Abstract 5247: Use of two markers of hypoxia to study migration, re-oxygenation and repopulation of originally hypoxic cells in MCF-7 tumor xenografts following chemotherapy
Authors: Saggar Jasdeep K. , Tannock Ian F. 
Published: August 2012

Background: Hypoxia occurs in solid tumors: chronic diffusion-limited hypoxia occurs distal from functional blood vessels, while acute hypoxia may occur because of interruptions in blood flow. Well-nourished tumor cells close to blood vessels tend to be rapidly proliferating but hypoxic cells located farther away are slowly-proliferating. Chemotherapy may spare hypoxic cells because of poor drug distribution to them and because most drugs are selectively toxic to proliferating cells. Intervals between administration of chemotherapy allow for recovery of normal tissues (e.g. repopulation of bone marrow) but might allow re-oxygenation and resumed proliferation of formerly hypoxic cells due to better supply of nutrients to them, as is observed during radiotherapy. Therefore, it is important to establish a technique whereby hypoxia can be studied allowing for the assessment of different hypoxia-modulating therapies that can be used to prevent tumor relapse. Methods: Two specific markers of hypoxic cells (pimonidazole [pimo] and EF5) were injected into mice bearing MCF7 tumor xenografts, and tumor cells labeled with one or both markers were recognized in tumor sections (in relation to DioC7+ve functional tumor blood vessels) using appropriate fluorescence-labeled antibodies and imunohistochemistry. Proliferating cells were identified by an antibody to Ki67. Mice were treated with pimo and then either doxorubicin or saline one hour later; EF5 was given after a variable interval of 24, 48, 72, 96 or 120 hours; mice were killed two hours after the second injection. Changes in the location, proliferation and oxygen status of formerly hypoxic (pimo+ve) cells were quantified by their distance from blood vessels, Ki67 status and uptake of EF5 as a function of time. Results: Following treatment with doxorubicin, the proportion of hypoxic cells in the entire tumor decreased from1.5% (pimo+ve) prior to injection to 0.7% (EF5+ve) at 24 hours. The proportion of pimo+ve formerly hypoxic cells that are no longer hypoxic (i.e. EF5 - ve) at 24 hours was 75% after treatment compared to 18% in controls indicating rescue of previous hypoxic cells that would have died in the absence of treatment. The proportion of these pimo+ve cells that were cycling (Ki67+ve) increased from 4.7% to 15.0% at 24 Hours and then slowly decreased. Conclusions: There is a decrease in the proportion of hypoxic cells in MCF7 xenografts following treatment with doxorubicin. Originally hypoxic cells (that may have died in the absence of treatment) can move closer to blood vessels, re-oxygenate, and repopulate the tumor. Treatment directed to killing hypoxic cells (e.g. with hypoxia activated pro-drugs) has the potential to improve the outcome of chemotherapy by inhibiting tumor cell repopulation. Supported by are search grant from the Canadian Institutes for Health Research.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5247. doi:1538-7445.AM2012-5247