Wenxia Zhu

In Vivo Quantification of Myelin Changes in the Vertebrate Nervous System

Destruction or changes associated with myelin membranes in the CNS play a key role in the pathogenesis of multiple sclerosis and other related neurodegenerative disorders. A long-standing goal has been to detect and quantify myelin content in vivo. For this reason, we have developed a myelin-imaging technique based on positron emission tomography (PET). PET is a quantitative imaging modality that has been widely used in clinical settings for direct assessment of biological processes at the molecular level. However, lack of myelin-imaging probes has hampered the use of PET for imaging of myelination in the CNS. Here, we report a myelin-imaging agent, termed Case Imaging Compound (CIC) that readily penetrates the blood–brain barrier and preferentially localizes to myelinated regions of the brain. After radiolabeling with positron-emitting carbon-11, [11C]CIC–PET was conducted in longitudinal studies using a lysolethicin-induced rat model of focal demyelination and subsequent remyelination. Quantitative analysis showed that the retention of [11C]CIC correlates with the level of demyelination/remyelination. These studies indicate that, for the first time, [11C]CIC–PET can be used as an imaging marker of myelination, which has the potential to be translated into clinical studies in multiple sclerosis and other myelin-related diseases for early diagnosis, subtyping, and efficacy evaluation of therapeutic treatments aimed at myelin repair.

In vivo positron emission tomography (PET) imaging of mesenchymal-epithelial transition (MET) receptor.

We report the radiosynthesis and evaluation of 3-[3,5-dimethyl-4-(4-[11C]methylpiperazinecarbonyl)-1H-pyrrol-2-ylmethylene]-2-oxo-2,3-dihydro-1H-indole-5-sulfonic acid (3-chlorophenyl)methylamide, termed [11C]SU11274 ([11C]14) for in vivo imaging of mesenchymal-epithelial transition (MET) receptor by positron emission tomography (PET). Following the synthesis of the precursor (13) that was achieved in 10 steps with a total yield of 9.7%, [11C]14 was obtained through radiomethylation in a range of 5-10% radiochemical yield and over 95% radiochemical purity. For in vivo PET studies, two human lung cancer xenograft models were established using MET-positive NCI-H1975 and MET-negative NCI-H520 cell lines. Quantitative [11C]14-PET studies showed that the tumor uptake of [11C]14 in the NCI-H1975 xenografts was significantly higher than that in the NCI-H520 xenografts, which is consistent with their corresponding immunohistochemical tissue staining patterns of MET receptors from the same animals. These studies demonstrated that [11C]14-PET is an appropriate imaging marker for quantification of MET receptor in vivo, which can facilitate efficacy evaluation in the clinical development of MET-targeted cancer therapeutics.

Direct detection and quantification of abasic sites for in vivo studies of DNA damage and repair

Use of chemotherapeutic agents to induce cytotoxic DNA damage and programmed cell death is a key strategy in cancer treatments. However, the efficacy of DNA-targeted agents such as temozolomide is often compromised by intrinsic cellular responses such as DNA base excision repair (BER). Previous studies have shown that BER pathway resulted in formation of abasic or apurinic/apyrimidinic (AP) sites, and blockage of AP sites led to a significant enhancement of drug sensitivity due to reduction of DNA base excision repair. Since a number of chemotherapeutic agents also induce formation of AP sites, monitoring of these sites as a clinical correlate of drug effect will provide a useful tool in the development of DNA-targeted chemotherapies aimed at blocking abasic sites from repair. Here we report an imaging technique based on positron emission tomography (PET) that allows for direct quantification of AP sites in vivo. For this purpose, positron-emitting carbon-11 has been incorporated into methoxyamine ([(11)C]MX) that binds covalently to AP sites with high specificity. The binding specificity of [(11)C]MX for AP sites was demonstrated by in vivo blocking experiments. Using [(11)C]MX as a radiotracer, animal PET studies have been conducted in melanoma and glioma xenografts for quantification of AP sites. Following induction of AP sites by temozolomide, both tumor models showed significant increase of [(11)C]MX uptake in tumor regions in terms of radioactivity concentration as a function of time, which correlates well with conventional aldehyde reactive probe (ARP)-based bioassays for AP sites.

In vivo Quantification of Abasic Sites for Efficacious Evaluation of DNA Targeted Chemotherapies

DNA damage and repair represent important biological processes that are targets of various chemotherapies against cancer. In many ways, chemotherapeutic agents can induce DNA damage in cancerous as well as normal cells. However, DNA damage induced by chemotherapeutic agents can be intrinsically repaired by normal physiologic responses, which hampers inhibition of tumor growth and cause drug-resistance. Base excision repair (BER) is one such physiologic process that is important in the cellular response to many chemotherapeutic agents, specifically those agents that target DNAs. Once the BER pathway is triggered, damaged DNA bases undergo a series of chemical modifications resulting in the formation of abasic or apurinic/apyrimidinic (AP) site, which serves as key intermediates in the excision of damaged DNA bases and restoration of regular bases. To monitor BER- conferred intrinsic drug-resistance to chemotherapeutic agents such as DNA-alternating temozolomide (TMZ), pemetrexed (Alimta ® ), and fludarabine, we have developed a F-18 labeled fluoroethoxyamine ([ 11 C]FEX) as an imaging agent for positron emission tomography (PET) imaging of DNA damage and repair in vivo . In this work, we report the synthesis, radiolabeling, and evaluation of [ 18 F]FEX in vivo in mice. We have shown that [ 18 F]FEX- PET can be used to monitor DNA damage and repair in tumor xenograft mouse models including an uracil DNA glycosylase (UDG)-knockout tumor mouse model of non-small cell lung cancer (NSCLC).

Abstract 836: In vivo PET imaging of MET receptor in human cancer using [ 11 C]SU11274

The mesenchymal-epithelial transition (MET) receptor plays prominent role in human tumorigenesis, tumor progression and metastasis. Overexpression or mutation of MET results in activation of oncogenic signaling pathways and leads to upregulation of diverse tumor cell functions. Mounting evidence has validated MET receptor as novel target for anticancer therapy. For efficacy evaluation of various MET-targeted anticancer therapies currently under development, it is critical to develop an imaging tool that permits detection and quantification of MET expression in vivo. To date, a number of small molecule MET inhibitors have been developed, including SU11274, which is a specific, reversible MET inhibitor that induces apoptosis in cells transformed by activated MET kinase. Here, we report the radiosynthesis and evaluation of a molecular imaging probe, [ 11 C]SU11274, for quantification of MET receptors in human cancers in vivo. The radiolabeling with C-11 was accomplished through radiomethylation using 3-[3,5-Dimethyl-4-(piperazine-1-carbonyl)-1H-pyrrol-2-ylmethylene]-2-oxo-2,3-dihyro-1H-indole-5-sulfonic acid (3-chlorophenyl)-methyl amide hydrochloride as a key precursor. Following the synthesis of the precursor that was achieved in 10 steps with a total yield of 9.7%, [ 11 C]SU11274 was obtained through radiomethylation in a range of 5-10 % radiochemical yield and over 95% radiochemical purity. For in vivo PET studies, two human lung adenocarcinoma cancer xenograft models were established using MET-positive H1975 and MET-negative H520 cell lines. The time-radioactivity curves showed that the uptake of [ 11 C]SU11274 in the H1975 xenograft was significantly higher than in the H520 xenograft (p = 0.00019). Compared to H520, the H1975 xenograft mice exhibited an increased tumor uptake of [ 11 C]SU11274 throughout the entire measurement period of 90 min. At 65 min post-injection, the radioactivity concentration ratio of H1975/H520 reached a statistically significant and enhanced ratio of 2.1, and peaked at 2.6 at 80 min. The PET imaging results were consistent with their corresponding immunohistochemical tissue staining patterns of MET receptors from the same animals. Hence, our results suggest that [ 11 C]SU11274-PET is a specific imaging marker that can differentiate and quantify MET receptor expression in vivo. These studies demonstrated that [ 11 C]SU11274-PET can facilitate the clinical development of MET-targeted cancer therapeutics. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 836.

Abstract LB-138: In vivo imaging of DNA damage and repair

Objectives: To directly quantify abasic sites induced by DNA-targeted chemotherapeutic agents for in vivo studies of base excision repair pathway. Background: Current cancer treatments rely heavily on chemotherapeutic agents to induce cytotoxic DNA damages and programmed cell death in ancer cells. However, the efficacy of DNA-targeted agents such as temozolomide is often compromised by intrinsic cellular responses such as DNA base excision repair (BER). Previous studies have shown that BER pathway results in formation of abasic or apurinic/apyrimidinic (AP) sites and inhibition of AP sites leads to significant reduction of drug resistance and enhancement of drug sensitivity. Thus, AP-site formation has been identified as an important biomarker in DNA-targeted chemotherapies. Methods: Design, synthesis, and evaluation of PET imaging agents that bind to AP sites with high affinity and specificity. We date, we have developed positron-emitting [ 11 C]methoxyamine for positron emission tomography (PET) that allows for quantification of AP sites in vivo. Results: [ 11 C]methoxyamine has been synthesized with high radiochemical yield and purity. Following radiolabelling, microPET studies have been conducted to evaluate their pharmacokinetic profiles in melanoma and glioma xenograft tumor mouse models that are pre-treated with temozolomide to induce AP-site formation. Subsequent quantitative analysis showed that the radioactivity concentration were elevated in proportion to the AP sites induced in tumor regions pre-treated with temozolomide relative to tumor regions without any treatment. In vivo blocking studies based on microPET also showed that the agents bound to AP sites with high specificity. Conclusion : our studies demonstrated that PET imaging can be used to monitor BER pathway and evaluate efficacy of DNA-targeted therapeutic treatments in cancer. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr LB-138.