Michael F. Tweedle

Background: Male dogs can develop spontaneous prostate cancer, which is similar physiologically to human disease. Recently, Tweedle and coworkers have developed an orthotopic canine prostate model allowing implanted tumors and therapeutic agents to be tested in a more translational large animal model. We used the canine model to evaluate prostate‐specific membrane antigen (PSMA)‐targeted gold nanoparticles as a theranostic approach for fluorescence (FL) imaging and photodynamic therapy (PDT) of early stage prostate cancer.Methods: Dogs (four in total) were immunosuppressed with a cyclosporine‐based immunosuppressant regimen and their prostate glands were injected with Ace‐1‐hPSMA cells using transabdominal ultrasound (US) guidance. Intraprostatic tumors grew in 4–5 weeks and were monitored by ultrasound (US). When tumors reached an appropriate size, dogs were injected intravenously (iv) with PSMA‐targeted nano agents (AuNPs‐Pc158) and underwent surgery 24 h later to expose the prostate tumors for FL imaging and PDT. Ex vivo FL imaging and histopathological studies were performed to confirm PDT efficacy.Results: All dogs had tumor growth in the prostate gland as revealed by US. Twenty‐four hours after injection of PSMA‐targeted nano agents (AuNPs‐Pc158), the tumors were imaged using a Curadel FL imaging device. While normal prostate tissue had minimal fluorescent signal, the prostate tumors had significantly increased FL. PDT was activated by irradiating specific fluorescent tumor areas with laser light (672 nm). PDT bleached the FL signal, while fluorescent signals from the other unexposed tumor tissues were unaffected. Histological analysis of tumors and adjacent prostate revealed that PDT damaged the irradiated areas to a depth of 1–2 mms with the presence of necrosis, hemorrhage, secondary inflammation, and occasional focal thrombosis. The nonirradiated areas showed no visible damages by PDT.Conclusion: We have successfully established a PSMA‐expressing canine orthotopic prostate tumor model and used the model to evaluate the PSMA‐targeted nano agents (AuNPs‐Pc158) in the application of FL imaging and PDT. It was demonstrated that the nano agents allowed visualization of the cancer cells and enabled their destruction when they were irradiated with a specific wavelength of light.

Objectives: Preclinical assessments were performed according to the US Food and Drug Administration guidelines to determine the physicochemical properties, pharmacokinetics, clearance, safety, and tumor-specific magnetic resonance (MR) imaging of MT218, a peptidic gadolinium-based MR imaging agent targeting to extradomain B fibronectin for MR molecular imaging of aggressive tumors. Materials and Methods: Relaxivity, chelation stability, binding affinity, safety-related target profiling, and effects on CYP450 enzymes and transporters were evaluated in vitro. Magnetic resonance imaging was performed with rats bearing prostate cancer xenografts, immunocompetent mice bearing murine pancreatic cancer allografts, and mice bearing lung cancer xenografts at different doses of MT218. Pharmacological effects on cardiovascular, respiratory, and central nervous systems were determined in rats and conscious beagle dogs. Pharmacokinetics were tested in rats and dogs. Biodistribution and excretion were studied in rats. Single and repeated dosing toxicity was evaluated in rats and dogs. In vitro and in vivo genotoxicity, in vitro hemolysis, and anaphylactic reactivity were also performed. Results: At 1.4 T, the r ₁ and r ₂ relaxivities of MT218 were 5.43 and 7.40 mM⁻¹ s⁻¹ in pure water, 6.58 and 8.87 mM⁻¹ s⁻¹ in phosphate-buffered saline, and 6.54 and 8.70 mM⁻¹ s⁻¹ in aqueous solution of human serum albumin, respectively. The binding affinity of MT218 to extradomain B fragment is 3.45 μM. MT218 exhibited no dissociation of the Gd(III) chelates under physiological conditions. The peptide degradation half-life (t _1/2) of MT218 was 1.63, 5.85, and 2.63 hours in rat, dog, and human plasma, respectively. It had little effect on CYP450 enzymes and transporters. MT218 produced up to 7-fold increase of contrast-to-noise ratios in the extradomain B fibronectin–rich tumors with a dose of 0.04 mmol/kg for at least 30 minutes. MT218 had little pharmacological effect on central nervous, cardiovascular, or respiratory systems. MT218 had a mean plasma elimination half-life (t _1/2) of 0.31 and 0.89 hours in rats and dogs at 0.1 mmol/kg, respectively. No detectable Gd deposition was observed in the brain at 6 hours postinjection of MT218 at 0.1 mmol/kg in rats. MT218 was not mutagenic and had no mortality or morbidity in the rats or dogs up to 1.39 and 0.70 mmol/kg/d, respectively. The no observed adverse effect level of MT218 in Sprague-Dawley rats was 1.39 mmol/kg for single dosing and 0.46 mmol/kg/d for repeated dosing. The no observed adverse effect level in dogs was 0.07 mmol/kg/d. MT218 exhibited no genotoxicity, hemolysis, and anaphylactic reactivity. Conclusion: The preclinical assessments showed that the targeted contrast agent MT218 has high r ₁ and r ₂ relaxivities, satisfactory physicochemical properties, pharmacokinetic, and safety profiles and produces effective tumor enhancement in multiple cancer types in rats and mice at reduced doses.

Gadolinium-based contrast agents have been used in hundreds of millions of patients in the past 30 years, with an exemplary safety record. However, assumptions made at their inception have been recently challenged, rekindling innovation efforts. This critical review outlines the motivations, technical obstacles, problems, and the most recent published progress toward the creation of alternatives to the existing gadolinium-based contrast agent.

Head and neck squamous cell carcinoma (HNSCC) survival rates have not improved in a decade, with a 63% 5-year recurrence rate after surgery, making HNSCC a compelling indication for optical surgical navigation (OSN). A promising peptide, HN1, targeted and internalized in human HNSCC cells in multiple laboratories, but was slow (24 h) to accumulate. We modified HN1 and explored structural variables to improve the uptake kinetics and create IRdye800 adducts useful for OSN. Eleven new molecules were synthesized and characterized chemically, in human HNSCC cells (Cal 27), and in HNSCC xenograft mice. Cal 27 flank xenografts in Balb/c nude mice were imaged for 3–48 h after 40 nmol intravenous doses of IR800-labeled molecules. Cell uptake kinetics in the 1–2 h window incubated at 1–10 μM were independent of the dye label (FITC, Cy5, or IR800), but increased markedly with additional N-terminal lipophilic substitution, and after resequencing the peptide to separate polar amino acids and move the lysine-dye more centrally. Microscopy confirmed the strong Cal 27 cell binding and demonstrated primarily cytosolic and membrane localization of the fastest peptide, 4Iphf-HN17. 4Iph-HN17-IR800 showed 26-fold greater rate of uptake in cells than HN1-IR800, and far stronger OSN imaging intensity and tumor to background contrast in mice, suggesting that the new peptide is a promising candidate for OSN of HNSCC.

The prostate-specific membrane antigen (PSMA) and gastrin-releasing peptide receptor (GRPR) are identified as important targets on prostate cancer. Receptor-targeting radiolabeled imaging pharmaceuticals with high affinity and specificity are useful in studying and monitoring biological processes and responses. Two potential imaging pharmaceuticals, AMBA agonist (where AMBA = DO3A-CH2CO-G-[4-aminobenzyl]- Gln-Trp-Ala-Val-Gly-His-Leu-Met-NH₂) and RM1 antagonist (where RM1 = DO3A-CH₂CO-G-[4-aminobenzyl]-D-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH₂), have demonstrated high binding affinity (IC₅₀) to GRP receptors and high tumor uptake. Antagonists, despite the poor tumor cell internalization properties, can show clearer images and pharmacokinetic profiles by virtue of their higher tumor uptake in animal models compared to agonists. For characterization, development, and translation of a potential imaging pharmaceutical into the clinic, it must be evaluated in a series of tests, including in vitro cell binding assays, in vitro buffer and serum stability studies, the biodistribution of the radiolabeled material, and finally imaging studies in preclinical animal models. Data related to acetate buffer, mouse, canine, and human sera stability of ¹⁷⁷Lu-labeled RM1 are presented here and compared with the acetate buffer and sera stability data of AMBA agonist. The samples of ¹⁷⁷Lu-labeled RM1 with a high radioconcentration degrade faster than low-radioconcentration samples upon storage at 2–8 °C. Addition of stabilizers, ascorbic acid and gentisic acid, improve the stability of ¹⁷⁷Lu-labeled RM1 significantly with gentisic acid being more efficient than ascorbic acid as a stabilizer. The degradation kinetics of ¹⁷⁷Lu-labeled AMBA and RM1 in sera follow the order (fastest to slowest): mouse > canine > human sera. Finally, ¹⁷⁷Lu-labeled RM1 antagonist is slower to degrade in mouse, canine, and human sera than ¹⁷⁷Lu-labeled AMBA agonist, further suggesting that an antagonist is a more promising candidate than agonist for the positron emission tomography (PET) imaging and therapy of prostate cancer patients.

Background: Ace‐1 canine prostate cancer cells grow orthotopically in cyclosporine immunosuppressed laboratory beagles. We previously transfected (human Gastrin‐Releasing Peptide Receptor, huGRPr) into Ace‐1 cells and demonstrated receptor‐targeted NIRF imaging with IR800‐G‐Abz4‐t‐BBN, an agonist to huGRPr. Herein, we used the new cell line to develop the first canine prostate cancer model expressing a human growth factor receptor.Methods: Dogs were immunosuppressed with cyclosporine, azathioprine, prednisolone, and methylprednisolone. Their prostate glands were implanted with Ace‐1^huGRPr cells. The implantation wounds were sealed with a cyanoacrylic adhesive to prevent extraprostatic tumor growth. Intraprostatic tumors grew in 4‐5 week. A lobar prostatic artery was then catheterized via the carotid artery and 25‐100 nmol IR800‐Abz4‐t‐BBN was infused in 2 mL followed by euthanasia in dogs 1‐2, and recovery for 24 h before euthanasia in dogs 3‐6. Excised tissues were imaged optically imaged, and histopathology performed.Results: Dog1 grew no tumors with cyclosporine alone. Using the four drug protocol, Dogs 2‐6 grew abundant 1‐2 mm intracapsular and 1‐2 cm intraglandular tumors. Tumors grew >5 cm when the prostate cancer cells became extracapsular. Dogs 4‐6 with sealed prostatic capsule implantation sites had growth of intracapsular and intraglandular tumors and LN metastases at 5 weeks. High tumor to background BPH signal in the NIRF images of sectioned prostate glands resulted from the 100 nmol dose (∼8 nmol/kg) in dogs 2‐4 and 50 nmol dose in dog 5, but not from the 25 nmol dose in Dog 6. Imaging of mouse Ace‐1^huGRPr tumors required an intravenous dose of 500 nmol/kg body wt. A lymph node that drained the prostate gland was detectable in Dog 4. Histologic findings confirmed the imaging data.Conclusion: Ace‐1^huGRPr cells created viable, huGRPr‐expressing tumors when implanted orthotopically into immune‐suppressed dogs. Local delivery of an imaging agent through the prostatic artery allowed a very low imaging dose, suggesting that therapeutic agents could be used safely for treatment of early localized intraglandular prostate cancer as adjuvant therapy for active surveillance or focal ablation therapies, or for treating multifocal intraglandular disease where focal ablation therapies are not indicated or ineffective.

Boehm-Sturm et al pose a possible paradigm shift in MR imaging: the substitution of iron chelates for gadolinium chelates as paramagnetic contrast agents. SummaryBoehm-Sturm et al (1) pose a possible paradigm shift in magnetic resonance (MR) imaging: the substitution of iron chelates for gadolinium chelates as paramagnetic contrast agents. The advent...