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How to generate in-human data on antibodies with microdosing

Pieter Steinkamp, MD, PhD cand.

The labelling of (novel drugs) with fluorophores and radioisotopes is an attractive approach to investigate drug distribution on a macro- and microscopic level. For example, with labelled therapeutic antibodies in medical oncology. Mainly due to relatively easy labeling strategies, that do not have a direct impact on the drug mechanism of action itself, the accumulation of an antibody at the cellular level can already be tested in early clinical trials, even before Phase I. Additionally, with a low, non-pharmacological, drug dose the potential efficacy of the drug can already be predicted. This so-called microdosing strategy is used in innovative, Proof of Concept (PoC) studies to assess biodistribution and plasma pharmacokinetics.

The advantages of microdosing.

Microdosing is associated with multiple advantages over time. Firstly, a microdosing strategy is associated with a minimized risk of (serious) adverse events for participants in a medical trial due to very low dosing of the antibody. Secondly, efficient macro- and microscopic biodistribution data can be obtained. This data provides efficient information about drug concentration in a small number of patients. Thirdly, by conducting a PoC study with microdosing in a minimal number of participants the overall clinical trial costs decrease. Namely, often large animal models can be skipped, and you already possess in-human data.

Moreover, microdosing is encouraged and approved by the EMA and FDA. They recognize it as a step-up for the next clinical phases using a standardized regulatory framework. In the end, microdosing prevents unnecessary exposure of novel drugs to a large number of participants. Furthermore, it helps drug developers avoid from investing in non-effective clinical trials.

Microdosing with antibodies – example studies.

Worldwide, multiple optical imaging trials have been performed using the microdosing strategy in labelling antibodies for imaging purposes. A commonly used antibody is bevacizumab conjugated to a fluorescent near-infrared dye (IRDye-800CW). Bevacizumab  targets vascular endothelial growth factor A (VEGF-A). VEGF-A induces angiogenesis and is overexpressed in a variety of solid tumors. For example, a study by Lamberts investigated its feasibility in breast cancer.1 A non-therapeutic amount of 4.5 mg bevacizumab-800CW was considered safe in all tumor types. Furthermore, sarcomas, colorectal cancer and esophageal cancer were identified as safe and feasible target tissue to improve decision making.2

Endothelial growth factor (EGFR) targeting in head-and neck cancer patients is used in a variety of large Phase II studies. Cetuximab labeled with IRDye-800CW and panatunimab-800CW are used in an intra-operative setting to identify tumor tissue and residual disease. All these studies, with the intention to decrease the number of tumor-positive margins, could be easily translated towards a study workup with novel antibodies conjugated to IRDye-800CW or ICG.

Data analysis of antibody distribution.

When following a standard imaging workflow, combining intra-operative imaging with ex vivo tissue imaging, as described by Koller et al., standardized data of antibody distribution can be obtained.3 There are a few pillars which are investigated in this workflow. In the first pillar, ex vivo imaging data is correlated to standard histopathology to investigate whether the drug reaches the proposed target. Moreover, we are able to characterize tracer distribution and differentiation between malignant and benign tissue expression both in- and ex vivo.

For drug development purposes, this pillar is extremely important to determine whether a degree of target engagement is delivered. Namely, this could eventually lead to therapeutic effect of antibodies. Most important, this molecular imaging strategy provides quantitative drug behavior data in a very early stage of drug development. This quantitative data consists of fluorescence intensities and hence tumor-to-background ratios both in- and ex vivo (second pillar) and pharmacokinetics (third pillar) using blood samples obtained before and after tracer administration.

When using this strategy in novel drug development, for example for novel immune modulatory antibodies, the translation towards therapeutic concentrations and therapeutic effects can be managed with the backbone of a cost-effective microdosing pathway. In order to do so, a standardized imaging and processing protocol is key towards effective implementation in medical research.

Microdosing with antibodies – a standard in future clinical trials.

A microdosing strategy can eventually replace extensive animal and healthy volunteer testing during drug development. Namely, by immediately evaluating the antibody of interest in the intended target population. Labeling antibodies following Good Manufacturing Practice (GMP) allows for early clinical studies to assess the pharmacological targeting and biodistribution aspect of the novel drugs. When results show a non-sufficient targeting, a fast decision can be made about proceeding with the drug into the next clinical phases.

At TRACER we provide a standardized working method with experienced researchers, which allows pharmaceuticals to be evaluated for fast inclusion of patients in a variety of (pre-) malignant diseases. Contact us for more information.


  1. Lamberts, L. E. et al. Tumor-specific uptake of fluorescent bevacizumab-IRDye800CW microdosing in patients with primary breast cancer: A phase I feasibility study. Clinical Cancer Research 23, (2017).
  2. Steinkamp, P. J. et al. Fluorescence-guided visualization of soft tissue sarcomas by targeting vascular endothelial growth factor-A: a phase 1 single-center clinical trial. Journal of Nuclear Medicine (2020) doi:10.2967/jnumed.120.245696.
  3. Koller, M. et al. Implementation and benchmarking of a novel analytical framework to clinically evaluate tumor-specific fluorescent tracers. Nature Communications 9, (2018).

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