Hence, it has been assumed that molecular information found in a tumor biopsy (e.g., mutations, DNA copy number changes, DNA rearrangements) recapitulates the molecular events of the whole neoplasm. This concept has been challenged by Gerlinger et al., who reported intratumor heterogeneity in primary renal cell carcinoma and associated metastasis

by testing dense scrutiny of mutations using next-generation sequencing. Sampling included nine specimens from the primary tumor and additional specimens from two metastatic sites, all from the same individual. They identified 128 nonsynonymous mutations with different regional distribution. The results included 40 mutations ubiquitous to all specimens, 59 shared by several HDAC inhibitor but not all regions and 29 unique to specific specimens (so-called private mutations). Thus, most somatic mutations (∼65%) were not detected across every tumor region explored. One such target was the mutation of mammalian target of rapamycin (mTOR) affecting the kinase domain (L2431P), which correlated with mTOR pathway activation in human samples and in experimental models of renal cancer. This finding suggests that genetic intratumor heterogeneity was also inducing functional heterogeneity. Interestingly, one of the samples from the primary tumor shared mutations with the

metastatic sites. The gene expression data revealed that this same specimen also shared a gene signature with the metastasis, pointing MK-2206 in vitro toward a possible location of metastasis-enabling cells within the primary tumor. Based on these 上海皓元 data, authors inferred ancestral relationships and were able to construct a phylogenetic tree with all tumor specimens from the same individual. These findings are in line with the hypothesis of clonal evolution,11 a

model that applies Darwinian selection rules to justify constant evolutionary changes in cancers and provides a general mechanistic framework to explain tumor heterogeneity and drug resistance.12 Additional evidence in other malignancies suggests frequent intra-individual heterogeneity in advanced cancer stages. For instance, a study analyzing mutations in different lesions from a patient with metastatic pancreatic cancer found a mixture of cellular subclones in the primary tumor that correlated with molecular changes in metastasis, an additional clue for the presence of metastasis-enabling cells in the primary tumor.13 Data from a similar report focusing on chromosomal aberrations also showed considerable intratumor heterogeneity in pancreatic cancer, probably responsible for independent metastasis.14 Strikingly, sophisticated mathematical modeling of pancreatic metastasis kinetics indicates that all patients are expected to harbor subclones of metastasis-enabling cells in the primary tumor at the time of diagnosis, even when tumor size is fairly small.