Leveraging Programmatic Collaboration for a Radiopharmaceutical Clinic.

Radiation oncologists, radiopharmacists, nuclear medicine physicians, and medical oncologists have seen a renewed clinical interest in radiopharmaceuticals for the curative or the palliative treatment of cancer. To allow for the discovery and the clinical advancement of targeted radiopharmaceuticals, these stakeholders have reformed their trial efforts and remodeled their facilities to accommodate the obligations of a program centered upon radioactive investigational drug products. Now considered informally as drugs and not beam radiotherapy, radiopharmaceuticals can be more easily studied in the traditional clinical trial enterprise ranging from phase 0-I to phase III studies. Resources and physical facilities allocated to radiopharmaceuticals have brought forth new logistics and patient experience for safe and satisfactory drug delivery. The clinical use of theranostic agents-that is, diagnostic and therapeutic radionuclide pairs-has accelerated radiopharmaceutical development.

Practical Considerations for Implementation of 177Lu-DOTATATE Neuroendocrine Tumor Treatment Programs.

The 2018 FDA approval of 177Lu-DOTATATE for the treatment of somatostatin receptor-positive (SSTR) neuroendocrine tumors (NETs) represents a paradigm shifting approach to cancer treatments around the globe. Gastroenteropancreatic (GEP) NETs overexpress the somatostatin subtype receptor 2, which is now exploited for receptor-based imaging and therapy, thus generating significant progress in the diagnosis and treatment of this orphan disease. The recent FDA approval of receptor-based PET radiopharmaceuticals and a new peptide receptor radiopharmaceutical therapy (PRRT), 177Lu-DOTATATE, has dramatically impacted NET patient management. The focus of this paper is to review clinical considerations associated with implementing a 177Lu-DOTATATE program. We review receptor-based NET radiopharmaceuticals, 177Lu-DOTATATE patient selection criteria, administration methods, clinical, regulatory, and radiation safety considerations, technical factors, tissue dosimetry, and reimbursement guidelines.

"Click"-cyclized (68)Ga-labeled peptides for molecular imaging and therapy: synthesis and preliminary in vitro and in vivo evaluation in a melanoma model system.

Cyclization techniques are used often to impart higher in vivo stability and binding affinity to peptide targeting vectors for molecular imaging and therapy. The two most often used techniques to impart these qualities are lactam bridge construction and disulfide bond formation. While these techniques have been demonstrated to be effective, orthogonal protection/deprotection steps can limit achievable product yields. In the work described in this chapter, new α-melanocyte stimulating hormone (α-MSH) peptide analogs were synthesized and cyclized by copper-catalyzed terminal azide-alkyne cycloaddition "click" chemistry techniques. The α-MSH peptide and its cognate receptor (melanocortin receptor subtype 1, MC1R) represent a well-characterized model system to examine the effect of the triazole linkage for peptide cyclization on receptor binding in vitro and in vivo. Four new DOTA-conjugated α-MSH analogs were cyclized and evaluated by in vitro competitive binding assays, serum stability testing, and in vivo imaging by positron emission tomography (PET) of tumor-bearing mice. These new DOTA-conjugated click-cyclized analogs exhibited selective high binding affinity (<2 nM) for MC1R on melanoma cells in vitro, high stability in human serum, and produced high-contrast PET/CT images of tumor xenografts. (68)Ga-labeled DOTA bioconjugates displayed rapid pharmacokinetics with receptor-mediated tumor accumulation of up to 16 ± 5% ID/g. The results indicate that the triazole ring is an effective bioisosteric replacement for the standard lactam bridge assemblage for peptide cyclization. Radiolabeling results confirm that Cu catalyst is sufficiently removed prior to DOTA chelator addition to enable insertion of radio metals or stable metals for molecular imaging and therapy. Thus, these click-chemistry-cyclized variants show promise as agents for melanocortin receptor-targeted imaging and radionuclide therapy.

Improved synthesis and biological evaluation of chelator-modified α-MSH analogs prepared by copper-free click chemistry.

Radionuclide chelators (DOTA, NOTA) functionalized with a monofluorocyclooctyne group were prepared. These materials reacted rapidly and in high yield with a fully deprotected azide-modified peptide via Cu-free click chemistry under mild reaction conditions (aqueous solution, room temperature). The resulting bioconjugates bind with high affinity and specificity to their cell-surface receptor targets in vitro and appear stable to degradation in mouse serum over 3h of incubation at 37°C.

A DOTA-peptide conjugate by copper-free click chemistry.

Attachment of DOTA to a novel monofluoro-cyclooctyne facilitates bioconjugation to an azide-modified peptide via Cu-free click chemistry. The resulting conjugate was radiolabeled with (111)In to afford a potential targeted molecular imaging agent with high specific activity and an excellent radiochemical purity.

A novel class of intrinsic proteasome inhibitory gene transfer peptides.

Proteasomes are multisubunit enzymes responsible for the degradation of many cytosolic proteins. The inhibition of the proteasome has been the subject of intense interest in the development of drug therapies. We have previously demonstrated that simultaneous administration of a tripeptide aldehyde proteasome inhibitor (MG115 or MG132) with a peptide (Cys-Trp-Lys18) DNA condensate boosted gene expression by 30-fold in cell culture. In the present study, we have developed a convergent synthesis to allow the incorporation of a proteasome inhibitor tripeptide into the C-terminal end of a gene delivery peptide. The resulting peptides formed DNA condensates that mediated a 100-fold enhancement in gene expression over a control peptide lacking all or part of the tripeptide inhibitor. Gene transfer peptides possessing intrinsic proteasome inhibitors were also found to be nontoxic to cells in culture. These results suggest that intrinsic proteasome inhibition may also be used to boost the efficiency of peptide-mediated nonviral gene delivery systems in vivo.

Peptide-guided gene delivery.

Although currently less efficient than their viral counterparts, nonviral vectors are under intense investigation as a safer alternative for gene therapy. For successful delivery, the nonviral vector must be able to overcome many barriers to protect DNA and specifically deliver it for efficient gene expression in target cells. The use of peptides as gene delivery vectors is advantageous over other nonviral agents in that they are able to achieve all of these goals. This review will focus on the application of peptides to mediate nonviral gene delivery. By examining the literature over the past 20 years, it becomes clear that no other class of biomolecules are simultaneously capable of DNA condensation, blocking metabolism, endosomal escape, nuclear localization, and receptor targeting. Based on virtually limitless diversity of peptide sequence and function information from nature, it is increasingly clear that peptide-guided gene delivery is still in its infancy.

Synthetic PEGylated glycoproteins and their utility in gene delivery.

PEGylated glycoproteins (PGPs) were synthesized by copolymerizing a Cys-terminated PEG-peptide, glycopeptide, and melittin peptide. Compositionally unique PGPs were prepared by varying the ratio of PEG-peptide (20-90%) and melittin (0-70%) with a constant amount of glycopeptide (10%). The PGPs were purified by RP-HPLC, and characterized for molecular weight and polydispersity by GPC-HPLC and SDS-PAGE and for composition by RP-HPLC following reduction to form monomeric peptides. PGPs formed DNA condensates of 200-300 nm in diameter that were administered to mice via the tail vein. Biodistribution studies confirmed their primary targeting to liver hepatocytes with a DNA metabolic half-life of 1 h. Upon stimulation by hydrodynamic dosing with saline, PGP DNA (5 microg) mediated luciferase expression in the liver detected by bioluminescence imaging (BLI) after 24 h. The level of gene expression mediated by PGP DNA was 5000-fold less than direct hydrodynamic dosing of an equivalent amount of DNA and was independent of the mol percent of melittin incorporated into the polymer, but dependent on the presence of galactose on PGP. The results establish the ability to prepare three-component gene delivery polymers that function in vivo. Further design improvements in fusogenic peptides for gene delivery and for the simultaneous use of a nuclear targeting strategy will be necessary to approach levels of expression mediated by the direct hydrodynamic dosing of DNA.