Table 2. Transcriptional heterogeneity in CTCs.
| #CTC | #Pts | Isolation | Analysis | Targets | Heterogeneity | Ref. |
|---|---|---|---|---|---|---|
| Breast cancer | ||||||
| n.s. | 17 | HBCTC-Chip | RNA-ish; RNA-seq-DGE | EMT markers | Heterogeneous fractions of Epithelial (E), Mesenchymal (M), and EM-CTCs; In TNBC more homogeneous pools of M-CTCs. Temporal heterogeneity: at progressive disease, 10 patients harbored emerging numbers of M-CTCs. | 81 |
| 105 | 35 | MagSweeper + MM | qRT-PCR | 87 cancer-associated genes | Two major subgroups of CTCs, i.e. high expression of EMTgenes and high metastasis-associated genes. Heterogeneity based on CTCs not clustering by patient-ID and 8 patients having CTCs in both clusters. | 132 |
| 15 pools + 14 clusters | 10 | CTC-iChip + MM | RNA-Seq | Whole transcriptome | Based on global gene expression level, all isolated CTCs clustered closely by patient of origin. Based on JUP and 31 cluster-associated genes, CTC-clusters could be differentiated from pooled single CTCs. | 124 |
| ~400 | 20 | IM (Maintrac) + AP | PCR + gelelectro-phoresis | HER2, EpCAM, Vimentin, and NANOG | Expression patterns changed after surgery, with emerging of a sub-population of EpCAM positive CTC expressing NANOG and/or vimentin. | 130 |
| Prostate cancer | ||||||
| 77 | 13 | negCTC-iChip + MM | RNA-seq | Whole transcriptome | Single CTCs from nine individual patient with at least 3 CTCs analyzed, showed considerably higher intra-patient heterogeneity in their transcriptional profiles compared to single cells from prostate cancer cell lines. | 126 |
| 20 | 4 | MagSweeper + MM | RNA-seq | Whole transcriptome | All CTCs, except two, cluster in a patient specific manner. 181 cancer-specific genes were overexpressed in the CTCs, compared to normal tissue. Specific transcripts, e.g. related to CRPC or ERG-fusion, were detected homogeneously within the same patients. | 131 |
| 48 | 2 | MagSweeper + Nanowell | RNA-seq | KLK3 (PSA) mRNA | KLK3 expression was variable between the 26 individual CTCs, for which a sufficient number of genes including KLK3 were covered. | 34 |
| 38 | 8 | MF + MM | qRT-PCR | 84 EMT-related genes | Heterogeneous upregulation of EMT-associated gene expression, especially in CRPC. | 129 |
| pools | 21 | IM (AdnaTest) | qRT-PCR | AR full length + AR-V7 | Temporal heterogeneity: 1 out 9 patients converted to AR-V7 positive, at progression on Taxane. While 7 out 12 patient who were at baseline AR-V7 positive became negative at progression. | 73 |
| Pancreatic cancer | ||||||
| 265 | 15 | HBCTC-Chip | RNA-ish; RNA-Seq-DGE | WNT2 | RNA-ish showed heterogeneity of WNT2 expression in CTCs and the primary tumor. This was confirmed by RNA-seq with DGE, showing rare WNT2 RNA reads in the enriched CTC sample and the primary tumor. | 37 |
| Melanoma | ||||||
| 6 | 1 | MagSweeper + MM | RNA-seq | Whole transcriptome | CTCs show a uniform upregulation of melanoma markers, including MAGE as well as uniform up- or downregulation of certain plasma membrane proteins. | 128 |
| Multiple cancers | ||||||
| 7, 29, 77 | n.s. | negCTC-iChip + MM | RNA-seq | Whole transcriptome | High expression of stromal-derived ECM proteins in > 15% of CTC samples. One glycoprotein was expressed in 100% of pancreatic CTCs compared to 31% of breast and 9% of prostate CTCs. | 125 |
Abbreviations: CTC, circulating tumor cell; DGE, digital gene extraction; ECM, extracellular matrix; EMT, epithelial-to-mesenchymal transition; IM, immunomagnetic enrichment; MF, microfiltration; MM, micromanipulation; qRT-PCR, quantitative reverse transcription polyclonal chain reaction; RNA-ish, RNA in situ hybridization; RNA-seq, RNA sequencing; n.s., not specified.