Immune-checkpoint blockade (ICB) combined with neoadjuvant chemotherapy improves pathological complete response in breast cancer (BC). To understand why only a subset of tumors respond to ICB, patients with hormone receptor-positive or triple-negative BC were treated with anti-PD1 prior to surgery. Paired pre-versus on-treatment biopsies from treatment-naïve patients receiving anti-PD-1 (n=29) or patients receiving neoadjuvant chemotherapy prior to anti-PD1 (n=11) were subjected to single-cell transcriptome, T-cell receptor and proteome profiling. One-third of tumors contained PD1-expressing T-cells, which clonally expanded upon anti-PD1 treatment irrespective of tumor subtype. Expansion mainly involved CD8 + T-cells with pronounced expression of cytotoxic-activity (PRF1, GZMB), immune-cell homing (CXCL13) and exhaustion markers (HAVCR2, LAG3), and CD4 + T-cells characterized by expression of T-helper-1 (IFNG) and follicular-helper (BCL6, CXCR5) markers. In pre-treatment biopsies, the relative frequency of immunoregulatory dendritic cells (PD-L1), specific macrophage phenotypes (CCR2 or MMP9) and cancer cells exhibiting MHC class I/II expression correlated positively with T-cell expansion. Conversely, undifferentiated preeffector/memory T-cells (TCF7, GZMK) or inhibitory macrophages (CXCR3, C3) were inversely correlated. Collectively, our data identify various immunophenotypes and associated gene sets that are positively or negatively correlated with T-cell expansion following anti-PD1. We shed light on the heterogeneity in treatment response to anti-PD1 in breast cancer.

Most common breast cancer susceptibility variants have been identified through genome-wide association studies (GWAS) of predominantly estrogen receptor (ER)-positive disease1. We conducted a GWAS using 21,468 ER-negative cases and 100,594 controls combined with 18,908 BRCA1 mutation carriers (9,414 with breast cancer), all of European origin. We identified independent associations at P < 5 × 10−8 with ten variants at nine new loci. At P < 0.05, we replicated associations with 10 of 11 variants previously reported in ER-negative disease or BRCA1 mutation carrier GWAS and observed consistent associations with ER-negative disease for 105 susceptibility variants identified by other studies. These 125 variants explain approximately 14% of the familial risk of this breast cancer subtype. There was high genetic correlation (0.72) between risk of ER-negative breast cancer and breast cancer risk for BRCA1 mutation carriers. These findings may lead to improved risk prediction and inform further fine-mapping and functional work to better understand the biological basis of ER-negative breast cancer.

Genome-wide association studies (GWAS) have identified more than 170 breast cancer susceptibility loci. Here we hypothesize that some risk-associated variants might act in non-breast tissues, specifically adipose tissue and immune cells from blood and spleen. Using expression quantitative trait loci (eQTL) reported in these tissues, we identify 26 previously unreported, likely target genes of overall breast cancer risk variants, and 17 for estrogen receptor (ER)-negative breast cancer, several with a known immune function. We determine the directional effect of gene expression on disease risk measured based on single and multiple eQTL. In addition, using a gene-based test of association that considers eQTL from multiple tissues, we identify seven (and four) regions with variants associated with overall (and ER-negative) breast cancer risk, which were not reported in previous GWAS. Further investigation of the function of the implicated genes in breast and immune cells may provide insights into the etiology of breast cancer.

In most mammalian species force of contraction of cardiac muscle increases with increasing rate of stimulation, i.e. a positive force‐frequency relationship. In single mouse ventricular cells, both positive and negative relationships have been described and little is known about the underlying mechanisms. We studied enzymatically isolated single ventricular mouse myocytes, at 30 °C. During field stimulation, amplitude of unloaded cell shortening increased with increasing frequency of stimulation (0.04 ± 0.01 ΔL/L0 at 1 Hz to 0.07 ± 0.01 ΔL/L0 at 4 Hz, n= 12, P < 0.05). During whole cell voltage clamp with 50 μM [K5‐fluo‐3]pip, both peak and baseline [Ca2+]i increased at higher stimulation frequencies, but the net Δ[Ca2+]i increased only modestly from 1.59 ± 0.08 ΔF/F0 at 1 Hz, to 1.71 ± 0.11 ΔF/F0 at 4 Hz (n= 17, P < 0.05). When a 1 s pause was interposed during stimulation at 2 and 4 Hz, [Ca2+]i transients were significantly larger (at 4 Hz, peak F/F0 increased by 78 ± 2 %, n= 5). SR Ca2+ content assessed during caffeine application, significantly increased from 91 ± 24 μmol l−1 at 1 Hz to 173 ± 20 μmol l−1 at 4 Hz (n= 5, P < 0.05). Peak ICa,L decreased at higher frequencies (by 28 ± 6 % at 2 Hz, and 45 ± 8 % at 4 Hz), due to slow recovery from inactivation. This loss of ICa,L resulted in reduced fractional release. Thus, in mouse ventricular myocytes the [Ca2+]i‐frequency response depends on a balance between the increase in SR content and the loss of trigger ICa,L. Small changes in this balance may contribute to variability in frequency‐dependent behaviour. In addition, there may be a regulation of the contractile response downstream of [Ca2+]i.

Background BRCA1 interacting protein C-terminal helicase 1 (BRIP1) is one of the Fanconi Anaemia Complementation (FANC) group family of DNA repair proteins. Biallelic mutations in BRIP1 are responsible for FANC group J, and previous studies have also suggested that rare protein truncating variants in BRIP1 are associated with an increased risk of breast cancer. These studies have led to inclusion of BRIP1 on targeted sequencing panels for breast cancer risk prediction. Methods We evaluated a truncating variant, p.Arg798Ter (rs137852986), and 10 missense variants of BRIP1, in 48 144 cases and 43 607 controls of European origin, drawn from 41 studies participating in the Breast Cancer Association Consortium (BCAC). Additionally, we sequenced the coding regions of BRIP1 in 13 213 cases and 5242 controls from the UK, 1313 cases and 1123 controls from three population-based studies as part of the Breast Cancer Family Registry, and 1853 familial cases and 2001 controls from Australia. Results The rare truncating allele of rs137852986 was observed in 23 cases and 18 controls in Europeans in BCAC (OR 1.09, 95% CI 0.58 to 2.03, p=0.79). Truncating variants were found in the sequencing studies in 34 cases (0.21%) and 19 controls (0.23%) (combined OR 0.90, 95% CI 0.48 to 1.70, p=0.75). Conclusions These results suggest that truncating variants in BRIP1, and in particular p.Arg798Ter, are not associated with a substantial increase in breast cancer risk. Such observations have important implications for the reporting of results from breast cancer screening panels.