovered to some extent, although B and CD4+ T cells remained significantly depleted even 9 months post-chemotherapy (p < 0.001). Phenotypes of repopulating B and CD4+ T cells were significantly different from, and showed no sign of returning to pre-chemotherapy profiles. Repopulating B cells were highly depleted in memory cells, with proportions of memory cells falling from 38 % to 10 % (p < 0.001). Conversely, repopulating CD4+ T cells were enriched in memory cells, which increased from 63 % to 75 % (p < 0.001). Differences in chemotherapy regimen and patient smoking were associated with significant differences in depletion extent or repopulation dynamics. Titres of anti-pneumococcal and anti-tetanus antibodies were both significantly reduced post-chemotherapy and did not recover during the study (p < 0.001). Conclusion: Breast cancer chemotherapy is associated with long-term changes in immune parameters that should be considered during clinical management