Abstract PD8-08: Development of humanized immune DCIS models using patient peripheral blood derived hematopoietic stem cells (CD34+)
Authors: Behbod F , Harper H , Hansford H , Limback D , Hong Y , Elsarraj H , Ricci LR , Fan F , Tawfik O , May L , Cusick T , Inciardi M , Redick M , Gatewood J , Winblad O , Fields TA , Fabian C , Godwin AK , Fields PE , Meierotto R , Perry J 
Published: May 2019

Background:

Ductal carcinoma in situ (DCIS) is the most common form of non-invasive breast cancer. To accurately study the natural progression of DCIS lesions in mice, we devised the mouse-intraductal (MIND) animal model, which involves intraductal injection of human DCIS epithelial cells into the mammary ducts of immunocompromised mice. To improve the translational application of the MIND model, we aimed to mimic the natural microenvironment of human DCIS with patient-derived immune cells and assess the role of engrafted immune cells on human DCIS progression. In order to achieve successful engraftment of the entire immune system in mice, we utilized MISTRG mice. These mice were developed by Rongvaux et al., on an immunodeficient (Rag2-/-IL2rγ-/-) background. The genes encoding human M-CSF (M), human IL-3 (I), SIRP1α (S), human thrombopoietin (TPO)(TR), and GM-CSF (G) were knocked into their respective mouse loci. As such, MISTRG mice are highly permissive for human hematopoiesis, supporting the development and function of lymphocytes, monocytes, and natural killer (NK) cells. In contrast, previous studies have used the humanized CD34+ NOD-SCID IL2rγ-/- mice (CD34+NSG), which are unable to support myeloid cell differentiation due to lack of expression of human-specific cytokines. Moreover, prior xenograft studies in the CD34+NSG mice have not used immune cells derived from the same patient as the tumor.

Results:

Human CD34+ cells derived from patients' peripheral blood were expanded ex vivo ˜100-fold using a novel formulation of culture medium. Transplantation of ex vivo expanded CD34+ cells via tail vein injection of MISTRG mice resulted in the successful engraftment of human immune cells as early as 4 weeks following injection. Successful engraftment was confirmed by flow cytometry using human specific antibodies that recognize human leukocytes (anti-CD45), T cells (anti-CD3), B cells (anti-CD20), and myeloid cells (anti-CD33) in spleen, bone marrow, and peripheral blood of MISTRG mice. Once engraftment was confirmed, DCIS epithelial cells from the same DCIS patients or DCIS cell lines were injected intraductally. Recruitment of patient-derived immune cells to the DCIS lesions was confirmed by immunofluorescence using human-specific antibodies that recognize neutrophils (anti-myeloperoxidase), macrophages (anti-CD68), M2-polarized macrophages (anti-c-MAF), natural killer cells (anti-CD56), dendritic cells (anti-CD21), T cells (anti-CD3) and B cells (anti-CD20).

Conclusion:

This model represents the first to enable the study of mechanisms of DCIS progression in a manner that fully represents the heterogeneity of human disease, including the influence of the patients' own immune cells on DCIS progression.

Citation Format: Behbod F, Harper H, Hansford H, Limback D, Hong Y, Elsarraj H, Ricci LR, Fan F, Tawfik O, May L, Cusick T, Inciardi M, Redick M, Gatewood J, Winblad O, Fields TA, Fabian C, Godwin AK, Fields PE, Meierotto R, Perry J. Development of humanized immune DCIS models using patient peripheral blood derived hematopoietic stem cells (CD34+) [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr PD8-08.