Measuring the Likelihood of Disease Recurrence in Lymph Nodes
Profiling Tumor-Draining Lymph Nodes Using Multispectral Imaging - Predicting Breast Cancer Recurrence
For the majority of human solid cancer, regional draining lymph nodes (LNs) are the initial site for metastases and the LN status serves as a highly significant prognostic indicator for clinical outcome. Impaired immune functions in the sentinel lymph node (SLN) may facilitate early metastatic events or may reflect overall host response to the cancer.
Kohrt, Lee and colleagues at Stanford have recently showed that decreases in T cells and dendritic cells within tumor-draining lymph nodes correlate strongly with the probability of recurrence of metastatic breast cancer and with 5-year disease-free survival (DFS) in early-stage breast cancer, regardless of whether or not the tumor had actually spread to the draining lymph nodes.
Immunohistochemical analysis using the Nuance multispectral imaging system can resolve and measure multiple molecular targets that are spectrally and/or spatially overlapping while preserving the architectural context of the lymph node. This allows for a detailed architectural and functional analysis of lymph nodes removed from breast cancer patients and may provide an accurate predictor of disease-free survival time.
In one study, multispectral imaging was employed to characterize the anatomical distribution and molecular phenotype of immune cells within TDLNs and tumor of 29 breast cancer patients, aged 29-76 years, and correlated to five-year clinical outcome. Immunohistochemistry (IHC) staining was performed on 3-µm thick tissue sections cut from formalin-fixed, paraffin-embedded nodes. Antigenic targets included CD4, CD8, CD1a, CD83, granzyme B, FOXP3, and AE1/AE3. The samples were imaged, using the Nuance spectral imaging system (CRi, Woburn, MA) and the resulting images analyzed using CRI’s spectral unmixing algorithms and custom image analysis software.
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Figure 1: Breast-cancer-draining lymph node tissue section immunohistochemically labeled with antibodies to CD4, CD8, CD1a, CD83, granzyme B, FOXP3, and AE1/AE3. Images taken with Nuance (left to right): a conventional RGB image of field 1; Nuance composite image comprised of multiple and separate component images of field 1, a conventional RGB image of field 3; Nuance composite image comprised of multiple and separate component images of field 3.
The results, still under review, suggest that breast cancer can alter draining lymph nodes by inhibiting effective anti-tumor immune response. Because the staining and analysis involved chromogenic labeling of multiple antigens on individual cells, the project required the capabilities of CRi’s multispectral detection, unmixing and analysis hardware and software.
Grant support :
Supported in part by NCI SBIR grant R44 CA088684 to CRI
Holbrook E. Kohrt1, A. Francesca Setiadi1, Edina B. Levic1, Adam Kapelner1, Rudy Angeles3, Daniel Winer2, Susan Holmes3, Chris M. van der Loos4, Erich Schwartz2, Peter P. Lee1
1Department of Medicine, 2Department of Pathology, 3Department of Statistics, Stanford University, Stanford, CA; 4Academic Medical Center, Department of Cardiovascular Pathology, Amsterdam, The Netherlands.
