Prior chemotherapy leaves lasting genetic and gene-expression marks in metastatic colorectal cancer organoids
Lab-grown tumor organoids from 35 patients reveal how prior chemo shapes mutations and drug resistance, with oxaliplatin resistance proving especially durable.
Riepilogo
Treatment-resistant metastatic colorectal cancer is hard to study because prior chemotherapy changes tumors in ways that are poorly understood. Researchers compared 35 patient-derived organoids from chemonaive (17) and heavily pretreated (18) patients using drug screening, RNA sequencing, and whole-genome sequencing. Pretreated organoids carried more mutations and structural variants, plus chemotherapy-linked mutational signatures that tracked with the number of treatment cycles. Oxaliplatin resistance persisted in organoids, tied to ZNF300, TGM2, and Hedgehog signaling. Irinotecan (SN-38) resistance was linked to deep deletions at common fragile sites and certain signatures, while 5-FU resistance was only partly captured. The findings suggest organoids preserve some, but not all, treatment-induced resistance mechanisms.
Riepilogo Dettagliato
Drug resistance is the main obstacle in treating metastatic colorectal cancer (mCRC), and most drug-development models come from chemonaive tumors. That leaves open how years of chemotherapy reshape tumor cells, and whether those changes can be modeled in the lab. This matters particularly for heavily pretreated patients, who are often enrolled in early-phase trials that mostly fail for lack of efficacy.
The team analyzed 35 patient-derived metastatic organoids (PDOs), mostly from liver metastases: 17 from chemonaive patients biopsied before first-line therapy and 18 from patients previously exposed to 5-FU, oxaliplatin, and irinotecan. They screened sensitivity to SN-38, 5-FU, oxaliplatin, and four experimental drugs, and paired this with bulk RNA sequencing and whole-genome sequencing. Organoids matched their original tumors closely (99% driver-gene concordance in six pairs).
Pretreated PDOs had higher mutational load (median 132 vs 97), burden (7.8 vs 5.4 per Mb), and structural variants (229 vs 124). Chemotherapy-associated signatures (SBS17a/b for 5-FU, SBS35 and DBS5 for oxaliplatin) were common in pretreated samples and rare in chemonaive ones; SBS17a/b and DBS5 correlated with the number of cycles received. A larger WGS cohort (78 chemonaive, 138 pretreated) confirmed the higher mutation burden. Deep deletions in common fragile sites were more frequent in pretreated PDOs (44% vs 29%) and, unexpectedly, were associated with greater SN-38 resistance (median GRAUC 0.77 vs 0.49), alongside higher BIRC3 and FBXO44 expression. SBS17a was also linked to SN-38 resistance.
Oxaliplatin resistance was clearly retained: organoids from clinically refractory patients were less sensitive (GRAUC 0.67 vs 0.44), with upregulated ZNF300 and TGM2 and Hedgehog pathway enrichment. Gene-expression analyses indicated oxaliplatin-resistance programs (e.g., L1CAM, an EMT/hypoxia/KRAS-enriched module) were present regardless of treatment history, whereas 5-FU and SN-38 resistance-associated expression appeared mainly in pretreated PDOs. Several identified genes (ETS1, L1CAM, MIR31HG, NRIP3, NDEL1) tracked with worse survival in an independent dataset.
Together, the results suggest PDOs capture chemotherapy-induced changes differently by drug, reflecting both stable tumor-intrinsic properties and dynamic cell states. Caveats include the small cohort, differing clinical profiles between groups (e.g., sidedness, survival), correlative findings that cannot establish whether alterations cause or merely follow resistance, and incomplete modeling of 5-FU resistance in vitro.
Risultati Principali
- Pretreated organoids had higher mutational load (132 vs 97), burden, and structural variants (229 vs 124) than chemonaive organoids.
- Chemotherapy-linked mutational signatures (SBS17a/b, DBS5) were common after treatment and correlated with 5-FU and oxaliplatin cycle numbers.
- Oxaliplatin resistance persisted in organoids, with ZNF300 and TGM2 upregulation and Hedgehog pathway enrichment.
- Deep deletions at common fragile sites and SBS17a were associated with greater SN-38 (irinotecan) resistance in pretreated organoids.
- Oxaliplatin-resistance expression programs appeared across all organoids, while 5-FU and SN-38 patterns were mostly limited to pretreated ones.
Metodologia
Researchers compared 35 patient-derived metastatic CRC organoids (17 chemonaive, 18 pretreated), integrating drug-sensitivity screening (GRAUC) with bulk RNA-seq and whole-genome sequencing, including mutational signature analysis, differential expression, and WGCNA. Findings were cross-checked against a larger WGS cohort, DepMap cell lines, a public survival dataset, and immunohistochemistry.
Limitazioni dello Studio
The cohort is small, and the chemonaive and pretreated groups differ clinically (e.g., tumor sidedness, survival), which may confound comparisons. Associations between genomic or expression changes and resistance are correlational, and 5-FU and irinotecan acquired resistance were only partially captured in organoids, which lack the tumor microenvironment.
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