Melanoma is a cancer with increasing incidence and there is a need for alternatives to immunotherapy within effective approaches to treatment of metastatic melanoma. We performed comparative radioimmunotherapy (RIT) of experimental B16-F10 melanoma with novel humanized IgG to melanin h8C3 labeled with a beta emitter, ¹⁷⁷Lu, and an alpha-emitter, ²¹³Bi, as well as biodistribution, microSPECT/CT imaging, and mouse and human dosimetry calculations. microSPECT/CT imaging showed that a humanized antibody that targets “free” melanin in the tumor microenvironment had high tumor uptake in B16F10 murine melanoma in C57Bl/6 mice, with little to no uptake in naturally melanized tissues. Extrapolation of the mouse dosimetry data to an adult human demonstrated that doses delivered to major organs and the whole body by ¹⁷⁷Lu-h8C3 would be approximately two times higher than those delivered by ²¹³Bi-h8C3, while the doses to the tumor would be almost similar. RIT results indicated that ²¹³Bi-h8C3 was more effective in slowing down the tumor growth than ¹⁷⁷Lu-h8C3, while both radiolabeled antibodies did not produce significant hematologic or systemic side effects. We concluded that h8C3 antibody labeled with ²¹³Bi is a promising reagent for translation into a clinical trial in patients with metastatic melanoma.

Melanoma incidence continues to rise, and while therapeutic approaches for early stage cases are effective, metastatic melanoma continues to be associated with high mortality. Immune checkpoint blockade (ICB) has demonstrated clinical success with approved drugs in cohorts of patients with metastatic melanoma and targeted radionuclide therapy strategies showed promise in several clinical trials against various cancers including metastatic melanoma. This led our group to investigate the combination of these two treatments which could be potentially offered to patients with metastatic melanoma not responsive to ICB alone. Previously, we have demonstrated that a combination of humanized anti-melanin antibody conjugated to 213Bismuth and anti-PD-1 ICB reduced tumor growth and increased survival in the Cloudman S91 murine melanoma DBA/2 mouse model. In the current study, we sought to improve the tumoricidal effect by using the long-lived radionuclides 177Lutetium and 225Actinium. Male Cloudman S91-bearing DBA/2 mice were treated intraperitoneally with PBS (Sham), unlabeled antibody to melanin, anti-PD-1 ICB, 177Lutetium or 225Actinium RIT, or a combination of ICB and RIT. Treatment with anti-PD-1 alone or low-dose 177Lutetium RIT alone resulted in modest tumor reduction, while their combination significantly reduced tumor growth and increased survival, suggesting synergy. 225Actinium RIT, alone or in combination with ICB, showed no therapeutic benefit, suggesting that the two radionuclides with different energetic properties work in distinct ways. We did not detect an increase in tumor-infiltrating T cells in the tumor microenvironment, which suggests the involvement of alternative mechanisms that improve the effect of combination therapy beyond that observed in the single therapies.

(1) Background: Monoclonal antibodies are used in the treatment of multiple conditions including cancer, autoimmune disorders, and infectious diseases. One of the initial steps in the selection of an antibody candidate for further pre-clinical development is determining its pharmacokinetics in small animal models. The use of mass spectrometry and other techniques to determine the fate of these antibodies is laborious and expensive. Here we describe a straightforward and highly reproducible methodology for utilizing radiolabeled antibodies for pharmacokinetics studies. (2) Methods: Commercially available bifunctional linker CHXA” and ¹¹¹Indium radionuclide were used. A melanin-specific chimeric antibody A1 and an isotype matching irrelevant control A2 were conjugated with the CHXA”, and then radiolabeled with ¹¹¹In. The biodistribution was performed at 4 and 24 h time points in melanoma tumor-bearing and healthy C57BL/6 female mice. (3) The biodistribution of the melanin-binding antibody showed the significant uptake in the tumor, which increased with time, and very low uptake in healthy melanin-containing tissues such as the retina of the eye and melanized skin. This biodistribution pattern in healthy tissues was very close to that of the isotype matching control antibody. (4) Conclusions: The biodistribution experiment allows us to assess the pharmacokinetics of both antibodies side by side and to make a conclusion regarding the suitability of specific antibodies for further development.

Background: the primary challenge with implant infections is the formation of biofilm, which harbors dormant bacteria that reduce the effectiveness of antibiotics and amplify antibiotic resistance, exacerbating the global antimicrobial resistance crisis. A potential novel treatment strategy is radioimmunotherapy, which uses antibodies linked to radioisotopes to deliver targeted radiation to the bacteria and biofilm. We describe the first in vivo use of targeted radiation therapy, employing Actinium-225 (α-radiation) and Lutetium-177 (β-radiation) labeled antibodies to treat a Staphylococcus aureus biofilm-associated intramedullary implant infection. Untargeted radiation in the form of unbound radionuclide treatment was also evaluated. Methods: to assess therapeutic efficacy, bacterial counts were performed on implant and surrounding bone after seven days of follow-up. Biodistribution was evaluated using SPECT/CT and ex vivo gamma counting. Results: radioimmunotherapy using an antibody against wall teichoic acid which was labeled with Actinium-225 and Lutetium-177 achieved bacterial reductions between 45% and 93% on the implant and surrounding bone. Surprisingly, a similar antimicrobial effect was observed with unbound Actinium-225 treatment reducing the bacterial load by 80% on the implant and 98% in the surrounding bone. Indications of maximum tolerated dose (MTD) with Lutetium-177 labeled antibodies were observed through hepatic and renal function evaluations. Conclusions: These results should be interpreted in the context of the study’s constraints, particularly the limited animal sample size. Nonetheless, the results suggest that in vivo applied radiation may help reduce a biofilm-associated infection at the implant site as well as in the surrounding bone. These findings encourage further investigation into the use of targeted and non-targeted radiation, potentially combined with antibiotics, to develop effective strategies for eradicating biofilm-associated implant infections.

CCR8 chemokine receptor is a selective marker of tumor-infiltrating regulatory T cells (ti-Tregs) which interfere with the efficacy of checkpoint inhibitor immunotherapy (ICI) in many types of cancer. Eliminating CCR8+ ti-Tregs dramatically improves the results of subsequent ICI. We have recently reported using 225Actinium-labeled anti-CCR8 IgG for killing CCR8+ ti-Tregs in murine colorectal tumors which synergized with subsequent anti-CTLA4 ICI. Here, we aimed to compare the in vivo behavior of anti-CCR8 full-sized IgG and its Fab fragments to select the best antibody format for the pre-clinical development of this combination modality. Anti-CCR8 Fab fragments were generated by papain digest of the whole IgG. The whole IgG and Fab were conjugated to bifunctional chelating agent DOTA and labeled with 111Indium (¹¹¹In). MC8 and CT6 murine colorectal tumor-bearing C57Bl6 and Balb/c mice, respectively, were administered ¹¹¹In-DOTA-IgG or ¹¹¹In-DOTA-Fab and imaged with microSPECT/CT at 2–72 h post-injection. A biodistribution was performed after the last imaging time point. Both ¹¹¹In-DOTA-IgG and ¹¹¹In-DOTA-Fab demonstrated high tumor uptake in both MC38 and CT26 tumors, with ¹¹¹In-DOTA-IgG uptake being significantly higher from the 24 h time point and onwards. ¹¹¹In-DOTA-Fab also exhibited pronounced kidney uptake which persisted even at 72 h. The kidney clearance and retention of ¹¹¹In-DOTA-Fab might represent a problem during therapy employing 225Actimium or other long-lived therapeutic radionuclides by potentially causing a dose-limiting kidney toxicity. This imaging/biodistribution evaluation not only determined that full-size anti-CCR8 IgG is the optimal antibody format for pre-clinical development but also informed on the timing of immunotherapy administration in future radioimmunotherapy and immunotherapy combination studies.