Abstract
Patient‐derived in vitro cultures of colorectal cancer (CRC) may help guide treatment strategies prior to patient treatment. However, most previous studies have been performed on a single biopsy per tumor. The purpose of this study was to analyze multiple spatially distinct biopsies from CRCs and see how well intratumor heterogeneity (ITH) was recapitulated in matching patient‐derived spheroids. Three to five biopsies were collected from six CRC tumors. Each biopsy was split in two; one half was used for spheroid culturing, while the other half was used for DNA and RNA purification. For two patients, lymph node metastases were analyzed. Somatic mutations were called from whole exome sequencing data. Each tumor contained mutations shared across all biopsies and spheroids, including major CRC drivers such as APC, KRAS, and TP53. At the same time, all tumors exhibited ITH on both mutation and copy number level. The concordance between biopsies and spheroids ranged between 40 and 70% for coding mutations. For three patients, the biopsy and spheroid from matching areas clustered together, meaning that the spheroid resembled the area of origin more than the other areas. However, all biopsies and spheroids contained private mutations. Therefore, multiple cultures from spatially distinct sites of the tumor increase the insight into the genetic profile of the entire tumor. Molecular subtypes were called from RNA sequencing data. When based on transcripts from both cancer and noncancerous cells, the subtypes were largely independent of sampling site. In contrast, subtyping based on cancer cell transcripts alone was dependent on sample site and genetic ITH. In conclusion, all examined CRC tumors showed genetic ITH. Spheroid cultures partly reflected this ITH, and having multiple cultures from distinct tumor sites improved the representation of the genetic tumor subclones. This should be taken into account when establishing patient‐derived models for drug screening.
Highlights
Colorectal cancer (CRC) is among the leading causes of cancer-related deaths in the Western World
Regional lymph node metastasis (LNM) were identified during pathological examination of the resected tumor specimen
The cancer percentage of each biopsy was assessed by histology, and all samples with a cancer percentage below 60% were purified by laser capture microdissection (LCM) before DNA and RNA extraction
Summary
Colorectal cancer (CRC) is among the leading causes of cancer-related deaths in the Western World. Primary models of CRC such as cancer tissue-originated spheroids and patient-derived organoids (PDOs) are being established with increasing success rate (Ashley et al, 2014; Jeppesen et al, 2017; Kondo et al, 2011; Sato et al, 2011; Schu€tte et al, 2017; van de Wetering et al, 2015). These 3D culturing systems increase the success rate of primary cultures and resemble the primary tumor better than traditional onedimensional cell culturing (Weiswald et al, 2015). The aim of this study was to characterize the ITH within CRC and investigate how well it is reflected in matching spheroids derived from multiple spatially distinct sites of the primary tumor
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