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. 2015 Mar;22(3):866-73.
doi: 10.1245/s10434-014-4174-8. Epub 2014 Oct 18.

A novel animal model for locally advanced breast cancer

Maria V Bogachek  1 , Jung Min Park, James P De Andrade, Mikhail V Kulak, Jeffrey R White, Tong Wu, Philip M Spanheimer, Thomas B Bair, Alicia K Olivier, Ronald J Weigel
Affiliations

A novel animal model for locally advanced breast cancer

Maria V Bogachek et al. Ann Surg Oncol. 2015 Mar.

Abstract

Background: Locally advanced breast cancer (LABC) poses complex management issues due to failure of response to chemotherapy and progression to local complications such as skin erosion, superinfection, and lymphedema. Most cell line and animal models are not adequate to study LABC.

Methods: A patient-derived xenograft (IOWA-1T) from a patient with LABC was characterized for expression profile, short tandem repeat profile, oncogenic mutations, xenograft growth, and response to therapy.

Results: Short tandem repeat profile authenticated the cell line as derived from a human woman. The primary tumor and derived xenografts were weakly estrogen receptor alpha positive (<5%), progesterone receptor negative, and HER2 nonamplified. Expression array profile compared to MCF-7 and BT-549 cell lines indicate that IOWA-1T was more closely related to basal breast cancer. IOWA-1T harbors a homozygous R248Q mutation of the TP53 gene; in vitro invasion assay was comparable to BT-549 and greater than MCF-7. IOWA-1T xenografts developed palpable tumors in 9.6 ± 1.6 days, compared to 49 ± 13 days for parallel experiments with BT-20 cells (p < 0.002). Tumor xenografts became locally advanced, growing to >2 cm in 21.6 ± 2 days, characterized by skin erosion necessitating euthanasia. The SUMO inhibitor anacardic acid inhibited the outgrowth of IOWA-1T xenografts, while doxorubicin had no effect on tumorigenesis.

Conclusions: IOWA-1T is a novel cell line with an expression pattern consistent with basal breast cancer. Xenografts recapitulated LABC and provide a novel model for testing therapeutic drugs that may be effective in cases resistant to conventional chemotherapy.

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Conflict of interest statement

Conflict of Interest: The authors declare that they have no conflict of interest.

Figures

Figure 1
Figure 1. IOWA-1T Cell Culture and Expression Array
A. Shown is a photomicrograph of the IOWA-1T cell line morphology by light microscopy. The cells do not adhere to plastic and grow in spheroids and cell aggregates. B. Expression array for the IOWA-1T cell line in comparison to MCF-7 and BT-549 with whole genome array with genes that differ by a factor of 2. C. Selected genes characteristic of luminal (ESR1, FOXA1, KRT8, MUC1, RET, GATA3, GREB1, MYB) and basal (MMP14, GSTP1, SFRP1) expression profiles.
Figure 2
Figure 2. RNA Expression of Luminal and Basal Gene Expression
RT-qPCR analysis of RNA expression is shown for selected luminal (ESR1, FOXA1, KRT8, MUC1, RET, GATA3, GREB1, MYB) and basal (MMP14, GSTP1, SFRP1) genes for MCF-7, IOWA-1T and BT-549 cell lines. Relative expression was normalized to MCF-7. The findings confirm the expression array data and indicate that the IOWA-1T cells have a basal-like expression pattern. Statistical significance of differences compared to MCF-7 are noted by an asterisk, * p < 0.05.
Figure 3
Figure 3. Western blots for Luminal and Basal Genes
Western blot data for protein expression of ESR1/ERα, FOXA1, KRT8, MUC1, RET, GATA3, GREB1, MYB, MMP14, GSTP1, SFRP1 for MCF-7, IOWA-1T and BT-549 cell lines demonstrate that the IOWA-1T cells have a basal-like phenotype by protein expression.
Figure 4
Figure 4. FACS Analysis with CD44 and CD24 and Immunohistochemistry
A. Flow cytometry is shown for CD44 and CD24 staining for MCF7, IOWA-1T and BT-549 cells. The data demonstrates that 93% of the cells in the IOWA-1T cell line are contained within the CD44+/hi/CD24-/low cancer stem cell population. By comparison the BT-549 cell line had 98.6% and MCF-7 had 0% cells in the CD44+/hi/CD24-/low population. B. Immunochemistry of the primary tumor from the original core biopsy (top) and from IOWA-1T xenografts (bottom) stained for ERα, PgR and HER2. Both the primary tumor and IOWA-1T xenografts were ERα weakly positive (<10%), PgR-negative and HER2-negative.
Figure 5
Figure 5. Characterization of IOWA-1T Invasion and Xenograft Growth
A. In vitro invasion assay comparing MCF-7, IOWA-1T and BT-549 cells. For comparisons, * p < 0.05 and NS: not significantly different. B. Example of IOWA-1T xenograft showing large tumor with skin erosion. C. Tumor free survival with time from tumor cell inoculation for IOWA-1T vs. BT-20 cells. D. Overall survival for IOWA-1T vs. BT-20 cells from time of tumor cell inoculation. E. Tumor-free survival for IOWA-1T xenografts with mice treated with vehicle (control), doxorubicin by tail vein injection (Doxorubicin) or anacardic acid via gavage (Anacardic) demonstrates increased TFS with anacardic acid and no significant effect of doxorubicin.
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