BOB Ped Surg 2023 - Bernadette Jeremiasse, IPSO - Presentation

Achieving complete tumor resections without major complications remains challenging because some tumors are difficult to distinguish from healthy tissue and in other tumors the margin is very narrow.

Fluorescence guided surgery with tumor-specific probes can visually assist the surgeon in discriminating tumors from healthy tissue.

Tumor-specific probes are currently screened one by one and per tumor type.

Organoids maintain molecular tumor characteristics and can be held in culture for a long time.

For neuroblastoma, kidney organoids were chosen as control organoids based on the tissue surrounding the tumor.

For breast cancer, healthy breast organoids were chosen as control organoids.

Surface markers upregulated in neuroblastoma or breast cancer were identified based on RNA profiling, the human protein atlas, and literature review.

Probes targeting surface markers were conjugated to six different fluorophores ranging between 488 and 647 nanometers.

Seven-colored 3D live imaging using confocal microscopy was performed on living organoids using six probes plus the general marker E fluor.

Segmentation analysis was performed using the staple pipeline developed in the lab, and fluorescent signals were quantified.

The three most promising probes for neuroblastoma were validated in vivo using a mouse xenograft model with tumors originating from neuroblastoma organoids.

Probe fluorescence varies between patients and even within a patient in neuroblastoma organoid lines.

Different probes work on tumor organoids compared to healthy control organoids.

The three most promising probes tested in vivo were GD2, L1CAM, and NCAM.

An organoid was determined to be positive when the intensity of the tumor organoid was at least twice the fluorescence of the healthy control organoid.

The three validated probes resulted in tumor-to-background ratios above 2, indicating good intraoperative visibility.

In vitro imaging patterns resemble the efficacy in vivo, with L1CAM showing higher signal than GD2 in both settings.

Thousands of organoids were quantified using the segmentation and fluorescence intensity methodology.

Four markers showed promise for neuroblastoma based on high percentages of positive organoids and high fluorescent intensities.

The order of neuroblastoma organoid lines by probe efficacy was similar in vitro and in vivo, with line 129 being highest and line 67 being lowest.

One of the tested probes is going into a clinical trial this year.