Introduction: It is widely understood that the T-lymphocyte is closely linked to
prognosis and treatment response in certain solid organ malignancies including
breast. B-Lymphocytes have, however, received comparatively little attention in this
field and it remains unclear what role, if any, B-lymphocyte subtypes play in
carcinogenesis or prognosis in breast cancers. Likewise, it is unknown what effect
systemic treatments such as chemotherapy have on B-cell immunity, or the role
these lymphocyte subtypes may have in the side-effect profile of such therapies.
Cancer related fatigue (CRF) is one such side-effect of cancer and its treatments,
the pathogenesis of which has been linked to impaired immunity including altered
lymphocyte phenotypes.
My aim in this work was to perform detailed phenotypical analyses of B- and Tlymphocytes
in breast cancer, as well as phenotypical alterations driven by cancer
treatments, to provide insight into the function of B-lymphocytes in this disease
process, adding to the growing body of knowledge driving immunotherapeutic
development.
Methods: 43 primary breast cancer patients, scheduled to receive adjuvant
chemotherapy, were recruited following resection surgery alongside 10 age- and
sex-matched healthy controls. 27 Chronic fatigue syndrome (CFS) patients were
recruited at diagnosis, as a comparator group for assessments relating to CRF. 15
further patients with primary breast cancer were recruited prior to resection surgery
for analyses of fresh tumour tissue and peripheral blood. Phenotypes of circulating
lymphocytes were analysed using multicolour flow cytometry at various time-points
in the larger breast cancer cohort and its associated comparator groups (CFS and
healthy controls), along with general health and well-being screening questionnaires
as used in the diagnosis of CFS. For the fresh tissue study, tumour tissue was
biopsied and analyses of tissue-resident B- and T-lymphocytes were performed,
along with circulating analyses as previously, focusing on regulatory B and T
subtypes. Results were analysed using paired T-tests, 2-way ANOVA, and
correlation analysis.
Results: Lymphocyte phenotype was vastly altered by chemotherapy. Within the Tcell
pool, the naïve subset is diminished with proportional increases to the memory
cell pool and the overall expansion of HLA-DR surface expression on CD4+ T-cells.
Regulatory T-cells were unchanged. Within the B-cell pool, memory cells were
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largely reduced by chemotherapy, the resulting phenotype being naïve and
transitional dominant. Pro-inflammatory cytokine expression by regulatory B-cells
was diminished, yet IL-10 expression remains unchanged.
The CD20+CD27+ regulatory Memory B-cell subset is demonstrated as a critical Bcell
phenotype in breast cancer prognosis and in fatigue. CD27+ regulatory memory
B-cell dominates the phenotype in tumour tissue where these subsets, in addition to
naïve T-cells, correlated positively with poor prognostic indicators in breast cancer
both in peripheral blood and tumour tissue. Additionally, expression of effector
regulatory cytokines IL-10 and TNF-α from the memory B-cell subsets correlated
with prognosis and fatigue.
Conclusions: Detailed B-lymphocyte phenotypes have been clearly demonstrated
in breast cancer and in fatigue, with the regulatory CD20+CD27+ memory B-cell
subset prevailing as the dominant cell type relating to breast cancer prognosis, both
in tumour and peripheral blood, as well as fatigue scores. This first insight into the
phenotype and function of B-cells and B-regs in breast cancer highlights the
memory B-cell subset as a driver in the pathogenesis of this disease, and provides a
target for further research and potential novel lines of investigation in the on-going
search for immunotherapies.
