Denise G. Hemmings

Metastasis remains a major challenge in treating breast cancer. Breast tumors metastasize to organ-specific locations such as the brain, lungs, and bone, but why some organs are favored over others remains unclear. Breast tumors also show heterogeneity, plasticity, and distinct microenvironments. This contributes to treatment failure and relapse. The interaction of breast cancer cells with their metastatic microenvironment has led to the concept that primary breast cancer cells act as seeds, whereas the metastatic tissue microenvironment (TME) is the soil. Improving our understanding of this interaction could lead to better treatment strategies for metastatic breast cancer. Targeted treatments for different subtypes of breast cancers have improved overall patient survival, even with metastasis. However, these targeted treatments are based upon the biology of the primary tumor and often these patients’ relapse, after therapy, with metastatic tumors. The advent of immunotherapy allowed the immune system to target metastatic tumors. Unfortunately, immunotherapy has not been as effective in metastatic breast cancer relative to other cancers with metastases, such as melanoma. This review will describe the heterogeneic nature of breast cancer cells and their microenvironments. The distinct properties of metastatic breast cancer cells and their microenvironments that allow interactions, especially in bone and brain metastasis, will also be described. Finally, we will review immunotherapy approaches to treat metastatic breast tumors and discuss future therapeutic approaches to improve treatments for metastatic breast cancer.

Preeclampsia (PE) involves inadequate placental function. This can occur due to elevated pro-inflammatory tumor necrosis factor-α (TNF-α). In other tissues, TNF-α signals via sphingosine kinase 1 (SphK1). SphK1 hinders syncytial formation. Whether this occurs downstream of TNF-α signaling is unclear. We hypothesized that placental SphK1 levels are higher in PE and elevated TNF-α decreases syncytial function, increases syncytial shedding, and increases cytokine/factor release via SphK1 activity. Term placental biopsies were analyzed for SphK1 using immunofluorescence and qRT-PCR. Term placental explants were treated after 4 days of culture, at the start of syncytial regeneration, with TNF-α and/or SphK1 inhibitors, PF-543. Syncytialization was assessed by measuring fusion and chorionic gonadotropin release. Cell death and shedding were measured by lactate dehydrogenase release and placental alkaline phosphatase-positive shed particles. Forty-two cytokines were measured using multiplex assays. Placental SphK1 was increased in PE. Increased cell death, shedding, interferon-α2, IFN-γ-induced protein 10, fibroblast growth factor 2, and platelet-derived growth factor-AA release induced by TNF-α were reversed upon SphK1 inhibition. TNF-α increased the release of 26 cytokines independently of SphK1. TNF-α decreased IL-10 release and inhibiting SphK1 reversed this effect. Inhibiting SphK1 alone decreased TNF-α release. Hence, SphK1 partially mediates the TNF-α-induced PE placental phenotype, primarily through cell damage, shedding, and specific cytokine release.

Cytomegalovirus (CMV) infects 40–70% of women, but infection has been reported in >95% of breast cancer patients. We investigated the consequences of these observations by infecting mice with mCMV or a negative control medium for 4 days, 11 days or 10 weeks to establish active, intermediate or latent infections, respectively. Syngeneic 4T1 or E0771 breast cancer cells were then injected into a mammary fat pad of BALB/c or C57BL/6 mice, respectively. Infection did not affect tumor growth in these conditions, but latently infected BALB/c mice developed more lung metastases. The latent mCMV infection of MMTV-PyVT mice, which develop spontaneous breast tumors, also did not affect the number or sizes of breast tumors. However, there were more tumors that were multilobed with greater blood content, which had enhanced vasculature and decreased collagen content. Most significantly, mCMV infection also increased the number and size of lung metastases, which showed a higher cell proliferation. Viral DNA was detected in breast tumors and lung nodules although viral mRNA was not. These novel results have important clinical implications since an increased metastasis is prognostic of decreased survival. This work provides evidence that treating or preventing HCMV infections may increase the life expectancy of breast cancer patients by decreasing metastasis.

Recruitment is a challenge in developing population-representative pregnancy and birth cohorts. We developed a collaborative recruitment infrastructure (CRI) to recruit pregnant women for 4 pregnancy cohorts using: faxes from obstetrical offices, in-clinic recruiters, university and funder-driven free-media events, paid-media, and attendance at relevant tradeshows. Recruitment rates and demographic differences were compared between recruitment methods. We received 5008 referrals over 40 months. Compared to fax, free-media referrals were 13 times more likely to be recruited (OR 13.0, 95% CI 4.2, 40.4: p < 0.001) and paid-media referrals were 4 times more likely to be recruited (OR 4.6, 95% CI 2.1, 10.3: p < 0.001). Among paid-media advertisements, free-to-read print (e.g. Metro) was the most effective (OR 3.3, 95% CI 2.3, 4.5: p < 0.05). Several demographic differences were identified between recruitment methods and against a reference population. Between recruitment methods, media recruits had a similar proportion families with incomes ≥ $40,000 (paid-media: 94.4%; free-media: 93.3%) compared to fax recruits (95.7%), while in-clinic recruits were less likely to have family incomes ≥ $40,000 (88.8%, p < 0.05). Maternal recruits from fax and in-clinic were more likely to attend university (Fax: 92.6%, in-clinic 89.8%) versus the reference population (52.0%; p < 0.05 for both) and both were less likely to smoke (Fax: 6.8%, in-clinic 4.2%) versus reference (18.6%; p < 0.05 for both). However, while fax referrals were more likely to be Caucasian (85.9% versus reference 77.5%; p < 0.05), in-clinic referrals were not significantly different (78.2%; P > 0.05). Recruitment methods result in different recruitment rates and participant demographics. A variety of methods are required to recruit a generalizable sample.