General

• Mitosis and apoptosis were rarely identified in the 8 pediatric chordomas and showed no correlation with MIB-1 LI. [1]
• The clinical benefit observed in chordoma patients treated with imatinib seems to be attributable to the switching off of all three receptors . [2]

Cytokine Receptor

• Cells in a tissue microarray of 70 chordomas samples showed nuclear staining for phosphorylated signal transducers and activators of transcription (Stat3). The Stat3 pathway is constitutively activated in chordomas. [3]
• MTT assay showed that the growth of 3 out of 3 chordoma cell lines (UCH1, CH 8 and GB 60) was inhibited by SD-1029, an inhibitor of Stat3 activation. Treatment with SD-1029 inhibited expression of Stat3 signaling cascade (Western blot), phosphorylation of Stat3 in chordoma cells in vitro, proliferation in three-dimensional culture (immunofluorescence), and antiapoptotic proteins Bcl-xL and MCL-1. [3]
• Phosphorylated isoforms of p44/42 mitogen-activated protein kinase, Akt and STAT3, indicative of tyrosine kinase activity, were detected in 86%, 76% and 67% of cases, respectively. [4]
• The cytotoxicity of the combination of SD-1029 and chemotherapeutic drugs cisplatin or doxorubicin is significantly better than either agent alone. Cytotoxicity assay showed that the growth of 3 out of 3 chordoma cell lines was significantly inhibited after treatment with the combination of SD-1029 and cisplatin or doxorubicin (P < 0.01). [3]

Receptor Tyrosine Kinase

• Western blot analysis showed phosphorylation of FRS2a and ERK1/2 in 6 out of 6 chordomas that were also reactive by immunohistochemistry for at least one of the FGFRs : FGFR1, FGFR2, FGFR3, and FGFR4. [5]
• IHC analysis of 21 cases of chordoma for expression of proteins involved in signal transduction from RTKs indicated platelet-derived growth factor receptor-b (PDGFR-b), epidermal growth factor receptor (EGFR), KIT and HER2 were detected in 100%, 67%, 33% and 0% of cases, respectively. [4]
• Correlation between EGFR expression and p-EGFR was not statistically significant. Positive staining for p-EGFR correlated with p-STAT3 (McNemar’s test; P = 0.0235), but not with p-MAPK or p-Akt. [4]
• Phosphorylated PDGFRA, PDGFRB and KIT were detected 12/12, 18/18 and 12/14 chordomas, respectively. PDGFRB was detected in all of these samples at levels higher than synovial sarcoma specimen used as a positive control, while PDGFRA and KIT were less highly expressed than the GIST sample used as a positive control. [6]
• No gain-of-function mutations were found in PDGFRA, PDGFRB or KIT. [6]
• PDGFA, PDGFB, and SCF mRNA were detected in 31 of 31 chordomas, suggesting that activation of their corresponding receptors is via the the autocrine/paracrine loop. [2]
• S6 was expressed in 14 of 22 chordomas. Phosphorylation of S6 Ser240–244 was detected to a low / very low extent in 11 of the 14 S6 positive tumors. Phosphorylation of S6 Ser235–236 was detected in 8 of 13 chordomas analyzed, suggesting mTOR-independent activation of S6 in some cases via the RAS/MAPK pathway. Erk1/2 (p44 and p42) was highly expressed and phosphorylated (Thr202 / Tyr204) in 22 of 22 chordomas by western blot. [7]
• 6/8 chordoma samples that were immunoreactive for at least one FGFR also showed phosphorylation FRS2-alpha and ERK1/2. [8]
• 2/8 chordoma samples that were not immunoreactive for any FGFR were also negative by Western blot analysis for p-FRS2 a and positive for p-ERK1/2. [8]
• 24/49 non-skull-based and 22/48 skull-based chordoma samples showed membranous or cytoplasmic p-EGFR immunoreactivity. [8]
• RTK antibody array membranes showed p-EGFR, MSPR, and EphB2 to be highly expressed in the U-CH1 cell line and three primary chordoma tumors. [8]

PI3K

• Among 5 chordomas, most demonstrated activation of the Akt/mTOR cascade including expression of p-Akt (4/5), mTOR (5/5), p-mTOR (5/5), p-S6K(5/5) and 4E-BP1 (4/5). [9]
• mTORC1 signaling in chordoma-derived cell lines is deregulated in response to growth factor deprivation, but remains sensitive to amino acid availability. mTORC1 signaling is hyperactivated in sporadic sacral chordomas. [4]
• Serum-starved U-CH1 cells exhibited high levels of pAkt, pTSC2, and pPRAS40. Akt signaling in U-CH1 cells is also constitutively activated in a growth factor-independent manner. ERK phosphorylation was also high in serum-starved U-CH1 cells. [9]
• PTEN expression was not observed in U-CH1 cells and was significantly reduced in Ch1 cells. These results suggest that constitutively high Akt activity, due to PTEN loss, may be responsible for hyperactivation of mTORC1 signaling through inactivation of TSC2 and PRAS40 in U-CH1 cells. Partial or complete deficiency of PTEN could be responsible for hyperactivation of mTORC1 signaling in at least a subset of chordomas. [9]
• The p85 subunit of PI3K co-immunoprecipitated with activated PDGFRB in 22 of 22 samples, confirming binding of these two signaling molecules. [7]
• Rapamycin inhibited mTORC1 activation and suppressed proliferation of chordoma-derived cell line. [9]
• Immunohistochemistry for AKT/TSC/mTOR pathway molecules performed on tissue microarray slides from sacro-ccocygeal chordomas showed that 45/49 samples were positive for p-AKT (Ser 473 ), 22/48 samples were positive for TSC1, 49/49 samples were positive for TSC2, 47/49 samples were positive for p-TSC2 (Thr 1462 ), 13/48 samples were positive for p-mTOR (Ser 2448 ), 33/44 samples were positive for mTOR, 29/47 samples were positive for p-p70S6K (Thr 389 ), 50/50 samples were positive for S6K, 11/49 samples were positive p-RPS6 (Ser 235/236 ), 22/45 samples were positive for RPS6, 46/48 samples were positive for p-4E-BP1 (Thr 70 ), 47/48 samples were positive for eIF-4E, 37/43 samples were positive for PTEN, and 2/48 samples were positive for CDKN2A. [8]
• Immunohistochemistry for AKT/TSC/mTOR pathway molecules performed on tissue microarray slides from skull-based chordomas showed that 30/48 samples were positive for p-AKT (Ser 473 ), 33/48 samples were positive for p-TSC2 (Thr 1462 ), 26/48 samples were positive for p-mTOR (Ser 2448 ), 25/48 samples were positive for p-p70S6K (Thr 389 ), 30/48 samples were positive for p-RPS6 (Ser 235/236 ), 46/48 samples were positive for p-4E-BP1 (Thr 70 ), 44/48 samples were positive for eIF-4E, and 42/50 samples were positive for PTEN. [8]
• 13/13 chordoma samples showed phosphorylation of 4E-BP1 and expression of eIF-4E. 12/13 showed immunoreactivity for p-mTOR and total mTOR and demonstrated two copies of mTOR locus by FISH. 11 of these 12 showed activation of p70S6K and 7 of these 12 showed immunoreactivity for p-RPS6. Chordomas negative for p70S6K were also negative for p-RPS6. [8]
• 35/48 chordoma samples were negative for p-mTOR. 21/35 of these were immunoreactive for total mTOR. 11/21 of these showed two mTOR alleles and 4/21 of these showed loss of one allele (only 15/21 were analyzable by FISH). 5 of the 9 cases negative for both p-mTOR and total mTOR showed loss of one mTOR allele. ~50% of p-mTOR-negative chordomas showed activation of neither p70S6K nor RPS6; the other ~50% were positive for p-p70S6K. [8]
• 38/49 chordomas were negative for p-RPS6. [8]
• The PI3K/AKT/TSC/mTOR pathway was found to be activated in at least 65% of the tumors examined from samples originating from 50 sacro-coccygeal and 50-skull-based human chordomas and the cell line U-CH1. [8]
• Western blot analysis of 13 tissue samples from patients diagnosed and surgically treated for chordoma showed positive staining for P-PDK-1, PDK-1, P-AKT, AKT, P-mTOR, mTOR, P-S6, and S6 in the majority of samples, demonstrating activation of the PI3K/AKT/mTOR pathway in chordoma. [10]

References