Current Role at Stanford
Director of the Cyclotron and Radiochemistry Facility
The CRF team is excited to be designing our second cyclotron facility where we will expand production into radiometals.
Education & Certifications
PhD, Imperial College London, Chemistry (2017)
MSc, University of Bristol, Chemistry (2012)
PET imaging of focused-ultrasound enhanced delivery of AAVs into the murine brain.
2023; 13 (15): 5151-5169
Rationale: Despite recent advances in the use of adeno-associated viruses (AAVs) as potential vehicles for genetic intervention of central and peripheral nervous system-associated disorders, gene therapy for the treatment of neuropathology in adults has not been approved to date. The currently FDA-approved AAV-vector based gene therapies rely on naturally occurring serotypes, such as AAV2 or AAV9, which display limited or no transport across the blood-brain barrier (BBB) if systemically administered. Recently developed engineered AAV variants have shown broad brain transduction and reduced off-target liver toxicity in non-human primates (NHPs). However, these vectors lack spatial selectivity for targeted gene delivery, a potentially critical limitation for delivering therapeutic doses in defined areas of the brain. The use of microbubbles, in conjunction with focused ultrasound (FUS), can enhance regional brain AAV transduction, but methods to assess transduction in vivo are needed. Methods: In a murine model, we combined positron emission tomography (PET) and optical imaging of reporter gene payloads to non-invasively assess the spatial distribution and transduction efficiency of systemically administered AAV9 after FUS and microbubble treatment. Capsid and reporter probe accumulation are reported as percent injected dose per cubic centimeter (%ID/cc) for in vivo PET quantification, whereas results for ex vivo assays are reported as percent injected dose per gram (%ID/g). Results: In a study spanning accumulation and transduction, mean AAV9 accumulation within the brain was 0.29 %ID/cc without FUS, whereas in the insonified region of interest of FUS-treated mice, the spatial mean and maximum reached ~2.3 %ID/cc and 4.3 %ID/cc, respectively. Transgene expression assessed in vivo by PET reporter gene imaging employing the pyruvate kinase M2 (PKM2)/[18F]DASA-10 reporter system increased up to 10-fold in the FUS-treated regions, as compared to mice receiving AAVs without FUS. Systemic injection of AAV9 packaging the EF1A-PKM2 transgene followed by FUS in one hemisphere resulted in 1) an average 102-fold increase in PKM2 mRNA concentration compared to mice treated with AAVs only and 2) a 12.5-fold increase in the insonified compared to the contralateral hemisphere of FUS-treated mice. Conclusion: Combining microbubbles with US-guided treatment facilitated a multi-hour BBB disruption and stable AAV transduction in targeted areas of the murine brain. This unique platform has the potential to provide insight and aid in the translation of AAV-based therapies for the treatment of neuropathologies.
View details for DOI 10.7150/thno.85549
View details for PubMedID 37908737
View details for PubMedCentralID PMC10614693
Preclinical evaluation of 89Zr-Panitumumab for biology-guided radiotherapy.
International journal of radiation oncology, biology, physics
Biology-guided radiotherapy (BgRT) uses real-time line-of-response data from on-board PET detectors to guide beamlet delivery during therapeutic radiation. The current workflow requires 18F-fluorodeoxyglucose (FDG) administration daily prior to each treatment fraction. However, there are advantages to reducing the number of tracer injections by using a PET tracer with a longer decay time. In this context, we investigated 89Zr-Panitumumab (89Zr-Pan), an antibody PET tracer with a half-life of 78 hours that can be imaged for up to 9 days using PET.The BgRT workflow was evaluated pre-clinically in mouse colorectal cancer xenografts (HCT116) using small-animal PET/CT for imaging, and image-guided kilovoltage conformal irradiation for therapy. Mice (n=5 per group) received 7 MBq of 89Zr-Pan as a single dose 2 weeks after tumor induction, with or without fractionated radiation therapy (RT; 6×6.6 Gy) to the tumor region. The mice were imaged longitudinally to assess the kinetics of the tracer over 9 days. PET images were then analyzed to determine the stability of the PET signal in irradiated tumors over time.Mice in the treatment group experienced complete tumor regression, whereas those in the control group were sacrificed due to tumor burden. PET imaging of 89Zr-Pan showed well-delineated tumors with minimal background in both groups. On day 9 post-injection, tumor uptake of 89Zr-Pan was 7.2 ± 1.7 in the control group vs 5.2 ± 0.5 in the treatment group (mean %ID/g ± SD; P = 0.07), both significantly higher than FDG uptake (1.1 ± 0.5 %ID/g) 1 hour post injection. To assess BgRT feasibility, the clinical eligibility criteria was computed using human-equivalent uptake values that were extrapolated from preclinical PET data. Based on this semiquantitative analysis, BgRT may be feasible for 5 consecutive days following a single 740 MBq injection of 89Zr-Pan.This study indicates the potential of long-lived antibody-based PET tracers for guiding clinical BgRT.
View details for DOI 10.1016/j.ijrobp.2023.01.007
View details for PubMedID 36669541
Multimodal imaging of capsid and cargo reveals differential brain targeting and liver detargeting of systemically-administered AAVs.
The development of gene delivery vehicles with high organ specificity when administered systemically is a critical goal for gene therapy. We combine optical and positron emission tomography (PET) imaging of 1) reporter genes and 2) capsid tags to assess the temporal and spatial distribution and transduction of adeno-associated viruses (AAVs). AAV9 and two engineered AAV vectors (PHP.eB and CAP-B10) that are noteworthy for maximizing blood-brain barrier transport were compared. CAP-B10 shares a modification in the 588 loop with PHP.eB, but also has a modification in the 455 loop, added with the goal of reducing off-target transduction. PET and optical imaging revealed that the additional modifications retained brain receptor affinity. In the liver, the accumulation of AAV9 and the engineered AAV capsids was similar (15% of the injected dose per cc and not significantly different between capsids at 21h). However, the engineered capsids were primarily internalized by Kupffer cells rather than hepatocytes, and liver transduction was greatly reduced. PET reporter gene imaging after engineered AAV systemic injection provided a non-invasive method to monitor AAV-mediated protein expression over time. Through comparison with capsid tagging, differences between brain localization and transduction were revealed. In summary, AAV capsids bearing imaging tags and reporter gene payloads create a unique and powerful platform to assay the pharmacokinetics, cellular specificity and protein expression kinetics of AAV vectors in vivo, a key enabler for the field of gene therapy.
View details for DOI 10.1016/j.biomaterials.2022.121701
View details for PubMedID 35985893
Radiosynthesis and initial preclinical evaluation of [11C]AZD1283 as a potential P2Y12R PET radiotracer.
Nuclear medicine and biology
INTRO: Chronic neuroinflammation and microglial dysfunction are key features of many neurological diseases, including Alzheimer's Disease and multiple sclerosis. While there is unfortunately a dearth of highly selective molecular imaging biomarkers/probes for studying microglia in vivo, P2Y12R has emerged as an attractive candidate PET biomarker being explored for this purpose. Importantly, P2Y12R is selectively expressed on microglia in the CNS and undergoes dynamic changes in expression according to inflammatory context (e.g., toxic versus beneficial/healing states), thus having the potential to reveal functional information about microglia in living subjects. Herein, we identified a high affinity, small molecule P2Y12R antagonist (AZD1283) to radiolabel and assess as a candidate radiotracer through in vitro assays and in vivo positron emission tomography (PET) imaging of both wild-type and total knockout mice and a non-human primate.METHODS: First, we evaluated the metabolic stability and passive permeability of non-radioactive AZD1283 in vitro. Next, we radiolabeled [11C]AZD1283 with radioactive precursor [11C]NH4CN and determined stability in formulation and human plasma. Finally, we investigated the in vivo stability and kinetics of [11C]AZD1283 via dynamic PET imaging of naive wild-type mice, P2Y12R knockout mouse, and a rhesus macaque.RESULTS: We determined the half-life of AZD1283 in mouse and human liver microsomes to be 37 and>160min, respectively, and predicted passive CNS uptake with a small amount of active efflux, using a Caco-2 assay. Our radiolabeling efforts afforded [11C]AZD1283 in an activity of 12.69±10.64mCi with high chemical and radiochemical purity (>99%) and molar activity of 1142.84±504.73mCi/mumol (average of n=3). Of note, we found [11C]AZD1283 to be highly stable in vitro, with >99% intact tracer present after 90min of incubation in formulation and 60min of incubation in human serum. PET imaging revealed negligible brain signal in healthy wild-type mice (n=3) and a P2Y12 knockout mouse (0.55±0.37%ID/g at 5min post injection). Strikingly, high signal was detected in the liver of all mice within the first 20min of administration (peak uptake=58.28±18.75%ID/g at 5min post injection) and persisted for the remaining duration of the scan. Ex vivo gamma counting of mouse tissues at 60min post-injection mirrored in vivo data with a mean %ID/g of 0.9%±0.40, 0.02%±0.01, and 106±29.70% in the blood, brain, and liver, respectively (n=4). High performance liquid chromatography (HPLC) analysis of murine blood and liver metabolite samples revealed a single radioactive peak (relative area under peak: 100%), representing intact tracer. Finally, PET imaging of a rhesus macaque also revealed negligible CNS uptake/binding in monkey brain (peak uptake=0.37 Standard Uptake Values (SUV)).CONCLUSION: Despite our initial encouraging liver microsome and Caco-2 monolayer data, in addition to the observed high stability of [11C]AZD1283 in formulation and human serum, in vivo brain uptake was negligible and rapid accumulation was observed in the liver of both naive wildtype and P2Y12R knockout mice. Liver signal appeared to be independent of both metabolism and P2Y12R expression due to the confirmation of intact tracer in this tissue for both wildtype and P2Y12R knockout mice. In Rhesus Macaque, negligible uptake of [11C]AZD1283 brain indicates a lack of potential for translation or its further investigation in vivo. P2Y12R is an extremely promising potential PET biomarker, and the data presented here suggests encouraging metabolic stability for this scaffold; however, the mechanism of liver uptake in mice should be elucidated prior to further analogue development.
View details for DOI 10.1016/j.nucmedbio.2022.05.001
View details for PubMedID 35680502
Synthesis and Characterization of 9-(4-[18F]Fluoro-3-(hydroxymethyl)butyl)-2-(phenylthio)-6-oxopurine as a Novel PET Agent for Mutant Herpes Simplex Virus Type 1 Thymidine Kinase Reporter Gene Imaging.
Molecular imaging and biology
PURPOSE: [18F]FHBG has been used as a positron emission tomography (PET) imaging tracer for the monitoring of herpes simplex virus type 1 thymidine kinase (HSV1-tk), a reporter gene for cell and gene therapy in humans. However, this tracer shows inadequate blood-brain barrier (BBB) penetration and, therefore, would be limited for accurate quantification of reporter gene expression in the brain. Here, we report the synthesis and evaluation of 9-(4-[18F]fluoro-3-(hydroxymethyl)butyl)-2(phenylthio)-6-oxopurine ([18F]FHBT) as a new PET tracer for imaging reporter gene expression of HSV1-tk and its mutant HSV1-sr39tk, with the aim of improved BBB penetration.PROCEDURES: [18F]FHBT was prepared by using a tosylate precursor and [18F]KF. The cellular uptake of [18F]FHBT was performed in HSV1-sr39tk-positive (+) or HSV1-sr39tk-negative (-) MDA-MB-231 breast cancer cells. The specificity of [18F]FHBT to assess HSV1-sr39tk expression was evaluated by in vitro blocking studies using 1mM of ganciclovir (GCV). Penetration of [18F]FHBT and [18F]FHBG across the BBB was assessed by dynamic PET imaging studies in normal mice.RESULTS: The tosylate precursor reacted with [18F]KF using Kryptofix2.2.2 followed by deprotection to give [18F]FHBT in 10% radiochemical yield (decay-corrected). The uptake of [18F]FHBT in HSV1-sr39tk (+) cells was significantly higher than that of HSV1-sr39tk (-) cells. In the presence of GCV (1mM), the uptake of [18F]FHBT was significantly decreased, indicating that [18F]FHBT serves as a selective substrate of HSV1-sr39TK. PET images and time-activity curves of [18F]FHBT in the brain regions showed similar initial brain uptakes (~12.75min) as [18F]FHBG (P>0.855). Slower washout of [18F]FHBT was observed at the later time points (17.75 - 57.75 min, P >0.207).CONCLUSIONS: Although [18F]FHBT showed no statistically significant improvement of BBB permeability compared with [18F]FHBG, we have demonstrated that the 2-(phenylthio)-6-oxopurine backbone can serve as a novel scaffold for developing HSV1-tk/HSV1-sr39tk reporter gene imaging agents for additional research in the future.
View details for DOI 10.1007/s11307-020-01517-5
View details for PubMedID 32691392
Synthesis and validation of [F-18]6 ''-fluromaltotriose, a radiotracer for imaging bacterial infections
WILEY. 2019: S315–S316
View details for Web of Science ID 000468965200248
Synthesis and characterization of 9-(4fluoro-3-(hydroxymethyl)butyl)-2-(phenylthio)6-oxopurine ([F-18]FHBT) as a PET tracer for HSV1-tk reporter gene imaging
WILEY. 2019: S537–S538
View details for Web of Science ID 000468965200450
The discovery of quinoline-3-carboxamides as hematopoietic prostaglandin D synthase (H-PGDS) inhibitors.
Bioorganic & medicinal chemistry
With the goal of discovering more selective anti-inflammatory drugs, than COX inhibitors, to attenuate prostaglandin signaling, a fragment-based screen of hematopoietic prostaglandin D synthase was performed. The 76 crystallographic hits were sorted into similar groups, with the 3-cyano-quinoline 1a (FP IC50 = 220,000 nM, LE = 0.43) being a potent member of the 6,6-fused heterocyclic cluster. Employing SAR insights gained from structural comparisons of other H-PGDS fragment binding mode clusters, the initial hit 1a was converted into the 70-fold more potent quinoline 1d (IC50 = 3,100 nM, LE = 0.49). A systematic substitution of the amine moiety of 1d, utilizing structural information and array chemistry, with modifications to improve inhibitor stability, resulted in the identification of the 300-fold more active H-PGDS inhibitor tool compound 1bv (IC50 = 9.9 nM, LE = 0.42). This selective inhibitor exhibited good murine pharmacokinetics, dose-dependently attenuated PGD2 production in a mast cell degranulation assay and should be suitable to further explore H-PGDS biology.
View details for DOI 10.1016/j.bmc.2019.02.017
View details for PubMedID 30858025
Electrochemical [(11)C]CO2 to [(11)C]CO conversion for PET imaging.
Chemical communications (Cambridge, England)
2017; 53 (20): 2982-2985
The development of a novel electrochemical methodology to generate carbon-11 carbon monoxide ([(11)C]CO) from cyclotron-produced carbon-11 carbon dioxide ([(11)C]CO2) using Ni(cyclam) and Zn(cyclen) complexes is described. This methodology allows up to 10% yields of [(11)C]CO from [(11)C]CO2. Produced [(11)C]CO was subsequently converted to [(11)C]N-benzylbenzamide under mild conditions with a radiochemical purity (RCP) of >98%.
View details for DOI 10.1039/c7cc00319f
View details for PubMedID 28234400