Kyle Kuszpit

[18F]FLT–PET Imaging Does Not Always “Light Up” Proliferating Tumor Cells

Purpose: [18F]FLT (3′-Fluoro-3′ deoxythymidine)–PET imaging was proposed as a tool for measuring in vivo tumor cell proliferation. The aim of this article was to validate the use of [18F]FLT–PET imaging for measuring xenograft proliferation and subsequent monitoring of targeted therapy. Experimental Design: In exponentially growing xenografts, factors that could impact the outcome of [18F]FLT–PET imaging, such as nucleoside transporters, thymidine kinase 1, the relative contribution of DNA salvage pathway, and the ratio of FLT to thymidine, were evaluated. The [18F]FLT tracer avidity was compared with other proliferation markers. Results: In a panel of proliferating xenografts, [18F]FLT or [3H]thymidine tracer avidity failed to reflect the tumor growth rate across different tumor types, despite the high expressions of Ki67 and TK1. When FLT was injected at the same dose level as used in the preclinical [18F]FLT–PET imaging, the plasma exposure ratio of FLT to thymidine was approximately 1:200. Thymidine levels in different tumor types seemed to be variable and exhibited an inverse relationship with the FLT tracer avidity. In contrast, high-dose administration of bromdeoxyuridine (BrdUrd; 50 mg/kg) yielded a plasma exposure of more than 4-fold higher than thymidine and leads to a strong correlation between the BrdUrd uptake and the tumor proliferation rate. In FLT tracer-avid models, [18F]FLT–PET imaging as a surrogate biomarker predicted the therapeutic response of CDK4/6 inhibitor PD-0332991. Conclusions: Tumor thymidine level is one of the factors that impact the correlation between [18F]FLT uptake and tumor cell proliferation. With careful validation, [18F]FLT–PET imaging can be used to monitor antiproliferative therapies in tracer-avid malignancies. Clin Cancer Res; 18(5); 1303–12. ©2011 AACR.

A Methodology for Registration of a Histological Slide and in vivo MRI Volume based on Optimizing Mutual Information

We present a method for registering histology and in vivo imaging that requires minimal microtoming and is automatic following the users initialization. In this demonstration, we register a single hematoxylin-and-eosin-stained histological slide of a coronal section of a rat brain harboring a 9L gliosarcoma with an in vivo 7T MR image volume of the same brain. Because the spatial resolution of the in vivo MRI is limited, we add the step of obtaining a high spatial resolution, ex vivo MRI in situ for intermediate registration. The approach taken was to maximize mutual information in order to optimize the registration between all pairings of image data whether the sources are MRI, tissue block photograph, or stained sample photograph. The warping interpolant used was thin plate splines with the appropriate basis function for either 2-D or 3-D applications. All registrations were implemented by user initialization of the approximate pose between the two data sets, followed by automatic optimization based on maximizing mutual information. Only the higher quality anatomical images were used in the registration process; however, the spatial transformation was directly applied to a quantitative diffusion image. Quantitative diffusion maps from the registered location appeared highly correlated with the H&E slide. Overall, this approach provides a robust method for coregistration of in vivo images with histological sections and will have broad applications in the field of functional and molecular imaging.

PF-03732010: a fully human monoclonal antibody against P-cadherin with antitumor and antimetastatic activity.

Purpose: P-cadherin is a membrane glycoprotein that functionally mediates tumor cell adhesion, proliferation, and invasiveness. We characterized the biological properties of PF-03732010, a human monoclonal antibody against P-cadherin, in cell-based assays and tumor models. Experimental Design: The affinity, selectivity, and cellular inhibitory activity of PF-03732010 were tested in vitro. Multiple orthotopic and metastatic tumor models were used for assessing the antitumor and antimetastatic activities of PF-03732010. Treatment-associated pharmacodynamic changes were also investigated. Results: PF-03732010 selectively inhibits P-cadherin–mediated cell adhesion and aggregation in vitro. In the P-cadherin–overexpressing tumor models, including MDA-MB-231-CDH3, 4T1-CDH3, MDA-MB-435HAL-CDH3, HCT116, H1650, PC3M-CDH3, and DU145, PF-03732010 inhibited the growth of primary tumors and metastatic progression, as determined by bioluminescence imaging. Computed tomography imaging, H&E stain, and quantitative PCR analysis confirmed the antimetastatic activity of PF-03732010. In contrast, PF-03732010 did not show antitumor and antimetastatic efficacy in the counterpart tumor models exhibiting low P-cadherin expression. Mechanistic studies via immunofluorescence, immunohistochemical analyses, and 3′-[18F]fluoro-3′-deoxythymidine–positron emission tomography imaging revealed that PF-03732010 suppressed P-cadherin levels, caused degradation of membrane β-catenin, and concurrently suppressed cytoplasmic vimentin, resulting in diminished metastatic capacity. Changes in the levels of Ki67, caspase-3, and 3′-[18F]fluoro-3′-deoxythymidine tracer uptake also indicated antiproliferative activity and increased apoptosis in the tested xenografts. Conclusions: These findings suggest that interrupting the P-cadherin signaling pathway may be a novel therapeutic approach for cancer therapy. PF-03732010 is presently undergoing evaluation in Phase 1 clinical trials. Clin Cancer Res; 16(21); 5177–88. ©2010 AACR.

PF-00477736 Mediates Checkpoint Kinase 1 Signaling Pathway and Potentiates Docetaxel-Induced Efficacy in Xenografts

Purpose: Checkpoint kinase 1 (Chk1) plays a critical role in the activation of mitotic spindle checkpoint and DNA damage checkpoint. We examined the preclinical use of the Chk1 inhibitor PF-00477736 as a docetaxel-sensitizing agent. Specifically, we investigated the correlation between PF-00477736–mediated modulation of biomarkers and the sensitization of docetaxel efficacy. Experimental Design:In vitro and in vivo studies using COLO205 and other cell lines were done to assess PF-00477736–induced enhancement of docetaxel efficacy and effects on associated biomarkers. Results: PF-00477736 significantly enhanced the docetaxel-induced efficacy in tumor cells and xenografts. Docetaxel induced dose- and time-dependent increase in the levels of phosphorylated Chk1 (Ser345), phosphorylated histone H3 (Ser10), and γH2AX foci and promoted the cytoplasmic localization of phosphorylated Cdc25C (Ser216). PF-00477736 cotreatment suppressed docetaxel-induced changes in phosphorylated histone H3 and cytoplasmic phosphorylated Cdc25C (Ser216) levels and concurrently sensitized the docetaxel-induced apoptosis. Docetaxel alone or in combination with PF-00477736 induced significant antiproliferative activity in xenografts, shown via [18F]FLT-PET imaging. However, changes in [18F]FLT uptake did not reflect the potentiation of docetaxel efficacy. In contrast, bioluminescence imaging showed that PF-00477736 sensitized docetaxel-induced suppression of tumor survival. Conclusions: Docetaxel triggers mitotic spindle checkpoint activation at low concentrations and activates both the DNA damage checkpoint and the spindle checkpoint at high concentrations. In combination with docetaxel, PF-00477736 abrogates the mitotic checkpoint, as well as the DNA damage checkpoint, and results in sensitization to docetaxel. Chk1 inhibitor PF-00477736 offers a therapeutic potential for the enhancement of taxane therapy.

Rapid and differential losses of in vivo dopamine transporter (DAT) and vesicular monoamine transporter (VMAT2) radioligand binding in MPTP-treated mice

The dose‐ and time‐dependent changes of in vivo radioligand binding to the neuronal membrane dopamine transporter (DAT) and vesicular monoamine transporter type 2 (VMAT2) were examined in mouse brain after MPTP (1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine) administrations. Regional brain distribution studies were done in male C57BL/6 mice using simultaneous injections of d‐threo‐[3H]methylphenidate (DAT) and (+)‐α‐[11C]dihydrotetrabenazine (VMAT2). Single (55 mg/kg i.p.) or multiple (4 × 10 mg/kg i.p., 1‐hour intervals) administration of MPTP caused significant reductions in [3H]methylphenidate and [11C]dihydrotetrabenazine specific striatal binding, measured 14 days later. The single high dose of MPTP produced greater losses of [11C]dihydrotetrabenazine binding than did the multiple MPTP dosing regimen. Using the single high dose of MPTP, changes of in vivo binding of the two radioligands were determined at 1, 3, and 14 days after neurotoxin injection. At 1 day, there are large losses of [3H]methylphenidate binding (DAT) but no changes in [11C]dihydrotetrabenazine binding to the VMAT2 site in the striatum. At 3 and 14 days, there were >50% losses of binding of both bot radioligands, but significantly (P < 0.001) greater losses of VMAT2 binding of [11C]dihydrotetrabenazine. These studies indicate that the losses of the neuronal membrane and vesicular transporters are not always equal, and do not occur in the same time frame, after administration of the neurotoxin MPTP. Synapse 35:250–255, 2000.

Evaluation of Treatment-Associated Inflammatory Response on Diffusion-Weighted Magnetic Resonance Imaging and 2-[18F]-Fluoro-2-Deoxy-d-Glucose-Positron Emission Tomography Imaging Biomarkers

Purpose: Functional imaging biomarkers of cancer treatment response offer the potential for early determination of outcome through the assessment of biochemical, physiologic, and microenvironmental readouts. Cell death may result in an immunologic response, thus complicating the interpretation of biomarker readouts. This study evaluated the temporal effect of treatment-associated inflammatory activity on diffusion magnetic resonance imaging and 2-[18F]-fluoro-2-deoxy-d-glucose-positron emission tomography imaging (FDG-PET) biomarkers to delineate the effects of the inflammatory response on imaging readouts. Experimental Design: Rats with intracerebral 9L gliosarcomas were separated into four groups consisting of control, an immunosuppressive agent dexamethasone (Dex), 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU), and BCNU+Dex. Animals were imaged using diffusion-weighted magnetic resonance imaging and FDG-PET at 0, 3, and 7 days posttreatment. Results: In the BCNU- and BCNU+Dex-treated animal groups, diffusion values increased progressively over the 7-day study period to ∼23% over baseline. The FDG percentage change of standard uptake value decreased at day 3 (−30.9%) but increased over baseline levels at day 7 (+20.1%). FDG-PET of BCNU+Dex-treated animals were found to have percentage of standard uptake value reductions of −31.4% and −24.7% at days 3 and 7, respectively, following treatment. Activated macrophages were observed on day 7 in the BCNU treatment group with much fewer found in the BCNU+Dex group. Conclusions: Results revealed that treatment-associated inflammatory response following tumor therapy resulted in the accentuation of tumor diffusion response along with a corresponding increase in tumor FDG uptake due to the presence of glucose-consuming activated macrophages. The dynamics and magnitude of potential inflammatory response should be considered when interpreting imaging biomarker results. Clin Cancer Res; 16(5); 1542–52