Patent Publication Number: US-2019167628-A1

Title: Methods for treating sarcoma

Description:
CROSS REFERENCE TO RELATED APPLICATIONS 
     This application claims priority under 35 U.S.C. § 119(e) from U.S. Provisional Patent Application Ser. No. 62/593,670, filed Dec. 1, 2017, the entire contents of which are hereby incorporated by reference. 
    
    
     FIELD OF THE INVENTION 
     Improved methods for treating sarcoma are provided. 
     BACKGROUND 
     Sarcomas are a heterogeneous group of connective tissue cancers. Examples of sarcomas include osteosarcoma, Ewing&#39;s sarcoma, leiomyosarcoma, liposarcoma, and rhabdomyosarcoma. Anthracyclines are often the first line of treatment for the various sarcomas, but gemcitabine, taxane and other agents have shown activity. Treatment regimens for sarcoma have not to date used mechanism-based insights where treatment has been informed by molecular markers of response to tumor therapy. The methods described below describe measurements of molecular markers of response to various chemotherapies in tumor samples of 5 sarcoma subtypes. The methods were used to identify effective treatment regimens based on expression of chemotherapy biomarkers. 
     SUMMARY OF THE INVENTION 
     What are provided are methods of treating leiomyosarcoma, which include treating a patient suffering from leiomyosarcoma with a regimen that includes an effective amount of a tubulin inhibitor and/or an effective amount of a platinum-based agent. The tubulin inhibitor may be a taxane. The platinum-based agent may be cisplatin or carboplatin. In these methods the regimen may include an effective amount of a tubulin inhibitor and an effective amount of a platinum-based agent. 
     Further provided are methods of treating liposarcoma, which include treating a patient suffering from liposarcoma with a regimen that includes an effective amount of a pyrimidine antagonist and/or an effective amount of a topoisomerase inhibitor. Also provided are methods of treating osteosarcoma, which include treating a patient suffering from osteosarcoma with a regimen that includes an effective amount of a pyrimidine antagonist and/or an effective amount of a topoisomerase inhibitor. The pyrimidine antagonist may be, for example, Gemcitabine. The topoisomerase inhibitor may be, for example, irinotecan. In these methods, the regimen may include an effective amount of a pyrimidine antagonist and an effective amount of a topoisomerase inhibitor. Proteomic analysis of tumor tissue from sarcoma patients was carried out to identify tumor molecular characteristics that are predictive of response or resistance to chemotherapies. The results of the analysis were surprising and were used to design new and improved methods of treatment for sarcoma patients. 
    
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         FIG. 1  shows the proteomic landscape of various chemotherapy markers in osteosarcoma. 
         FIG. 2  shows the proteomic landscape of various chemotherapy markers in Ewing&#39;s sarcoma. 
         FIG. 3  shows the proteomic landscape of various chemotherapy markers in leiomyosarcoma. 
         FIG. 4  shows the proteomic landscape of various chemotherapy markers in liposarcoma. 
         FIG. 5  shows the proteomic landscape of various chemotherapy markers in rhabdomyosarcoma. 
         FIG. 6  shows proteogenomic (protein and RNA-seq) correlation of the chemotherapy marker ERCC1. For this marker there was an inverse correlation (r=−0.36). 
         FIG. 7  shows proteogenomic correlation of the chemotherapy marker TOPO2A. This marker has a positive correlation (r=0.86). 
         FIG. 8  shows proteogenomic correlation of the chemotherapy marker hENT1. This marker has a positive correlation (r=0.45 
         FIG. 9  shows proteogenomic correlation of the chemotherapy marker RRM1. This marker has a positive correlation (r=0.37) 
         FIG. 10  shows that 93% of osteosarcoma samples (n=14) expressed the response marker for gemcitabine (hENT1), while only 14% expressed RRM1 protein at a level conferring gemcitabine resistance. 
     
    
    
     DETAILED DESCRIPTION 
     75 sarcoma samples were analyzed with the GPS Cancer™ (Genomic Proteomic Spectrometry) Cancer molecular diagnostic test (NantHealth, Inc., described at http://www.gpscancer.com/) The GPS test employs whole genome sequencing, whole transcriptome (RNA) sequencing, and quantitative proteomics based on mass spectrometry-based Selected Reaction Monitoring (SRM). Microdissected sarcoma tumor tissues were solubilized using the Liquid Tissue® protocol and reagents available from Expression Pathology (Rockville, Md.) and the resulting lysates injected into a mass spectrometer for quantitation of 30 biomarker proteins. 
     The GPS Cancer™ test includes whole genome sequencing (WGS), RNA-sequencing and targeted proteomic analysis. For the nucleic acid sequencing, tumor tissue was biopsied, and sent along with blood for preparation for omics analysis. The sample was microdissected, and DNA &amp; RNA were extracted. For the proteomic analysis, tumor cells from formalin-fixed paraffin-embedded slides were identified by a pathologist. The cells were microdissected by non-contact laser-based dissection using DIRECTOR® slides, followed by Liquid Tissue® processing to release protein fragments into solution. Multi-protein quantitation was carried out by mass spectrometry. 
     Table 1 below shows the genomic features of the 75 clinical samples by sarcoma subtype: 
     
       
         
           
               
             
               
                 TABLE 1 
               
             
            
               
                   
               
               
                 Tumor characteristics 
               
            
           
           
               
               
               
               
            
               
                   
                 Tumor type 
                 No. of sample 
                 Genomic features 
               
               
                   
                   
               
            
           
           
               
               
               
               
            
               
                   
                 Osteosarcoma 
                 14 
                 MKRN2-RAF1 
               
               
                   
                 Ewing&#39;s sarcoma 
                 12 
                 EWSR1-FLI1 
               
               
                   
                   
                   
                 FLI1-EWSR1 
               
               
                   
                 Leiomyosarcoma 
                 25 
                 FAM9A-BRD3 
               
               
                   
                   
                   
                 MSH2-FEZ2 
               
               
                   
                 Liposarcoma 
                 13 
                 MSH2-FEZ2 
               
               
                   
                   
                   
                 NTRK3-TOP1 
               
               
                   
                   
                   
                 TMCO1-NTRK3 
               
               
                   
                   
                   
                 FUS-DDIT3 
               
               
                   
                 Rhabdomyosarcoma 
                 11 
                 PAX3-FOXO1 
               
               
                   
                   
               
            
           
         
       
     
     Analysis of the expression of the proteins ERCC1, TUBB3, TOP2A, TOP1, hENT1/RRM1, and MGMT was carried out on the tissue. Each of these proteins has been previously suggested as being indicative of either sensitivity or resistance to treatment regimens, as summarized in Table 1 below: The molecular assessment of these potential biomarkers in sarcoma cancer subtypes was then used to guide new and improved methods and treatments compared to current standard of care regimens. 
     
       
         
           
               
               
               
             
               
                   
                 TABLE 2 
               
               
                   
                   
               
               
                   
                 Marker 
                 Agents 
               
               
                   
                   
               
             
            
               
                   
               
            
           
           
               
            
               
                 Sensitivity Markers 
               
            
           
           
               
               
               
            
               
                   
                 hENT1 
                 Gemcitabine 
               
               
                   
                 TOPO2A 
                 Anthracyclines (Doxorubicin, Epirubicin), 
               
               
                   
                   
                 Etoposide 
               
               
                   
                 TOPO1 
                 Irinotecan, Topotecan 
               
            
           
           
               
            
               
                 Resistance Markers 
               
            
           
           
               
               
               
            
               
                   
                 RRM1 
                 Gemcitabine 
               
               
                   
                 MGMT 
                 Temzolomide 
               
               
                   
                 TUBB3 
                 Taxanes 
               
               
                   
                 ERCC1 
                 Platinum salts 
               
               
                   
                   
               
            
           
         
       
     
     Results: 
       FIGS. 1-5  show the proteomic landscape of chemotherapy (sensitivity and resistance) markers for each of the sarcoma subtypes (osteosarcoma, Ewing&#39;s sarcoma, leiomyosarcoma. Liposarcoma and rhabdomyosarcoma, respectively).  FIGS. 6-9  show proteogenomic correlations for the chemotherapy markers ERCC1, TOPO2A, hENT1 and RRM1 respectively. Protein and RNA-seq correlation of selected chemotherapy biomarkers are shown. Some targets were inversely correlated (e.g., ERCC1; r=−0.36), while others had positive correlation (TOPO2A; r=0.86). The data were used to analyze the likely response of patients to chemotherapy agents. Table 3 below shows the percentage of patients likely to respond to chemotherapy agents based on proteomic biomarker expression, by sarcoma subtype. 
     
       
         
           
               
               
               
               
               
               
             
               
                   
                 TABLE 3 
               
               
                   
                   
               
               
                   
                   
                 Ewing&#39;s 
                   
                   
                   
               
               
                   
                 Osteosarcoma 
                 Sarcoma 
                 Leiomyosarcoma 
                 Liposarcoma 
                 Rhabdomyosarcoma 
               
               
                   
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
               
               
            
               
                 Topotecan/ 
                 7 
                 33 
                 4 
                 15 
                 36 
               
               
                 Irinotecan 
               
               
                 Anthracycline 
                 7 
                 8 
                 13 
                 0 
                 27 
               
               
                 Gemcitabine 
                 79 
                 25 
                 32 
                 38 
                 9 
               
               
                 Platinum 
                 54 
                 17 
                 72 
                 46 
                 36 
               
               
                 Taxane 
                 43 
                 50 
                 68 
                 38 
                 55 
               
               
                 Temozolomide 
                 33 
                 12 
                 42 
                 33 
                 12 
               
               
                   
               
            
           
         
       
     
     Osteosarcoma 
     TOPO1 protein was overexpressed in 12 of 14 (86%) osteosarcomas. hENT1 is a nucleoside transporter which enables gemcitabine to enter the cell. The effect of gemcitabine is reduced by high levels of RRM1. 93% of osteosarcoma samples (n=14) expressed the response marker for gemcitabine (hENT1), while only 14% expressed RRM1 protein at a level conferring gemcitabine resistance. Overall, ˜80% of osteosarcoma shows sensitivity to gemcitabine. 
     The vast majority of these samples expressed hENT1 protein and lacked expression of RRM1 protein. This proteomic signature indicates that these patients will benefit from a regimen including TOP1 inhibitors (irinotecan) and gemcitabine. The data are summarized in  FIG. 10 . The threshold for expression of hENT1 that conferred sensitivity to gemcitabine was &gt;100 amol/μg, while the threshold of RRM1 that conferred resistance to gemcitabine was &gt;700 amol/μg 
     The current standard of care treatment for osteosarcoma is a combination of a platinum-based agent plus and an anthracycline. The results described herein surprisingly indicate that treatment with Gemcitabine and Irinotecan is effective. 
     Ewing&#39;s Sarcoma 
     Patients with Ewing&#39;s sarcoma (n=12) overexpressed TOPO1 (92%) and TOPO2A (58%) proteins, indicating susceptibility to Irinotecan and anthracyclines. Temozolomide has been studied as a putative treatment for Ewing&#39;s sarcoma, the results described herein suggest that this treatment will be ineffective because the majority (88%) of patients expressed MGMT, a marker for temozolomide resistance. 
     The current standard of care for Ewing&#39;s sarcoma is a combination of Irinotecan and an anthracycline. The results described herein indicate no change from this standard of care 
     Leiomyosarcoma 
     18 of 25 (72%) patients with leiomyosarcoma (LMS) did not express ERCC1 protein, a DNA-repairing endonuclease, indicating susceptibility to DNA damaging agents (e.g., cisplatin, carboplatin). Similarly, 17 of 25 (68%) samples of LMS did not express TUBB3, a resistance marker for tubulin inhibitors (e.g., taxane). 
     The current standard of care treatment for leiomyosarcoma is a combination of Gemcitabine and an anthracycline agent. The results described herein surprisingly indicate that treatment with a taxane+a platinum-based agent is effective. 
     Liposarcoma 
     7/13 (54%) dedifferentiated liposarcoma (DDLPS) samples expressed TOPO1 protein indicating that these patients will benefit from irinotecan-based therapy. 
     The current standard of care treatment for liposarcoma is Gemcitabine and an anthracycline agent. The results described herein surprisingly indicate that treatment with Gemcitabine and Irinotecan is effective. 
     Rhabdomyosarcoma 
     In patients with rhabdomyosarcoma (n=11), TOPO1 and TOPO2A proteins were overexpressed in 91% and 82% of samples, respectively. TOPO1 and TOPO2A are response markers for irinotecan and doxorubicin. In addition, 55% of samples lacked expression of the taxane resistance marker TUBB3. Markers for gemcitabine (hENT1/RRM1) pointed to resistance, with 73% of patients overexpressing RRM1. 
     The current standard of care for rhabdomyosarcoma is a combination of a taxane, an anthracycline and Irinotecan and the results described herein indicate no change from this standard of care. 
     These data are summarized below in Table 4, which shows a comparison of standard of care and potentially effective treatments based on tumor biology. 
     
       
         
           
               
               
               
               
               
               
             
               
                   
                 TABLE 4 
               
               
                   
                   
               
               
                   
                 Osteosarcoma 
                 Ewings 
                 Leiomyosarcoma 
                 Liposarcoma 
                 Rhabdomyosarcoma 
               
               
                   
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
               
               
            
               
                 Standard 
                 methotrexate, 
                 vincristine, 
                 gemcitabine, 
                 gemcitabine, 
                 taxane, 
               
               
                 of care 
                 anthracycline, 
                 ifosfamide, 
                 anthracycline, 
                 anthracycline, 
                 anthracycline, 
               
               
                   
                 platinum 
                 anthracycline, 
                 trabectedin 
                 trabectedin 
                 irinotecan 
               
               
                   
                   
                 Etoposide 
               
               
                 Potentially 
                 gemcitabine, 
                 taxane 
                 taxane, 
                 gemcitabine, 
                 taxane, 
               
               
                 effective 
                 platinum, 
                   
                 platinum, 
                 platinum, 
                 anthracycline, 
               
               
                   
                 trabectedin 
                   
                 trabectedin 
                 trabectedin 
                 irinotecan