Eyal Mishani

Potential 18F-labeled biomarkers for epidermal growth factor receptor tyrosine kinase

Abstract As PET candidate tracers for EGFr-TK, five 4-(anilino)quinazoline derivatives, each fluorinated in the aniline moiety, were prepared. Each was tested in vitro for inhibition of EGFr autophosphorylation in A431 cell line. The leading compounds were then radiolabeled with 18 F and cell binding experiments, biodistribution and PET studies in A431 tumor-bearing mice were performed. Metabolic studies were carried out in a mice control group. From our results, we concluded that while in vitro experiments indicates efficacy of 4-(anilino)quinazoline compounds, kinetic factors and rapid blood clearance make them unsuitable as tracers for nuclear medicine imaging of EGFr-TK.

68Ga-DOTA-NOC PET/CT Imaging of Neuroendocrine Tumors: Comparison with 111In-DTPA-Octreotide (OctreoScan®)

PurposeRecent data have indicated that 68Ga-DOTA-NOC positron emission tomography/X-ray computed tomography (PET/CT) may yield improved images in a shorter acquisition protocol than 111In-DTPA-octreotide (OctreoScan®, OCT). Therefore, we performed a prospective comparison of 68Ga-DOTA-NOC and OCT for the detection of neuroendocrine tumors (NETs).MethodsNineteen patients (eight carcinoid, nine pancreatic NETs, and two NE carcinoma of unknown origin) with previous positive OCT scans underwent 68Ga-DOTA-NOC PET/CT and OCT single-photon emission computed tomography imaging for staging or follow-up. Findings were compared by region and verified with conventional imaging.ResultsAll images of both modalities demonstrated focal uptake, often at multiple sites. 68Ga-DOTA-NOC images were clearer than OCT images, facilitating interpretation. Similar foci were identified with both modalities in 41 regions, with additional foci on 68Ga-DOTA-NOC in 21 and on OCT in 15 regions. CT, magnetic resonance imaging, or ultrasound confirmed the concordant findings in 31 of 41 regions and findings seen with 68Ga-DOTA-NOC only in 15 of 21 regions. Findings seen with OCT only were less clear and were only confirmed in 4 of 15 regions. 68Ga-DOTA-NOC had impact on staging in four patients and on management in three patients.ConclusionsAlthough 68Ga-DOTA-NOC and OCT images were similar, in this study, 68Ga-DOTA-NOC demonstrated more true positive tumor foci and was better tolerated by patients. This direct comparison supports replacement of OCT with 68Ga-DOTA-NOC-PET/CT in the evaluation of NETs.

Blood vessels restrain pancreas branching, differentiation and growth

How organ size and form are controlled during development is a major question in biology. Blood vessels have been shown to be essential for early development of the liver and pancreas, and are fundamental to normal and pathological tissue growth. Here, we report that, surprisingly, non-nutritional signals from blood vessels act to restrain pancreas growth. Elimination of endothelial cells increases the size of embryonic pancreatic buds. Conversely, VEGF-induced hypervascularization decreases pancreas size. The growth phenotype results from vascular restriction of pancreatic tip cell formation, lateral branching and differentiation of the pancreatic epithelium into endocrine and acinar cells. The effects are seen both in vivo and ex vivo, indicating a perfusion-independent mechanism. Thus, the vasculature controls pancreas morphogenesis and growth by reducing branching and differentiation of primitive epithelial cells.

A perfusion-independent role of blood vessels in determining branching stereotypy of lung airways

Blood vessels have been shown to play perfusion-independent roles in organogenesis. Here, we examined whether blood vessels determine branching stereotypy of the mouse lung airways in which coordinated branching of epithelial and vascular tubes culminates in their co-alignment. Using different ablative strategies to eliminate the lung vasculature, both in vivo and in lung explants, we show that proximity to the vasculature is indeed essential for patterning airway branching. Remarkably, although epithelial branching per se proceeded at a nearly normal rate, branching stereotypy was dramatically perturbed following vascular ablation. Specifically, branching events requiring a rotation to change the branching plane were selectively affected. This was evidenced by either the complete absence or the shallow angle of their projections, with both events contributing to an overall flat lung morphology. Vascular ablation also led to a high frequency of ectopic branching. Regain of vascularization fully rescued arrested airway branching and restored normal lung size and its three-dimensional architecture. This role of the vasculature is independent of perfusion, flow or blood-borne substances. Inhibition of normal branching resulting from vascular loss could be explained in part by perturbing the unique spatial expression pattern of the key branching mediator FGF10 and by misregulated expression of the branching regulators Shh and sprouty2. Together, these findings uncovered a novel role of the vasculature in organogenesis, namely, determining stereotypy of epithelial branching morphogenesis.

Labeled EGFr-TK irreversible inhibitor (ML03): In vitro and in vivo properties, potential as PET biomarker for cancer and feasibility as anticancer drug

Radiosynthesis of ML03 (N‐{4‐[(4,5‐dichloro‐2‐fluorophenyl)amino]quinazolin‐6‐yl}acrylamide), an irreversible EGFr‐TK inhibitor, was developed. Its in vitro and in vivo properties, its potential as PET biomarker in cancer and the feasibility of this type of compounds to be used as anticancer drug agents were evaluated. The compound was labeled with carbon‐11 at the acryloyl amide group, via automated method with high yield, chemical and radiochemical purities. ELISA carried out with A431 lysate showed high potency of ML03 with an apparent IC50 of 0.037 nM. The irreversible binding nature of ML03 was studied and 97.5% EGFr‐TK autophosphorylation inhibition was observed in intact A431 cells 8 hr post incubation with the inhibitor. Specific binding (67%) of [11C]ML03 was obtained in cells. An A431 tumor‐bearing rat model was developed and the validity of the model was tested. In biodistribution studies carried out with tumor‐bearing rats, moderate uptake was observed in tumor and high uptake in liver, kidney and intestine. In metabolic studies, fast degradation of [11C]ML03 was observed in liver and blood indicating a short half‐life of the compound in the body. PET scan with tumor‐bearing rats confirmed the results obtained in the ex vivo biodistribution studies. Although in vitro experiments may indicate efficacy of ML03, non‐specific binding, ligand delivery and degradation in vivo make ML03 ineffective as PET bioprobe. Derivatives of ML03 with lower metabolic clearance rate and higher bioavailability should be synthesized and their potential as anticancer drugs and PET bioprobes evaluated.

Imaging of EGFR and EGFR tyrosine kinase overexpression in tumors by nuclear medicine modalities.

Protein tyrosine kinases (PTKs) play a pivotal role in signal transduction pathways and in the development and maintenance of various cancers. They are involved in multiple processes such as transcription, cell cycle progression, proliferation, angiogenesis and inhibition of apoptosis. Among the PTKs, the EGFR is one of the most widely studied and has emerged as a promising key target for the treatment of cancer. Indeed, several drugs directed at this receptor are FDA-approved and many others are at various stages of development. However, thus far, the therapeutic outcome of EGFR-targeted therapy is suboptimal and needs to be refined. Quantitative PET molecular imaging coupled with selective labelled biomarkers may facilitate in vivo EGFR-targeted drug efficacy by noninvasively assessing the expression of EGFR in tumor, guiding dose and regime by measuring target drug binding and receptor occupancy as well as potentially detecting the existence of a primary or secondary mutation leading to either drug interaction or failure of EGFR recognition by the drug. This review describes the attempts to develop labelled EGFR molecular imaging agents that are based either on low molecular weight tyrosine kinase inhibitors or monoclonal antibodies directed to the extracellular binding domain of the receptor to be used in nuclear medicine modalities.

Strategies for Molecular Imaging of Epidermal Growth Factor Receptor Tyrosine Kinase in Cancer

A wealth of research has focused on developing targeted cancer therapies by specifically inhibiting epidermal growth factor receptor tyrosine kinase (EGFR-TK). However, the outcome of most EGFR-TK–targeted drugs that were approved by the Food and Drug Administration or entered clinical trials has been only moderate. Enhancement of EGFR-targeted therapy hinges on a reliable in vivo quantitative molecular imaging method. Such a method would enable monitoring of receptor drug binding and receptor occupancy in vivo; determination of the duration of EGFR inhibition in vivo; and, potentially, identification of a primary or secondary mutation in EGFR leading to drug interaction or loss of EGFR recognition by the drug. This review analyzes the most recent strategies to visualize and quantify EGFR-TK in cancer by nuclear medicine imaging and describes future directions.

Cancer molecular imaging: radionuclide-based biomarkers of the epidermal growth factor receptor (EGFR).

Overexpression of the epidermal growth factor receptor tyrosine kinase (EGFR-TK) has been documented in numerous human cancers of epithelial origin, and was found to correlate with resistance to treatment and poor prognosis. Recognizing the central role that this receptor plays in cancer development and progression, various approaches have been developed to target it in order to more specifically eradicate cancer cells. These methods include, among others, low-molecular weight inhibitors of the TK domain that are commonly designed to treat those tumors that overexpress the EGFR. Nevertheless, no currently available assay provides non-invasive, longitudinal and sensitive quantitation of receptor levels in tumors so as to better identify candidate patients for EGFR-targeted therapies. Hence, attempts have been made to develop radiolabeled molecular imaging agents as potential bioprobes to quantitatively monitor treatment efficiency. Such EGFR-targeted bioprobes could not only improve patient selection and treatment monitoring, but also allow a direct delivery of radionuclides for radiotherapy. In this review, the role that EGFR plays in cancer development and therapy is briefly presented, followed by a short review of prominent milestones in the development of EGFR-TK inhibitors. These inhibitors constitute the fundamental core structure for the development of radiolabeled probes to visualize the EGFR in vivo. The considerations that need to be taken into account for the development of such probes will be presented, along with a critical examination on the progress that has been made thus far in the field.

PolyIC GE11 polyplex inhibits EGFR‐overexpressing tumors

Phage display has identified the dodecapeptide YHWYGYTPQNVI (GE11) as a ligand that binds to the epidermal growth factor receptor (EGFR) but does not activate the receptor. Here, we compare the EGFR binding affinities of GE11, EGF, and their polyethyleneimine‐polyethyleneglycol (PEI‐PEG) conjugates. We found that although GE11 by itself does not exhibit measurable affinity to the EGFR, tethering it to PEI‐PEG increases its affinity markedly, and complex formation with polyinosine/cytosine (polyIC) further enhances the affinity to the submicromolar range. PolyIC/PPGE11 has a similar strong antitumor effect against EGFR overexpressing tumors in vitro and in vivo, as polyIC/polyethyleneimine‐polyetheleneglycol‐EGF (polyIC/PP‐EGF). Absence of EGFR activation, as previously shown by us and easier production of GE11 and GE11 conjugates, confer polyIC/PPGE11 a significant advantage over similar EGF‐based polyplexes as a potential therapy of EGFR overexpressing tumors.

11C-acetate PET/CT in bladder urothelial carcinoma: intraindividual comparison with 11C-choline.

Objective: We present a first study of the use of 11C-acetate (ACET) positron emission tomography (PET)/computed tomography (CT) in bladder urothelial carcinoma (UC) and an intraindividual comparison with 11C- choline (CHOL) PET/CT. Methods: Fourteen patients with biopsy-proven UC (11 T2, 3 T1 refractory to treatment) were prospectively evaluated before radical cystectomy and excision of pelvic lymph nodes (LNs), with ACET and CHOL PET/CT scans performed within 1 week. Image acquisition started 5 minutes after intravenous injection of 12 to 14 mCi for both tracers. Standardized uptake values (SUVs) and tumor-to-background ratios (TBR) were calculated for all tumor and nodal findings and correlated with histopathology and follow-up. Results: ACET and CHOL were taken up in all UCs, involved LNs, and prostate pathology. SUVs were on average slightly, nonsignificantly higher for CHOL uptake (SUV) in UCs and significantly higher for ACET in LNs. TBR was nonsignificantly higher with CHOL for UC and significantly higher for LNs. Conclusions: In this preliminary series, 11C-ACET and 11C-CHOL PET/CT showed equivalent results in the preoperative evaluation of UC. Both tracers have the potential to contribute to selecting patients who would benefit from combined treatment—neoadjuvant chemotherapy and surgery—by identifying pathologic LNs or from surgery only, thanks to their high negative predictive value for LN involvement.