Elastin-like polypeptides are not synthetic polymers in the conventional sense. They are proteins built by recombinant DNA technology, typically consisting of many repeats of the pentapeptide sequence VPGXG, where the guest residue X can be swapped for almost any amino acid. This simple design rule gives researchers extraordinary control. By choosing the guest residue and the number of repeats, they can tune the polymer’s molecular weight, its hydrophobicity, and, most importantly, its phase-transition temperature. Below this temperature the chains stay dissolved in water; above it they collapse into coacervates, micelles, or gel-like depots. In effect, researchers can encode a material that behaves like a soluble drug in the bloodstream but turns into an insoluble depot the moment it encounters body temperature or a mild heat source applied to a tumor.

This thermoresponsive behavior is what makes ELPs interesting for radiopharmaceutical delivery. Radiopharmaceuticals work by attaching radionuclides, which emit alpha particles, beta particles, or Auger electrons, to carrier molecules that guide them to diseased tissue. The carrier determines biodistribution, target accumulation, local retention, and clearance, while the radionuclide determines half-life, emission energy, and tissue range. Effective treatment requires aligning the physical half-life of the radionuclide with the biological kinetics of the carrier. Small molecules and short peptides penetrate tissues rapidly and provide favorable imaging kinetics, but they clear so quickly that lesion retention and sustained dose deposition suffer. Antibodies offer high affinity and specificity but circulate slowly, potentially increasing radiation exposure to bone marrow, liver, and spleen. Nanoparticles carry high payloads but suffer from reticuloendothelial uptake, persistent tissue retention, manufacturing complexity, and batch variability. ELPs, the review argues, sit at a different point on this spectrum: they are biodegradable, genetically programmable, and capable of self-assembly, which may support more defined production than conventional nanocarriers.

The review identifies three main labelling strategies. Direct radioiodination exploits tyrosine residues in the ELP sequence, allowing isotopes such as iodine-123, iodine-124, iodine-125, and iodine-131 to be introduced by electrophilic substitution on the aromatic ring. This approach avoids an additional chelator and may preserve the ELP architecture, phase transition, and self-assembly, which is why it has been used for local injectable depots and short-term in vivo tracking. Chelator-mediated labelling, by contrast, attaches bifunctional chelators such as DOTA, NOTA, or DTPA to coordinate radiometals including gallium-68, copper-64, indium-111, lutetium-177, yttrium-90, and actinium-225. Site-specific labelling, using cysteine, lysine, tyrosine, or noncanonical amino acids positioned genetically outside the repetitive domain, offers better control over conjugation site and stoichiometry and may preserve ELP structure and function. Beyond covalent conjugation, amphiphilic ELP block copolymers can self-assemble into micelles, vesicles, and nanoparticles whose hydrophobic domains can accommodate radiolabelled compounds or co-therapeutic agents, although direct radiopharmaceutical evidence for physically loaded ELP assemblies remains limited.

The review then examines three delivery modes. Systemic delivery uses soluble ELP chains, macromolecular conjugates, or defined nanostructures, with molecular weight, conformation, surface charge, and hydrophilic-hydrophobic balance influencing renal clearance, circulation, and organ uptake. Fluorescence studies of ELPs ranging from 25 to 110 kilodaltons reported longer terminal plasma half-lives with increasing size and substantial size-dependent renal deposition, but the review cautions that these non-radioactive findings are indirect and cannot define an optimal radiopharmaceutical size, because longer residence also changes integrated exposure in blood, kidney, liver, and marrow. Thermally triggered delivery exploits the lower critical solution temperature transition: suitable constructs remain soluble in circulation and undergo local phase transition when a lesion is exposed to clinically achievable mild hyperthermia. In an early direct study, radioiodinated ELPs were used to test whether mild local hyperthermia increased tumour accumulation after intravenous administration, and one hour of heating produced approximately twofold greater accumulation in SKOV-3 and D-54MG xenografts, with higher uptake than a soluble control. Local in situ depot delivery, the third mode, aims to retain radionuclides within accessible lesions and currently provides the strongest direct therapeutic evidence. After intratumoural or image-guided administration, selected ELPs can undergo phase separation, self-assembly, or gelation to form a radioactive depot, and preclinical radiolabelled depots have delayed tumour growth, although efficacy varied between models and complete spatial dose information was generally unavailable.

The review is refreshingly honest about the limits of current evidence. Direct evidence for ELP radiopharmaceuticals remains limited and comes mainly from local or intratumoural depot studies. Systemic therapy and thermally triggered targeting rely mostly on radiotracer studies or non-radioactive ELP research. Available studies differ in ELP design, radionuclide, administration route, tumour model, and evaluation methods. Complete spatial dose information is uncommon, and no clinical comparison has established superiority over existing carriers or local radiation methods. Claims of improved tumour uptake, lower normal-organ exposure, multimodal synergy, or personalised theranostics should therefore remain cautious, the authors write. They also note that the heterogeneous enhanced permeability and retention effect, often invoked to justify passive tumour targeting of macromolecules, should be treated as a patient- and lesion-specific factor rather than a general claim, since human lesions are generally more heterogeneous than rapidly growing xenografts and average nanoparticle delivery remains low.

Several technical challenges stand between the laboratory and the clinic. The transition temperature of an ELP is not a fixed material constant; it depends on guest-residue composition, molecular mass, polymer concentration, ionic strength, co-solutes, attached cargo, and the heating or cooling protocol. Measurements of an unlabelled precursor do not show whether the radioconjugate will remain injectable or form a depot at 37 degrees Celsius, so the transition profile should be measured for the final labelled product at the administered concentration and in the intended vehicle. Radiolysis may cause oxidation, fragmentation, or crosslinking that is absent from the non-radioactive precursor, and these changes can alter phase behaviour and biological clearance. Radioiodinated ELPs may undergo in vivo deiodination, leading to activity in iodine-avid tissues such as the thyroid and stomach, while high labelling density may alter hydrophobicity, conformation, transition temperature, and biodistribution. For radiometals, stable chelation can reduce radionuclide dissociation, but performance depends on the metal-chelator pair, conjugation chemistry, labelling conditions, and in vivo stability, all of which must be optimised without disrupting ELP phase behaviour or pharmacokinetics.

Biocompatibility offers some reassurance. ELPs are derived from conserved motifs in natural elastin and have shown favourable biocompatibility and potential biodegradability. Unlike nondegradable inorganic materials and some synthetic polymers, ELPs are protein-based and can enter normal metabolic pathways, with endogenous proteases gradually degrading ELP chains into smaller peptides and amino acids that are subsequently eliminated. This behaviour may reduce risks associated with persistent tissue accumulation, chronic inflammation, and delayed toxicity, although safety must be established for each final formulation. Genetic engineering also enables production of ELPs with predetermined sequences and relatively uniform molecular structures, which may reduce broad molecular-weight distributions, batch variability, and residual synthetic impurities associated with some conventional polymers, supporting reproducibility, quality control, safety assessment, and clinical translation.

The review outlines a practical workflow for future development. It begins with a target product profile linking the administration route, lesion geometry, radionuclide properties, ELP kinetics, and required absorbed dose. Candidate products should then undergo staged evaluation of sequence, transition temperature, radiochemical purity, stability, assembly state, biodistribution, imaging, dosimetry, and safety, with predefined advancement criteria including batch reproducibility, manufacturability, sterility, stability, and evidence that radiolabelling preserves ELP function. For systemic formulations, measurements of blood clearance, lesion penetration and retention, and normal-organ exposure are essential, since longer circulation does not necessarily improve tumour delivery. Thermally triggered systems require reliable temperature monitoring and controls for heating-induced changes in blood flow. Local depots require three-dimensional activity maps, assessment of needle-track leakage, and adequate coverage of irregular lesions. Clinical translation will also require scalable production, purification, endotoxin control, identity and potency specifications, rapid release testing, and stability during use, alongside long-term degradation, clearance, immunogenicity, repeat-dose safety, regulatory classification, and comparison with established treatments.

At present, the authors conclude, ELPs are best regarded as a promising early-stage engineering platform rather than an established clinical delivery system. Their flexibility does not establish more effective or safer radionuclide delivery, because changing one feature may also alter solubility, assembly, clearance, or label stability. The final radiolabelled product must therefore be evaluated directly, under relevant biological conditions, with quantitative imaging and dosimetric modelling to estimate lesion and organ absorbed doses. Continued optimisation may support the development of ELP-based systems for controlled radionuclide delivery and cancer theranostics, but the burden of proof now rests on demonstrating, for each radiolabelled construct, that programmability translates into measurable benefit for patients. For a field where precision is everything, that is a reasonable place to start.

Subject of Research: Biomedical Engineering

Article Title: Elastin-like polypeptides in radiopharmaceutical delivery: current evidence, design principles, and translational challenges

Article References: Tian, H., Yang, K., Li, H., & Xiao, J. (2026). Elastin-like polypeptides in radiopharmaceutical delivery: current evidence, design principles, and translational challenges. Frontiers in Biomaterials Science, 5, Article 1911811. https://doi.org/10.3389/fbiom.2026.1911811

Image Credits: AI Generated

DOI: 10.3389/fbiom.2026.1911811

Keywords: elastin-like polypeptides, radiopharmaceuticals, radionuclide delivery, cancer theranostics, thermoresponsive phase transition, nuclear medicine, drug delivery, Elastin-like, polypeptides, radiopharmaceutical, delivery, current

The rationale for targeting macrophages directly is rooted in their complicated role in cancer biology. Macrophages are versatile immune cells that maintain tissue homeostasis and defend against pathogens, but within the tumor microenvironment they are notoriously plastic and heterogeneous, adopting functions that range from pro-inflammatory to strongly immunosuppressive. Dense infiltration of tumors by CD68-positive macrophages has been linked in breast cancer to larger tumor size, higher histologic grade, hormone-receptor negativity, lymphovascular invasion, and reduced overall and progression-free survival. Existing imaging probes have struggled to capture this picture accurately. Some tracers, such as those targeting CD163 or CD206, preferentially label the immunosuppressive M2-like subset, while probes aimed at the translocator protein TSPO are not exclusive to macrophages and are affected by genetic polymorphisms that vary between individuals. A copper-64-labeled CD68-Fc fusion protein has been explored, but it targets scavenger receptors indirectly rather than macrophages themselves.

To fill this gap, the researchers turned to phage display, a technique that screens enormous libraries of antibody fragments for those that bind a chosen target. They used a synthetic Fab library, designated Library F, that was deliberately designed without lysine residues in its complementarity-determining regions, the loops that make contact with the antigen. Lysines were replaced with arginines, which preserve the basic character of the loops but lack the reactive nucleophilic amine that would otherwise be chemically modified during radiolabeling. Framework lysines elsewhere in the molecule were retained to allow controlled conjugation to a chelator without disturbing the antigen-binding surface. Screening this library against immobilized recombinant human CD68 yielded 14 unique sequences, of which five, named Fab3, Fab4, Fab7, Fab8, and Fab14, could be expressed successfully in bacterial cultures and purified on Protein A columns.

Characterization of the five candidates revealed important differences. Enzyme-linked immunosorbent assays showed that all five bound human CD68 with high significance compared with a bovine serum albumin control, but Fab3 also bound strongly to BSA itself and cross-reacted with mouse CD68, so it was eliminated. The remaining four Fabs bound human CD68 in a concentration-dependent manner with half-maximal effective concentrations ranging from 1.7 to 15 nanomolar, an affinity range well suited to immunoPET probes, and showed no detectable binding to BSA or mouse CD68. Production metrics then became the deciding factor. Fab4 and Fab7, despite their favorable affinities, expressed inconsistently and lost purity during storage, retaining only 71 and 75 percent monomer after 60 days at 4 degrees Celsius. Fab14 expressed at high yields but with purities of only 60 to 70 percent. Fab8, by contrast, delivered consistent yields of about 1.7 milligrams per liter, the highest purity of 80 to 90 percent straight from Protein A purification, and 96 percent monomer after two months of storage, making it the clear lead candidate.

Converting Fab8 into a PET tracer required attaching a chelator that could hold a positron-emitting radionuclide. The team selected p-SCN-Bn-deferoxamine, or DFO, and zirconium-89, a pairing with a well-established chemistry and a track record of clinical translation. After optimizing the conjugation stoichiometry to a 4:1 ratio of DFO to Fab, which minimized free chelator without inducing protein aggregation, the researchers radiolabeled the conjugate at a specific activity of 7.4 kilobecquerels per nanomole, achieving a radiochemical yield of 99 percent. Stability testing showed that the resulting tracer, [89Zr]Zr-DFO-Fab8, remained more than 95 percent intact monomer in phosphate-buffered saline over 48 hours, and about 80 percent intact in human serum at 37 degrees Celsius over the same period, with the remainder of the radioactivity associated with serum proteins.

In vitro experiments confirmed that the tracer behaved as intended on real cells. The human monocytic leukemia line THP-1 was differentiated into macrophages, which expressed human CD68 on roughly 25 percent of cells compared with about 4 percent of undifferentiated THP-1 cells, as measured by flow cytometry. When fixed macrophages were incubated with the tracer, binding reached about 10 percent of the added activity, five times higher than an irrelevant radiolabeled Fab control and roughly three times higher than in undifferentiated THP-1 cells. Importantly, neither the unlabeled conjugate nor the radiolabeled tracer compromised macrophage viability at the concentrations tested, with only a modest 19 percent decrease at the highest radiotracer concentration of 125 nanomolar, far above the estimated nanomolar circulating levels in the animal studies.

The first in vivo test used a cleverly simple model. Because Fab8 does not cross-react with mouse CD68, the team implanted subcutaneous plugs of Matrigel mixed with either recombinant human CD68 protein or BSA into opposite shoulders of immunocompetent BALB/c mice. PET/CT imaging at 1 and 4 hours after tracer injection showed significantly higher uptake in the human CD68 plugs than in the BSA control plugs, with plug-to-muscle ratios of 11 and 13 versus roughly 5 and 6, respectively. Voxel-level histogram analysis confirmed that the spatial distribution of tracer within the antigen plugs differed significantly from the controls over time. Biodistribution measurements correlated strongly with the imaging data, with a Pearson correlation coefficient of 0.97, and showed the expected renal dominance typical of antibody fragments of this size.

The more biologically demanding test came next. The researchers grew bilateral MDA-MB-231 triple-negative breast cancer xenografts, a tumor line that forms highly vascularized tumors and does not express CD68, in immunodeficient NSG mice. Once the tumors had developed vasculature, they injected differentiated THP-1 macrophages directly into one tumor and undifferentiated THP-1 cells into the other, then administered the tracer systemically. PET imaging at 1 and 6 hours revealed significantly greater uptake in the macrophage-containing tumors, with tumor-to-muscle ratios of about 5.2 and 5.9 compared with 2.7 and 3.6 in the control tumors. Immunofluorescence staining of the excised tumors provided the decisive validation: macrophage-injected tumors contained roughly 660 CD68-positive cells per area versus only 16 in the controls, and the CD68 density correlated with the PET standardized uptake values with a Pearson coefficient of 0.9663. This is the study’s central achievement, a direct, quantitative bridge between a noninvasive PET signal and the histopathological gold standard for pan-macrophage burden.

The authors are candid about the limitations that must be addressed before clinical translation. Kidney uptake was high, as expected for a 50-kilodalton Fab, and about 10 percent aggregation in serum may have contributed to elevated liver and spleen signal through sequestration by the reticuloendothelial system. Because the target uptake in the xenograft model was inherently low, background signal was comparatively prominent, and tumor-to-heart ratios below one suggested incomplete blood clearance at the early imaging timepoints used. Future work will explore polyethylene glycol modification to reduce renal retention, an approach already validated clinically by the approved drug certolizumab pegol, along with better animal models of human macrophage infiltration. Even so, the study establishes a complete pipeline, from phage display discovery through radiopharmaceutical engineering to correlated PET and pathology, for a first-generation human CD68 immunoPET tracer. Such a probe could eventually help select patients for macrophage-targeting therapies, monitor treatment response, and predict resistance to immunotherapy, all from a single noninvasive scan.

Subject of Research: Development of a zirconium-89-labeled anti-CD68 Fab immunoPET tracer for imaging human macrophages

Article Title: A FABulous approach to image human macrophages

Article References: Roohani, B., Vaughn Embs, A., Kumar, R., Lee, S., Katz, S. R., La Prairie, C., Nash, D., Dangarwala, M., Kluger, H. M., Klein, D. E., Zhang, P., Larimer, B. M., Nelson, B., & Marquez-Nostra, B. (2026). A FABulous approach to image human macrophages. European Journal of Nuclear Medicine and Molecular Imaging. https://doi.org/10.1007/s00259-026-08180-y

Image Credits: AI Generated

DOI: 10.1007/s00259-026-08180-y

Keywords: CD68, macrophages, immunoPET, phage display, Fab, zirconium-89, tumor-associated macrophages, PET imaging, triple-negative breast cancer, molecular imaging, radiopharmaceutical, tumor microenvironment