Abstract 2988: Microenvironmental distribution of trastuzumab is heterogeneous and decreases sharply when administered following a single dose of bevacizumab in Her2+ve xenografts and metastases models
Authors: Baker Jennifer H. E. , Kyle Alastair H. , Reinsberg Stefan A. , Moosvi Firas , Cran Jordan , Hafeli Urs , Saatchi Katayoun , Minchinton Andrew I. 
Published: May 2015

Abstract
We have previously shown that trastuzumab distributes heterogeneously within MDA-435-Her2 xenografts. Though peak accumulation occurs 24h after administration & trastuzumab was detected greater than 150 µm from many vessels, there remained substantial Her2+ve areas that were negative for bound drug.
Methods: Her2+ve human carcinoma xenografts were grown in NOD/SCID mice as solid tumors or as metastatic models following ip or iv injection. Tumors were MR-imaged (T1-weighted) with a 500kDa Gd-labeled hyperbranched polyglycerol (HPG) contrast agent (CA). Treatments included bevacizumab (BvMAb, 10 mg/kg), trastuzumab (TsMab, 10 mg/kg) and human IgG (10 mg/kg). Multiplexed staining and imaging of whole tumor sections was used to evaluate the distribution of TsMAb in the context of the tumor microenvironment.
Findings: BT474, MDA361 and SKOV3 primary tumor xenografts all show highly heterogeneous distribution of TsMAb at 24h, where some areas are highly saturated with TsMAb and others contain unbound Her2+ tissue. Small metastases in the peritoneal cavity and lung also exhibit substantial proportions of tissue that are negative for TsMAb. CD31-stained vessels dual-labeled for a perfusion marker are often seen to have little or no bound TsMAb on perivascular tumor cells. Pretreatment with BvMAb for 24-72h results in significant reductions in tumor accumulation of TsMAb, with tumors showing 60-90% of bound TsMAb relative to IgG-treated controls (p&lt;0.05). BvMAb -treated xenografts had reduced fractions of perfused vessels and reduced measures of vascular function assessed using DCE-MRI (49% reduction in CA, p&lt;0.05). Corresponding parameter maps of MR-imaged high vascular function and histological images of bound TsMAb do not consistently correlate. Detailed maps of the tumor microenvironment show that even with VEGF blockade, the distribution of TsMAb still does not correspond to vessels that are perfused. Significant numbers of perfused vessels persist with no perivascular bound TsMAb despite the presence of Her2+ cells; saturation with bound TsMAb is also not consistently correlated with the presence or absence of smooth muscle, collagen or ZO-1 stained tight junctions, examined using immunohistochemistry.
Summary: The extravascular distribution of TsMAb in Her2-overexpressing xenografts, including micrometastases, is heterogeneous and is not explained simply by the presence or absence of vascular function. Pre-treatment with BvMAb for even short exposures of 24-72h worsens the distribution of TsMAb within the tumor microenvironment. The persistence of tissues that are unbound for trastuzumab suggests the possibility of inadequate drug exposure as a mechanism for drug resistance that may be exacerbated by pre-treatment with a VEGF blockade.
Citation Format: Jennifer H. E. Baker, Alastair H. Kyle, Stefan A. Reinsberg, Firas Moosvi, Jordan Cran, Urs Hafeli, Katayoun Saatchi, Andrew I. Minchinton. Microenvironmental distribution of trastuzumab is heterogeneous and decreases sharply when administered following a single dose of bevacizumab in Her2+ve xenografts and metastases models. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2988. doi:10.1158/1538-7445.AM2014-2988