WO2015184066A1

WO2015184066A1 - Differential expression of genes in benign and malignant lipomatous tumors

Info

Publication number: WO2015184066A1
Application number: PCT/US2015/032831
Authority: WO
Inventors: Brad A. BRYAN; Kristin Ward
Original assignee: Texas Tech University System
Priority date: 2014-05-28
Filing date: 2015-05-28
Publication date: 2015-12-03

Abstract

The present invention includes method for diagnosis and treatment of a malignant liposarcoma comprising obtaining a biological sample from a subject suspected of having a malignant liposarcoma; detecting a differential expression of a Class II Major Histocompatibility Complex Transactivator (CIITA), Sex Determining Region Y-Box 30 (SOX30); ATP -Binding Cassette, Sub-Family B (MDR/TAP), Member 11(ABCB11); or Synuclein, gamma (breast cancer-specific protein 1) (SNCG) obtained from the biological sample, wherein the differential expression of the CIITA, SOX30, ABCBl11, or SNCG is a prognostic marker of disease progression and metastatic potential in tumors and can to distinguish between benign or early stage liposarcomas and well-differentiated, aggressive liposarcoma; and modifying the type and extent of immunotherapy treatment depending on the differentiated disease prognosis and metastatic potential of the well-differentiated, aggressive liposarcoma.

Description

DIFFERENTIAL EXPRESSION OF GENES IN BENIGN AND MALIGNANT

LIPOMATOUS TUMORS

TECHNICAL FIELD OF THE INVENTION

The present invention relates in general to the field of cancer detection and treatment, and more particularly, to the use of differential expression of the Class II Major Histocompatibility Complex Trans activator (CIITA), Sex Determining Region Y-Box 30 (SOX30); ATP-Binding Cassette, Sub-Family B (MDR/TAP), Member 1 1 (ABCB1 1); or Synuclein, gamma (breast cancer-specific protein 1) (SNCG) to distinguish between benign and malignant lipomatous tumors. BACKGROUND OF THE INVENTION

Without limiting the scope of the invention, its background is described in connection with the identification and treatment of lipomas.

Benign and malignant lipomatous tumors are a common group of mesenchymal lesions characterized by an overgrowth of adipose cells. Lipomas are the most common benign form of soft tissue tumor. Malignant liposarcomas are the second most common soft tissue sarcoma (composing 18% of all soft tissue sarcomas) with five-year survival rates of 100% to 39% based on histological subtype. Well-differentiated and dedifferentiated liposarcomas are largely characterized by the presence of several massive abnormal chromosomes called neochromosomes, which carry dozens of copies of the oncogenes MDM2 and CDK4 (Pedeutou Genes Chromosomes Cancer 1994; Pilotti J Pathol 1998). Myxoid liposarcomas are malignant tumors displaying an immature adipogenic status, with aberrantly regulated ERK/MAPK and PI3K/AKT pathways and translocations resulting in a fusion in sarcoma-DNA-damage-inducible transcript 3 (FUS-DDIT3) fusion gene (Cheng Hum Pathol 2009; Rubin Histopathology 1997). Unlike other well-defined lipomatous tumors, a common molecular aberration does not underlie pleomorphic liposarcoma tumorigenesis, as these highly aggressive tumors harbor diverse chromosomal rearrangements and genomic profiles without unifying molecular alterations (Guillou Virchows Arch 2010). Lipomas, which account for nearly one-half of all benign lesions, display an abnormal karyotype with more than two-thirds of these aberrations involving recombination of chromosome segment 12ql3-15. The most common genomic rearrangement in lipomas is a chimeric gene fusion between the high mobility group protein gene HMGIC and the LIM protein gene LPP (Petit Genomics 1996). While cytogenetic studies are often effective at affirming the separate nature of the many variants of lipomatous tumors, histological characterization can sometimes lead to incorrect diagnosis. For instance, many benign lipomas display unusual features such as the presence of hibernomas or tumor pleomorphism, and may be confused with malignant liposarcomas (Weiss Monogr Pathol 1996). Many liposarcomas display transitional features of low-grade to high- grade lesions that vary not only by anatomical location but may show heterogeneity even within the tumor itself (Weiss Mongr Pathol 1996). For instance, many dedifferentiated liposarcoma tumors contain regions consisting of well-differentiated areas associated with non-lipogenic sarcoma that often resembles malignant fibrous histiocytomas or fibrosarcoma (Weiss Mongr Pathol 1996). Thus sensitive and specific biomarkers that can easily differentiate benign from malignant lipomatous tumors are needed.

SUMMARY OF THE INVENTION

In one embodiment, the present invention includes a method for diagnosis and treatment of a malignant liposarcoma comprising: obtaining a biological sample from a subject suspected of having a malignant liposarcoma; detecting a differential expression of at least one of Class II Major Histocompatibility Complex Trans activator (CIITA), Sex Determining Region Y-Box 30 (SOX30); ATP-Binding Cassette, Sub-Family B (MDR/TAP), Member 1 1 (ABCB 11); or Synuclein, gamma (breast cancer-specific protein 1) (SNCG) obtained from the biological sample, wherein the differential expression of the CIITA, SOX30, ABCB 11, or SNCG is a prognostic marker of disease progression and metastatic potential in tumors and can to distinguish between benign lipomas or early stage liposarcomas and well-differentiated, aggressive liposarcoma; and modifying the type and extent of immunotherapy treatment depending on the differentiated disease prognosis and metastatic potential of the well- differentiated, aggressive liposarcoma. In one aspect, the biological sample is a blood, a plasma or a tissue biopsy. In another aspect, the well-differentiated, aggressive liposarcoma. In another aspect, the subject is a mammal. In one aspect, the treatment for the lipoma is no treatment or surgical removal. In one aspect, the treatment the well differentiated liposarcoma treatments is radiotherapy, chemotherapy, or surgical removal. In another aspect, the subject is a mammal, e.g., a human. In another aspect, the expression of CIITA is measured by detecting protein expression, which is detected by at least one of immunohistochemically, enzyme linked immunosorbent assay, Western blot, or fluorescence activated cell sorting. In another aspect, the level of expression of CIITA, SOX30, ABCB1 1, or SNCG nucleic acids is measured by at least one of fluorescence detection, chemiluminescence detection, electrochemiluminescence detection and patterned arrays, reverse transcriptase-polymerase chain reaction, fluorescence activated sorting, detectable bead sorting, microarrays, allele specific primer extension, target specific primer extension, solid-phase binding arrays, liquid phase binding arrays, fluorescent resonance transfer, or radioactive labeling. In another aspect, the level of CIITA, SOX30, ABCBl l, or SNCG is measured at a first and a second timepoint, wherein an increase in the level of detection of CIITA, SOX30, or ABCB l l, or a decrease in SNCG levels between the first and second timepoint is indicative of increased liposarcoma aggressiveness. In another aspect, the method further comprises the step of detecting the level of expression of CIITA, SOX30, ABCBl l, or SNCG at two or more points and time, and if there is an increase in the expression of CIITA, SOX30, ABCB l l, or a decrease in SNCG levels initiating a more aggressive chemotherapeutic regimen against the liposarcoma, surgical removal of the liposarcoma or both. In another aspect, the immunotherapy is at least one of T-cell specific for malignant liposarcoma, an antibody specific for CIITA, SOX30, ABCB l l, or SNCG, an immunotoxin that is specific for CIITA, or a bivalent or multivalent antibody that targets a CIITA-MHC complex.

In another embodiment the present invention includes a method for treatment of a malignant liposarcoma comprising: detecting a differential expression of at least one of Class II Major Histocompatibility Complex Transactivator (CIITA), Sex Determining Region Y-Box 30 (SOX30); ATP-Binding Cassette, Sub-Family B (MDR/TAP), Member 1 1 (ABCB l l); or Synuclein, gamma (breast cancer-specific protein 1) (SNCG) expression obtained from the biological sample obtained from a subject suspected of having a liposarcoma or lipoma, wherein the differential expression of the CIITA, SOX30, ABCBl l, or SNCG is a prognostic marker of disease progression and metastatic potential and is used to distinguish between benign or early stage liposarcomas and well-differentiated, aggressive liposarcoma; and directing an immunotherapy against the well-differentiated, aggressive liposarcoma. In one aspect, the immunotherapy is at least one of T-cell specific for malignant lipomas, an antibody specific for CIITA, SOX30, ABCBl l, or SNCG, an immunotoxin that is specific for CIITA, SOX30, ABCBl l, or SNCG, or a bivalent or multivalent antibody that targets a CIITA-MHC complex. In another aspect, the biological sample is a blood, a plasma, or a tissue biopsy. In another aspect, the well-differentiated, aggressive. In another aspect, the subject is a mammal, e.g., a human. In another aspect, the level of expression of CIITA, SOX30, ABCBl l, or SNCG is detected at the mRNA or protein level. In another aspect, wherein the expression of CIITA, SOX30, ABCB l l, or SNCG is measured by detecting protein expression, which is detected by at least one of immunohistochemically, by enzyme linked immunosorbent assay, by Western blot, the fluorescence activated cell sorting. In another aspect, the level of expression of CIITA, SOX30, ABCB11, or SNCG nucleic acids is measured by at least one of fluorescence detection, chemiluminescence detection, electrochemiluminescence detection and patterned arrays, reverse transcriptase-polymerase chain reaction, fluorescence activated sorting, detectable bead sorting, microarrays, allele specific primer extension, target specific primer extension, solid-phase binding arrays, liquid phase binding arrays, fluorescent resonance transfer, or radioactive labeling. In another aspect, the increase in the level of detection of CIITA, SOX30, ABCB1 1, or a decrease in level SNCG is indicative increased aggressiveness of a liposarcoma. In another aspect, the method further comprises the step of detecting the level of expression of CIITA, SOX30, ABCB11, or SNCG at two or more points and time, and if there is an increase in the expression of CIITA, SOX30, ABCB1 1, or a decrease in SNCG expression initiating a more aggressive chemotherapeutic regimen against the liposarcoma, surgical removal of the liposarcoma or both.

In another embodiment, the present invention also includes a method of evaluating a candidate drug believed to be useful in treating malignant liposarcomas, the method comprising: (a) measuring the expression of detecting a differential expression of at least one of Class II Major Histocompatibility Complex Transactivator (CIITA), Sex Determining Region Y-Box 30 (SOX30); ATP-Binding Cassette, Sub-Family B (MDR/TAP), Member 1 1 (ABCB 11); or Synuclein, gamma (breast cancer-specific protein 1) (SNCG) from tissue or cells suspected of being a malignant liposarcoma from a first set of tissues of cells; (b) administering a candidate drug to a first subset of the tissue or cells, and a placebo to a second subset of normal or benign lipoma tissue or cells; (c) repeating step a) after the administration of the candidate drug or the placebo; and (d) determining if the candidate drug reduces the number of cells in the first set of cells that have an change in the expression of CIITA SOX30, ABCB1 1, or SNCG that is statistically significant as compared to any increase or reduction occurring in the second subset of cells, wherein a statistically significant reduction indicates that the candidate drug is useful in treating a well-differentiated malignant liposarcoma. In one aspect, the well-differentiated, aggressive liposarcoma is at least one of a dedifferentiated liposarcomas, myxoid, or has stromal infiltration. In another aspect, the expression of CIITA, SOX30, ABCB1 1, or SNCG is measured by detecting protein expression, which is detected by at least one of immunohistochemically, by enzyme linked immunosorbent assay, by Western blot, the fluorescence activated cell sorting. In another aspect, the level of expression of CIITA, SOX30, ABCB1 1, or SNCG nucleic acids is measured by at least one of fluorescence detection, chemiluminescence detection, electrochemiluminescence detection and patterned arrays, reverse transcriptase-polymerase chain reaction, fluorescence activated sorting, detectable bead sorting, microarrays, allele specific primer extension, target specific primer extension, solid-phase binding arrays, liquid phase binding arrays, fluorescent resonance transfer, or radioactive labeling.

In another embodiment, the present invention include a method for detecting a benign, an early stage liposarcomas, or a well-differentiated, aggressive liposarcoma cell comprising: obtaining a biological sample having a cell from a subject suspected of having the benign, the early stage liposarcomas, or the well-differentiated, aggressive liposarcoma cell; detecting a differential expression of at least one of Class II Major Histocompatibility Complex Trans activator (CIITA), Sex Determining Region Y-Box 30 (SOX30); ATP-Binding Cassette, Sub-Family B (MDR/TAP), Member 11(ABCB 11); or Synuclein, gamma (breast cancer-specific protein 1) (SNCG) obtained from the biological sample, wherein the differential expression of the CIITA, SOX30, ABCBl l, or SNCG is used to distinguish between the benign, the early stage liposarcomas, or the well-differentiated, aggressive liposarcoma; and determining if the cell is benign, early stage liposarcomas, or well-differentiated, aggressive liposarcoma based on an increase in the expression of CIITA, SOX30, ABCB l l, or a decrease in the expression of SNCG. In one aspect, the biological sample is a blood, a plasma or a tissue biopsy. In another aspect, the method distinguishes between a lipoma and the well-differentiated, aggressive liposarcoma cell. In another aspect, the subject is a mammal or a human. In another aspect, the expression of CIITA, SOX30, ABCB l l, or SNCG is measured by detecting protein expression, which is detected by at least one of immunohistochemically, enzyme linked immunosorbent assay, Western blot, or fluorescence activated cell sorting. In another aspect, the level of expression of CIITA, SOX30, ABCBl l, or SNCG nucleic acids is measured by at least one of fluorescence detection, chemiluminescence detection, electrochemiluminescence detection and patterned arrays, reverse transcriptase-polymerase chain reaction, fluorescence activated sorting, detectable bead sorting, microarrays, allele specific primer extension, target specific primer extension, solid-phase binding arrays, liquid phase binding arrays, fluorescent resonance transfer, or radioactive labeling. In another aspect, the level of CIITA, SOX30, ABCBl l, or SNCG is measured at a first and a second timepoint, wherein an increase in the level of detection of CIITA, SOX30, ABCB l l, or a decrease in SNCG levels between the first and second timepoint is indicative of increased liposarcoma aggressiveness. In another aspect, the method further comprises the step of detecting the level of expression of CIITA at two or more points and time, and if there is an increase in the expression of CIITA, SOX30, ABCB1 1, or a decrease in expression of SNCG initiating at least one of a more aggressive chemotherapeutic regimen against the liposarcoma, surgical removal of the liposarcoma, or both.

BRIEF DESCRIPTION OF THE DRAWINGS

For a more complete understanding of the features and advantages of the present invention, reference is now made to the detailed description of the invention along with the accompanying figures and in which:

Figure 1 is a graph that shows a meta-analysis of CIITA gene expression across benign and malignant lipomatous tumors. Meta-analysis of CIITA gene expression obtained from the Gene Expression Omnibus dataset GSE6481 by Nakayama et al (Mod Pathol 2007). Data displayed are the mean normalized CIITA gene expression values from lipoma (n=3), well-differentiated liposarcoma (n=3), myxoid liposarcoma (n=19), and dedifferentiated liposarcoma (n=15) plus or minus the standard error of the mean. Statistical significance was determined using Student's t-test (asterisk indicates p<0.05 relative to the lipoma values).

Figures 2A to 2F are micrographs that show immunohistochemical staining for CIITA protein expression in lipomatous tumors (100X magnification). CIITA immunostaining (CIITA staining is red) in formalin fixed paraffin embedded tissue from (Figure 2A) lipoma, (Figure 2B) well- differentiated liposarcoma, (Figure 2C) myxoid liposarcoma, (Figure 2D) pleomorphic liposarcoma, and (Figure 2E) dedifferentiated liposarcoma. (Figure 2F) Control staining without primary antibody. (lOOx total magnification).

Figures 3 A to 3 F are micrographs that show immunohistochemical staining for CIITA protein expression in lipomatous tumors (600X magnification). CIITA immunostaining (CIITA staining is red) in formalin fixed paraffin embedded tissue from (Figure 3A) lipoma, (Figure 3B) well- differentiated liposarcoma, (Figure 3C) myxoid liposarcoma, (Figure 3D) pleomorphic liposarcoma, and (Figure 3E) dedifferentiated liposarcoma. (Figure 3F) Control staining without primary antibody. (600x total magnification).

Figures 4A and 4B are micrographs that show immunohistochemical staining for CIITA protein expression in well-differentiated liposarcomas. CIITA immunostaining (CIITA staining is red) in formalin fixed paraffin embedded tissue from well-differentiated liposarcomas. Figure 4A is a representative image of a well-differentiated liposarcoma lacking immunoreactivity against CIITA. Dedifferentiation is minimal to absent in this image. Figure 4B is a representative image of a well-differentiated liposarcoma with positive immunoreactivity against CIITA in the regions of dedifferentiation (indicated with black arrows). Figure 4C is a graph that shows a meta-analysis of adipogenic gene expression obtained from the Gene Expression Omnibus dataset GSE6481 by Nakayama et al (Mod Pathol 2007). Data displayed in log¹⁰ scale are the mean normalized gene expression values from lipoma (n=3), well-differentiated liposarcoma (n=3), myxoid liposarcoma (n=19), and dedifferentiated liposarcoma (n=15). The values written above each gene symbol on the x-axis are the R values calculated using Pearson's correlation coefficient.

DETAILED DESCRIPTION OF THE INVENTION

While the making and using of various embodiments of the present invention are discussed in detail below, it should be appreciated that the present invention provides many applicable inventive concepts that can be embodied in a wide variety of specific contexts. The specific embodiments discussed herein are merely illustrative of specific ways to make and use the invention and do not delimit the scope of the invention.

To facilitate the understanding of this invention, a number of terms are defined below. Terms defined herein have meanings as commonly understood by a person of ordinary skill in the areas relevant to the present invention. Terms such as "a", "an" and "the" are not intended to refer to only a singular entity, but include the general class of which a specific example may be used for illustration. The terminology herein is used to describe specific embodiments of the invention, but their usage does not delimit the invention, except as outlined in the claims.

Lipomatous tumors, including benign lipomas and malignant liposarcomas, are a diverse group of mesenchymal tumors characterized by an overgrowth of adipose cells. Lipomas are highly prevalent in humans (1% of the general population) and are the most common benign form of soft tissue tumor. In contrast, liposarcomas are relatively rare (approximately 2.5 cases per million people), but are still the second most common of all soft tissue sarcomas with histological classifications including well-differentiated liposarcomas, myxoid/round cell liposarcomas, pleomorphic liposarcomas, and de-differentiated liposarcomas. These malignant tumors exhibit five-year survival rates as low as 39% depending on the particular histological subtype.

Many molecular and clinical similarities exist between lipomas and liposarcomas (Dei Tos Ann Diagn Pathol 2000), thus a diagnostic dilemma sometimes exists in differentiating these tumor types. When comparing lipomas and low grade liposarcomas, patients are misdiagnosed approximately 30-40% of the time following radiological detection (O'Donnell Sarcoma 2013; Gaskin Am J Roentgenology 2004) and in 7- 17% of histological evaluations (Hasegawa Human Pathology 2002). Even fluorescent in situ hybridization (FISH) for MDM2-CDK4 amplification (considered the gold standard and relatively costly diagnostic marker distinguishing lipomas from low grade and de-differentiated liposarcomas) is inaccurate and/or provides uninterpretable results in 10% of cases (Sirvent Am J Surg Pathol 2007; Shimada Hum Pathol 2006). Diagnostic accuracy is further complicated due to morphological heterogeneity within particular lipomatous tumors. For instance, benign pleomorphic lipomas display unusual features such as the presence of hibernomas or tumor pleomorphism, and may be confused with malignant liposarcomas (Weiss 1996). Additionally, many liposarcomas display transitional features of low to high-grade lesions or consist of well differentiated areas associated with non-lipogenic sarcoma often resembling malignant fibrous histiocytomas or fibrosarcoma (Weiss 1996).

Accurate, timely, and cost effective diagnosis of lipomatous tumors is essential for proper patient treatment and increased long term survival. A combination of bioinformatic and immunohistochemistry techniques were used herein to accurately and sensitively identify biomarkers that effectively distinguish benign from malignant lipomatous tumors.

As used herein the following human genes and proteins are discussed and used: (1) CIITA-Class II, Major Histocompatibility Complex, Transactivator; (2) SOX30-SRY (Sex Determining Region Y)-Box 30; (3) ABCB 11 -ATP-Binding Cassette, Sub-Family B (MDR/TAP), Member 1 1; and (4) SNCG-Synuclein, gamma (breast cancer-specific protein 1).

Immune cell recognition of MHC class II histocompatibility complexes has been shown to directly mediate cytotoxicity against tumor cells. CIITA, encoded by the C2TA gene on chromosome 16p 13 , is a positive regulator of class II major histocompatibility complex gene transcription and is referred to as the "master control factor" for the expression of these genes. Prompted by a metaanalysis, the present inventors show that CIITA is differentially expressed between benign lipomas and metastatic liposarcomas. Thus, the use of CIITA as an immunohistochemical marker differentiating benign from malignant lipomatous tumors was determined. Formalin-fixed, paraffin embedded tumor sections were obtained for the following lipomatous tumors (n=52) lipomas (n=ll), well-differentiated liposarcomas (n=20), myxoid liposarcomas (n=15), pleomorphic liposarcomas (n=3), and dedifferentiated liposarcomas (n=3). Utilizing standard immunohistochemistry protocols, these sections were stained with an anti-CIITA antibody and immunopositivity was assessed based on signal intensity. CIITA immunoreactivity was observed in all cases of myxoid (15/15), pleiomorphic (3/3), and dedifferentiated (3/3) liposarcomas and in dedifferentiated regions in the large majority of well- differentiated liposarcomas (15/20). CIITA staining was absent in all benign lipomas tested. It is demonstrated herein that histological diversity of lipomatous tumors correlates with the expression of the positive regulator of class II major histocompatibility complex gene transcription, CIITA.

Meta-analysis of CIITA expression. The normalized intensity values of CIITA, TNFRSF9, GLUT4, ZIC1, SLC27A8, CIDEA, and LEP were evaluated from the Gene Expression Omnibus reference series GSE6481, deposited by Nakayama et al (Mod Pathol 2007). This dataset compared the global gene expression profiles of various soft tissue sarcomas including lipomas (n=3), well-differentiated liposarcomas (n=3), myxoid liposarcomas (n=19), and dedifferentiated liposarcomas (n=15). Data is represented as the mean intensity values from each tumor type plus or minus the standard error of the mean.

Case Material. Tissue arrays of formalin-fixed, paraffin embedded lipomatous tumor blocks were obtained from Super Bio Chips (www.tissue-array.com). These clinically characterized tumor samples consisted of 2 mm diameter cores with a section thickness of 4 microns and were composed of lipomas (n=ll), well-differentiated liposarcomas (n=20), myxoid liposarcomas (n=15), pleomorphic liposarcomas (n=3), and dedifferentiated liposarcomas (n=3). The cases were reviewed by a pathologist and the diagnoses were confirmed by histomorphology per established morphological criteria (Weiss, Goldblum, Liposarcoma— Weiss's Soft Tissue Tumors 5^th ed, 2008).

Immunohistochemistry. Sections were deparaffinized, rehydrated, and treated for antigen retrieval using Trilogy (Cell Marque; Cat. No. 920P-10). To block nonspecific binding, sections were incubated in Background Block solution (Cell Marque; Cat. No. 927B-05) at room temperature for 10 minutes before application of the anti-CIITA H-300 primary antibody (sc- 48797; Santa Cruz Biotechnology). Sections were then washed in phosphate buffered saline immunohistochemistry wash buffer with tween 20 (Cell Marque; Cat. No. 934B-09) three times for 5 minutes each and then incubated with the CytoScan Alkaline Phos Detection System (Cell Marque; Cat. No. 952D-20). The immunostaining was carried out using Permanent Red Chromogen detection kit (Cell Marque; Cat. No. 960D-20) and counterstained with Hematoxylin.

Quantitation of and Statistical Analysis of Immunohistochemistry. Immunopositivity was scored semi-quantitatively for the percentage of tumor cells staining and intensity (0=negative, + = weak, ++ = moderate, +++ = strong). For statistical analysis, scoring was converted to numerical values (0 = 0, + = 1; ++ = 2; +++ = 3) and mean values plus or minus the standard error of the mean were calculated using two tailed t-tests. Pearson's correlation test and Fisher's exact test were applied to assess the correlation between immunopositivity and lipomatous tumor malignancy. Significance thresholds for all statistical analysis were set at p<0.05.

Meta-analysis of lipomatous tumor genomic data.

Nakayama et al (Mod Pathol 2007) previously used global gene expression profiling to compare the transcriptomes of various soft tissue sarcomas, and included in these datasets were expression 40 profiles from benign and malignant lipomatous including lipomas and well differentiated, de-differentiated, and myxoid liposarcoma tumors. This data was publically deposited in the Gene Expression Omnibus (www.ncbi.nlm.nih.gov/geo/), reference GSE6481. In order to develop a list of genes whose expression was differentially expressed at statistically significant levels between benign and malignant lipomatous tumors, the inventors used metaanalysis strictly on the lipomatous tumor data obtained from this dataset to identify 3684 genes (2800 genes upregulated and 884 genes downregulated in malignant verses benign lipomatous tumors) whose mRNA expression levels were differentially expressed at statistically significant levels (2 fold or greater fold change, p<0.05) between benign lipomas and malignant liposarcomas (data not shown). Systems level analysis using Metacore signaling network software identified the process networks differing between benign and malignant lipomatous tumors with the highest statistical significance and included genes involved in muscle development and contraction (206 genes), neurogenesis (182 genes), and cytoskeletal regulation (64 genes) (data not shown).

Though the number of candidate genes was overwhelming, a confirmatory analysis was used on a subset of four putative genes including CIITA, SOX30, ABCB 11, and SNCG whose expression differences in the malignant verses the benign lipomatous tumors based on the present inventors' meta-analysis of the Nakayama et al data. The expression levels of CIITA, SOX30, and ABCB1 1 were significantly increased in malignant compared to benign lipomatous tumors. In contrast, SNCG was significantly decreased in malignant compared to benign lipomatous tumors. As malignant lipomatous tumors have been shown to display characteristics of de-differentiation relative to lipomas (Riester Plos Comput Biol 2010), a Pearson's correlation analysis was used to measure the strength of the linear relationship between the expression levels of CIITA, SOX30, ABCB1 1, and SNCG against the expression levels of four genes whose expression has been previously reported to be tightly correlated to the degree of lipogenic differentiation including SLC27A5, GLUT4, LEP, and ZIC1 (Figure 1). Strong correlations between the expression of the candidate genes and the expression of known lipogenic genes would increase our confidence that these genes could effectively serve as markers that differentiate benign from malignant lipomatous tumors. Correlation coefficients above 0.40 or below -0.40 were considered strong positive or negative relationships, respectively. Strong correlations were observed between CIITA and all four established lipogenic markers, while moderate to strong correlations were observed between SOX30, ABCB1 1, and SNCG for three of the four established lipogenic markers. These data show that the steady state mRNA expression levels of CIITA, SOX30, ABCB 11, and SNCG are correlated to adipogenic gene transcription signatures and may serve as effective markers differentiating benign from malignant lipomatous tumors.

Independent confirmation of lipomatous tumor mRNA expression at the protein level.

While the data provided by Nakayama et al (Mod Pathol 2007) was useful in forming a hypothesis regarding the use of these four putative markers of malignancy in lipomatous tumors, we sought to corroborate our data using an independent panel of clinically defined lipomatous tumors at the steady state protein level. To accomplish this, the inventors examined the expression of CIITA, SOX30, ABCB1 1, and SNCG proteins on a panel of 1 1 benign lipomas and 41 liposarcomas (including well differentiated, de-differentiated, myxoid, and pleiomorphic liposarcomas) using immunohistochemistry. Immunopositivity of these proteins across the lipomatous tumor panel is shown in Table 1 and Table 2, with staining for CIITA, SOX30, ABCB1 1, and SNCG protein detected in 0%, 36%, 0%, and 64% of lipomas, and 90%, 71%, 71%, and 100% of malignant tumors, respectively. Additionally, it was very clear that intensity of staining increased in the malignant liposarcomas compared to the benign lipomas. This data clearly matched our genomics meta-analysis for each protein with the exception of SNCG expression. Interestingly, the immunohistochemical analysis used herein revealed that SNCG protein expression was inversely proportional to its mRNA expression determined from our meta-analysis, suggesting some level of post-translational regulation may be involved for this gene in these tumor types.

A statistical analysis was used to determine if either the presence of staining or the intensity of staining for each of these tested proteins could be used as biomarkers to distinguish benign from malignant lipomatous tumors. The inventors sought to determine if simply the presence (not intensity) of CIITA, SOX30, ABCB 11, or SNCG protein could statistically distinguish benign from malignant lipomatous tumors. To accomplish this, the lipomatous tumors were rated using a binary system as either immuno-negative or immune-positive based on antigen detection and we employed Fisher's exact test to generate contingency tables for examination of the significance of the association between antigen expression and clinical diagnosis (benign vs malignant). With a significance cut-off at p<0.05, the Fisher exact test statistic values of CIITA and ABCB1 1 in benign verses malignant tumors were virtually zero, suggesting that expression of these two antigens could be statistically useful in differentiating these tumor types (Table 3). Using this same statistical test, differential expression of SOX30 and SNCG based solely on immunopositivity, but not staining intensity levels, was not at significant levels to differentiate benign from malignant lipomatous tumors (Table 3).

Next, it was determined if staining intensity for each of these tested proteins could be used as biomarkers to distinguish benign from malignant lipomatous tumors. As opposed to a binomial rating system as used above, staining intensity ratings (0, +, ++, or +++) were converted directly to a numerical system (0, 1, 2, or 3). The mean plus or minus the standard error of the mean was calculated for benign and malignant lipomatous tumors (Table 4), revealing that the level of staining intensity for CIITA, SOX30, ABCB 11, and SNCG was significantly increased in malignant liposarcomas compared to benign lipomas. This indicates that staining intensity levels for each of the proteins were indicative of benign verses malignant liposarcomas.

Table 1. Immunopositivity of lipomatous tumors for each protein.

Table 2. Rating of antigen staining by tumor type and clinical characteristics.

Age Gender Location Diagnosis SubTissue CIITA SOX3 ABCB1 SNCG (yrs) type Type Score 0 1 Score

Score Score

37 F Neck Lip B 0 0 0 0

56 M Back Lip B 0 + 0 + F Back Lip B 0 + 0 0

F Shoulder Lip B 0 0 0 0

F Neck Lip B 0 0 0 +

M Breast Lip B 0 0 0 +

F Shoulder Lip B 0 + 0 +

M Axilla Lip B 0 0 0 +

F Thigh Lip B 0 0 0 0

F Breast Lip B 0 + 0 ++

F Scalp Lip B 0 0 0 +

M etroperitoneum LS WD M 0 + 0 +

F Shoulder LS WD M 0 0 0 +

F Thigh LS WD M 0 0 + +

F Axilla LS WD M 0 0 0 +

F Thigh LS WD M + 0 0 +

F Thigh LS WD M ++ +++ ++ ++

M Thigh LS WD M + + + ++

F Retroperitoneum LS WD M + + + +

F Axilla LS WD M + + + ++

F Retroperitoneal LS WD M + 0 0 +

cavity

M Neck LS WD M + 0 0 ++

M Calf LS WD M + 0 0 ++

M Retroperitoneum LS WD M + 0 0 +

F Axilla LS WD M + 0 0 +

F Buttock LS WD M + 0 0 +

F Mesentery LS WD M ++ ++ ++ ++

F Mediastinum LS WD M + + + +

F Retroperitoneum LS WD M ++ ++ ++ ++

M Thigh LS WD M + + + +

M Abdominal cavity LS WD M + 0 0 +

M Abdomen LS Myx M ++ ++ ++ ++

F Retroperitoneum LS Myx M + ++ ++ ++

F Retroperitoneum LS Myx M + + + +

F Perirenal LS Myx M ++ + ++ +++

M Intraabdominal LS Myx M + + + +

M Forearm LS Myx M ++ +++ ++ ++

F Pelvic cavity LS Myx M ++ ++ ++ +

M Retroperitoneum LS Myx M + ++ +++ +++

M Axilla LS Myx M + ++ ++ ++ 78 M Thigh LS Myx M +^■ 4-4- 4- +

64 M Abdominal cavity LS Myx M +^■ 4-4-4- 4-4- 4-4-

45 M Abdomen LS Myx M 4- +

74 F Thigh LS Myx M +^■ 4-4- 4-4- 4-4-4-

43 M Thigh LS Myx M +^■ +4- 4-4-4- 4-4- 4-4-

41 M Peritoneal cavity LS Myx M 4 -4- 0 0 +

59 M Chest LS PM M 4 -4- 4-4- 4-4-4- 4-4-

66 F Abdominal cavity LS PM M 4 -4- 4-4- 4- +

70 F Abdominal cavity LS PM M 4-4- 4-4-

53 M etroperitoneum LS DD M +^■ +4- 4-4- 4-4- +

61 F Abdominal cavity LS DD M 4- 4-4-

61 M Abdominal cavity LS DD M 4- 4-4-

F = female; M = male

Lip = lipoma; LS = liposarcoma

WD = well differentiated; Myx = myxoid, PM = pleomorphic; DD = dedifferentiated

B = benign; M = malignant

0 = no detectable staining; 4- = weak staining; 4-4- = moderate staining; 4-4-4- = strong staining

Statistical analysis of immunopositivity between benign and malignant lipomati

Protein Fisher exact test value Significant

CIITA 0.00 Yes

SOX30 0.07 No

ABCB1 1 0.00 Yes

SNCG 0.10 No

Table 4. Statistical analysis of antigen staining intensity between benign and malignant lipomatous tumors.

The variability of CIITA staining in the well-differentiated liposarcomas where a portion of these particular tumors displayed no immunoreactivity against this antigen was determined. While myxoid and pleomorphic liposarcomas are highly dedifferentiated tumors, well- differentiated liposarcomas often express a time-dependent gradient of differentiation. For instance, pathologists refer to the most pure form of well-differentiated liposarcoma as an atypical lipomatous tumor, indicating that these low-grade tumors do not metastasize and reflect a highly adipocytic phenotype. Over time, these low-grade well-differentiated tumors progressively dedifferentiate and change to dedifferentiated liposarcomas that exhibit increased mortality rates in patients. As illustrated in Figure 4A, a visual comparison of the negative verses positive stained well-differentiated liposarcomas very clearly revealed that the level of CIITA expression was in proportion to the amount of tumor cell dedifferentiation present in the tissue. Well-differentiated liposarcomas that would classified as very low-grade atypical lipomatous tumors lacked immunoreactivity against CIITA, however progressively dedifferentiating tumors were positive for this protein. The inventors calculated the Pearson's correlation coefficient between CIITA gene expression and known markers of adipogenic lineage/maturity using the microarray data previously deposited in the Gene Expression Omnibus by Nakayama et al (Mod Pathol 2007). The inventors' analysis revealed moderate to strong direct and indirect correlations between the expression of CIITA and several known adipogenic genes (Figure 4B), indeed suggesting that CIITA expression is directly tied to adipogenic gene transcription signatures.

These data demonstrate that CIITA is expressed in the large majority of less aggressive well- differentiated liposarcomas and all aggressive liposarcomas tested. The expression of this protein was undetectable in all benign lipomas and several well-differentiated liposarcomas, with CIITA expression indirectly correlating to the degree of differentiation in the tumor. Given that the risk of distant liposarcoma metastasis as well as patient survival relate to the degree of histological differentiation in the tumor, CIITA may serve as an excellent molecular marker correlating to the degree of tumor dedifferentiation and effectively distinguishing lipomas and less aggressive well-differentiated liposarcomas from the more aggressive lipomatous tumors.

In addition to identification of CIITA as a differentially expressed biomarker, this study provides support for the potential use of immunotherapy against malignant liposarcomas that highly express CIITA. CD4+ T cell recognition of MHC class II histocompatibility complexes, which are induced by CIITA activity, have been shown to directly mediate cytotoxicity against tumor cells (Quezada and Peggs Immunotherapy 201 1), and impairments in the presentation or recognition of MHC class II tumor associated antigens enhances the evasion of tumor cells from immunosurveillance. Indeed, several reports have linked defective CIITA activity to the regulation of tumorigenesis. CIITA is a recurrent gene fusion partner in lymphoid cancers, occurring in 38% of primary mediastinal B-cell lymphomas and 15% of classic Hodgkin lymphomas, leading to downregulation of surface HLA class II expression that may impact antitumor immune responses in cancer (Steidl Nature 2011). Methylation and subsequent silencing of CIITA allows some gastrointestinal cancer cells to escape immune surveillance (Satoh Oncogene 2004). Overexpression of the CIITA gene in melanoma cells produces exosomes exhibiting enhanced anti-tumor effects in part due to induction of higher amounts of Thl- polarized immune responses such as Thl-type IgG2a antibodies and IFN-gamma cytokines and TRP2-specific CD8+ T cells (Lee Exp Mol Med 2011). If enhanced CIITA expression in liposarcoma cells directly translates to enhanced presentation of immunogenic MHC class II epitopes, then malignant liposarcomas can be targeted for immunotherapy.

It is contemplated that any embodiment discussed in this specification can be implemented with respect to any method, kit, reagent, or composition of the invention, and vice versa. Furthermore, compositions of the invention can be used to achieve methods of the invention.

It will be understood that particular embodiments described herein are shown by way of illustration and not as limitations of the invention. The principal features of this invention can be employed in various embodiments without departing from the scope of the invention. Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, numerous equivalents to the specific procedures described herein. Such equivalents are considered to be within the scope of this invention and are covered by the claims.

All publications and patent applications mentioned in the specification are indicative of the level of skill of those skilled in the art to which this invention pertains. All publications and patent applications are herein incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference.

The use of the word "a" or "an" when used in conjunction with the term "comprising" in the claims and/or the specification may mean "one," but it is also consistent with the meaning of "one or more," "at least one," and "one or more than one." The use of the term "or" in the claims is used to mean "and/or" unless explicitly indicated to refer to alternatives only or the alternatives are mutually exclusive, although the disclosure supports a definition that refers to only alternatives and "and/or." Throughout this application, the term "about" is used to indicate that a value includes the inherent variation of error for the device, the method being employed to determine the value, or the variation that exists among the study subjects. As used in this specification and claim(s), the words "comprising" (and any form of comprising, such as "comprise" and "comprises"), "having" (and any form of having, such as "have" and "has"), "including" (and any form of including, such as "includes" and "include") or "containing" (and any form of containing, such as "contains" and "contain") are inclusive or open-ended and do not exclude additional, unrecited elements or method steps. In embodiments of any of the compositions and methods provided herein, "comprising" may be replaced with "consisting essentially of or "consisting of. As used herein, the phrase "consisting essentially of requires the specified integer(s) or steps as well as those that do not materially affect the character or function of the claimed invention. As used herein, the term "consisting" is used to indicate the presence of the recited integer (e.g., a feature, an element, a characteristic, a property, a method/process step or a limitation) or group of integers (e.g., feature(s), element(s), characteristic(s), propertie(s), method/process steps or limitation(s)) only.

The term "or combinations thereof as used herein refers to all permutations and combinations of the listed items preceding the term. For example, "A, B, C, or combinations thereof is intended to include at least one of: A, B, C, AB, AC, BC, or ABC, and if order is important in a particular context, also BA, CA, CB, CBA, BCA, ACB, BAC, or CAB. Continuing with this example, expressly included are combinations that contain repeats of one or more item or term, such as BB, AAA, AB, BBC, AAABCCCC, CBBAAA, CABABB, and so forth. The skilled artisan will understand that typically there is no limit on the number of items or terms in any combination, unless otherwise apparent from the context.

As used herein, words of approximation such as, without limitation, "about", "substantial" or "substantially" refers to a condition that when so modified is understood to not necessarily be absolute or perfect but would be considered close enough to those of ordinary skill in the art to warrant designating the condition as being present. The extent to which the description may vary will depend on how great a change can be instituted and still have one of ordinary skilled in the art recognize the modified feature as still having the required characteristics and capabilities of the unmodified feature. In general, but subject to the preceding discussion, a numerical value herein that is modified by a word of approximation such as "about" may vary from the stated value by at least ±1, 2, 3, 4, 5, 6, 7, 10, 12 or 15%.

All of the compositions and/or methods disclosed and claimed herein can be made and executed without undue experimentation in light of the present disclosure. While the compositions and methods of this invention have been described in terms of preferred embodiments, it will be apparent to those of skill in the art that variations may be applied to the compositions and/or methods and in the steps or in the sequence of steps of the method described herein without departing from the concept, spirit and scope of the invention. All such similar substitutes and modifications apparent to those skilled in the art are deemed to be within the spirit, scope and concept of the invention as defined by the appended claims. REFERENCES

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Binh MB, Sastre-Garau X, Guillou L, et al. MDM2 and CDK4 immunostainings are useful adjuncts in diagnosing well-differentiated and dedifferentiated liposarcoma subtypes: a comparative analysis of 559 soft tissue neoplasms with genetic data. Am J Surg Pathol 2005;29(10): 1340-7 doi: 00000478-200510000-0001 1 [published Online First: Epub Date]|.

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Claims

What is claimed is:

1. A method for diagnosis and treatment of a malignant liposarcoma comprising:

obtaining a biological sample from a subject suspected of having a malignant liposarcoma;

detecting a differential expression of at least one of Class II Major Histocompatibility

Complex Trans activator (CIITA), Sex Determining Region Y-Box 30 (SOX30); ATP-Binding Cassette, Sub-Family B (MDR/TAP), Member 1 1(ABCB1 1); or Synuclein, gamma (breast cancer-specific protein 1) (SNCG) obtained from the biological sample, wherein the differential expression of the CIITA SOX30, ABCB 11, or SNCG is a prognostic marker of disease progression and metastatic potential in tumors and can to distinguish between benign or early stage liposarcomas and well-differentiated, aggressive liposarcoma; and

modifying the type and extent of immunotherapy treatment depending on the differentiated disease prognosis and metastatic potential of the well-differentiated, aggressive liposarcoma.

2. The method of claim 1, wherein the biological sample is a blood, a plasma or a tissue biopsy.

3. The method of claim 1 , wherein the treatment for the lipoma is no treatment or surgical removal.

4. The method of claim 1, wherein the treatment the well differentiated liposarcoma treatments is radiotherapy, chemotherapy, or surgical removal.

5. The method of claim 1, wherein the method distinguishes between lipomas and well- differentiated, aggressive liposarcoma .

6. The method of claim 1, wherein the subject is a mammal.

7. The method of claim 1, wherein the subject is a human.

8. The method of claim 1, wherein the expression of CIITA, SOX30, ABCB1 1, or SNCG is measured by detecting protein expression, which is detected by at least one of

immunohistochemically, enzyme linked immunosorbent assay, Western blot, or fluorescence activated cell sorting.

9. The method of claim 1, wherein the level of expression of CIITA, SOX30, ABCB 11, or SNCG nucleic acids is measured by at least one of fluorescence detection, chemiluminescence detection, electrochemiluminescence detection and patterned arrays, reverse transcriptase- polymerase chain reaction, fluorescence activated sorting, detectable bead sorting, microarrays, allele specific primer extension, target specific primer extension, solid-phase binding arrays, liquid phase binding arrays, fluorescent resonance transfer, or radioactive labeling.

10. The method of claim 1, wherein the level of CIITA, SOX30, ABCB1 1, or SNCG is measured at a first and a second timepoint, wherein an increase in the level of detection of

CIITA, SOX30, ABCB11, or a decrease in SNCG levels between the first and second timepoint is indicative of increased liposarcoma aggressiveness.

1 1. The method of claim 1, further comprising the step of detecting the level of expression of CIITA, SOX30, ABCB 11, SNCG at two or more points and time, and if there is an increase in the expression of CIITA, SOX30, ABCB1 1, or a decrease in the expression of SNCG initiating at least one of a more aggressive chemotherapeutic regimen against the liposarcoma, surgical removal of the liposarcoma, or both.

12. The method of claim 1, wherein the immunotherapy is at least one of T-cell specific for malignant liposarcoma, an antibody specific for CIITA, SOX30, ABCB1 1, or SNCG, an immunotoxin that is specific for CIITA, SOX30, ABCB 11, or SNCG or a bivalent or multivalent antibody that targets a CIITA-MHC complex.

13. A method for treatment of a malignant liposarcoma comprising:

detecting a differential expression of at least one of Class II Major Histocompatibility Complex Trans activator (CIITA), Sex Determining Region Y-Box 30 (SOX30); ATP-Binding Cassette, Sub-Family B (MDR/TAP), Member 1 1(ABCB1 1); or Synuclein, gamma (breast cancer-specific protein 1) (SNCG) expression obtained from the biological sample obtained from a subject suspected of having a liposarcoma or lipoma, wherein the differential expression of the CIITA, SOX30, ABCB 11, or SNCG is a prognostic marker of disease progression and metastatic potential and is used to distinguish between benign or early stage liposarcomas and well-differentiated, aggressive liposarcoma; and

directing an immunotherapy against the well-differentiated, aggressive liposarcoma.

14. The method of claim 13, wherein the immunotherapy is at least one of T-cell specific for malignant lipomas, an antibody specific for CIITA, SOX30, ABCB1 1, or SNCG, an

immunotoxin that is specific for CIITA, SOX30, ABCB1 1, or SNCG or a bivalent or multivalent antibody that targets a CIITA-MHC complex.

15. The method of claim 13, wherein the biological sample is a blood, a plasma, or a tissue biopsy.

16. The method of claim 13, wherein the method distinguishes between lipomas and well- differentiated, aggressive liposarcoma.

17. The method of claim 13, wherein the subject is a mammal.

18. The method of claim 13, wherein the subject is a human.

19. The method of claim 13, wherein the level of expression of CIITA, SOX30, ABCB1 1, or SNCG is detected at the mRNA or protein level.

20. The method of claim 13, wherein the expression of CIITA, SOX30, ABCB 11 , or SNCG is measured by detecting protein expression, which is detected by at least one of

21. The method of claim 13, wherein the level of expression of CIITA, SOX30, ABCB1 1, or SNCG nucleic acids is measured by at least one of fluorescence detection, chemiluminescence detection, electrochemiluminescence detection and patterned arrays, reverse transcriptase- polymerase chain reaction, fluorescence activated sorting, detectable bead sorting, microarrays, allele specific primer extension, target specific primer extension, solid-phase binding arrays, liquid phase binding arrays, fluorescent resonance transfer, or radioactive labeling.

22. The method of claim 13, wherein an increase in the level of detection of CIITA, SOX30, ABCB 1 1, or a decrease in the level of expression of SNCG is indicative increased

aggressiveness of a liposarcoma.

23. The method of claim 13, further comprising the step of detecting the level of expression of CIITA at two or more points and time, and if there is an increase in the expression of CIITA, SOX30, ABCB 1 1, or a decrease in the expression of SNCG initiating at least one of a more aggressive chemotherapeutic regimen against the liposarcoma, surgical removal of the liposarcoma, or both.

24. A method of evaluating a candidate drug believed to be useful in treating malignant liposarcomas, the method comprising:

(a) measuring the expression of at least one of Class II Major Histocompatibility Complex Transactivator (CIITA), Sex Determining Region Y-Box 30 (SOX30); ATP -Binding Cassette, Sub-Family B (MDR/TAP), Member 1 1 (ABCB 1 1); or Synuclein, gamma (breast cancer-specific protein 1) (SNCG) from tissue or cells suspected of being a malignant liposarcoma from a first set of tissues of cells; (b) administering a candidate drug to a first subset of the tissue or cells, and a placebo to a second subset of normal or benign lipoma tissue or cells;

(c) repeating step a) after the administration of the candidate drug or the placebo; and

(d) determining if the candidate drug reduces the number of cells in the first set of cells that have an change in the expression of the CIITA, SOX30, ABCB1 1, or SNCG that is statistically significant as compared to any reduction occurring in the second subset of cells, wherein a statistically significant change indicates that the candidate drug is useful in treating a well-differentiated malignant liposarcoma.

25. The method of claim 24, wherein the method distinguishes between lipomas and well- differentiated, aggressive liposarcomas .

26. The method of claim 24, wherein the expression of CIITA, SOX30, ABCB 11 , or SNCG is measured by detecting protein expression, which is detected by at least one of

27. The method of claim 24, wherein the level of expression of CIITA, SOX30, ABCB 1 1, or SNCG nucleic acids is measured by at least one of fluorescence detection, chemiluminescence detection, electrochemiluminescence detection and patterned arrays, reverse transcriptase- polymerase chain reaction, fluorescence activated sorting, detectable bead sorting, microarrays, allele specific primer extension, target specific primer extension, solid-phase binding arrays, liquid phase binding arrays, fluorescent resonance transfer, or radioactive labeling.

28. A method for detecting a benign, an early stage liposarcomas, or a well-differentiated, aggressive liposarcoma cell comprising:

obtaining a biological sample having a cell from a subject suspected of having the benign, the early stage liposarcomas, or the well-differentiated, aggressive liposarcoma cell; detecting a differential expression of at least one of Class II Major Histocompatibility

Complex Trans activator (CIITA), Sex Determining Region Y-Box 30 (SOX30); ATP-Binding Cassette, Sub-Family B (MDR/TAP), Member 1 1 (ABCB 1 1); or Synuclein, gamma (breast cancer-specific protein 1) (SNCG) obtained from the biological sample, wherein the differential expression of the CIITA, SOX30, ABCB 1 1, or SNCG is used to distinguish between the benign, the early stage liposarcomas, or the well-differentiated, aggressive liposarcoma; and determining if the cell is benign, early stage liposarcomas, or well-differentiated, aggressive liposarcoma based on an increase in the expression of CIITA, SOX30, ABCBl 1, or a decrease in the expression of SNCG.

29. The method of claim 28, wherein the biological sample is a blood, a plasma or a tissue biopsy.

30. The method of claim 28, wherein the method distinguishes between a lipoma and the well-differentiated, aggressive liposarcoma cell.

31. The method of claim 28, wherein the subject is a mammal or a human.

32. The method of claim 28, wherein the expression of CIITA, SOX30, ABCB l 1, or SNCG is measured by detecting protein expression, which is detected by at least one of

33. The method of claim 28, wherein the level of expression of CIITA, SOX30, ABCBl 1, or SNCG nucleic acids is measured by at least one of fluorescence detection, chemiluminescence detection, electrochemiluminescence detection and patterned arrays, reverse transcriptase- polymerase chain reaction, fluorescence activated sorting, detectable bead sorting, microarrays, allele specific primer extension, target specific primer extension, solid-phase binding arrays, liquid phase binding arrays, fluorescent resonance transfer, or radioactive labeling.

34. The method of claim 28, wherein the level of CIITA, SOX30, ABCBl 1, or SNCG is measured at a first and a second timepoint, wherein an increase in the level of detection of

CIITA, SOX30, ABCBl 1, or a decrease in SNCG levels between the first and second timepoint is indicative of increased liposarcoma aggressiveness.

35. The method of claim 28, further comprising the step of detecting the level of expression of CIITA at two or more points and time, and if there is an increase in the expression of CIITA, SOX30, ABCBl 1, or a decrease in expression of SNCG initiating at least one of a more aggressive chemotherapeutic regimen against the liposarcoma, surgical removal of the liposarcoma, or both.