Axillary lymph node status together with estrogen and progesterone receptor status are important prognostic factors in breast cancer. In this study, the potential of using MR metabolomics for prediction of these prognostic factors was evaluated. Biopsies from breast cancer patients (n = 160) were excised during surgery and analyzed by high resolution magic angle spinning MR spectroscopy (HR MAS MRS). The spectral data were preprocessed and variable stability (VAST) scaled, and training and test sets were generated using the Kennard-Stone and SPXY sample selection algorithms. The data were analyzed by partial least-squares discriminant analysis (PLS-DA), probabilistic neural networks (PNNs) and Bayesian belief networks (BBNs), and blind samples (n = 50) were predicted for verification. Estrogen and progesterone receptor status was successfully predicted from the MR spectra, and were best predicted by PLS-DA with a correct classification of 44 of 50 and 39 of 50 samples, respectively. Lymph node status was best predicted by BBN with 34 of 50 samples correctly classified, indicating a relationship between metabolic profile and lymph node status. Thus, MR profiles contain prognostic information that may be of benefit in treatment planning, and MR metabolomics may become an important tool for diagnosis of breast cancer patients.

IntroductionDysregulated choline metabolism is a well-known feature of breast cancer, but the underlying mechanisms are not fully understood. In this study, the metabolomic and transcriptomic characteristics of a large panel of human breast cancer xenograft models were mapped, with focus on choline metabolism.MethodsTumor specimens from 34 patient-derived xenograft models were collected and divided in two. One part was examined using high-resolution magic angle spinning (HR-MAS) MR spectroscopy while another part was analyzed using gene expression microarrays. Expression data of genes encoding proteins in the choline metabolism pathway were analyzed and correlated to the levels of choline (Cho), phosphocholine (PCho) and glycerophosphocholine (GPC) using Pearson’s correlation analysis. For comparison purposes, metabolic and gene expression data were collected from human breast tumors belonging to corresponding molecular subgroups.ResultsMost of the xenograft models were classified as basal-like (N = 19) or luminal B (N = 7). These two subgroups showed significantly different choline metabolic and gene expression profiles. The luminal B xenografts were characterized by a high PCho/GPC ratio while the basal-like xenografts were characterized by highly variable PCho/GPC ratio. Also, Cho, PCho and GPC levels were correlated to expression of several genes encoding proteins in the choline metabolism pathway, including choline kinase alpha (CHKA) and glycerophosphodiester phosphodiesterase domain containing 5 (GDPD5). These characteristics were similar to those found in human tumor samples.ConclusionThe higher PCho/GPC ratio found in luminal B compared with most basal-like breast cancer xenograft models and human tissue samples do not correspond to results observed from in vitro studies. It is likely that microenvironmental factors play a role in the in vivo regulation of choline metabolism. Cho, PCho and GPC were correlated to different choline pathway-encoding genes in luminal B compared with basal-like xenografts, suggesting that regulation of choline metabolism may vary between different breast cancer subgroups. The concordance between the metabolic and gene expression profiles from xenograft models with breast cancer tissue samples from patients indicates that these xenografts are representative models of human breast cancer and represent relevant models to study tumor metabolism in vivo.

Cancer-associated fibroblasts (CAFs) are abundantly present in solid tumors and affect tumorigenesis and therapeutic responses. In the context of clinical radiotherapy, the impact of irradiated CAFs to treatment outcomes is largely unexplored. Aiming at improving radiotherapy efficacy, we have here explored the effect of radiation on the inherent pro-tumorigenic capacity of CAFs in animals. Ionizing radiation was delivered to cultured CAFs as single-high or fractionated doses. Tumor development was compared in mice receiving A549 lung tumor cells admixed with irradiated or control CAFs. Biological mechanisms behind tumor growth regulation were investigated by quantitative histology and immunohistochemistry. Viability assessments confirmed that irradiated CAFs are fully functional prior to implantation. However, the enhanced tumorigenic effect observed in tumors co-implanted with control CAFs was abrogated in tumors established with irradiated CAFs. Experiments to ascertain fate of implanted fibroblasts showed that exogenously administered CAFs reside at the implantation site for few days, suggesting that tumor growth regulation from admixed CAFs take place during initial tumor formation. Our work demonstrate that irradiated CAFs lose their pro-tumorigenic potential in vivo, affecting angiogenesis and tumor engraftment. This finding propose a previously unknown advantageous effect induced by radiotherapy, adding to the direct cytotoxic effects on transformed epithelial cells.