SWI/SNF complex heterogeneity is related to polyphenotypic differentiation, prognosis, and immune response in rhabdoid tumors.

BACKGROUND: Rhabdoid tumors (RTs) arise within (atypical teratoid/rhabdoid tumor [AT/RT]) or outside the brain (extra [e]CNS-RT) and are driven mainly by inactivation of the SWItch/sucrose nonfermentable (SWI/SNF) complex subunit SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily B member 1 (SMARCB1). A pathognomonic hallmark of RTs is heterogeneous multilineage differentiation, including anomalous neuronal differentiation in some eCNS-RTs. Because remodeling of the SWI/SNF complex regulates differentiation, we hypothesized that SWI/SNF Brahma-associated factors (BAF) and polybromo-associated BAF (PBAF) complex heterogeneity are related to both multilineage differentiation and clinical outcome. METHODS: We performed an integrated analysis of SWI/SNF complex alterations in the developing kidney and cerebellum (most common regions of RT origin) in comparison to eCNS-RT (n = 14) and AT/RT (n = 25) tumors. RT samples were interrogated using immunohistochemistry, DNA methylation, and gene expression analyses. RESULTS: The SWI/SNF BAF paralogs actin-like protein (ACTL)6A and ACTL6B were expressed in a mutually exclusive manner in the developing cerebellum and kidney. In contrast, a subset of eCNS-RTs lost mutual exclusivity and coexpressed both subunits. These tumors showed aberrant DNA methylation of genes that regulate neuronal and renal development and demonstrated immunohistochemical evidence of neuronal differentiation. In addition, low expression of the PBAF subunit polybromo-1 (PBRM1) identified a group of AT/RTs in younger children with better overall prognosis. PBRM1-low AT/RT and eCNS-RTs showed altered DNA methylation and gene expression in immune-related genes. PBRM1 knockdown resulted in lowering immunosuppressive cytokines, and PBRM1 levels in tumor samples showed an inverse relationship with cluster of differentiation (CD)8 cytotoxic T-cell infiltration. CONCLUSIONS: Heterogeneity in SWI/SNF BAF (ACTL6A/ACTL6B) and PBAF (PBRM1) subunits is related to histogenesis, contributes to the immune microenvironment and prognosis in RTs, and may inform opportunities to develop immunotherapies.

Claudin-6 is a nonspecific marker for malignant rhabdoid and other pediatric tumors.

Claudins are tight junction proteins with claudin-6 (CLDN6) expression mostly restricted to embryonic and fetal life. Previously reported gene expression microarray analysis showed an increased level of CLDN6 in atypical teratoid rhabdoid tumors (AT/RT) compared with other central nervous system (CNS) tumors and sarcomas. However, there exist conflicting data on expression of CLDN6 as assessed by immunohistochemistry in CNS tumors. We established membranous staining as a specific and reproducible method for evaluating CLDN6 expression based on fetal and adolescent controls. We then evaluated a large group (257) of pediatric tumors using tissue microarrays, including: 47 malignant rhabdoid tumors (MRTs), (31 AT/RTs and 16 non-CNS MRTs); 67 small, round, blue cell tumors (10 Wilms tumors, 10 embryonal rhabdomyosarcomas, 10 neuroblastomas (NBs), 10 synovial sarcomas (SSs), 9 hepatoblastomas (HBs), 9 alveolar rhabdomyosarcomas, and 9 Ewings sarcomas); and 143 CNS tumors (24 medulloblastomas, 21 pilocytic astrocytomas, 14 astrocytomas grade II/III, 13 gangliogliomas, 12 glioblastomas, 12 ependymal tumors, 11 choroid plexus tumors, 10 meningiomas, 8 dysembryoplastic neuroepithelial tumors, 8 oligodendrogliomas, 4 craniopharyngiomas, 2 germinomas, 2 primitive neuroectodermal tumors (PNET), and 2 central neurocytomas). CLDN6 expression was seen in 12 of 31 (39%) AT/RTs, 7 of 16 (44%) non-CNS MRTs, 5 of 10 (50%) Wilms tumors, 1 of 9 (11%) HBs, 2 of 2 (100%) germinomas, 1 of 2 (50%) CNS PNETs, 1 of 24 (4%) medulloblastomas, and 1 of 10 (10%) meningiomas. Ten of 11 (91%) choroid plexus tumors showed apical staining but no concentric membranous staining. Although CLDN6 is expressed in both AT/RTs and MRTs, it is not a specific biomarker as it is expressed in a variety of other pediatric CNS and soft tissue tumors.

Malignant rhabdoid tumors express stem cell factors, which relate to the expression of EZH2 and Id proteins.

Malignant rhabdoid tumors (MRTs) are highly aggressive pediatric tumors associated with loss of expression of SMARCB1, commonly occurring in the central nervous system [referred to as atypical teratoid/rhabdoid tumors (AT/RTs)] and in the kidney and soft tissues. Histologically, MRTs are characterized by immunohistochemical evidence of primitive neuroectodermal, mesenchymal, and epithelial differentiation. The ability of MRTs to differentiate along multiple lines, as evidenced by both histologic features and polyphenotypic immunohistochemical staining, and the proliferative nature of MRT cells are characteristics shared with the self-renewal and plasticity of embryonic stem cells (ES). To test the hypothesis that MRTs share similarities with ES, we used immunohistochemistry to evaluate the expression of various stem cell markers in a tissue microarray containing 26 AT/RTs and 16 non-central nervous system MRTs (NCMRTs). Staining intensity was scored as negative (0), low (1+), moderate (2+), and strong (3+) and was multiplied by the percentage of positive tumor cells to establish a semiquantitative measure for each marker. In AT/RT, strong-to-low expression was noted with glypican-3 (20 of 26, 77%), Sall4 (23 of 26, 88%), T-cell leukemia/lymphoma 1 (25 of 26, 96%), and undifferentiated embryonic cell transcription factor 1 (19 of 26, 73%). Markers that showed low expression in AT/RT were Sox2 (8 of 26, 31%), Nanog (7 of 26, 27%), Klf4 (10 of 26, 38%), Zfp206 (5 of 26, 19%), and musashi-1 (21 of 26, 81%). Similarly, in NCMRT, expression was noted with glypican-3 (12 of 16, 75%), Sall4 (13 of 16, 81%), T-cell leukemia/lymphoma 1 (16 of 16, 100%), undifferentiated embryonic cell transcription factor 1 (12 of 16, 75%), Sox2 (5 of 16, 31%), Nanog (8 of 16, 50%), Klf4 (8 of 16, 50%), Zfp206 (13 of 16, 81%), and musashi-1 (11 of 16, 75%). Placental alkaline phosphatase, Oct4, c-KIT, CD30, α-fetoprotein, and ß- -human chorionic gonadotrophin were not expressed in all cases. Markers that regulate the expression of stem cell transcription factors were also expressed in MRT. AT/RT cases showed expression of Id proteins: Id1 (17 of 26, 65%), Id2 (24 of 26, 92%), Id3 (22 of 26, 85%), and Id4 (22 of 26, 85%). Low expression was observed with EZH2 (15 of 26, 58%). Similarly, NCMRT cases showed expression of Id1 (15 of 16, 94%), Id2 (16 of 16, 100%), Id3 (16 of 16, 100%), Id4 (13 of 16, 81%), and EZH2 (13 of 16, 81%). Finally, regression analysis revealed a significant relationship between the expression of stem cell markers and EZH2 (P<0.0001), Id1 (P=0.0087), Id2 (P=0.0002), Id3 (P=0.0033), and Id4 (P<0.0001). These data suggest that MRTs express many stem cell-associated transcription factors, which may be regulated by the expression of EZH2 and the Id family of proteins. This study underscores similarities between MRTs and stem cells and may help elucidate common biologic pathways that could serve in advancing more effective therapeutic strategies to treat MRTs.

p16INK4A and p14ARF tumor suppressor pathways are deregulated in malignant rhabdoid tumors.

Malignant rhabdoid tumors (MRTs) are aggressive tumors associated with mutations in the SMARCB1 gene. In experimental systems, the loss of SMARCB1 is hypothesized to alter p16(INK4A) pathways resulting in the repression of tumor suppressors. To determine whether these pathways are deregulated in human MRT, we used immunohistochemistry on tissue microarrays to evaluate p16(INK4A)/E2F1/RB and p14(ARF)/MDM2/p53 pathways in 25 atypical teratoid/rhabdoid tumors (AT/RT) and 11 non-CNS MRT. p16(INK4A) was negative or showed focal weak expression. p16(INK4A) downstream targets CDK4/cyclin D1/ppRB were variably expressed at moderate to low levels; E2F1 was negative. Unexpectedly, p14(ARF) expression was seen in many cases, which correlated positively with p53 and inversely with MDM2 immunostaining in AT/RT. TP53 mutational analysis in 19 of 25 AT/RT and in 8 of 11 non-CNS MRT cases showed point mutations in only 3 AT/RT cases, suggesting that p53 expression was driven mainly by p14(ARF). Finally, nucleophosmin, a protein that stabilizes p53, was positive in most cases and colocalized with p53. Together, these data suggest that, in MRT, there is deregulation not only of p16(INK4A) but also of the p14(ARF) pathway. These results provide insights into cell cycle deregulation in the pathogenesis of human MRT and may aid in the design and evaluation of potential therapies for these tumors.

Spontaneous rupture of the left common iliac vein.

We report a case of spontaneous rupture of the left common iliac vein in a 58-year-old woman. Because of hemorrhagic shock, the intervention had to be performed under emergency circumstances, precluding repair of the external iliac vein, which was ligated. The postoperative course was uneventful. We also review the 17 prior cases found in the literature.