Soft Tissue Sarcoma In Children Research Articles

Pediatric rhabdomyosarcoma incidence and survival in the United States: An assessment of 5656 cases, 2001-2017.

While rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in children and adolescents, past epidemiology studies of this malignancy used data that covered <30% of the US population. Therefore, we evaluated RMS incidence using data from U.S. Cancer Statistics (USCS) and survival trends using the National Program of Cancer Registries (NPCR), which covers 100% and 94% of the U.S. population, respectively. Incidence and survival were assessed for pediatric patients diagnosed with RMS during 2003-2017 and 2001-2016, respectively. Both demographic and clinical variables were evaluated. Age-adjusted incidence rates, average annual percent change (AAPC), and 5-year relative survival (RS) were calculated, all with corresponding 95% confidence intervals (CIs). Cox regression models were used to evaluate the impact of demographic and clinical variables on survival. We identified 5656 primary RMS cases in USCS during 2003-2017. The age-adjusted incidence rate was 4.58 per 1 million (95% CI: 4.46-4.70) with an AAPC of 0.3% (95% CI: -0.7 to 1.2%). In NPCR, 5-year RS for all cases was 68.0% (95% CI: 66.6-69.3%). In multivariable analyses, non-Hispanic (NH) Black cases had worse survival compared with NH White cases (hazard ratio [HR]=1.16, 95% CI: 1.01-1.33). The incidence and survival rates were stable in the largest and most comprehensive population-based analysis for pediatric RMS cases in the U.S. Additionally, we observed a survival disparity among NH Black cases. Findings from this study could inform interventions to address disparities, risk stratification strategies, and clinical trial design.

Prognostic factors of pediatric pelvic and genitourinary rhabdomyosarcoma: An analysis based on SEER database.

BackgroundRhabdomyosarcoma (RMS) is the most common soft-tissue sarcomas in children. This study aimed to investigate the prognostic factors of pelvic and genitourinary RMS in children and evaluate the survival outcomes of these children treated with or without radiation therapy (RT).MethodsThe Surveillance, Epidemiology, and End Results Program (SEER) database was required for children with pelvic and genitourinary RMS. Overall survival (OS) and cancer-specific survival (CSS) were analyzed using the Kaplan-Meier method, log-rank test, Cox proportional hazards models, and propensity score-matched analyses.ResultsFor the 262 patients analyzed, the most common biological subtypes were embryonic (n=209, 79.8%) and alveolar (n=29, 11.1%). Patients with alveolar RMS had the worst prognosis (P < 0.05). The testis (n=122, 46.6%) was the most common location, followed by the urinary bladder (n=57, 21.8%) and prostate (n=48, 18.3%). Uterus RMS had the highest survival rate, followed by testis, urinary bladder, and prostate RMS. Favorable prognostic factors were age at diagnosis < 15 years, non-alveolar histological subtype, early tumor stage (localized/regional), specific sites (uterus and testis), and treatment (cancer-directed surgery and chemotherapy) (P < 0.05). Propensity score-matched analyses comparing the cohorts of patients treated with or without RT demonstrated no significant differences in prognostic survival (OS: P=0.872, CSS: P=0.713).ConclusionThe nomogram constructed based on independent prognostic factors may accurately predict survival rates at 1 and 5 years. Surgery and adjuvant chemotherapy can be effective treatments, but RT fails to guarantee a survival benefit. Therefore, prospective trials evaluating RT for pediatric pelvic and genitourinary RMS are warranted.

Rhabdomyosarcoma in children: Retrospective analysis from a single tertiary care center in Saudi Arabia.

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in children occurring most commonly in the head and neck region. The treatment involves using a multimodality approach including chemotherapy, surgery, and radiation therapy. Survival for patients with localized disease has improved markedly, but the treatment of advanced disease remains a challenge. We report the clinical characteristics and outcome for patients treated at a tertiary care center in Saudi Arabia. Patients aged 0-14 years diagnosed with RMS between 2005 and 2018 were included. Statistical analysis was performed using SPSS software. Kaplan-Meier method was used to calculate overall and event free survival. Cox proportional hazards model was used for multivariate analysis. One hundred and twenty-four patients were analyzed. The median age was 5.7 years with male predominance (2.4:1). The most common primary sites were head/neck (30%) and the genitourinary tract (25%). Embryonal RMS was present in 81%; alveolar in 19%. Most patients had intermediate risk disease (60%). The 5-year overall and event free survivals were 64.3% and 53.3%, respectively. Survival was influenced by primary tumor site, histology, and clinical risk group. Unfavorable primary site, high risk stratification, and poor initial response to therapy predicted a poor outcome. This study provides an insight on the current management outcomes for our patients with RMS. Cytogenetics and molecular diagnostics need to be incorporated as standard of care in the therapeutic approach of our patients. In addition, there is a need for national collaborative efforts to improve the outcome of RMS in children and adolescents.

Mesenchymal tumor organoid models recapitulate rhabdomyosarcoma subtypes

Rhabdomyosarcomas (RMS) are mesenchyme‐derived tumors and the most common childhood soft tissue sarcomas. Treatment is intense, with a nevertheless poor prognosis for high‐risk patients. Discovery of new therapies would benefit from additional preclinical models. Here, we describe the generation of a collection of 19 pediatric RMS tumor organoid (tumoroid) models (success rate of 41%) comprising all major subtypes. For aggressive tumors, tumoroid models can often be established within 4–8 weeks, indicating the feasibility of personalized drug screening. Molecular, genetic, and histological characterization show that the models closely resemble the original tumors, with genetic stability over extended culture periods of up to 6 months. Importantly, drug screening reflects established sensitivities and the models can be modified by CRISPR/Cas9 with TP53 knockout in an embryonal RMS model resulting in replicative stress drug sensitivity. Tumors of mesenchymal origin can therefore be used to generate organoid models, relevant for a variety of preclinical and clinical research questions.

Targeting KDM4 for treating PAX3-FOXO1-driven alveolar rhabdomyosarcoma.

Chimeric transcription factors drive lineage-specific oncogenesis but are notoriously difficult to target. Alveolar rhabdomyosarcoma (RMS) is an aggressive childhood soft tissue sarcoma transformed by the pathognomonic Paired Box 3-Forkhead Box O1 (PAX3-FOXO1) fusion protein, which governs a core regulatory circuitry transcription factor network. Here, we show that the histone lysine demethylase 4B (KDM4B) is a therapeutic vulnerability for PAX3-FOXO1+ RMS. Genetic and pharmacologic inhibition of KDM4B substantially delayed tumor growth. Suppression of KDM4 proteins inhibited the expression of core oncogenic transcription factors and caused epigenetic alterations of PAX3-FOXO1-governed superenhancers. Combining KDM4 inhibition with cytotoxic chemotherapy led to tumor regression in preclinical PAX3-FOXO1+ RMS subcutaneous xenograft models. In summary, we identified a targetable mechanism required for maintenance of the PAX3-FOXO1-related transcription factor network, which may translate to a therapeutic approach for fusion-positive RMS.

Rhabdomyosarcoma of the gallbladder

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in the pediatric population, for 45% of soft tissue sarcomas in children. Commonly located in the head and neck, urogenital system, extremities and torso, RMS of the biliary system is rare. We present a specific case of RMS in the gallbladder of a 4-year-old girl who presented with epigastric abdominal pain and vomiting. Her work-up revealed transaminitis and tumefactive sludge in the gallbladder neck. After she had persistent symptoms and repeated work-up, the patient underwent a laparoscopic cholecystectomy and was found to have cholecystitis intraoperatively with cholelithiasis. However, pathology revealed botryoid embryonal RMS with positive microscopic margins. This case demonstrates the difficulty in diagnosing gallbladder RMS and the importance of considering malignancy in pediatric biliary disease.

Post-traumatic Embryonal Rhabdomyosarcoma: An Unfortunate Red Herring

Rhabdomyosarcoma (RMS) is a relatively rare malignancy but the most common cause of soft tissue sarcoma in children. It can present anywhere in the body, but most seen in the extremities, head, neck and genitourinary organs. The two most common types of RMS are embryonal RMS and the more aggressive kind, the alveolar RMS. Development of RMS occurring after trauma is uncommon but as children are readily prone for lumps and bumps due to their adventurous and playful nature, it is much important to always keep in mind the possibility of malicious causes of swelling as any delay in diagnosis directly affects the outcome of the disease. Here we present an infant who presented with symptoms suggestive of post trauma swelling but later was confirmed as embryonal RMS. The child underwent chemotherapy as the treatment of option.

Primary intra-testicular rhabdomyosarcoma: Case report.

RMS is a highly aggressive neoplasm that represents the most common soft tissue sarcoma in children and adolescents. It occurs most frequently in the head, neck, and genitourinary tract. Intra-testicular localization remains exceptional and very few cases have been reported in the literature. We represent a case of a 16-year-old patient, with no medical and surgical history, who presented with a painless right scrotal mass. Imaging revealed a suspicious testicular mass with no other secondary location. The patient underwent a right inguinal orchidectomy and the anatomopathological study confirmed the diagnosis of intratesticular rhabdomyosarcoma. The patient subsequently was referred to the oncology department for chemotherapy (Agha et al., 2020). Intra-testicular rhabdomyosarcoma is a very rare malignant mesenchymal tumor showing skeletal muscle differentiation. They are subclassified into four major histological subtypes: Embryonal, alveolar, pleomorphic and Spindle cell/sclerosing rhabdomyosarcoma. ERMS is the most common subtype. Diagnosis of ITRMS is based on anatomopathological examination. Surgery and chemotherapy remain the mainstay of treatment. Radiotherapy is useful for local recurrence and metastasis. In this report we aim to describe the clinical, pathological and immunohistochemical features of this rare entity, and to highlight the importance of an early diagnosis followed by treatment combining Radical inguinal orchiectomy and chemotherapy and its role in improving prognosis. • Intra-testicular localization of rhabdomyosarcoma remains exceptional and few cases have been reported in the literature. • Anatomopathological examination is the gold standard for diagnosis and for confirming the intratesticular origin • Radical inguinal orchiectomy followed by chemotherapy is the treatment of choice.

Abstract 700: Targeting transcriptional co-activators YAP1/TAZ in fusion-positive rhabdomyosarcoma demonstrates a novel strategy in ablating fusion-driven sarcoma transcriptional programing

Abstract Background: Approximately one third of all sarcomas are characterized by recurrent chromosomal translocations, and these fusion-positive sarcomas remain among the most difficult to treat of all malignancies. Rhabdomyosarcoma is the most common soft tissue sarcoma of childhood and a prototype fusion-oncogene-driven tumor. Fusion-positive rhabdomyosarcoma (FP-RMS) is driven by a chromosomal translocation encoding for the pathognomonic chimeric fusion product PAX3-FOXO1 (P3F). Because it is a large inherently disordered transcription factor lacking drug-binding sites, conventional efforts to directly target P3F have thus far proven futile, and 5-yr overall survival rates remain below 50%. Objective: We thus sought to identify therapeutically tractable co-activators or co-regulators that represent vulnerabilities to P3F transcriptional programing. Methods/Results: Interrogation of transcriptomic data and immunohistochemical (IHC) staining of human tissue microarrays demonstrated the transcriptional co-activators YAP1 and TAZ are highly abundant in FP-RMS tumors. We performed genetic gain- and loss-of-function experiments using RNAi and CRISPR/Cas9 to determine that YAP1/TAZ regulate many FP-RMS cancer phenotypes, including proliferation and xenograft tumor growth. Mechanistic studies using co-immunoprecipitation (co-IP) and co-IP-coupled mass spectrometry revealed a YAP1/TAZ-P3F physical interaction and that YAP1/TAZ and P3F share an enrichment for co-immunoprecipitated proteins involved in chromatin remodeling and DNA binding as well as transcriptional regulation. Functional studies using qRT-PCR, immunoblots, and reporter assays demonstrated YAP1/TAZ are positive regulators of P3F-mediated transcriptional activity. Unbiased approaches using RNA-Seq and quantitative tandem mass tag proteomics also demonstrated that YAP1/TAZ positively regulate the differential expression of P3F’s targets and gene signature. Pharmacologically targeting the YAP1/TAZ-P3F axis using novel NUAK1/2 kinase inhibitors that lead to YAP1/TAZ nuclear exclusion cause cell cycle arrest at the G2/M checkpoint as well as a decrease in FP-RMS cell viability through induction of apoptosis. Main Conclusions: Transcriptional co-activators YAP1/TAZ are highly abundant in FP-RMS tumors and are required for numerous FP-RMS cancer phenotypes. YAP1/TAZ physically associate with P3F to regulate P3F-mediated transcriptional activity. Therefore, while directly targeting P3F is not currently viable, we have identified YAP1/TAZ as therapeutically tractable dependencies to P3F transcriptional programing. Citation Format: Adam F. Pecoraro, Tooba Rashid, Breanne A. Burgess, Corinne M. Linardic, Michael D. Deel. Targeting transcriptional co-activators YAP1/TAZ in fusion-positive rhabdomyosarcoma demonstrates a novel strategy in ablating fusion-driven sarcoma transcriptional programing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 700.

Abstract 552: FGFR4 and CD276 dualtargeting CAR-T cells for treating rhabdomyosarcoma and other solid tumors

Abstract Background: Chimeric antigen receptor T-cell therapies (CAR-T) have shown success in treating refractory and relapsed leukemia and lymphoma, while they perform poorly in solid tumors due to heterogenous expression of tumor-associated antigens (TAAs), limited T cell persistence and propensity for exhaustion. The receptor tyrosine kinase FGFR4 and immune checkpoint molecule CD276 are highly and heterogeneously expressed in some solid tumors, including Rhabdomyosarcoma (RMS), a most common soft tissue sarcoma of childhood, and human hepatocellular carcinoma (HCC). However, their expression is usually low in normal human tissues. These features make FGFR4 and CD276 promising therapeutic targets for CAR-T therapy for RMS and HCC. We have developed a FGFR4 targeting CAR construct (3A11-BBz) with a CD8 hinge (H) and a transmembrane domain (TM) infused with a 4-1BB intracellular domain (ICD). 3A11-BBz CAR can efficiently eliminate low RMS disease burden in metastatic models, but less effectively for bulky disease in RMS intramuscular (I.M.) xenograft models. Testing of a CD276 targeting CAR T-cells showed significant shrinking of tumors in RMS I.M. xenograft models. Methods: To improve the CAR-T cells efficacy, we first modified the H/TM and ICD of 3A11-BBz CAR to CD28 (3A11-CD28z). To overcome tumor heterogeneity, we also created Bicistronic CARs (BiCisCARs) combining the complete FGFR4 and CD276 CAR into a single construct allowing co-expression of both constructs on the same T cells. We then tested the efficacy of these CARs in-vitro and in-vivo using intramuscular FP-RMS xenograft (RH30) or HCC intraperitoneal models. Results and Conclusions: We found either FGFR4 targeting CARs or dual targeting BiCisCARs, showed similar in-vitro cytotoxicity against RMS cells and HCC cells. However, CARs with CD28 ICD released more IL-2 than those with 4-1BB ICD when co-cultured with target cells. In RMS I.M. xenograft model, 3A11-CD28z CAR-T cells shrank and eliminated the tumor in 5/8 mice whereas 3A11-BBz only suppressed tumor growth. Furthermore, 3A11-CD28z BiCisCAR eradicated tumor cells in 8/8 mice, whereas 3A11-BBz BiCisCAR showed very poor efficacy. Moreover, there are more 3A11-CD28z BiCisCAR T-cells persisting in blood and spleen than the other bicistronic or single CAR-T cells, suggesting this BiCisCAR-T cells have prolonged persistence. Therefore, we have developed a potent BiCisCAR dual targeting both FGFR4 and CD276 that overcomes RMS heterogeneity and effectively eliminates tumors in-vivo, which will be developed as a future therapeutic CAR for clinical trials. Citation Format: Meijie Tian, Adam Cheuk, David Milewski, Jun S. Wei, Hsien-Chao Chou, Yong Yean Kim, Young K. Song, Brad St. Croix, Mitchell Ho, Javed Khan. FGFR4 and CD276 dualtargeting CAR-T cells for treating rhabdomyosarcoma and other solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 552.

Abstract 1667: Synthetic essentiality between PTEN and core dependency factor PAX7 dictates rhabdomyosarcoma identity

Abstract Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in children with no improvements in treatment options for RMS patients over the past four decades. Therefore, it is critical to understand the fundamental processes underlying RMS tumorigenesis. RMS is divided into two major histologic subtypes - alveolar and embryonal RMS. Nearly all alveolar RMS express oncogenic fusions of PAX3-FOXO1 or PAX7-FOXO1 whereas embryonal RMS are not driven by these fusion proteins. Instead, embryonal or fusion-negative (FN-RMS) are molecularly heterogeneous. Approximately one-third of fusion-negative RMS (FN-RMS) patients have copy number loss of PTEN (phosphatase and tensin homolog), and approximately 90% of tumors are hypermethylated at the PTEN promoter leading to decreased PTEN expression. This indicates a near universal role for PTEN loss in FN-RMS, but the functional role of PTEN is still unclear. To determine PTEN’s function in FN-RMS, we bred Ptenflox alleles into our aP2-Cre;SmoM2 (ASPWT) FN-RMS mouse model to obtain aP2-Cre;SmoM2;Ptenflox/flox mice (ASPcKO). Conditional Pten deletion accelerated tumorigenesis and produced a tumor with a less differentiated histological appearance, much like human FN-RMS. Interestingly, in PtenWT tumors, we found predominant PTEN immunoreactivity within the nucleus suggesting a role for nuclear PTEN in FN-RMS. Transcriptome analyses revealed robust gene expression changes between the ASPWT and ASPcKO tumors. The top overexpressed gene in ASPcKO tumors was Dbx1 (Developing brain homeobox 1), a homeobox transcription factor with no known cancer function but involved in innate behavioral processes such as breathing. We found FN-RMS patient-derived xenografts are dependent on DBX1 expression, and that DBX1 expression is controlled by PAX7 (Paired Box 7). PAX7 is a transcription factor expressed in satellite cells and maintains a de-differentiated state in FN-RMS. PAX7 expression is also increased in our ASPcKO tumors, and we show that human FN-RMS cells are dependent on PAX7 expression for proliferation. This suggests PTEN loss in FN-RMS engages a new transcriptional program necessary for FN-RMS survival. To determine if Pax7 loss can rescue the deleterious effects of Pten loss in our murine FN-RMS model, we deleted both Pten and Pax7 in our aP2-Cre;SmoM2 mice (ASPcKOP7cKO). ASPcKOP7cKO tumor onset kinetics resembled tumors with wild-type PTEN and were negative for skeletal muscle markers MYOD1 and MYOGENIN. However, these tumors were positive for leiomyosarcoma markers smooth muscle actin and CALDESMON. Together, our data suggests PTEN and PAX7 have a synthetic essential relationship in FN-RMS and that PAX7 is a proliferative driver and lineage dependency for FN-RMS tumors. This work also illustrates the power of murine models to unravel the genetic dependencies underlying both tumor maintenance and identity. Citation Format: Casey G. Langdon, Katherine E. Gadek, Matthew R. Garcia, Myron K. Evans, Kristin B. Reed, Madeline Bush, Jason A. Hanna, Catherine J. Drummond, Matthew C. Maguire, Patrick J. Leavey, David Finkelstein, Hongjian Jin, Patrick A. Schreiner, Jerold E. Rehg, Mark E. Hatley. Synthetic essentiality between PTEN and core dependency factor PAX7 dictates rhabdomyosarcoma identity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1667.

Abstract 683: Identification of common germline variants associated with pediatric rhabdomyosarcoma survival: A report from the Children's Oncology Group (COG)

Abstract Background: Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in children and has one of the poorest survival rates among pediatric cancers, underscoring the need to identify factors which may be leveraged to improve therapeutic options for these individuals. Methods: We carried out a genome-wide association study of overall survival (OS) and event-free survival (EFS) in 920 RMS patients from COG protocols and randomly divided them into discovery (n=642) and replication (n=278) cohorts. Genotyping was conducted using the Illumina OmniExpress or Global Screening Array and imputed using the Haplotype Reference Consortium. We used Cox proportional hazards regression to calculate an adjusted hazard ratio (aHR) and P value for each common variant (minor allele frequency [MAF]&amp;gt;5%) for OS and EFS while adjusting for age at diagnosis, tumor stage, histological subtype, and the top five principal components. Analyses were also conducted by histological subtype: embryonal RMS (ERMS, n=544) and alveolar RMS (ARMS, n=268). Finally, we performed a meta-analysis of the results from the discovery and replication cohorts to generate a summary aHR and P value for each single nucleotide polymorphism (SNP). Results: We identified an intergenic SNP at chr8q21.13 associated with worse RMS EFS across subtypes (aHR=2.08, P=2.80x10-9), which had consistent effects across the discovery (aHR=1.91, P=5.05x10-6) and replication (aHR=2.62, P=7.16x10-5) cohorts. This SNP lies in a region which spans the genomic binding site for GATA2 and GATA3, transcription factors that are recognized to contribute to cancer development. We also identified a significant association between a SNP at chr12q21.1 and worse EFS (aHR=2.04, P=3.35x10-8) with consistent effects across the discovery and replication cohorts. Based on data from the Genotype-Tissue Expression project (GTEx), this SNP is associated with expression of SLCO1B1, a gene which encodes a liver anion transporter linked to RMS treatment-related toxicities. In subtype-specific analyses, we identified a SNP at chr17q21.32 that was significantly associated with worse ARMS OS (129 events; aHR=3.18, P=3.12x10-8; discovery: aHR=3.19, P=6.23x10-4; replication: aHR=3.16, P=1.43x10-3). In GTEx, this SNP is associated with expression and splicing of genes including PITPNM3, KIAA0753, and MED31 across various tissues. No SNPs were significantly associated with ERMS OS or EFS. Conclusion: In the first GWAS of RMS survival outcomes, we identified two SNPs that were significantly associated with worse EFS across RMS subtypes. Further, we identified a SNP that was associated with OS in ARMS patients, a subtype that is associated with worse outcomes. Additional investigation of the impact of these SNPs may further support their consideration for novel risk stratification protocols. Citation Format: Bailey A. Martin-Giacalone, Michael E. Scheurer, Javed Khan, Stephen J. Chanock, Shengchao Alfred Li, Meredith Yeager, Deborah A. Marquez-Do, Donald A. Barkauskas, David Hall, Matthew T. McEvoy, Melissa A. Richard, Pagna Sok, Austin L. Brown, Aniko Sabo, Stephen X. Skapek, Douglas S. Hawkins, Rajkumar Venkatramani, Lisa Mirabello, Philip J. Lupo. Identification of common germline variants associated with pediatric rhabdomyosarcoma survival: A report from the Children's Oncology Group (COG) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 683.

Abstract 2020: Investigation of CYP450 enzymes as therapeutic targets in rhabdomyosarcoma

Abstract Introduction: Rhabdomyosarcoma (RMS) is a rare soft tissue sarcoma (STS) that predominantly affects children and teenagers. It is the most frequent STS in children and accounts for around 5-8% of total childhood cancers. The improvement of chemotherapeutic regimens has raised RMS survival chances over the last decades. However, the mortality rate remains exceptionally high in high-risk RMS patients. Duocarmycins have arisen as an opportunity for targeted therapy in cancer. These ultrapotent anticancer agents have been reengineered to generate bioprecursors that are selectively activated bycertain cytochrome P450 (CYP) isoforms (e.g. CYP1A1, CYP1B1, CYP2W1). The overexpression of these isoforms in tumors provides the possibility to locoregionally bioactivate duocarmycin prodrugs only in the tumor site. Previous studies have shown that duocarmycins can be chemically modified and deactivated by removal of a hydroxyl group in the pharmacophore, which can be reinstalled by CYPsto generate a potent DNA-targeted cytotoxic metabolite. In the past, a consistent CYP2W1overexpression in patient-derived RMS biopsies as compared to matched, non-transformed tissue was revealed. Results: Ongoing studies are focused on measuring CYP1A1, CYP1B1 and CYP2W1 expression levels in RMS cell lines and in three-dimensional spheroids as well as evaluating the antiproliferative activity of duocarmycin bioprecursors in these models. With the exception of RH30 cells (IC50=0.2µM forICT2700) the IC50 for CYP1A1- and CYP2W1-bioactivated duocarmycins (ICT2700 and ICT2706,respectively) lie in the low micromolar range in RMS monolayer cultures, suggesting that CYP1A1 andCYP2W1 might not be a suitable target in these 2D cancer models. Consistently, only RH30 cells displayed a high CYP1A1 expression in monolayer cultures among the RMS cell line panel, and noCYP2W1 expression could be identified. However, preliminary data suggests that CYP1A1 and CYP2W1enzymes might be restored when spheroids are generated from RMS cell lines. A functional 7-ethoxyresorufin-O-deethylase (EROD) assay has been validated to analyze CYP1A1- and CYP1B1-specific enzymatic activity in RMS cell lines. Vorinostat (SAHA) is an epigenetic drug currently used in the clinic for cancer treatment, and it has been associated with modulatory effects on CYP1expression. Our data suggests cell line-dependent synergistic or antagonistic effects of SAHA when combined with ICT2700, and these correlate with SAHA-induced CYP1A1 upregulation and downregulation respectively at both gene and protein levels. Conclusion: Evaluation of CYP target expression in RMS is being established to assess if duocarmycin bioprecursorscan be progressed as a novel type of therapy in RMS. Interestingly, CYP450 expression in RMS cell lines seems to vary depending on the complexity of the in vitro model used. Citation Format: Enric Arasanz Picher, Klaus Pors. Investigation of CYP450 enzymes as therapeutic targets in rhabdomyosarcoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2020.

Clinical characteristics and outcomes of adult alveolar rhabdomyosarcoma (ARMS) patients on front-line therapies: An MD Anderson Cancer Center (MDACC) case series.

e23548 Background: Rhabdomyosarcomas are the most common soft tissue sarcoma in children, and the prognosis of pediatric ARMS has improved with the use of multi-modality therapies. However, ARMS in adults is rare, and long-term outcomes continue to be poor. This study aimed to evaluate clinical outcomes of an adult ARMS population on different front-line systemic chemotherapies, particularly the vincristine/doxorubicin/ifosfamide (VDI) regimen. Methods: Adult fusion-positive confirmed ARMS patients over the age of 18 years (y) treated at MDACC from 2004 to 2018 were identified in our patient registry. Descriptive clinical statistics including staging, front-line chemotherapy, multimodal therapy usage, and survival analyses were performed. Results: 49 patients were identified, with mean age of 34.9 y (range 18y - 67y), and 53% were male. Most patients were white (53%, 26 pts), and the most common primary tumor site was the parameningeal space (63%; 31 pts). Patients were either intermediate (67%) or high clinical risk (33%). Most patients were IRSG clinical group IIIa (36%), IIIb (20%) or IV (33%) and were classified clinical stage 3 (49%) or 4 (33%). Of all patients at diagnosis, 71% had nodal disease and 32% were metastatic. Radiotherapy and surgery were given with upfront chemotherapy in 33 pts (67%) and 24 pts (49%) respectively, with 19 patients receiving both. Median OS for the entire cohort was 3.6 years. Doxorubicin containing chemotherapy regimens trended to worse OS than non-doxorubicin containing regimens (2.3 yrs vs 4.0 yrs, p = 0.355). Comparing patients who received VDI (19 pts) vs non-VDI (30 pts; 13 received Actinomycin D, 12 received doxorubicin in different regimens, and 5 received neither), median OS was 1.8 yrs vs 3.8 yrs (p = 0.283) respectively. There were similar number of front-line chemotherapy cycles (8.5 vs 9.5 cycles), high clinical risk (26% vs 37%) and metastatic disease (21% vs 36%) in the VDI vs non-VDI cohorts. Patients receiving upfront radiation had improved survival (3.7 vs 1.5 yr, p = 0.01), but this is likely confounded by those with metastases being less likely to receive upfront radiation. Conclusions: In this single center retrospective analysis of adult ARMS patients, survival outcomes continue to be similar to historical outcomes. There was no statistically significant OS difference in patients who did or did not receive doxorubicin containing front-line chemotherapy regimens, or in particular VDI therapy, although there was a trend to decreased OS. However, limitations to this study include limited sample size, non-randomization to treatment selection, and possible biases in patient selection for different chemo regimens. Based on these observations, randomized prospective studies are necessary to delineate which frontline chemotherapy regimen is most beneficial in this rare tumor in adults.

A phase 1 dose-escalation/expansion clinical trial of mocetinostat in combination with vinorelbine in adolescents and young adults with refractory and/or recurrent rhabdomyosarcoma: Interim results.

11553 Background: Rhabdomyosarcoma(RMS) is the most common soft tissue sarcoma in children. Relapsed/refractory(R/R) RMS has a poor outcome and remains an area of unmet need. Histone deacetylase (HDAC) inhibitors have been shown to have activity in preclinical models of RMS. Mocetinostat (Mirati Pharmaceuticals) is an investigational oral HDAC inhibitor, that targets HDACs 1, 2, 3 and 11. Mocetinostat displayed high activity in in vitro RMS cell lines, RMS xenograft models and exerted synergistic activity in combination with vinorelbine. We report early interim results of the Phase 1 trial of mocetinostat with vinorelbine in R/R RMS. Methods: An investigator initiated Phase 1,open-label, dose escalation/expansion clinical trial. A modified intent to treat approach is used for efficacy analysis for a target accrual of 20 subjects in the dose expansion cohort. Subjects with R/R RMS were age ≥ 18 years old(yo) for the phase 1 dose escalation cohort and ≥12 yo for the phase 1 dose expansion. Mocetinostat 40mg, 70mg or 90mg was taken orally 3 times weekly with vinorelbine 25mg/m2 IV on day 1,8, and 15 in 21 day cycles. Maximum tolerated dose of mocetinostat in the dose escalation phase was 40mg, which was selected for dose expansion. Subjects were treated until disease progression by RECIST 1.1 or unacceptable toxicity. Results: A total of 8(6 FOXO translocation(+), 1 (FOXO-), and 1 unknown) have been enrolled at time of submission. 5 in dose escalation cohort, and 3 in dose expansion. Median age was 19 yo(range 16-63), Median prior treatment regimens were 2(range 1-4). All patients had measurable metastatic disease. 6 of 8 subjects had prior exposure to vinorelbine in prior salvage chemotherapy or maintenance chemotherapy. As of 20JAN2022 safety cutoff, most common AEs (all grades) observed in 7 evaluable treated patients include neutropenia (n = 5), anemia(n = 5), and nausea(n = 4). The only grade 3 or 4 treatment related AEs were neutropenia, lymphopenia and anemia. Myelosuppression was transient, reversible and responsive to growth factors. No SAEs related to mocetinostat and/or vinorelbine have been reported. As of efficacy cutoff 20JAN2022, 7 of 8 patients are evaluable for response. 4 subjects had a partial response (PR), 2 subjects had stable disease (SD) and 1 subject had progressive disease for a clinical benefit rate of 86% (CR+PR+SD). Rapid responses were seen in the majority of patients at median of 1.5 months(mos). Of the 6 responders, 4 had duration of responses (DOR) &gt; 6 mo with a median DOR of 8 mos (range 4-16mo). Conclusions: In this interim analysis, Mocetinostat plus vinorelbine shows high efficacy and acceptable safety profile in this heavily pretreated group of R/R RMS patients. This study is open to accrual and enrollment is ongoing. Clinical trial information: NCT04299113.

Optimizing Rhabdomyosarcoma Treatment in Adolescents and Young Adults.

Simple SummaryDue to their rarity, we still know comparatively little about rhabdomyosarcomas (RMS) in adolescents and young adults (AYA). Factors responsible for the significantly poor outcomes in AYA-RMS are tumor biology, physiological factors specific to the age group concerned, refractoriness to multimodal treatments, and various psychosocial and medical care issues. The present review aims to examine these issues and offers possible solutions based on integrating new developments and findings with older, fundamental evidence regarding pediatric RMS.Rhabdomyosarcoma (RMS) is the most common form of soft tissue sarcoma in children, but can also develop in adolescents and young adults (AYA). The mainstay of treatment is multi-agent chemotherapy, ideally with concomitant local treatment, including surgical resection and/or radiation therapy. Although most treatment decisions for RMS in AYA are based on scientific evidence accumulated through clinical studies of pediatric RMS, treatment outcomes are significantly inferior in AYA patients than in children. Factors responsible for the significantly poor outcomes in AYA are tumor biology, the physiology specific to the age group concerned, refractoriness to multimodal treatments, and various psychosocial and medical care issues. The present review aims to examine the various issues involved in the treatment and care of AYA patients with RMS, discuss possible solutions, and provide an overview of the literature on the topic with several observations from the author’s own experience. Clinical trials for RMS in AYA are the best way to develop an optimal treatment. However, a well-designed clinical trial requires a great deal of time and resources, especially when targeting such a rare population. Until clinical trials are designed and implemented, and their findings duly analyzed, we must provide the best possible practice for RMS treatment in AYA patients based on our own expertise in manipulating the dosage schedules of various chemotherapeutic agents and administering local treatments in a manner appropriate for each patient. Precision medicine based on state-of-the-art cancer genomics will also form an integral part of this personalized approach. In the current situation, the only way to realize such a holistic treatment approach is to integrate new developments and findings, such as gene-based diagnostics and treatments, with older, fundamental evidence that can be selectively applied to individual cases.

Utilizing mpx:gfp transgenic zebrafish to assess eya3’s potential effects on neutrophils

Rhabdomyosarcoma (RMS) is a malignant cancer that develops in children’s muscles. Although RMS is one of the most common soft tissue sarcomas in children (NIH, 2020), there are a limited number of treatment options. As a potential new treatment target, we are investigating the roles of eya3, a transcriptional regulator, in normal muscle development. Previous research has shown a connection between eya3 and the innate immune system (Liu, 2012; Okabe, 2009), sometimes indicating eya3 is involved in innate immune responses, other times demonstrating that eya3 inhibits immune responses (Vartuli, 2018). Innate immune cells include neutrophils, which express mpx or myeloid‐specific peroxidase. To investigate how eya3 affects neutrophil location and/or levels in developing embryos, we utilized mpx:gfp transgenic zebrafish to visualize green fluorescent neutrophils via fluorescent imaging. After injecting eya3 morpholinos into wildtype zebrafish embryos to decrease eya3 expression, we used quantitative PCR (qPCR) to quantify the amount of neutrophils present. While our fluorescent imaging confirmed the presence of green fluorescent neutrophils in the mpx:gfp zebrafish at 96 hours post fertilization (hpf), we have yet to image neutrophils in our experimental group at the same timepoint. Our qPCR results suggest a decrease in mpx expression at 24 hpf with eya3 knockdown. We plan to confirm the potential decrease in neutrophil levels following eya3 knockdown with visualization by fluorescent microscopy. It will be interesting to determine if neutrophil levels are decreased in all locations, or depleted only from some locations in the zebrafish.

Biliary Tract Rhabdomyosarcoma: A Challenging Diagnosis

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in children. It is located mainly in the head and neck, extremities and trunk, rarely achieving the biliary tract (1%). The clinical presentation is characterized by obstructive jaundice, abdominal pain and palpable mass. The prognosis is favourable when being early diagnosed. The main differential diagnosis is the choledochal cyst. We bring a case report about a patient with biliary tract rhabdomyosarcoma, diagnosed and treated for 1 year as choledochal cyst, presenting obstructive jaundice for one year. A hepatic biopsy was done, showing hepatic fibrosis. He presented clinical worsening and was referred for liver transplant. An exploratory laparotomy was done before planning the transplant, which showed a biliary tract mass, followed by a biopsy. The anatomopathological result was RMS. Biliary tract RMS is a rare entity that should be included in the differential diagnosis of obstructive jaundice in children. Because of its similarity with choledochal cyst, pediatricians and surgeons should consider this hypothesis, for adequate investigation and treatment.

Rhabdomyosarcoma Presenting as Metastatic Abdominal or Chest Lumps: Report of 2 Cases

Rhabdomyosarcoma is a malignant mesenchymal tumor with skeletal muscle differentiation. It is the most common soft tissue sarcoma in children and adolescents. Its incidence is 4.5 cases per 1 million people aged 0-20 year. Embryonal, alveolar, pleomorphic and sclerosing /spindle cell types are the major variants of rhabdomyosarcoma. Embryonal rhabdomyosarcoma is the most common type. Rhabdomyosarcoma is highly aggressive and usually patient presented with pressure symptoms or metastasis. We are reporting 2 cases of rhabdomyosarcoma with one patient presented with abdominal wall nodules and other patient with bilateral breast lumps.

Dipyridophenazine iridium(III) complex as a phototoxic cancer stem cell selective, mitochondria targeting agent

In this work, the mechanism underlying the anticancer activity of a photoactivatable Ir(III) compound of the type [Ir(C^N)2(dppz)][PF6] where C^N = 1-methyl-2-(2′-thienyl)benzimidazole (complex 1) was investigated. Complex 1 photoactivated by visible light shows potent activity against highly aggressive and poorly treatable Rhabdomyosarcoma (RD) cells, the most frequent soft tissue sarcomas of children. This remarkable activity of 1 was observed not only in RD cells cultured in 2D monolayers but, more importantly, also in 3D spheroids, which resemble in many aspects solid tumors and serve as a promising model to mimic the in vivo situation. Importantly, photoactivated 1 kills not only differentiated RD cells but also even more effectively cancer stem cells (CSCs) of RD. One of the factors responsible for the activity of irradiated 1 in RD CSCs is its ability to produce ROS in these cells more effectively than in differentiated RD cells. Moreover, photoactivated 1 caused in RD differentiated cells and CSCs a significant decrease of mitochondrial membrane potential and promotes opening mitochondrial permeability transition pores in these cells, a mechanism that has never been demonstrated for any other metal-based anticancer complex. The results of this work give evidence that 1 has a potential for further evaluation using in vivo models as a promising chemotherapeutic agent for photodynamic therapy of hardly treatable human Rhabdomyosarcoma, particularly for its activity in both stem and differentiated cancer cells.