Because WDLS and DDLS respond poorly to systemic chemotherapy, it is essential that novel molecular targets be identified to provide new possibilities for therapies. WDLS and DDLS have been known for some time to be characterized by amplification of genes on chromosome 12q. Neochromosomes carry supernumerary copies of the recognized oncogenes HDM2, HMGA2
, and CDK4
, all of which have been implicated in liposarcomagenesis. shRNAs directed against MDM2 and CDK4 inhibit proliferation of liposarcoma cell lines in vitro
], and targeting of MDM2 and CDK4 is an active area of clinical research [18
MDM2 is essential for ubiquitination and degradation of the tumor suppressor p53. MDM2
amplification is, therefore, thought to result in reduced levels of p53 and thus to induce transformation of the progenitor cell [19
]. To modulate p53, MDM2 requires the function of a RING domain, which binds to p53 [21
]. Competitive inhibitors (e.g., nutlin) of the MDM2-p53 interaction have been developed [22
]. In DDLS cell lines, nutlin treatment increases both p53 levels and apoptotic rates [23
]. Given these data, MDM2 inhibitors are now in clinical trials for WDLS and DDLS; results are eagerly awaited.
Drugs that inhibit CDK4 are also being developed. This protein kinase modulates the G1/M cell cycle transition. In vitro
, inhibition of CDK4 protein leads to proliferation arrest [17
]. In clinical trials, CDK4 inhibitors have good safety profiles; however, efficacy has not yet been reported. CDK4 overexpression appears to correlate with a chromosome 12q amplicon distinct from that associated with gain of MDM2 and HMGA2, and a small set of WDLS with good prognosis do not have increased levels of CDK4 protein [24
]. Thus, CDK4 may be essential for progression, but not initiation, of liposarcoma, and inhibiting CDK4 may not be sufficient to induce tumor cell death. Instead, CDK4 inhibitors may slow tumor growth, be effective in only a subset of patients, or prove to be ineffective as single agents.
To continue the search for molecular targets in WDLS and DDLS, several groups are seeking to identify aberrantly regulated genes that are essential to liposarcomagenesis and whose protein products are potentially targetable by small-molecule inhibitors. To facilitate these studies, primary cell lines and tumor xenografts have been developed from WDLS and DDLS tumors. The receptor tyrosine kinases MET, AXL, KIT, and IGF-1R are upregulated in these cell lines as compared to adipocytes, and could be studied as potential drug targets [25
Gene expression profiling has been performed on primary tumor tissue to address the mechanisms of liposarcomagenesis [23
]. Sixty-nine liposarcomas and nine normal fat samples were profiled using Affymetrix U133A microarrays. This analysis identified 998 genes that had ≥2-fold difference in expression between WDLS and DDLS and normal fat. Through Ingenuity pathway analysis, these genes were found to affect numerous cellular pathways. One of the most significant of these was, not surprisingly, the cell cycle/checkpoint pathway including CDK4, MDM2, CDC2, CDC7, and cyclins B1, B2, and E2. It is possible that, in addition to CDK4 and MDM2, other components of this pathway could be targeted for liposarcoma therapies. Three of the most strongly overexpressed genes in DDLS and WDLS were TOP2A, RRM2,
. TOP2A (topoisomerase 2) and RRM2 (ribonucleotide reductase M2 polypeptide) can be targeted by currently available drug combinations. ZIC1
, which encodes a transcription factor normally expressed only in the adult cerebellum, was shown in subsequent experiments to be essential for liposarcomagenesis, suggesting that drugs directed against ZIC1 may likewise have therapeutic benefit.
A more extensive analysis of DDLS was recently performed as part of the Sarcoma Genome Project in collaboration between Memorial Sloan-Kettering Cancer Center and the Broad Institute at MIT [17
]. Fifty DDLS samples were examined to identify not only aberrantly expressed genes, but also copy number alterations and somatic mutations that could contribute to liposarcoma initiation and progression. Point mutations were identified in CTNNB1
(ephrin A1), and FBXW7
(a component of the ubiquitin protein ligase complex), each of which has potential oncogenic effects on the liposarcoma cell. Amplification on chromosome 12q was confirmed as a common feature in DDLS, but amplification was also observed to affect chromosomes 1q, 5p, and 20q. To identify previously unrecognized oncogenes essential for liposarcomagenesis, 385 genes from amplified regions were subjected to an shRNA screen in three DDLS cell lines. For 27 genes, including CDK4
, shRNA knockdown inhibited proliferation of cell lines, identifying these genes as potential oncogenes (). A function for MDM2 was not observed in the screen even though it is known to be essential for liposarcoma cell growth, demonstrating that even an extensive screen of this nature may underestimate the complexity of the genomic alterations that induce tumor formation.
Selected genes amplified in WDLS and DDLS and necessary for cell proliferation