Genomics of rhabdomyosarcoma

Embryonal rhabdosarcoma: Characterized by loss of heterozygosity at 11p15 and by a high frequency of variants in genes in the RAS pathway. For the purposes of this section, patients with embryonal rhabdomyosarcoma are considered negative for PAX3::FOXO1 and PAX7::FOXO1 gene fusions (i.e., fusion-negative rhabdomyosarcoma).

Alveolar rhabdomyosarcoma: Characterized by gene fusions involving FOXO1 with either PAX3 or PAX7 (i.e., FOXO1 fusion–positive rhabdomyosarcoma). Cases with alveolar rhabdomyosarcoma histology without FOXO1 gene fusions have clinical behavior, gene alteration patterns, and transcriptomic profiles like cases with embryonal rhabdomyosarcoma. Therefore, the discussion below focuses only on alveolar rhabdomyosarcoma with FOXO1 gene fusions.[49-53]

Spindle cell/sclerosing rhabdomyosarcoma: Characterized by variants of MYOD1 in older patients and by VGLL2 and NCOA2 gene rearrangements in young children.

Pleomorphic rhabdomyosarcoma: Characterized by complex karyotypes with numerical and unbalanced structural changes that are indistinguishable from those of undifferentiated pleomorphic sarcomas.

Fusion-negative rhabdomyosarcoma (embryonal histology): Embryonal rhabdomyosarcoma tumors often show loss of heterozygosity at 11p15 and gains on chromosome 8.[55-58] Embryonal tumors have a higher background variant rate and a higher single-nucleotide variant rate than do alveolar rhabdomyosarcoma tumors, and the number of somatic variants increases with older age at diagnosis.[58,59] The most common recurring variants include those in the RAS pathway (e.g., NRAS, KRAS, HRAS, and NF1), which together are observed in approximately one-half of cases.[54] Variants in NRAS are the most frequent RAS pathway gene variants beyond infancy, while variants in HRAS predominate during infancy.[54] The presence of a RAS variant does not confer prognostic significance. Among the RAS pathway genes, germline pathogenic variants in NF1 and HRAS predispose to rhabdomyosarcoma. In a study of 615 children with rhabdomyosarcoma, 347 had tumors with embryonal histology. Of these, nine patients had NF1 germline pathogenic variants, and five patients had HRAS germline pathogenic variants, representing 2.6% and 1.4% of embryonal histology cases, respectively.[60]Other genes with recurring variants in fusion-negative rhabdomyosarcoma tumors include FGFR4, PIK3CA, CTNNB1, FBXW7, and BCOR, all of which are present in fewer than 15% of cases.[54,58,59]TP53 variants:TP53 variants are observed in 10% to 15% of patients with fusion-negative rhabdomyosarcoma and occur less commonly (about 4%) in patients with alveolar rhabdomyosarcoma.[54] In other childhood cancers (e.g., Wilms tumor), TP53 variants are associated with anaplastic histology,[61] and the same is true for embryonal rhabdomyosarcoma. In a study of 146 rhabdomyosarcoma patients with known TP53 status, approximately two-thirds of tumors with TP53 variants showed anaplasia (69%), but only one-quarter of tumors with anaplasia had TP53 variants.[62]The presence of TP53 variants was associated with reduced EFS in both nonrisk-stratified and risk-stratified analyses for both a COG and a U.K. rhabdomyosarcoma cohort.[54] The poor prognosis associated with TP53 variants was observed for both embryonal and alveolar patients. Based on these results, the COG plans to consider TP53 variant as a high-risk defining characteristic in its upcoming trials.[63]Rhabdomyosarcoma is one of the childhood cancers associated with Li-Fraumeni syndrome. In a study of 614 pediatric patients with rhabdomyosarcoma, 11 patients (1.7%) had TP53 germline pathogenic variants. Variants were less common in patients with alveolar histology (0.6%), compared with patients with nonalveolar histologies (2.2%).[60] Rhabdomyosarcoma with nonalveolar anaplastic morphology may be a presenting feature for children with Li-Fraumeni syndrome and germline TP53 variants.[64] DICER1 variants in embryonal rhabdomyosarcoma:DICER1 variants are observed in a small subset of patients with embryonal rhabdomyosarcoma, most commonly arising in tumors of the female genitourinary tract.[54] More specifically, most cases of cervical embryonal rhabdomyosarcoma,[66-68] which most commonly occurs in adolescents and young adults,[69,70] have DICER1 variants. In contrast, DICER1 variants are rarely observed in patients with vaginal primary sites, an entity occurring primarily in girls younger than 2 or 3 years.[67,69] DICER1 variants are also common in embryonal rhabdomyosarcoma arising in the uterine corpus, but this presentation is primarily observed in adults.[67,71] Cervical rhabdomyosarcoma generally shows a sarcoma botryoides histological pattern, and many cases show areas of cartilaginous differentiation, a feature also observed in other tumor types with DICER1 variants.[69,70,72] In support of the distinctive biology of embryonal rhabdomyosarcoma with DICER1 variants, these cases have a DNA methylation pattern that is distinctive from that of other embryonal rhabdomyosarcoma cases.[68] A diagnosis of cervical rhabdomyosarcoma is an indication for genetic testing for DICER1 syndrome.[67,73]

Fusion-negative rhabdomyosarcoma (embryonal histology): Embryonal rhabdomyosarcoma tumors often show loss of heterozygosity at 11p15 and gains on chromosome 8.[55-58] Embryonal tumors have a higher background variant rate and a higher single-nucleotide variant rate than do alveolar rhabdomyosarcoma tumors, and the number of somatic variants increases with older age at diagnosis.[58,59] The most common recurring variants include those in the RAS pathway (e.g., NRAS, KRAS, HRAS, and NF1), which together are observed in approximately one-half of cases.[54] Variants in NRAS are the most frequent RAS pathway gene variants beyond infancy, while variants in HRAS predominate during infancy.[54] The presence of a RAS variant does not confer prognostic significance. Among the RAS pathway genes, germline pathogenic variants in NF1 and HRAS predispose to rhabdomyosarcoma. In a study of 615 children with rhabdomyosarcoma, 347 had tumors with embryonal histology. Of these, nine patients had NF1 germline pathogenic variants, and five patients had HRAS germline pathogenic variants, representing 2.6% and 1.4% of embryonal histology cases, respectively.[60]Other genes with recurring variants in fusion-negative rhabdomyosarcoma tumors include FGFR4, PIK3CA, CTNNB1, FBXW7, and BCOR, all of which are present in fewer than 15% of cases.[54,58,59]TP53 variants:TP53 variants are observed in 10% to 15% of patients with fusion-negative rhabdomyosarcoma and occur less commonly (about 4%) in patients with alveolar rhabdomyosarcoma.[54] In other childhood cancers (e.g., Wilms tumor), TP53 variants are associated with anaplastic histology,[61] and the same is true for embryonal rhabdomyosarcoma. In a study of 146 rhabdomyosarcoma patients with known TP53 status, approximately two-thirds of tumors with TP53 variants showed anaplasia (69%), but only one-quarter of tumors with anaplasia had TP53 variants.[62]The presence of TP53 variants was associated with reduced EFS in both nonrisk-stratified and risk-stratified analyses for both a COG and a U.K. rhabdomyosarcoma cohort.[54] The poor prognosis associated with TP53 variants was observed for both embryonal and alveolar patients. Based on these results, the COG plans to consider TP53 variant as a high-risk defining characteristic in its upcoming trials.[63]Rhabdomyosarcoma is one of the childhood cancers associated with Li-Fraumeni syndrome. In a study of 614 pediatric patients with rhabdomyosarcoma, 11 patients (1.7%) had TP53 germline pathogenic variants. Variants were less common in patients with alveolar histology (0.6%), compared with patients with nonalveolar histologies (2.2%).[60] Rhabdomyosarcoma with nonalveolar anaplastic morphology may be a presenting feature for children with Li-Fraumeni syndrome and germline TP53 variants.[64] DICER1 variants in embryonal rhabdomyosarcoma:DICER1 variants are observed in a small subset of patients with embryonal rhabdomyosarcoma, most commonly arising in tumors of the female genitourinary tract.[54] More specifically, most cases of cervical embryonal rhabdomyosarcoma,[66-68] which most commonly occurs in adolescents and young adults,[69,70] have DICER1 variants. In contrast, DICER1 variants are rarely observed in patients with vaginal primary sites, an entity occurring primarily in girls younger than 2 or 3 years.[67,69] DICER1 variants are also common in embryonal rhabdomyosarcoma arising in the uterine corpus, but this presentation is primarily observed in adults.[67,71] Cervical rhabdomyosarcoma generally shows a sarcoma botryoides histological pattern, and many cases show areas of cartilaginous differentiation, a feature also observed in other tumor types with DICER1 variants.[69,70,72] In support of the distinctive biology of embryonal rhabdomyosarcoma with DICER1 variants, these cases have a DNA methylation pattern that is distinctive from that of other embryonal rhabdomyosarcoma cases.[68] A diagnosis of cervical rhabdomyosarcoma is an indication for genetic testing for DICER1 syndrome.[67,73]

Among eight consecutively presenting children with rhabdomyosarcoma and TP53 germline pathogenic variants, all showed anaplastic morphology. Among an additional seven children with anaplastic rhabdomyosarcoma and unknown TP53 germline variant status, three of the seven children had functionally relevant TP53 germline pathogenic variants. The median age at diagnosis of the 11 children with TP53 germline pathogenic variant status was 40 months (range, 19–67 months).[64]

Fusion-negative rhabdomyosarcoma (embryonal histology): Embryonal rhabdomyosarcoma tumors often show loss of heterozygosity at 11p15 and gains on chromosome 8.[55-58] Embryonal tumors have a higher background variant rate and a higher single-nucleotide variant rate than do alveolar rhabdomyosarcoma tumors, and the number of somatic variants increases with older age at diagnosis.[58,59] The most common recurring variants include those in the RAS pathway (e.g., NRAS, KRAS, HRAS, and NF1), which together are observed in approximately one-half of cases.[54] Variants in NRAS are the most frequent RAS pathway gene variants beyond infancy, while variants in HRAS predominate during infancy.[54] The presence of a RAS variant does not confer prognostic significance. Among the RAS pathway genes, germline pathogenic variants in NF1 and HRAS predispose to rhabdomyosarcoma. In a study of 615 children with rhabdomyosarcoma, 347 had tumors with embryonal histology. Of these, nine patients had NF1 germline pathogenic variants, and five patients had HRAS germline pathogenic variants, representing 2.6% and 1.4% of embryonal histology cases, respectively.[60]Other genes with recurring variants in fusion-negative rhabdomyosarcoma tumors include FGFR4, PIK3CA, CTNNB1, FBXW7, and BCOR, all of which are present in fewer than 15% of cases.[54,58,59]TP53 variants:TP53 variants are observed in 10% to 15% of patients with fusion-negative rhabdomyosarcoma and occur less commonly (about 4%) in patients with alveolar rhabdomyosarcoma.[54] In other childhood cancers (e.g., Wilms tumor), TP53 variants are associated with anaplastic histology,[61] and the same is true for embryonal rhabdomyosarcoma. In a study of 146 rhabdomyosarcoma patients with known TP53 status, approximately two-thirds of tumors with TP53 variants showed anaplasia (69%), but only one-quarter of tumors with anaplasia had TP53 variants.[62]The presence of TP53 variants was associated with reduced EFS in both nonrisk-stratified and risk-stratified analyses for both a COG and a U.K. rhabdomyosarcoma cohort.[54] The poor prognosis associated with TP53 variants was observed for both embryonal and alveolar patients. Based on these results, the COG plans to consider TP53 variant as a high-risk defining characteristic in its upcoming trials.[63]Rhabdomyosarcoma is one of the childhood cancers associated with Li-Fraumeni syndrome. In a study of 614 pediatric patients with rhabdomyosarcoma, 11 patients (1.7%) had TP53 germline pathogenic variants. Variants were less common in patients with alveolar histology (0.6%), compared with patients with nonalveolar histologies (2.2%).[60] Rhabdomyosarcoma with nonalveolar anaplastic morphology may be a presenting feature for children with Li-Fraumeni syndrome and germline TP53 variants.[64] DICER1 variants in embryonal rhabdomyosarcoma:DICER1 variants are observed in a small subset of patients with embryonal rhabdomyosarcoma, most commonly arising in tumors of the female genitourinary tract.[54] More specifically, most cases of cervical embryonal rhabdomyosarcoma,[66-68] which most commonly occurs in adolescents and young adults,[69,70] have DICER1 variants. In contrast, DICER1 variants are rarely observed in patients with vaginal primary sites, an entity occurring primarily in girls younger than 2 or 3 years.[67,69] DICER1 variants are also common in embryonal rhabdomyosarcoma arising in the uterine corpus, but this presentation is primarily observed in adults.[67,71] Cervical rhabdomyosarcoma generally shows a sarcoma botryoides histological pattern, and many cases show areas of cartilaginous differentiation, a feature also observed in other tumor types with DICER1 variants.[69,70,72] In support of the distinctive biology of embryonal rhabdomyosarcoma with DICER1 variants, these cases have a DNA methylation pattern that is distinctive from that of other embryonal rhabdomyosarcoma cases.[68] A diagnosis of cervical rhabdomyosarcoma is an indication for genetic testing for DICER1 syndrome.[67,73]

In another series, 26 of 31 patients with germline TP53 pathogenic variants had tumors with embryonal histology. Of the 16 tumors that were submitted for central pathology review, 12 had focal or diffuse anaplasia. The median age of patients in this group was 2.3 years.[65]

Fusion-positive rhabdomyosarcoma (alveolar histology): About 70% to 80% of alveolar tumors are characterized by translocations between the FOXO1 gene on chromosome 13 and either the PAX3 gene on chromosome 2 (t(2;13)(q35;q14)) or the PAX7 gene on chromosome 1 (t(1;13)(p36;q14)).[55,74,75] Other rare fusions include PAX3::NCOA1 and PAX3::INO80D.[58] Translocations involving the PAX3 gene occur in approximately 60% of alveolar rhabdomyosarcoma cases, while the PAX7 gene appears to be involved in about 20% of cases.[74] Patients with solid-variant alveolar histology have a lower incidence of PAX::FOXO1 gene fusions than do patients showing classical alveolar histology.[76] The alveolar histology that is associated with the PAX7 gene in patients with or without metastatic disease appears to occur at a younger age and may be associated with longer EFS rates than those associated with PAX3 gene rearrangements.[77-82] Patients with alveolar histology and the PAX3 gene are older and have a higher incidence of invasive tumor (T2). Around 20% of cases showing alveolar histology have no detectable PAX gene translocation.[50,76] These patients have clinical behaviors, gene alteration patterns, and transcriptomic profiles that align with patients who have embryonal rhabdomyosarcoma and are now classified together with embryonal rhabdomyosarcoma, as fusion-negative rhabdomyosarcoma.[49-53]For the diagnosis of alveolar rhabdomyosarcoma, a FOXO1 gene rearrangement may be detected with good sensitivity and specificity using either fluorescence in situ hybridization or reverse transcription–polymerase chain reaction.[83] In addition to FOXO1 rearrangements, alveolar tumors are characterized by a lower mutational burden than are fusion-negative tumors, with fewer genes having recurring mutations.[58,59] The most frequently observed alterations in fusion-positive tumors are focal amplification of CDK4 (13%) or MYCN (10%), with small numbers of patients having recurring mutations in other genes (e.g., BCOR, 6%; NF1, 4%; TP53, 4%; and PIK3CA, 2%).[54] TP53 mutations in alveolar rhabdomyosarcoma appear to connote a high risk of treatment failure.[54]

Spindle cell/sclerosing histology: Spindle cell/sclerosing rhabdomyosarcoma has been proposed as a separate entity in the WHO Classification of Tumors of Soft Tissue and Bone.[84] Within the spindle cell/sclerosing rhabdomyosarcoma category, several entities have distinctive molecular and clinical characteristics, described below.Congenital/infantile spindle cell rhabdomyosarcoma: Several reports have described cases of congenital or infantile spindle cell rhabdomyosarcoma with gene fusions involving VGLL2 and NCOA2 (e.g., VGLL2::CITED2, TEAD1::NCOA2, VGLL2::NCOA2, SRF::NCOA2).[85,86]MYOD1-altered spindle cell/sclerosing rhabdomyosarcoma: In older children and adults with spindle cell/sclerosing rhabdomyosarcoma, a specific MYOD1 variant (p.L122R) has been observed in a large proportion of patients.[85,89-91] In the combined cohort of COG and U.K. rhabdomyosarcoma patients (n = 641), variants in MYOD1 were found in 3% (17 of 515) of all fusion-negative rhabdomyosarcoma cases and in no fusion-positive cases. The presenting age of patients with MYOD1 variants was 10.8 years.[54] Most cases in this cohort showed spindle or sclerosing features, but cases with densely packed cells that mimicked the dense pattern of embryonal rhabdomyosarcoma were also observed. Most cases in this cohort (15 of 17, 88%) had either head and neck or parameningeal region primary sites. Activating PIK3CA variants are seen in about one-half of cases with MYOD1 variants.[54,92] The presence of the MYOD1 variant is associated with a markedly increased risk of local and distant failure.[54,85,89,90] Intraosseous spindle cell rhabdomyosarcoma: Primary intraosseous rhabdomyosarcoma is a very uncommon presentation for rhabdomyosarcoma. Most cases present with gene rearrangements involving TFCP2, with either FUS or EWSR1.[93-97] Rhabdomyosarcoma with a FUS::TFCP2 or EWSR1::TFCP2 gene fusion most commonly presents in young adults, although cases in older children and adolescents have been reported.[93,96,97] Craniofacial bones are the most common primary tumor location, and positivity for ALK and cytokeratins by immunohistochemistry is commonly observed. Other characteristics of this entity include a complex genomic profile, with most cases showing deletion of the CDKN2A tumor suppressor gene.[96] Intraosseous spindle cell rhabdomyosarcoma with a FUS::TFCP2 or EWSR1::TFCP2 gene fusion shows an aggressive clinical course. In one study, the median OS was only 8 months.[96]

Spindle cell/sclerosing histology: Spindle cell/sclerosing rhabdomyosarcoma has been proposed as a separate entity in the WHO Classification of Tumors of Soft Tissue and Bone.[84] Within the spindle cell/sclerosing rhabdomyosarcoma category, several entities have distinctive molecular and clinical characteristics, described below.Congenital/infantile spindle cell rhabdomyosarcoma: Several reports have described cases of congenital or infantile spindle cell rhabdomyosarcoma with gene fusions involving VGLL2 and NCOA2 (e.g., VGLL2::CITED2, TEAD1::NCOA2, VGLL2::NCOA2, SRF::NCOA2).[85,86]MYOD1-altered spindle cell/sclerosing rhabdomyosarcoma: In older children and adults with spindle cell/sclerosing rhabdomyosarcoma, a specific MYOD1 variant (p.L122R) has been observed in a large proportion of patients.[85,89-91] In the combined cohort of COG and U.K. rhabdomyosarcoma patients (n = 641), variants in MYOD1 were found in 3% (17 of 515) of all fusion-negative rhabdomyosarcoma cases and in no fusion-positive cases. The presenting age of patients with MYOD1 variants was 10.8 years.[54] Most cases in this cohort showed spindle or sclerosing features, but cases with densely packed cells that mimicked the dense pattern of embryonal rhabdomyosarcoma were also observed. Most cases in this cohort (15 of 17, 88%) had either head and neck or parameningeal region primary sites. Activating PIK3CA variants are seen in about one-half of cases with MYOD1 variants.[54,92] The presence of the MYOD1 variant is associated with a markedly increased risk of local and distant failure.[54,85,89,90] Intraosseous spindle cell rhabdomyosarcoma: Primary intraosseous rhabdomyosarcoma is a very uncommon presentation for rhabdomyosarcoma. Most cases present with gene rearrangements involving TFCP2, with either FUS or EWSR1.[93-97] Rhabdomyosarcoma with a FUS::TFCP2 or EWSR1::TFCP2 gene fusion most commonly presents in young adults, although cases in older children and adolescents have been reported.[93,96,97] Craniofacial bones are the most common primary tumor location, and positivity for ALK and cytokeratins by immunohistochemistry is commonly observed. Other characteristics of this entity include a complex genomic profile, with most cases showing deletion of the CDKN2A tumor suppressor gene.[96] Intraosseous spindle cell rhabdomyosarcoma with a FUS::TFCP2 or EWSR1::TFCP2 gene fusion shows an aggressive clinical course. In one study, the median OS was only 8 months.[96]

For congenital/infantile spindle cell rhabdomyosarcoma, a study reported that 10 of 11 patients showed recurrent fusion genes. Most of these patients had truncal primary tumors, and there were no paratesticular tumors. Novel VGLL2 rearrangements were observed in seven patients (63%), including the VGLL2::CITED2 fusion in four patients and the VGLL2::NCOA2 fusion in two patients.[85] Three patients (27%) harbored different NCOA2 gene fusions, including TEAD1::NCOA2 in two patients and SRF::NCOA2 in one patient. In this report, all fusion-positive congenital/infantile spindle cell rhabdomyosarcoma patients with long-term follow-up data were alive and well, and no patients developed distant metastases.[85]

Spindle cell/sclerosing histology: Spindle cell/sclerosing rhabdomyosarcoma has been proposed as a separate entity in the WHO Classification of Tumors of Soft Tissue and Bone.[84] Within the spindle cell/sclerosing rhabdomyosarcoma category, several entities have distinctive molecular and clinical characteristics, described below.Congenital/infantile spindle cell rhabdomyosarcoma: Several reports have described cases of congenital or infantile spindle cell rhabdomyosarcoma with gene fusions involving VGLL2 and NCOA2 (e.g., VGLL2::CITED2, TEAD1::NCOA2, VGLL2::NCOA2, SRF::NCOA2).[85,86]MYOD1-altered spindle cell/sclerosing rhabdomyosarcoma: In older children and adults with spindle cell/sclerosing rhabdomyosarcoma, a specific MYOD1 variant (p.L122R) has been observed in a large proportion of patients.[85,89-91] In the combined cohort of COG and U.K. rhabdomyosarcoma patients (n = 641), variants in MYOD1 were found in 3% (17 of 515) of all fusion-negative rhabdomyosarcoma cases and in no fusion-positive cases. The presenting age of patients with MYOD1 variants was 10.8 years.[54] Most cases in this cohort showed spindle or sclerosing features, but cases with densely packed cells that mimicked the dense pattern of embryonal rhabdomyosarcoma were also observed. Most cases in this cohort (15 of 17, 88%) had either head and neck or parameningeal region primary sites. Activating PIK3CA variants are seen in about one-half of cases with MYOD1 variants.[54,92] The presence of the MYOD1 variant is associated with a markedly increased risk of local and distant failure.[54,85,89,90] Intraosseous spindle cell rhabdomyosarcoma: Primary intraosseous rhabdomyosarcoma is a very uncommon presentation for rhabdomyosarcoma. Most cases present with gene rearrangements involving TFCP2, with either FUS or EWSR1.[93-97] Rhabdomyosarcoma with a FUS::TFCP2 or EWSR1::TFCP2 gene fusion most commonly presents in young adults, although cases in older children and adolescents have been reported.[93,96,97] Craniofacial bones are the most common primary tumor location, and positivity for ALK and cytokeratins by immunohistochemistry is commonly observed. Other characteristics of this entity include a complex genomic profile, with most cases showing deletion of the CDKN2A tumor suppressor gene.[96] Intraosseous spindle cell rhabdomyosarcoma with a FUS::TFCP2 or EWSR1::TFCP2 gene fusion shows an aggressive clinical course. In one study, the median OS was only 8 months.[96]

While most studies of congenital/infantile spindle cell rhabdomyosarcoma have shown favorable outcomes, it was reported that four patients developed metastatic disease and two patients had fatal outcomes. Disease progression occurred a median of 3.5 years from diagnosis (range, 1–8 years).[87] All four patients had unresectable tumors and were treated with chemotherapy. However, most literature reported cases in which surgical resection was achieved. At disease progression, a tumor from one patient had a TP53 variant, and a tumor from another patient showed a homozygous CDKN2A and CDKN2B deletion.

Spindle cell/sclerosing histology: Spindle cell/sclerosing rhabdomyosarcoma has been proposed as a separate entity in the WHO Classification of Tumors of Soft Tissue and Bone.[84] Within the spindle cell/sclerosing rhabdomyosarcoma category, several entities have distinctive molecular and clinical characteristics, described below.Congenital/infantile spindle cell rhabdomyosarcoma: Several reports have described cases of congenital or infantile spindle cell rhabdomyosarcoma with gene fusions involving VGLL2 and NCOA2 (e.g., VGLL2::CITED2, TEAD1::NCOA2, VGLL2::NCOA2, SRF::NCOA2).[85,86]MYOD1-altered spindle cell/sclerosing rhabdomyosarcoma: In older children and adults with spindle cell/sclerosing rhabdomyosarcoma, a specific MYOD1 variant (p.L122R) has been observed in a large proportion of patients.[85,89-91] In the combined cohort of COG and U.K. rhabdomyosarcoma patients (n = 641), variants in MYOD1 were found in 3% (17 of 515) of all fusion-negative rhabdomyosarcoma cases and in no fusion-positive cases. The presenting age of patients with MYOD1 variants was 10.8 years.[54] Most cases in this cohort showed spindle or sclerosing features, but cases with densely packed cells that mimicked the dense pattern of embryonal rhabdomyosarcoma were also observed. Most cases in this cohort (15 of 17, 88%) had either head and neck or parameningeal region primary sites. Activating PIK3CA variants are seen in about one-half of cases with MYOD1 variants.[54,92] The presence of the MYOD1 variant is associated with a markedly increased risk of local and distant failure.[54,85,89,90] Intraosseous spindle cell rhabdomyosarcoma: Primary intraosseous rhabdomyosarcoma is a very uncommon presentation for rhabdomyosarcoma. Most cases present with gene rearrangements involving TFCP2, with either FUS or EWSR1.[93-97] Rhabdomyosarcoma with a FUS::TFCP2 or EWSR1::TFCP2 gene fusion most commonly presents in young adults, although cases in older children and adolescents have been reported.[93,96,97] Craniofacial bones are the most common primary tumor location, and positivity for ALK and cytokeratins by immunohistochemistry is commonly observed. Other characteristics of this entity include a complex genomic profile, with most cases showing deletion of the CDKN2A tumor suppressor gene.[96] Intraosseous spindle cell rhabdomyosarcoma with a FUS::TFCP2 or EWSR1::TFCP2 gene fusion shows an aggressive clinical course. In one study, the median OS was only 8 months.[96]

A study of 40 patients with congenital/infantile spindle cell rhabdomyosarcoma (defined by diagnosis at age ≤12 months) found that almost all patients had localized disease (n = 39) and that one-half of patients who underwent molecular testing (13 of 26) had rearrangements of NCOA2 and/or VGLL2.[88] Because testing was limited to NCOA2 and VGLL2, it is possible that more comprehensive genomic analysis would identify a higher proportion of patients with relevant gene fusions. The 5-year EFS rate for the 13 patients with either a VGLL2 and/or a NCOA2 fusion was 90% (95% CI, ±19%), and the overall survival (OS) rate was 100% (95% CI, ±9%).

Spindle cell/sclerosing histology: Spindle cell/sclerosing rhabdomyosarcoma has been proposed as a separate entity in the WHO Classification of Tumors of Soft Tissue and Bone.[84] Within the spindle cell/sclerosing rhabdomyosarcoma category, several entities have distinctive molecular and clinical characteristics, described below.Congenital/infantile spindle cell rhabdomyosarcoma: Several reports have described cases of congenital or infantile spindle cell rhabdomyosarcoma with gene fusions involving VGLL2 and NCOA2 (e.g., VGLL2::CITED2, TEAD1::NCOA2, VGLL2::NCOA2, SRF::NCOA2).[85,86]MYOD1-altered spindle cell/sclerosing rhabdomyosarcoma: In older children and adults with spindle cell/sclerosing rhabdomyosarcoma, a specific MYOD1 variant (p.L122R) has been observed in a large proportion of patients.[85,89-91] In the combined cohort of COG and U.K. rhabdomyosarcoma patients (n = 641), variants in MYOD1 were found in 3% (17 of 515) of all fusion-negative rhabdomyosarcoma cases and in no fusion-positive cases. The presenting age of patients with MYOD1 variants was 10.8 years.[54] Most cases in this cohort showed spindle or sclerosing features, but cases with densely packed cells that mimicked the dense pattern of embryonal rhabdomyosarcoma were also observed. Most cases in this cohort (15 of 17, 88%) had either head and neck or parameningeal region primary sites. Activating PIK3CA variants are seen in about one-half of cases with MYOD1 variants.[54,92] The presence of the MYOD1 variant is associated with a markedly increased risk of local and distant failure.[54,85,89,90] Intraosseous spindle cell rhabdomyosarcoma: Primary intraosseous rhabdomyosarcoma is a very uncommon presentation for rhabdomyosarcoma. Most cases present with gene rearrangements involving TFCP2, with either FUS or EWSR1.[93-97] Rhabdomyosarcoma with a FUS::TFCP2 or EWSR1::TFCP2 gene fusion most commonly presents in young adults, although cases in older children and adolescents have been reported.[93,96,97] Craniofacial bones are the most common primary tumor location, and positivity for ALK and cytokeratins by immunohistochemistry is commonly observed. Other characteristics of this entity include a complex genomic profile, with most cases showing deletion of the CDKN2A tumor suppressor gene.[96] Intraosseous spindle cell rhabdomyosarcoma with a FUS::TFCP2 or EWSR1::TFCP2 gene fusion shows an aggressive clinical course. In one study, the median OS was only 8 months.[96]

Further study is needed to better define the prevalence and prognostic significance of gene rearrangements in VGLL2, NCOA2, and other relevant genes in young children with congenital/infantile spindle cell rhabdomyosarcoma.