Assessment of the Pharmacokinetics, Disposition, and Duration of Action of the Tumour-Targeting Peptide CEND-1.

CEND-1 (iRGD) is a bifunctional cyclic peptide that can modulate the solid tumour microenvironment, enhancing the delivery and therapeutic index of co-administered anti-cancer agents. This study explored CEND-1's pharmacokinetic (PK) properties pre-clinically and clinically, and assessed CEND-1 distribution, tumour selectivity and duration of action in pre-clinical tumour models. Its PK properties were assessed after intravenous infusion of CEND-1 at various doses in animals (mice, rats, dogs and monkeys) and patients with metastatic pancreatic cancer. To assess tissue disposition, [3H]-CEND-1 radioligand was administered intravenously to mice bearing orthotopic 4T1 mammary carcinoma, followed by tissue measurement using quantitative whole-body autoradiography or quantitative radioactivity analysis. The duration of the tumour-penetrating effect of CEND-1 was evaluated by assessing tumour accumulation of Evans blue and gadolinium-based contrast agents in hepatocellular carcinoma (HCC) mouse models. The plasma half-life was approximately 25 min in mice and 2 h in patients following intravenous administration of CEND-1. [3H]-CEND-1 localised to the tumour and several healthy tissues shortly after administration but was cleared from most healthy tissues by 3 h. Despite the rapid systemic clearance, tumours retained significant [3H]-CEND-1 several hours post-administration. In mice with HCC, the tumour penetration activity remained elevated for at least 24 h after the injection of a single dose of CEND-1. These results indicate a favourable in vivo PK profile of CEND-1 and a specific and sustained tumour homing and tumour penetrability. Taken together, these data suggest that even single injections of CEND-1 may elicit long-lasting tumour PK improvements for co-administered anti-cancer agents.

Tissue Distribution of [14C]-Lefamulin into the Urogenital Tract in Rats.

Lefamulin, a semisynthetic pleuromutilin antibiotic approved in the United States, Canada, and Europe for intravenous and oral treatment of community-acquired bacterial pneumonia, is highly active in vitro against bacterial pathogens that cause sexually transmitted infections (STIs), including multidrug-resistant strains of Neisseria gonorrhoeae, Chlamydia trachomatis, and Mycoplasma genitalium. This nonclinical study used quantitative whole-body autoradiography (QWBA) and qualitative tape-transfer microautoradiography (MARG) to investigate lefamulin distribution into urogenital tract tissues down to a cellular level in male and female rats. A single intravenous dose (30 mg/kg) of [14C]-lefamulin was administered to 3 male and 3 female Sprague-Dawley rats. At 0.5, 6, and 24 h post dose, rats were euthanized and [14C]-lefamulin distribution was investigated using QWBA and MARG of sagittal planes. [14C]-lefamulin was well distributed throughout the carcasses of male and female rats, with the highest concentrations observed in male bulbourethral gland, urethra, prostate in female clitoral gland, uterus (particularly endometrium), and ovary. In these areas, concentrations were similar to or higher than those observed in the lungs. Concentrations peaked at 0.5 h post dose, remaining detectable in the urogenital tract up to 24 h post dose. [14C]-lefamulin in rats showed rapid, homogeneous distribution into urogenital tissues down to a cellular level, with high tissue:blood ratios in tissues relevant to STI treatment. These results, and the potent in vitro activity of lefamulin against multidrug-resistant bacteria known to cause STIs, will help inform further assessment of lefamulin, including potential clinical evaluation for treatment of STIs.

Matrix-assisted laser desorption/ionization-ion mobility separation-mass spectrometry imaging of vinblastine in whole body tissue sections.

During early-stage drug development, drug and metabolite distribution studies are carried out in animal tissues using a range of techniques, particularly whole body autoradiography (WBA). While widely employed, WBA has a number of limitations, including the following: expensive synthesis of radiolabeled drugs and analyte specificity and identification. WBA only images the radiolabel. MALDI MSI has been shown previously to be advantageous for imaging the distribution of a range of drugs and metabolites in whole body sections. Ion mobility separation (IMS) adds a further separation step to imaging experiments; demonstrated here is MALDI-IMS-MS whole body imaging of rats dosed at 6 mg/kg i.v. with an anticancer drug, vinblastine and shown is the distribution of the precursor ion m/z 811.4 and several product ions including m/z 793, 751, 733, 719, 691, 649, 524, and 355. The distribution of vinblastine within the ventricles of the brain is also depicted. Clearly demonstrated in these data are the removal of interfering isobaric ions within the images of m/z 811.4 and also of the transition m/z 811-751, resulting in a higher confidence in the imaging data. Within this work, IMS has shown to be advantageous in both MS and MS/MS imaging experiments by separating vinblastine from an endogenous isobaric lipid.

Quantitative biodistribution of biotherapeutics at whole body, organ and cellular levels by autoradiography.

AIM: Tools for mapping and quantifying monoclonal antibody (mAb) and peptide biotherapeutics distribumtion were evaluated by comparing data from three independent methods conducted at the whole body, organ or tissue, and cellular levels. MATERIALS & METHODS: [3H]-mAb1 and [3H]-peptide A were administered intravenously to rats followed by quantitative whole-body autoradiography, kidney macro-autoradiography and micro-autoradiography. RESULTS: [3H]-mAb1 and [3H]-peptide A concentrations were measured in anatomical regions ranging from whole body to whole organ to sub-organ level, such as the kidney glomerulus, with increasing resolution. The tissue/blood [3H]-mAb1 concentrations in selected kidney microenvironments were comparable among the three quantitative methods. CONCLUSION: Quantitative whole-body autoradiography, tissue macro-autoradiography and micro-autoradiography all provide useful tools for quantifying the concentrations of biotherapeutics at different anatomical levels in tissues, facilitating better predictions of efficacy and toxicity.

Quantitative whole-body autoradiography: past, present and future.

Traditional bioanalytical measurements determine concentrations of drug and metabolites in plasma; however, most drugs exert their effects in defined target tissues. As there is no clear relation between concentrations in plasma and those in tissue, alternative methods must be employed to study the absorption, distribution, metabolism and excretion properties of new therapeutic agents. Quantitative whole-body autoradiography is used in the drug development process to determine the distribution and concentrations of radiolabeled test compounds in laboratory animals. Quantitative whole-body autoradiography can provide information on tissue PKs, penetration, accumulation and retention. Although the technique is considered the industry standard for performing preclinical tissue distribution studies, it is perhaps timely, 60 years after the first reported use of the method, to re-assess the technique against modern alternatives.

Quantitative whole-body autoradiography, LC-MS/MS and MALDI for drug-distribution studies in biological samples: the ultimate matrix trilogy.

The drug-development process requires an understanding of the ADME properties of the novel therapeutic agent. Determination of drug concentrations and identity in excreta (urine and feces) examines the products of these processes. Similar measurements made on plasma, while accurately determining exposure, show only what is being transported around the body. Both activities fail to confirm the nature of components at the pharmacologically relevant matrix - the tissue. Attention is therefore being directed towards methods that can be employed to address this lack in our current methodologies, to provide better quality data on which risk assessments can be made, so that pharmacological models can be refined, and drug safety improved. In this article, we will look at the current methods used to obtain tissue drug and drug metabolite concentrations, and their potential use in drug discovery.

Recombinant fusion protein linking factor VIIa with albumin (rVIIa-FP): Tissue distribution in rats.

INTRODUCTION: A novel fusion protein linking coagulation factor VIIa with albumin (rVIIa-FP) is currently undergoing clinical investigations. OBJECTIVE: This study was conducted to examine the biodistribution of rVIIa-FP in comparison to recombinant factor VIIa (rFVIIa). MATERIALS AND METHODS: [(3)H]-rVIIa-FP (10mgkg(-1)) or [(3)H]-rFVIIa (1.6mgkg(-1)) were administered intravenously to rats, followed by quantitative whole-body and knee joint autoradiography for 24 ([(3)H]-rFVIIa) or 240 ([(3)H]-rVIIa-FP) hours post-dose. Pharmacokinetic and excretion balance analyses were performed. RESULTS: In contrast to [(3)H]-albumin, the tissue distributions of [(3)H]-rVIIa-FP and [(3)H]-rFVIIa were similar. Within the knee, both were rapidly present within synovial and mineralized regions. Importantly, rVIIa-FP- and albumin-derived radioactivity were detectable up to 72-120hours, whereas [(3)H]-rFVIIa signals were already close to detection limits at 24hours. The longest rVIIa-FP retention times were observed in bone marrow and endosteum, in which the retention times were up to 5 times longer for rVIIa-FP compared with rFVIIa. Up to 8hours post-dose, 100% of radioactivity was assigned to unchanged [(3)H]-rVIIa-FP. Elimination of both proteins occurred primarily via the urine. CONCLUSIONS: The data suggest that the FVIIa moiety is directing rVIIa-FP's tissue distribution while the albumin moiety is responsible for the prolonged tissue retention. Importantly, rVIIa-FP is highly concentrated and retained over a long period in the growth plate of the knee joint-a vulnerable site in haemophilia patients. Overall, these improved tissue distribution characteristics of rVIIa-FP may enhance compliance and allow a more convenient dosing frequency.

Biodistribution of the recombinant fusion protein linking coagulation factor IX with albumin (rIX-FP) in rats.

INTRODUCTION: The recombinant fusion protein linking coagulation factor IX with albumin (rIX-FP) is undergoing clinical trials for prophylaxis and on-demand treatment of haemophilia B patients. The aim of this study was to investigate the pharmacokinetics, whole-body and knee joint distribution of rIX-FP following intravenous administration to rats, compared with a marketed, non-fused rFIX and recombinant human albumin. MATERIAL AND METHODS: [(3)H]-rIX-FP, [(3)H]-rFIX or [(3)H]-albumin were administered to rats followed by quantitative whole-body autoradiography over 24 or 240 hours, and the tissue distribution as well as elimination of radioactivity were measured. RESULTS: Elimination of all radioactivity derived from the three proteins was shown to occur primarily via the urine. The tissue distribution of [(3)H]-rIX-FP and [(3)H]-rFIX (but not of [(3)H]-albumin) was comparable, both penetrating predominantly into bone, and well-perfused tissues, suggesting that the rIX moiety determines the distribution pattern of rIX-FP, while the albumin moity is responsible for the prolonged plasma and tissue retention. Detailed knee-joint analysis indicated rapid presence of [(3)H]-rIX-FP and [(3)H]-rFIX in synovial and mineralised bone tissue, mostly localised to the zone of calcified cartilage. Longest retention times were observed in the bone marrow and the endosteum of long bones. Intriguingly, [(3)H]-rIX-FP- and [(3)H]-albumin-derived radioactive signals were detectable up to 240 hours, while [(3)H]-rFIX-derived radioactivity rapidly declined after 1hour post-dosing correlating to the extended plasma half-life of [(3)H]-rIX-FP. CONCLUSION: The prolonged plasma and tissue retention of rIX-FP achieved by albumin fusion may allow a reduction in dosing frequency leading to increased therapeutic compliance and convenience.