Postpartum mammary gland involution is a tissue remodeling event that occurs in all mammals in the absence of nursing or after weaning to return the gland to the pre-pregnant state. The tissue microenvironment created by involution has proven to be tumor promotional. Here we report that the GPI-linked protein semaphorin 7a (SEMA7A) is expressed on mammary epithelial cells during involution and use preclinical models to demonstrate that tumors induced during involution express high levels of SEMA7A. Overexpression of SEMA7A promoted the presence of myeloid-derived podoplanin (PDPN)-expressing cells in the tumor microenvironment and during involution. SEMA7A drove the expression of PDPN in macrophages, which led to integrin- and PDPN-dependent motility and adherence to lymphatic endothelial cells to promote lymphangiogenesis. In support of this mechanism, mammary tissue from SEMA7A-knockout mice exhibited decreased myeloid-derived PDPN-expressing cells, PDPN-expressing endothelial cells, and lymphatic vessel density. Furthermore, co-expression of SEMA7A, PDPN, and macrophage marker CD68 predicted for decreased distant metastasis-free survival in a cohort of over 600 cases of breast cancer as well as in ovarian, lung, and gastric cancers. Together our results indicate that SEMA7A may orchestrate macrophage-mediated lymphatic vessel remodeling, which in turn drives metastasis in breast cancer

Blood-borne transmission of infectious prions during the symptomatic and asymptomatic stages of disease occurs for both human and animal transmissible spongiform encephalopathies (TSEs). The geographical distribution of the cervid TSE, chronic wasting disease (CWD), continues to spread across North America and the prospective number of individuals harboring an asymptomatic infection of human variant Creutzfeldt-Jakob Disease (vCJD) in the United Kingdom has been projected to be ~1 in 3000 residents. Thus, it is important to monitor cervid and human blood products to ensure herd health and human safety. Current methods for detecting blood-associated prions rely primarily upon bioassay in laboratory animals. While bioassay provides high sensitivity and specificity, it requires many months, animals, and it is costly. Here we report modification of the real time quaking-induced conversion (RT-QuIC) assay to detect blood-borne prions in whole blood from prion-infected preclinical white-tailed deer, muntjac deer, and Syrian hamsters, attaining sensitivity of >90% while maintaining 100% specificity. Our results indicate that RT-QuIC methodology as modified can provide consistent and reliable detection of blood-borne prions in preclinical and symptomatic stages of two animal TSEs, offering promise for prionemia detection in other species, including humans.

Infectious prions traverse epithelial barriers to gain access to the circulatory system, yet the temporal parameters of transepithelial transport and persistence in the blood over time remain unknown. We used whole-blood real-time quaking-induced conversion (wbRT-QuIC) to analyze whole blood collected from transmissible spongiform encephalopathy (TSE)-inoculated deer and hamsters throughout the incubation period for the presence of common prion protein-conversion competent amyloid (PrP C -CCA). We observed PrP C -CCA in the blood of TSE-inoculated hosts throughout the disease course from minutes postexposure to terminal disease.

Postpartum breast cancers are a highly metastatic subset of young women’s breast cancers defined as breast cancers diagnosed in the postpartum period or within 5 years of last child birth. Women diagnosed with postpartum breast cancer are nearly twice as likely to develop metastasis and to die from breast cancer when compared with nulliparous women. Additionally, epidemiological studies utilizing multiple cohorts also suggest that nearly half of all breast cancers in women aged <45 qualify as postpartum cases. Understanding the biology that underlies this increased risk for metastasis and death may lead to identification of targeted interventions that will benefit the large number of young women with breast cancer who fall into this subset. Preclinical mouse models of postpartum breast cancer have revealed that breast tumor cells become more aggressive if they are present during the normal physiologic process of postpartum mammary gland involution in mice. As involution appears to be a period of lymphatic growth and remodeling, and human postpartum breast cancers have high peritumor lymphatic vessel density (LVD) and increased incidence of lymph node metastasis (1, 2), we propose that novel insight into is to be gained through the study of the biological mechanisms driving normal postpartum mammary lymphangiogenesis as well as in the microenvironment of postpartum tumors.

Summary. A microelectrode was used to measure vaginal pH in a longitudinal study during pregnancy in 30 women, 28 of whom were delivered at term. Excluding pH readings made in the presence of pathogenic organisms, 120 readings were taken. The mean vaginal pH of all subjects was 4·16 (SD 0·41). Analysis of variance showed no significant within‐subject variation of pH during pregnancy. Two women had spontaneous preterm deliveries (<37 weeks) and their mean vaginal pH during pregnancy was 5·05 and 5·06. Eight women admitted in preterm labour, who then gave birth, had a mean pH of 5·43 (SD 0·94) which was significantly higher than the mean pH of 10 women in labour at term (4·58, SD 0·59). The relation between bacterial growth in vitro and the surrounding pH was also examined. Escherichia coli, a potential pathogen, behaved differently from both Lactobacillus (a commensal organism) and Candida albicans. The clinical observations show a trend towards higher vaginal pH values in association with preterm labour.

Post-partum breast cancer patients, or breast cancer patients diagnosed within 10 years of last childbirth, are ~3–5 times more likely to develop metastasis in comparison to non-post-partum, or nulliparous, patients. Additionally, post-partum patients have increased tumor-associated lymphatic vessels and LN involvement, including when controlled for size of the primary tumor. In pre-clinical, immune-competent , mouse mammary tumor models of post-partum breast cancer (PPBC), tumor growth and lymphogenous tumor cell spread occur more rapidly in post-partum hosts. Here we report on PD-L1 expression by lymphatic endothelial cells and CD11b+ cells in the microenvironment of post-partum tumors, which is accompanied by an increase in PD-1 expression by T cells. Additionally, we observed increases in PD-L1 and PD-1 in whole mammary tissues during post-partum mammary gland involution; a known driver of post-partum tumor growth, invasion, and metastasis in pre-clinical models. Importantly, implantation of murine mammary tumor cells during post-partum mammary gland involution elicits a CD8+ T cell population that expresses both the co-inhibitory receptors PD-1 and Lag-3. However, upon anti-PD-1 treatment, during post-partum mammary gland involution, the involution-initiated promotional effects on tumor growth are reversed and the PD-1, Lag-3 double positive population disappears. Consequently, we observed an expansion of poly-functional CD8+ T cells that produced both IFNγ and TNFα. Finally, lymphatic vessel frequency decreased significantly following anti-PD-1 suggesting that anti-PD-1/PD-L1 targeted therapies may have efficacy in reducing tumor growth and dissemination in post-partum breast cancer patients.

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Despite new targeted therapies and improved screening, breast cancer is the second leading cause of cancer-related death of women in the United States. This emphasizes the need to better understand the molecular pathways contributing to breast cancer progression and metastasis and the need for identification of novel therapeutic targets. This is especially true for triple-negative breast cancers (TNBCs), which in comparison to hormone receptor-positive (ER+/PR+) and HER2+ subtypes lack a targeted therapy. Without targeted therapy TNBC patients have limited options for treatment, resulting in decreased overall survival. Our lab identified semaphorin 7a (SEMA7A) as a novel protein that promotes TNBC progression. SEMA7A is a member of the semaphorin family, a group of proteins originally described for their developmental roles in axon guidance. SEMA7A is a unique member of the semaphorin family as it is the only GPI-linked semaphorin and can be shed into the extracellular space. Since breast cancer patients mainly die from metastatic disease, we aim to uncover the role of SEMA7A in TNBC cell invasion and metastasis. To study the role SEMA7A may have in promoting cellular invasion, we utilized an shRNA-mediated knockdown of SEMA7A in human TNBC cell lines. Our results show that SEMA7A plays an important role in many steps of the metastatic cascade, with shSEMA7A cells showing increased cell death in both attached and force suspension conditions (anoikis), reduced mesenchymal phenotypes, and less invasion in 3D matrices. We also observed decreased growth and invasion of shSEMA7A tumors in vivo and decreased synthesis of fibronectin and matrix-metalloproteinase 2 ex vivo from shSEMA7A tumors. Taken together, these results show that SEMA7A is a critical molecule for cellular invasion by modulating not only intrinsic cellular pathways, but also the tumor microenvironment. Citation Format: Sarah Elizabeth Tarullo, Alan Elder, Ryan Hill, Kirk Hansen, Traci Lyons. Investigating the role of semaphorin 7a in triple-negative breast cancer cell invasion [abstract]. In: Proceedings of the AACR Special Conference: Advances in Breast Cancer Research; 2017 Oct 7-10; Hollywood, CA. Philadelphia (PA): AACR; Mol Cancer Res 2018;16(8_Suppl):Abstract nr B55.