Abstract 734: Water diffusion decreased in low collagen containing hypoxic regions of breast cancer xenograft.
Authors: Kakkad Samata M. , Zhang Jiangyang , Akhbardeh Alireza , Solaiyappan Meiyappan , Raman Venu , Leibfritz Dieter , Glunde Kristine , Bhujwalla Zaver M. 
Published: August 2014

Abstract
Tumors display chaotic vasculature that leads to hypoxia. Hypoxia is associated with an aggressive phenotype and increased resistance to radiation and chemotherapy. Hypoxic tumor regions also contain very few collagen 1 fibers, which is a major component of the tumor extracellular matrix (ECM) and plays an important role in molecular movement through the ECM. Here we examined the effects of hypoxia on water diffusion in a breast cancer xenograft model engineered to fluoresce under hypoxia. We observed, for the first time, that in hypoxic regions water diffusion and diffusion anisotropy was lower than in normoxic regions that may be explained by the few collagen fibers in hypoxic regions.
Severe combined immunodeficient female mice were inoculated in the mammary fat pad with 2x106 MDA-MB-231 cells stably expressing red fluorescent protein (RFP) under the control of a hypoxia response element. Once tumor volume was approximately 300-400 mm3 the tumor was excised, fixed and imaged using high-resolution diffusion tensor imaging (DTI). The apparent diffusion coefficient (ADC) and fractional anisotropy (FA) maps were calculated. Following DTI, the tumor was sliced at 1 mm thick sections for optical imaging. Using a 1x lens to cover the entire tumor section, bright field and fluorescence optical images were acquired to detect the RFP-containing hypoxic regions. Multimodality registration was performed by feature extraction to co-register the optical images to the DTI images. Statistical dice similarity indices were calculated as a measure of the registration error. Hypoxic regions were identified from the optical fluorescence field images and the ADC and FA values were calculated in the corresponding hypoxic and normoxic regions.
ADC and FA value distributions were identified in the DTI data and red fluorescent hypoxic regions were identified from the optical images. Co-registration accuracy confirmed by dice similarity index of 0.88 indicates a good overlap of optical and DTI images. Overall, hypoxic regions had lower water diffusion (ADC) and diffusion anisotropy (FA) values than normoxic regions. We have consistently observed that collagen fibers are significantly decreased in hypoxic regions, which could explain the lower ADC and FA values observed in hypoxic regions. In separate studies we have also observed high ADC in high collagen fiber containing regions in human breast cancer tissue. Low ADC and FA values observed in low collagen containing hypoxic regions indicate a functional role of these fibers in molecular transport. Decreased diffusion of molecules due to fewer collagen fibers in hypoxic regions may also contribute to poor drug delivery and tumor recurrence in hypoxic regions. Increased cellularity is thought to be one reason why lower ADC and FA values are observed in tumors. An even lower ADC within this backdrop may be due to hypoxic tumor regions.
This work was supported by NIH P50CA103175 P30CA006973
Citation Format: Samata M. Kakkad, Jiangyang Zhang, Alireza Akhbardeh, Meiyappan Solaiyappan, Venu Raman, Dieter Leibfritz, Kristine Glunde, Zaver M. Bhujwalla. Water diffusion decreased in low collagen containing hypoxic regions of breast cancer xenograft. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 734. doi:10.1158/1538-7445.AM2013-734