Personalized chordoma organoids for drug discovery studies

Abstract

Chordomas are rare tumors of notochordal origin, most commonly arising in the sacrum or skull base. Chordomas are considered insensitive to conventional chemotherapy, and their rarity complicates running timely and adequately powered trials to identify effective treatments. Therefore, there is a need for discovery of novel therapeutic approaches. Patient-derived organoids can accelerate drug discovery and development studies and predict patient responses to therapy. In this proof-of-concept study, we successfully established organoids from seven chordoma tumor samples obtained from five patients presenting with tumors in different sites and stages of disease. The organoids recapitulated features of the original parent tumors and inter- as well as intrapatient heterogeneity. High-throughput screenings performed on the organoids highlighted targeted agents such as PI3K/mTOR, EGFR, and JAK2/STAT3 inhibitors among the most effective molecules. Pathway analysis underscored how the NF-κB and IGF-1R pathways are sensitive to perturbations and potential targets to pursue for combination therapy of chordoma.

Fig. 1.. Morphology of the PDOs established as visualized by brightfield imaging of maxi-rings in 24-well plates.

CHORD002 to CHORD005 were imaged daily over a 5-day incubation period, while CHORD001 was imaged for 4 days. The organoids displayed morphological features consistent with chordoma such as vacuolated cells arranged in clusters or nests. The sample shown for patient CHORD002 is CHORD002a. Scale bar, 50 μm.

Fig. 2.. Histology and immunohistopathology characterization of chordoma samples and derived organoids.

Formalin-fixed paraffin-embedded sections from both the parent tumor and organoid were stained with H&E, Ki-67, and brachyury. All organoids recapitulated features of the parent tumor. The sample shown for patient CHORD002 is CHORD002a. Scale bar, 20 μm.

Fig. 3.. Dot map of high-throughput screening results for 124 overlapping small molecules tested on all PDOs.

The size of each circle represents the Z score, with larger ones indicating a higher Z score value. The color of each point represents the normalized cell viability %. The drugs are clustered using the Jaccard distance based on common protein targets, and cases are clustered on the basis of their similarities in drug responses. Covariates included represent the type of tissue and location of each sample as well as FDA status for the various drugs.

Fig. 4.. Dot map of drug screening on all three samples procured from patient CHORD002.

(A and B) Drugs that induce 15% cell death in at least two of the samples are visualized (51 total). The size of each circle represents the Z score, with larger ones indicating a higher Z score value. The color of each point represents the normalized cell viability %. The drugs are clustered using the Jaccard distance based on common protein targets. Samples are ordered by date of acquisition. Covariates included represent the type of tissue, anatomic location of each sample, and the number of lines of treatment the patient was exposed to before procurement of sample as well as FDA status at the time of publication.

Fig. 5.. Pathway analysis for all chordoma PDOs.

The top 30 ranked pathways that are affected in at least two samples are visualized (53). For each dot, the amount of surface filled in (in black) corresponds to the percentage of proteins within a pathway that were targeted, while the size corresponds to the pathway score. The pathways are clustered using the distance based on their shared proteins. The samples are clustered by the Euclidian distance of their pathway scores. Covariates included represent the type of tissue and site of each sample.