Computerised scoring protocol for identification and quantification of different immune cell populations in breast tumour regions by the use of QuPath software.

AIMS: As important prognostic and predictive information can be obtained from the composition, functionality and spatial arrangement of different immune cell subtypes, this study aims at characterizing the immune infiltrate in breast tumours. METHODS AND RESULTS: Tumour-infiltrating lymphocytes (TILs) in 62 patients with luminal B-like breast cancer were characterised by immunohistochemical staining with standard markers, and were subsequently classified and quantified by the use of QuPath software. In different delineated tumour regions, the proportion and density of CD3+ , CD4+ , CD5+ , CD8+ , CD20+ and FOXP3+ cells were assessed. The results of the software analysis were compared with those of manual counting for CD8 and CD20 staining. The QuPath scoring protocol slightly overestimated positive, negative and total lymphocyte counts and density, while minimally underestimating the proportion of positively stained lymphocytes. However, for density and proportion, no real differences from manual counting were observed. For all markers, the density of positively stained immune cells was higher in the invasive front than in the tumour centre, pointing to an accumulation of immune cells near the tumour boundaries. When we looked at the proportion of immunohistochemically positive immune cells, we observed enrichment of CD5 (P = 0.025) and CD20 (P < 0.001) at the periphery, and FOXP3 enrichment in the centre (P < 0.001). CONCLUSION: The QuPath scoring protocol can adequately identify positively stained immune cells in breast tumours, and allows the evaluation of differences in immune cell proportion and density within different tumour regions. The entire tumour section can be quantitatively assessed quite rapidly, which is a major advantage over manual counting.

Age-related microRNAs in older breast cancer patients: biomarker potential and evolution during adjuvant chemotherapy.

BACKGROUND: MicroRNAs (miRNAs) are important regulators of cellular function and have been associated with both aging and cancer, but the impact of chemotherapy on age-related miRNAs has barely been studied. Our aim was to examine whether chemotherapy accelerates the aging process in elderly breast cancer patients using miRNA expression profiling. METHODS: We monitored age-related miRNAs in blood of women, aged 70 or older, receiving adjuvant chemotherapy (docetaxel and cyclophosphamide, TC) for invasive breast cancer (chemo group, CTG, n = 46). A control group of older breast cancer patients without chemotherapy was included for comparison (control group, CG, n = 43). All patients underwent geriatric assessment at inclusion (T0), after 3 months (T1) and 1 year (T2). Moreover, we analysed the serum expression of nine age-related miRNAs (miR-20a, miR-30b, miR-34a, miR-106b, miR-191, miR-301a, miR-320b, miR-374a, miR-378a) at each timepoint. RESULTS: Except for miR-106b, which behaved slightly different in CTG compared to CG, all miRNAs showed moderate fluctuations during the study course with no significant differences between groups. Several age-related miRNAs correlated with clinical frailty (miR-106b, miR-191, miR-301a, miR-320b, miR-374a), as well as with other biomarkers of aging, particularly Interleukin-6 (IL-6) and Monocyte Chemoattractant Protein-1 (MCP-1) (miR-106b, miR-301a, miR-374a-5p, miR-378a-3p). Moreover, based on their 'aging miRNA' profiles, patients clustered into two distinct groups exhibiting significantly different results for several biological/clinical aging parameters. CONCLUSIONS: These results further corroborate our earlier report, stating that adjuvant TC chemotherapy does not significantly boost aging progression in elderly breast cancer patients. Our findings also endorsed specific age-related miRNAs as promising aging/frailty biomarkers in oncogeriatric populations. TRIAL REGISTRATION: ClinicalTrials.gov, NCT00849758 . Registered on 20 February 2009. This clinical trial was registered prospectively.

Cancer and Aging: Two Tightly Interconnected Biological Processes.

Age is one of the main risk factors of cancer; several biological changes linked with the aging process can explain this. As our population is progressively aging, the proportion of older patients with cancer is increasing significantly. Due to the heterogeneity of general health and functional status amongst older persons, treatment of cancer is a major challenge in this vulnerable population. Older patients often experience more side effects of anticancer treatments. Over-treatment should be avoided to ensure an optimal quality of life. On the other hand, under-treatment due to fear of toxicity is a frequent problem and can lead to an increased risk of relapse and worse survival. There is a delicate balance between benefits of therapy and risk of toxicity. Robust biomarkers that reflect the body's biological age may aid in outlining optimal individual treatment regimens for older patients with cancer. In particular, the impact of age on systemic immunity and the tumor immune infiltrate should be considered, given the expanding role of immunotherapy in cancer treatment. In this review, we summarize current knowledge concerning the mechanistic connections between aging and cancer, as well as aging biomarkers that could be helpful in the field of geriatric oncology.

Blood Immunosenescence Signatures Reflecting Age, Frailty and Tumor Immune Infiltrate in Patients with Early Luminal Breast Cancer.

BACKGROUND: Immune/senescence-related host factors play a pivotal role in numerous biological and pathological process like aging, frailty and cancer. The assessment of these host factors via robust, non-invasive, and easy-to-measure blood biomarkers could improve insights in these processes. Here, we investigated in a series of breast cancer patients in which way single circulating biomarkers or biomarker panels relate to chronological age, frailty status, and tumor-associated inflammatory microenvironment. METHODS: An extensive panel of blood immune/senescence markers and the tumor immune infiltrate was studied in young, middle-aged, and old patients with an early invasive hormone-sensitive, HER2-negative breast cancer. In the old group, clinical frailty was estimated via the G8-scores. RESULTS: Several three-blood biomarker panels proved to be able to separate old chronological age from young age very efficiently. Clinically more important, several three-blood biomarker panels were strongly associated with clinical frailty. Performance of blood biomarker panels for prediction of the tumor immune infiltrate was lower. CONCLUSION: Immune/senescence blood biomarker panels strongly correlate with chronological age, and clinically more importantly with frailty status in early breast cancer patients. They require further investigation on their ability to provide a more complete picture on clinical frailty status and direct personalized therapy in older persons.

Comparison of the tumor immune microenvironment of primary hormone receptor-negative HER2-positive and triple negative breast cancer.

The vast majority of studies investigating immune checkpoint inhibition (ICI) in patients with breast cancer have focused on triple-negative breast cancer (TNBC). In this study, we compared the tumor immune microenvironment (TIME) between TNBC and hormone receptor-negative HER2-positive breast cancer based on a selection of immune markers at the protein level in an institutional retrospective series. Additionally, we performed a similar comparison using publicly available transcriptomics data. Altogether, the results show a comparable TIME in both groups, with possible implications for the use of ICI in patients with hormone receptor-negative HER2-positive breast tumors.

Age-related remodelling of the blood immunological portrait and the local tumor immune response in patients with luminal breast cancer.

OBJECTIVES: Aging is associated with altered immune function and chronic low-grade inflammation, referred to as immunosenescence. As breast cancer is an age-related disease, the impact of aging on tumor immune responses may have important consequences. However, effects of immunosenescence on breast tumor immune infiltration remain largely unknown. METHODS: This exploratory study investigated a broad panel of immune/senescence markers in peripheral blood and in the tumor microenvironment of young, middle-aged and old patients diagnosed with early invasive luminal (hormone-sensitive, HER2-negative) breast cancer. In the old group, G8-scores were computed as a correlate for clinical frailty. RESULTS: Significant age-related changes in plasma levels of several inflammatory mediators (IL-1α, IP-10, IL-8, MCP-1, CRP), immune checkpoint markers (Gal-9, sCD25, TIM-3, PD-L1), IGF-1 and circulating miRs (miR-18a, miR-19b, miR-20, miR-155, miR-195 and miR-326) were observed. Shifts were observed in distinct peripheral blood mononuclear cell populations, particularly naive CD8+ T-cells. At the tumor level, aging was associated with lower total lymphocytic infiltration, together with decreased abundance of several immune cell markers, especially CD8. The relative fractions of cell subsets in the immune infiltrate were also altered. Clinical frailty was associated with higher frequencies of exhausted/senescent (CD27-CD28- and/or CD57+) terminally differentiated CD8+ cells in the blood and with increased tumor infiltration by FOXP3+ cells. CONCLUSION: Aging and frailty are associated with profound changes of the blood and tumor immune profile in luminal breast cancer, pointing to a different interplay between tumor cells, immune cells and inflammatory mediators at higher age.