In vivo hyperspectral imaging of microvessel response to trastuzumab treatment in breast cancer xenografts

Abstract

HER2-amplified (HER2 + ) breast cancers are treated with the anti-HER2 monoclonal antibody trastuzumab. Although trastuzumab reduces production of the angiogenic factor VEGF in HER2 + tumors, the acute and sustained effects of trastuzumab on the tumor vasculature are not understood fully, particularly in trastuzumab-resistant tumors. We used mouse models of trastuzumab sensitive and trastuzumab-resistant HER2 + breast cancers to measure dynamic changes in tumor microvessel density and hemoglobin oxygenation (sO 2 ) in vivo using quantitative hyperspectral imaging at 2, 5, 9, and 14 days after antibody treatment. Further analysis quantified the distribution of microvessels into low and high oxygenation groups, and monitored changes in these distributions with trastuzumab treatment. Gold standard immunohistochemistry was performed to validate complementary markers of tumor cell and vascular response to treatment. Trastuzumab treatment in both responsive and resistant tumors resulted in decreased sO 2 5 days after initial treatment when compared to IgG-treated controls (p<0.05). Importantly, responsive tumors showed significantly higher vessel density and significantly lower sO 2 than all other groups at 5 days post-treatment (p<0.05). Distribution analysis of vessel sO 2 showed a significant (p<0.05) shift of highly oxygenated vessels towards lower oxygenation over the time-course in both trastuzumab-treated responsive and resistant tumors. This study suggests that longitudinal hyperspectral imaging of microvessel sO 2 and density could distinguish trastuzumabresponsive from trastuzumab-resistant tumors, a finding that could be exploited in the post-neoadjuvant setting to guide post-surgical treatment decisions. Udove, A. Ullrich, and et, "Studies of the HER-2/neu proto-oncogene in human breast and ovarian cancer," Science 244(4905), 707-712 (1989). 3. J. S. Ross and J. A. Fletcher, "The HER-2/neu oncogene in breast cancer: prognostic factor, predictive factor, and target for therapy," Stem Cells 16(6), 413-428 (1998) ©2014 Optical Society of America

Collagen density and alignment in responsive and resistant trastuzumab-treated breast cancer xenografts

“…Furthermore, our previous studies of Tz effects on the tumor vasculature of BT474 and HR6 tumors revealed significant decreases in vessel oxygen saturation with five days of Tz treatment in both tumor models. 24 Our studies and these previous results suggest that Tz treatment may affect the tumor microenvironment independent of its effects on the tumor cells.…”

“…BT474 tumors respond to Tz with increased cell death and decreased tumor volume, while Tz does not induce cell death or decrease growth in HR6 tumors. 22,24,25 Collagen SHG images were analyzed using CurveAlign to extract the relative fiber alignment and density of collagen positive pixels (Fig. 1).…”

“…We have also demonstrated that collagen SHG and histological trichrome staining analysis of collagen provide complementary endpoints of tumor collagen content and morphology. Collagen SHG imaging can be performed in vivo through window chambers for longitudinal studies, 24,37 while trichrome staining is inherently postmortem, potentially limiting the number of time points and increasing animal burden and expense. Trichrome staining revealed decreased collagen content in both Tz-responsive and nonresponsive tumors treated with Tz.…”

Dual HER2 Targeting with Trastuzumab and Liposomal-Encapsulated Doxorubicin (MM-302) Demonstrates Synergistic Antitumor Activity in Breast and Gastric Cancer

“…The increase in MM-302 deposition we observed when trastuzumab was coadministered was unexpected, especially so rapidly after injection of the drugs. Most published studies describe the effects of trastuzumab on tumor vessels days to weeks after treatment (rather than hours), although McCormack and colleagues describe changes in VEGF and CD31 in BT474 xenograft tumors 48 hours after administration of trastuzumab (39,41,42). We labeled CD31 to evaluate blood vessels in BT474-M3 tumors following treatment with trastuzumab and observed multiple subtle, yet significant, effects including reductions in vessel length, width and area (data not shown).…”

Vessel contrast enhancement in hyperspectral images

“…A few prior studies exist on vessel enhancement contrast enhancement in hyperspectral or multispectral images of tissue. McCormack et al 1 applied Beer's law and a Gabor filter 2 for extracting vessels from hyperspectral images of mice. However, these mice were imaged through a imaging window, which would naturally elucidate the vessels.…”