Potential conflict of interest: Dr. Takebe advises Healios KK.
Liver cancer is currently the second leading cause of cancer‐related death worldwide.1 Primary liver cancer (PLC) is a complex disease consisting of multiple cancer subtypes, each with a different clinical pathology and requiring different treatment. In addition to this, the coming of age of the genomics era has made it abundantly clear that significant genetic variations behind PLC on an individual level necessitate the need for personalized treatment regimens.
The field of cancer research is currently limited by lack of a tractable in vitro model. Most in vitro studies today are done in oversimplified two‐dimensional cell culture systems, which fail to recapitulate the patient phenotype and lead to a downstream bottleneck in drug discovery when studies are transitioned in vivo. Recently, three‐dimensional culture systems have been implemented, with the current state of the art being organoids that consist of multiple cell types and can mimic structural complexity on the organ level. Organoids use nontransformed cells and have been created using a variety of tissue types, and some studies have already looked at the creation of organoids from primary cancer tissues. For example, early cancer organoid models focused on relatively simple organs including esophagus and intestine2; however, more recent research has moved to increasingly complex tissue including pancreas and prostate.3 With more than 500 functions, the liver is one of the most complex organs in the body; and because it is the main site of drug metabolism, liver organoids offer great potential for the pharmaceutical industry.
The development of human liver organoid culture has been described by Huch and colleagues.5 In this follow‐up study, Broutier et al. published a paper in the December issue of Nature Medicine, wherein they develop a novel organoid culture system using primary tumors.6 These organoids derived from tumors (tumoroids) were created from primary patient samples and were able to be cultured long term. In order to grow tumoroids, an adapted version of the previous culture method defined for hepatic organoid development was used. In particular, the digestion time was increased to preclude nontumor tissue contamination, and a new PLC‐derived organoid isolation medium was used consisting of the classic isolation medium including dexamethasone and Rho kinase inhibitor but excluding R‐spondin, Noggin, and Wnt3a to enrich liver cancer organoids.
Upon these protocol optimizations, they created eight patient‐derived tumoroids from the three most common forms of PLC: hepatocellular carcinoma (HCC), cholangiocarcinoma (CC), and combined HCC/CC tumors. One of these organoids was unable to be maintained long term. However, the remaining seven tumoroids followed similar gene expression patterns, histology, and genetic mutations compared to the patients; and these phenotypes were maintained over time with limited additional alterations. Excitingly, not only were the transcriptome patterns seen in the tumoroids similar within subtypes of cancer but also global exome sequencing confirmed that specific genetic aberrations were maintained at >90% compared to parental tumor when organoids were cultured less than 2 months and >80% when cultured more than 4 months.
These tumoroids also maintained the metastatic capabilities of the original tumors in a xenograft model. Mice injected with tumoroids into the kidney capsule developed lung metastases. Finally, a proof‐of‐concept drug screen was performed. Tumoroids responded to 29 drugs with varying levels of sensitivity, further showing their use as a personalized drug screen platform. Interestingly, a new extracellular signal–regulated kinase (ERK) inhibitor drug was discovered that strongly inhibits the growth of tumoroids, despite lacking a mutation in the ERK gene. Most excitingly, this discovery translated in vivo as well; mice injected with tumoroids demonstrated consistent reduction in tumoroid size when treated with the ERK inhibitor.
Organoid‐related research faces a common challenge of variability and reproducibility that may apply to this research. For example, the authors claimed that omission of the Wnt agonist R‐spondin‐1 from normal organoid medium selectively amplify tumoroid lines; however, one line out of 7 patients, noted as CC‐1, requires the R‐spondin inclusion. This careful tune‐up process does not just complicate things from a technical aspect but could potentially lead to highly heterogeneous organoid formation, including normal organoids, which eventually outcompete tumoroids, as stated by the authors. To circumvent this, Broutier et al. used a manual pick‐out method to purify the cancer‐specific tumoroid, but this will potentially impose a reproducibility challenge by others without a highly experienced background. That said, further protocol standardization will be critical to assess the broader applicability of this approach.
While these studies form a basis for future cancer precision research, they are currently limited by a small number of patient samples, with only n = 2 for each liver cancer subtype regarding drug screen experiments. It will be interesting to see if larger sample sizes will still allow for exact matches between tumor samples and organoids or if subgroups with similar genotypes and expression patterns will arise. In addition to this, the current methods of organoid derivation are limited in their ability to derive organoids from very well‐differentiated tumors, narrowing the clinical applications to patients with more severe and advanced tumors. It also remains to be seen whether such a system will be able to identify drugs capable of treating secondary metastases.
The current landscape of genomic studies has shown an ever increasing number of potential cancer‐causing mutations and resulting mechanisms, suggesting that a personalized approach to cancer treatment is essential to treating the population at large.7 Broutier and colleagues show the immense potential of organoids to fill the emerging gap in personalized therapeutics as well as pharmaceutical drug screens. They have implemented a 384‐well format that will allow for large‐panel drug screens for individuals, as shown in Figure 1. This could also be translated to noncancerous organoids to look for potential side effects in healthy individuals, and the addition of stellate cells and macrophages to this system could allow the modeling of those with precancerous conditions such as fatty liver or fibrosis and cirrhosis. More broadly, a living biobank of cancer organoids is an exceedingly promising concept for establishing the basis of precision oncology. Indeed, breast cancer organoids have recently been reported following a similar experimental layout by using >100 primary and metastatic breast cancer samples.8
Overall, these studies form an important first step toward the use of liver cancer organoids in drug‐screening capabilities. This research shows a proof of concept with exciting potential as seen with the discovery of an ERK inhibitor as a new mechanism of drugs for inhibition of liver cancer growth. It is likely that using such a method will reveal novel pathways in cancer that will further speed up drug discovery and precision oncology.
1. Wong MCS, Jiang JY, Goggins WB, Liang M, Fang Y, Fung FDH, et al. International incidence and mortality trends of liver cancer: a global profile. Sci Rep 2017;7:45846.
2. Sato T, Stange DE, Ferrante M, Vries RGJ, Van Es JH, Van Den Brink S, et al. Long‐term expansion of epithelial organoids from human colon, adenoma, adenocarcinoma, and Barrett's epithelium. Gastroenterology 2011;141:1762‐1772.
3. Boj SF, Hwang CI, Baker LA, Chio II, Engle DD, Corbo V, et al. Organoid models of human and mouse ductal pancreatic cancer. Cell 2015;160:324‐338.
4. Gao D, Vela I, Sboner A, Iaquinta PJ, Karthaus WR, Gopalan A, et al. Organoid cultures derived from patients with advanced prostate cancer. Cell 2014;159:176‐187.
5. Huch M, Gehart H, Van Boxtel R, Hamer K, Blokzijl F, Verstegen MMA, et al. Long‐term culture of genome‐stable bipotent stem cells from adult human liver. Cell 2015;160:299‐312.
6. Broutier L, Mastrogiovanni G, Verstegen MM, Francies HE, Gavarró LM, Bradshaw CR, et al. Human primary liver cancer–derived organoid cultures for disease modeling and drug screening. Nat Med 2017;23:1424‐1435.
7. Zhang T, Hu Y, Wu X, Ma R, Jiang Q, Wang Y. Identifying liver cancer–related enhancer SNPs by integrating GWAS and histone modification ChIP‐seq data. Biomed Res Int 2016;2016:2395341.
8. Sachs N, de Ligt J, Kopper O, Gogola E, Bounova G, Weeber F, et al. A living biobank of breast cancer organoids captures disease heterogeneity. Cell 2018;172:373‐386.