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Angiogenesis-Promoting Gene Patterns in Alveolar Soft Part Sarcoma

Posted: Mon Jun 26, 2017 3:57 am
by D.ap
Purpose: We examined a cohort of patients with alveolar soft part sarcoma (ASPS) treated at our institution and showed the characteristic ASPSCR1-TFE3 fusion transcript in their tumors. Investigation of potential angiogenesis-modulating molecular determinants provided mechanistic and potentially therapeutically relevant insight into the enhanced vascularity characteristic of this unusual tumor.



http://clincancerres.aacrjournals.org/c ... full#ref-5

Re: Angiogenesis-Promoting Gene Patterns in Alveolar Soft Part Sarcoma

Posted: Mon Jun 26, 2017 4:02 am
by D.ap
Discussion

ASPS is rare, and most of our knowledge regarding its natural history is based on small retrospective clinical series. This study is one of only four from the past 20 years examining ASPS cohorts of >50 patients (5–7). Differing study designs and data analyses notwithstanding, all four show that ASPS behaves differently than other STS histologic subtypes. ASPS has a distinctive histology, specific molecular characteristics, and unique clinical behaviors. Compared with other STSs as a whole, ASPS generally develop in younger patients and occur more frequently in the lower extremity (5–7), and have a much lower incidence of local recurrence (20, 21). It should be noted that most sarcomas associated with known gene fusions generally occur in younger patients and ASPS is no exception to this observation. The high rate of ASPS metastasis (79% in this series) is much higher than the 50% overall rate of metastasis for all STS patients (22, 23), 10% of whom present with metastases, as compared with 20% to 65% of ASPS patients who are stage IV at presentation (5–7). The pattern of ASPS metastasis is also distinctive: as with other STS, the lung is the most common metastatic locus but brain and bone metastases, which rarely occur in other STSs, commonly occur in ASPS.

Our actuarial overall survival was similar to that previously published (5–7). It is encouraging that most patients with metastatic ASPS can survive long-term, however, these patients are not cured and many eventually succumb to ASPS, as suggested by the continuous decrease in survival even 20 or more years after metastasis. This is a crucial consideration for patients with ASPS, as most are in their 20s when initially diagnosed.

Currently, we lack effective systemic molecularly targeted therapies. There is limited knowledge of the biological processes and molecular determinants driving ASPS. The highly vascular nature of ASPS (8), possibly relevant to metastasis, combined with an emerging awareness that a specific molecular defect causes overexpression of the ASPSCR1-TFE3 chimeric transcription factor in ASPS (8, 14), merits focused attention. This chimeric protein seems to act as an aberrant transcription factor binding TFE3 promoter sites and causing dysregulation of gene expression. We showed that the ASPSCR1-TFE fusion transcript could be found in virtually all cases of ASPS in which high-quality RNA was obtained. The two negative cases could be false-negatives due to poor-quality RNA. It is possible that an abundant transcript such as β-actin may be detectable but the ASPSCR1-TFE3 fusion transcript, which is presumably less abundant, may escape amplification and detection in such cases. Some support to this idea of the universal presence of the fusion gene is supplied by the fact that all 33 FFPE ASPS cases in this series, including the 2 cases negative by PCR, expressed strong and diffuse nuclear staining for TFE3 following immunohistochemistry. This finding is strongly indicative of the diagnosis of ASPS and the presence of the fusion transcript (14). Our finding of only one reciprocal fusion product out of 16 fusion transcripts identified is consistent with previous results (8, 11, 24). Our results indicate that the fusion product is present in most ASPS cases and significantly expands our knowledge of the prevalence of the type 1 and 2 fusion transcripts. There is experimental data that the rarity of the balanced translocations may be due to the translocation occurring during the G2 phase of the cell cycle, with perhaps a selective advantage for tumor cells with gain of the telomeric p-arm of X or loss of the telomeric q-arm of 17 chromosomal material (11).

The ASPSCR1-TFE3 fusion transcript may underlie the expression of angiogenic factors in ASPS. The expression oligoarrays revealed a panel of angiogenesis-promoting genes that are reproducibly overexpressed in human ASPS, and interestingly, did not include those angiogenic molecules such as vascular endothelial growth factors, fibroblast growth factors, platelet-derived growth factors, angiopoietins, or matrix metalloproteases more commonly encountered in other malignancies and other sarcomas as well. It is tempting to postulate that the angiogenic profile is induced by the action of the ASPCR1-TEF3 fusion transcript and this idea is buttressed by our finding that several of the genes in our discovered angiogenic profile have putative TFE3 binding sites in their promoter regions. A recent report (15) described the effect of antiangiogenic, anti–vascular endothelial growth factor therapy in a patient with metastatic ASPS; although tumor regression was observed initially, disease progression occurred after 6 months. Based on our results, it is conceivable that inhibiting combinations of array-demonstrated overexpressed angiogenic factors might result in more sustained responses. It is noteworthy that three of the identified factors (neuropilin, midkine, and pleiotropin) are also associated with neuronal cells (25–27), perhaps of relevance to ASPS brain metastasis and meriting further investigation.

To validate our expression oligoarray results, we selected three protein products of the up-regulated genes (jag-1, midkine, and angiogenin), demonstrating that they were consistently up-regulated in an expanded cohort of human ASPS specimens. Although a regulatory role for TFE3 regarding any of the genes found in our array has not been previously shown, we were able to identify potential TFE3-binding sites in the promoters of the above three up-regulated genes. It is possible that the unique angiogenic gene profile of ASPS is related to this characteristic ASPS translocation, resulting in unregulated TFE3 overexpression. When these three factors were examined in additional sarcomas of other histologic types by both immunohistochemistry and expression oligoarrays, the proteins and the RNAs encoding them were present but lacked the simultaneous significant expression of all three factors characteristic of ASPS. It seems that ASPS is characterized by the ASPSCR1-TFE3 fusion transcript that likely induces the unique angiogenic profile. Overall, there seemed to be little variation in the overall angiogenic profile identified by oligoarray and expression of the three associated angiogenic proteins examined here in the larger ASPS tumor cohort. Although not shown for ASPS, data is emerging from other sarcoma mouse and other models that the characteristic fusion transcripts may sometimes be sufficient for the development of tumors when expressed at the right time and in the correct cellular compartment, although other genetic changes may also be required (28–34). Developing personalized molecular therapeutic strategies that might potentially block ASPS angiogenic promotion directly or indirectly via ASPSCR1-TFE3 inhibition (i.e., via TFE3-blocking short interfering RNA, or a combination of downstream effectors such as the angiogenic proteins identified here) may hopefully lead to increased curability for these all too young patients burdened by this unique and rare disease.

Gene expression profiling of ASPS

Posted: Tue Nov 14, 2017 7:04 pm
by D.ap
Abstract
Background

Alveolar soft-part sarcoma (ASPS) is an extremely rare, highly vascular soft tissue sarcoma affecting predominantly adolescents and young adults. In an attempt to gain insight into the pathobiology of this enigmatic tumor, we performed the first genome-wide gene expression profiling study.

Methods

For seven patients with confirmed primary or metastatic ASPS, RNA samples were isolated immediately following surgery, reverse transcribed to cDNA and each sample hybridized to duplicate high-density human U133 plus 2.0 microarrays. Array data was then analyzed relative to arrays hybridized to universal RNA to generate an unbiased transcriptome. Subsequent gene ontology analysis was used to identify transcripts with therapeutic or diagnostic potential. A subset of the most interesting genes was then validated using quantitative RT-PCR and immunohistochemistry.

Results

Analysis of patient array data versus universal RNA identified elevated expression of transcripts related to angiogenesis (ANGPTL2, HIF-1 alpha, MDK, c-MET, VEGF, TIMP-2), cell proliferation (PRL, IGFBP1, NTSR2, PCSK1), metastasis (ADAM9, ECM1, POSTN) and steroid biosynthesis (CYP17A1 and STS). A number of muscle-restricted transcripts (ITGB1BP3/MIBP, MYF5, MYF6 and TRIM63) were also identified, strengthening the case for a muscle cell progenitor as the origin of disease. Transcript differentials were validated using real-time PCR and subsequent immunohistochemical analysis confirmed protein expression for several of the most interesting changes (MDK, c-MET, VEGF, POSTN, CYP17A1, ITGB1BP3/MIBP and TRIM63).

Conclusion

Results from this first comprehensive study of ASPS gene expression identifies several targets involved in angiogenesis, metastasis and myogenic differentiation. These efforts represent the first step towards defining the cellular origin, pathogenesis and effective treatment strategies for this atypical



https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2635365/

Alveolar soft-part sarcoma (ASPS) resembles a mesenchymal stromal progenitor: evidence from meta-analysis of transcripto

Posted: Sun Dec 27, 2020 2:50 am
by D.ap
Alveolar soft-part sarcoma (ASPS) resembles a mesenchymal stromal progenitor: evidence from meta-analysis of transcriptomic data



Alveolar soft-part sarcoma (ASPS) is an extremely rare malignancy characterized by the unbalanced translocation der(17)t(X;17)(p11;q25). This translocation generates a fusion protein, ASPL-TFE3, that drives pathogenesis through aberrant transcriptional activity. Although considerable progress has been made in identifying ASPS therapeutic vulnerabilities (e.g., MET inhibitors), basic research efforts are hampered by the lack of appropriate in vitro reagents with which to study the disease. In this report, previously unmined microarray data for the ASPS cell line, ASPS-1, was analyzed relative to the NCI sarcoma cell line panel. These data were combined with meta- analysis of pre-existing ASPS patient microarray and RNA-seq data to derive a platform- independent ASPS transcriptome. Results demonstrated that ASPS-1, in the context of the NCI sarcoma cell panel, had some similarities to normal mesenchymal cells and connective tissue sarcomas. The cell line was characterized by high relative expression of transcripts such as CRYAB, MT1G, GCSAML, and SV2B. Notably, ASPS-1 lacked mRNA expression of myogenesis-related factors MYF5, MYF6, MYOD1, MYOG, PAX3, and PAX7. Furthermore, ASPS-1 had a predicted mRNA surfaceome resembling an undifferentiated mesenchymal stromal cell through expression of GPNMB, CD9 (TSPAN29), CD26 (DPP4), CD49C (ITGA3), CD54 (ICAM1), CD63 (TSPAN30), CD68 (SCARD1), CD130 (IL6ST), CD146 (MCAM), CD147 (BSG), CD151 (SFA-1), CD166 (ALCAM), CD222 (IGF2R), CD230 (PRP), CD236 (GPC), CD243 (ABCB1), and CD325 (CDHN). Subsequent re-analysis of ASPS patient data generated a consensus expression profile with considerable overlap between studies. In common with ASPS-1, elevated expression was noted for CTSK, DPP4, GPNMB, INHBE, LOXL4, PSG9, SLC20A1, STS, SULT1C2, SV2B, and UPP1. Transcripts over-expressed only in ASPS patient samples included ABCB5, CYP17A1, HIF1A, MDK, P4HB, PRL, and PSAP. These observations are consistent with that expected for a mesenchymal progenitor cell with adipogenic, osteogenic, or chondrogenic potential. In summary, the consensus data generated in this study highlight the unique and highly conserved nature of the ASPS transcriptome. Although the ability of the ASPL-TFE3 fusion to perturb mRNA expression must be acknowledged, the prevailing ASPS transcriptome resembles that of a mesenchymal stromal progenitor.


https://peerj.com/articles/9394.pdf

Re: Alveolar soft-part sarcoma (ASPS) resembles a mesenchymal stromal progenitor: evidence from meta-analysis of transcr

Posted: Sat Apr 13, 2024 6:59 am
by D.ap
**
D.ap wrote: Sun Dec 27, 2020 2:50 am Alveolar soft-part sarcoma (ASPS) resembles a mesenchymal stromal progenitor: evidence from meta-analysis of transcriptomic data



Alveolar soft-part sarcoma (ASPS) is an extremely rare malignancy characterized by the unbalanced translocation der(17)t(X;17)(p11;q25). This translocation generates a fusion protein, ASPL-TFE3, that drives pathogenesis through aberrant transcriptional activity. Although considerable progress has been made in identifying ASPS therapeutic vulnerabilities (e.g., MET inhibitors), basic research efforts are hampered by the lack of appropriate in vitro reagents with which to study the disease. In this report, previously unmined microarray data for the ASPS cell line, ASPS-1, was analyzed relative to the NCI sarcoma cell line panel. These data were combined with meta- analysis of pre-existing ASPS patient microarray and RNA-seq data to derive a platform- independent ASPS transcriptome. Results demonstrated that ASPS-1, in the context of the NCI sarcoma cell panel, had some similarities to normal mesenchymal cells and connective tissue sarcomas. The cell line was characterized by high relative expression of transcripts such as CRYAB, MT1G, GCSAML, and SV2B. Notably, ASPS-1 lacked mRNA expression of myogenesis-related factors MYF5, MYF6, MYOD1, MYOG, PAX3, and PAX7. Furthermore, ASPS-1 had a predicted mRNA surfaceome resembling an undifferentiated mesenchymal stromal cell through expression of GPNMB, CD9 (TSPAN29), CD26 (DPP4), CD49C (ITGA3), CD54 (ICAM1), CD63 (TSPAN30), CD68 (SCARD1), CD130 (IL6ST), CD146 (MCAM), CD147 (BSG), CD151 (SFA-1), CD166 (ALCAM), CD222 (IGF2R), CD230 (PRP), CD236 (GPC), CD243 (ABCB1), and CD325 (CDHN). Subsequent re-analysis of ASPS patient data generated a consensus expression profile with considerable overlap between studies. In common with ASPS-1, elevated expression was noted for CTSK, DPP4, GPNMB, INHBE, LOXL4, PSG9, SLC20A1, STS, SULT1C2, SV2B, and UPP1. Transcripts over-expressed only in ASPS patient samples included ABCB5, CYP17A1, HIF1A, MDK, P4HB, PRL, and PSAP.** These observations are consistent with that expected for a mesenchymal progenitor cell with adipogenic, osteogenic, or chondrogenic potential. .In summary, the consensus data generated in this study highlight the unique and highly conserved nature of the ASPS transcriptome. Although the ability of the ASPL-TFE3 fusion to perturb mRNA expression must be acknowledged, the prevailing ASPS transcriptome resembles that of a mesenchymal stromal progenitor.


https://peerj.com/articles/9394.pdf