Background Neoadjuvant chemotherapy (NAC) may alter the immune landscape of patients with early breast cancer (BC), potentially setting the scene for more effective implementation of checkpoint-targeted immunotherapy, albeit by largely unexplored mechanisms.

Methods This study investigated which alterations in the plasma concentrations of 16 soluble co-stimulatory and co-inhibitory, immune checkpoints were measured in a cohort of newly diagnosed, early BC patients (n=72) pre-treatment, post-NAC, and post-surgery using a Multiplex® bead array platform.

Results Relative to a group of healthy control subjects (n=45), the median pre-treatment levels (table 1) of 5 co-stimulatory (CD27, CD40, GITRL, ICOS, GITR) and 3 co-inhibitory (TIM-3, CTLA-4, PD-L1) soluble checkpoints were significantly lower in the BC patients vs. controls (p<0.021—p<0.0001; and p<0.008—p<0.00001, respectively). Following NAC (table 2), the plasma levels of 6 soluble co-stimulatory checkpoints (CD28, CD40, ICOS, CD27, CD80, GITR), all involved in activation of CD8⁺ cytotoxic T cells, were significantly increased relative to pre-treatment levels (p<0.04—p<0.00001), being comparable with control values and remaining at these levels post-surgery (table 3). Of the soluble co-inhibitory checkpoints, 3 (LAG-3, PD-L1, TIM-3) increased significantly post-NAC, while PD-1 was unchanged and BTLA and CTLA-4 were significantly decreased (p<0.03 and p<0.00001, respectively). A pathological complete response was documented in 65% of patients (mostly TNBC).

Conclusions Normalization of soluble co-stimulatory immune checkpoints is seemingly indicative of reversal of systemic immune dysregulation following administration of NAC in early BC, independent of response to treatment, while recovery of immune homeostasis may explain the increased levels of several negative checkpoint proteins, albeit with the exceptions of CTLA-4 and PD-1.

Ethics Approval Ethics approval was granted by The Research Ethics Committee, Faculty of Health Sciences, University of Pretoria (Ethics Committee Approval Numbers 517/2017.