Genomic landscape of cutaneous follicular lymphomas reveals 2 subgroups with clinically predictive molecular features.

Primary cutaneous follicle center lymphomas (PCFCLs) are indolent B-cell lymphomas that predominantly remain skin restricted and manageable with skin-directed therapy. Conversely, secondary cutaneous involvement by usual systemic follicular lymphoma (secondary cutaneous follicular lymphoma [SCFL]) has a worse prognosis and often necessitates systemic therapy. Unfortunately, no histopathologic or genetic features reliably differentiate PCFCL from SCFL at diagnosis. Imaging may miss low-burden internal disease in some cases of SCFLs, leading to misclassification as PCFCL. Whereas usual systemic FL is well characterized genetically, the genomic landscapes of PCFCL and SCFL are unknown. Herein, we analyzed clinicopathologic and immunophenotypic data from 30 cases of PCFCL and 10 of SCFL and performed whole-exome sequencing on 18 specimens of PCFCL and 6 of SCFL. During a median follow-up of 7 years, 26 (87%) of the PCFCLs remained skin restricted. In the remaining 4 cases, systemic disease developed within 3 years of diagnosis. Although the SCFLs universally expressed BCL2 and had BCL2 rearrangements, 73% of the PCFCLs lacked BCL2 expression, and only 8% of skin-restricted PCFCLs had BCL2 rearrangements. SCFLs showed low proliferation fractions, whereas 75% of PCFCLs had proliferation fractions >30%. Of the SCFLs, 67% had characteristic loss-of-function CREBBP or KMT2D mutations vs none in skin-restricted PCFCL. Both SCFL and skin-restricted PCFCL showed frequent TNFRSF14 loss-of-function mutations and copy number loss at chromosome 1p36. These data together establish PCFCL as a unique entity with biological features distinct from usual systemic FL and SCFL. We propose 3 criteria based on BCL2 rearrangement, chromatin-modifying gene mutations (CREBBP, KMT2D, EZH2, and EP300), and proliferation index to classify cutaneous FL specimens based on the likelihood of concurrent or future systemic spread.

Int22h1/Int22h2-mediated Xq28 duplication syndrome: de novo duplications, prenatal diagnoses, and additional phenotypic features.

Int22h1/Int22h2-mediated Xq28 duplication syndrome is a relatively new X-linked intellectual disability syndrome, arising from duplications of the subregion flanked by intron 22 homologous regions 1 and 2 on the q arm of chromosome X. Its primary manifestations include variable cognitive deficits, distinct facial dysmorphia, and neurobehavioral abnormalities that mainly include hyperactivity, irritability, and autistic behavior. Affected males are hemizygous for the duplication, which explains their often more severe manifestations compared with heterozygous females. In this report, we describe the cases of nine individuals recently identified having the syndrome, highlighting unique and previously unreported findings of this syndrome. Specifically, we report for the first time in this syndrome, two cases with de novo duplications, three receiving prenatal diagnosis with the syndrome, and three others having atypical versions of the duplication. Among the latter, one proband has a shortened version spanning only the centromeric half of the typical duplication, while the other two cases have a nearly identical length duplication as the classical duplication, with the exception that their duplication's breakpoints are telomerically shifted by about 0.2 Mb. Finally, we shed light on two new manifestations in this syndrome, vertebral anomalies and multiple malignancies, which possibly expand the phenotypic spectrum of the syndrome.

Pediatric acute myeloid leukemia with t(7;21)(p22;q22).

The t(7;21)(p22;q22) resulting in RUNX1-USP42 fusion, is a rare but recurrent cytogenetic abnormality associated with acute myeloid leukemia (AML) and myelodysplastic syndromes. The prognostic significance of this translocation has not been well established due to the limited number of patients. Herein, we report three pediatric AML patients with t(7;21)(p22;q22). All three patients presented with pancytopenia or leukopenia at diagnosis, accompanied by abnormal immunophenotypic expression of CD7 and CD56 on leukemic blasts. One patient had t(7;21)(p22;q22) as the sole abnormality, whereas the other two patients had additional numerical and structural aberrations including loss of 5q material. Fluorescence in situ hybridization analysis on interphase cells or sequential examination of metaphases showed the RUNX1 rearrangement and confirmed translocation 7;21. Genomic SNP microarray analysis, performed on DNA extracted from the bone marrow from the patient with isolated t(7;21)(p22;q22), showed a 32.2 Mb copy neutral loss of heterozygosity (cnLOH) within the short arm of chromosome 11. After 2-4 cycles of chemotherapy, all three patients underwent allogeneic hematopoietic stem cell transplantation (HSCT). One patient died due to complications related to viral reactivation and graft-versus-host disease. The other two patients achieved complete remission after HSCT. Our data displayed the accompanying cytogenetic abnormalities including del(5q) and cnLOH of 11p, the frequent pathological features shared with other reported cases, and clinical outcome in pediatric AML patients with t(7;21)(p22;q22). The heterogeneity in AML harboring similar cytogenetic alterations may be attributed to additional uncovered genetic lesions.

Identification of two 14q32 deletions involving DICER1 associated with the development of DICER1-related tumors.

DICER1 encodes an RNase III endonuclease protein that regulates the production of small non-coding RNAs. Germline mutations in DICER1 are associated with an autosomal dominant hereditary cancer predisposition syndrome that confers an increased risk for the development of several rare childhood and adult-onset tumors, the most frequent of which include pleuropulmonary blastoma, ovarian sex cord-stromal tumors, cystic nephroma, and thyroid gland neoplasia. The majority of reported germline DICER1 mutations are truncating sequence-level alterations, suggesting that a loss-of-function type mechanism drives tumor formation in DICER1 syndrome. However, reports of patients with germline DICER1 whole gene deletions are limited, and thus far, only two have reported an association with tumor development. Here we report the clinical findings of three patients from two unrelated families with 14q32 deletions that encompass the DICER1 locus. The deletion identified in Family I is 1.4 Mb and was initially identified in a 6-year-old male referred for developmental delay, hypotonia, macrocephaly, obesity, and behavioral problems. Subsequent testing revealed that this deletion was inherited from his mother, who had a clinical history that included bilateral multinodular goiter and papillary thyroid carcinoma. The second deletion is 5.0 Mb and was identified in a 15-year-old female who presented with autism, coarse facial features, Sertoli-Leydig cell tumor, and Wilms' tumor. These findings provide additional supportive evidence that germline deletion of DICER1 confers an increased risk for DICER1-related tumor development, and provide new insight into the clinical significance of deletions involving the 14q32 region.

Genomic Analyses Identify Recurrent Alterations in Immune Evasion Genes in Diffuse Large B-Cell Lymphoma, Leg Type.

Cutaneous diffuse large B-cell lymphomas (DLBCLs) are aggressive lymphomas with a poor prognosis. To elucidate their genetic bases, we analyzed exome sequencing of 37 cutaneous DLBCLs, including 31 DLBCLs, leg type (DLBCL-LT) and 6 cutaneous DLBCLs-not otherwise specified (DLBCL-NOS). As reported previously, 77% of DLBCL-LT harbor NF-κB-activating MYD88 mutations. In nearly all MYD88-wild-type DLBCL-LT, we found cancer-promoting mutations that either activate the NF-κB pathway through alternative genes (NFKBIE or REL) or activate other canonical cancer pathways (BRAF, MED12, PIK3R1, and STAT3). After NF-κB, the second most commonly mutated pathway putatively enables immune evasion via mutations predicted to downregulate antigen processing (B2M, CIITA, HLA) or T-cell co-stimulation (CD58). DLBCL-LT have little genetic overlap with the genetically heterogeneous DLBCL-NOS. Instead, they resemble primary central nervous system and testicular large B-cell lymphomas (primary central nervous system lymphomas and primary testicular lymphomas). Like primary central nervous system lymphomas/primary testicular lymphomas, 40% of DLBCL-LT (vs. 0% of DLBCLs-not otherwise specified) harbored PDL1/PDL2 translocations, which lead to overexpression of PD-L1 or PD-L2 in 50% of the cases. Collectively, these data broaden our understanding of cutaneous DLBCLs and suggest novel therapeutic approaches (e.g., BRAF or PI3K inhibitors). Additionally, they suggest novel treatment paradigms, wherein DLBCL-LT can be targeted with strategies (e.g., immune checkpoint blockers) currently being developed for genomically similar primary central nervous system lymphomas/primary testicular lymphomas.

Genetic evaluation of juvenile xanthogranuloma: genomic abnormalities are uncommon in solitary lesions, advanced cases may show more complexity.

Juvenile xanthogranuloma is a rare histiocytic proliferation primarily affecting infants and young children, characterized by aberrant infiltration of histiocyte-derived cells in the skin, soft tissues and more rarely, visceral organs. Juvenile xanthogranuloma is generally considered to be a benign disorder; most lesions are solitary cutaneous nodules that resolve spontaneously without treatment. However, cases with extracutaneous involvement, multiple lesions, and/or systemic disease often require aggressive therapy. Though molecular studies have provided evidence of clonality in juvenile xanthogranuloma, in support of a neoplastic process, little is known about the genetic profile of juvenile xanthogranuloma. We used molecular inversion probe array technology to evaluate the genomic characteristics (copy number alterations or copy neutral-loss of heterozygosity) of 21 archived cases of juvenile xanthogranuloma (19 solitary, 1 diffuse cutaneous, 1 systemic). Four cases (19%) showed acquired, clonal alterations. Two lesions from a case of diffuse cutaneous juvenile xanthogranuloma showed distinct profiles: JXG-1a contained trisomy 5 and 17 and JXG-1b contained loss of heterozygosity in 5q. The systemic juvenile xanthogranuloma (JXG-2) showed multiple genomic alterations. Only two of 19 solitary juvenile xanthogranulomas showed abnormal genomic profiles: JXG-3 showed gains on 1q and 11q and JXG-4 showed a 7.2 Mb loss in 3p. No recurrent abnormalities were observed among these cases. The presence of non-recurrent copy number alterations in a subset of samples implies that copy number changes are unlikely driving pathogenesis in juvenile xanthogranuloma, but may be acquired during disease progression. The presence of genomic abnormalities in more advanced cases (ie, systemic and diffuse cutaneous juvenile xanthogranuloma) supports this notion, particularly as the advanced cases of juvenile xanthogranuloma presented more genomic complexity.

Genomic analysis of follicular dendritic cell sarcoma by molecular inversion probe array reveals tumor suppressor-driven biology.

Follicular dendritic cell sarcoma is a rare malignant neoplasm of dendritic cell origin that is currently poorly characterized by genetic studies. To investigate whether recurrent genomic alterations may underlie the biology of follicular dendritic cell sarcoma and to identify potential contributory regions and genes, molecular inversion probe array analysis was performed on 14 independent formalin-fixed, paraffin-embedded samples. Abnormal genomic profiles were observed in 11 out of 14 (79%) cases. The majority showed extensive genomic complexity that was predominantly represented by hemizygous losses affecting multiple chromosomes. Alterations of chromosomal regions 1p (55%), 2p (55%), 3p (82%), 3q (45%), 6q (55%), 7q (73%), 8p (45%), 9p (64%), 11q (64%), 13q (91%), 14q (82%), 15q (64%), 17p (55%), 18q (64%), and 22q (55%) were recurrent across the 11 samples showing abnormal genomic profiles. Many recurrent genomic alterations in follicular dendritic cell sarcoma overlap deletions that are frequently observed across human cancers, suggesting selection, or an active role for these alterations in follicular dendritic cell sarcoma pathogenesis. In support of a tumor suppressor-driven biology, homozygous deletions involving tumor suppressor genes CDKN2A, RB1, BIRC3, and CYLD were also observed. Neither recurrent gains nor amplifications were observed. This genomic characterization provides new information regarding follicular dendritic cell sarcoma biology that may improve understanding about the underlying pathophysiology, provide better prognostication, and identify potential therapeutic markers for this rare disease.

Elements of the polycomb repressor SU(Z)12 needed for histone H3-K27 methylation, the interface with E(Z), and in vivo function.

Polycomb repressive complex 2 (PRC2) is an essential chromatin-modifying enzyme that implements gene silencing. PRC2 methylates histone H3 on lysine-27 and is conserved from plants to flies to humans. In Drosophila melanogaster, PRC2 contains four core subunits: E(Z), SU(Z)12, ESC, and NURF55. E(Z) bears a SET domain that houses the enzyme active site. However, PRC2 activity depends upon critical inputs from SU(Z)12 and ESC. The stimulatory mechanisms are not understood. We present here functional dissection of the SU(Z)12 subunit. SU(Z)12 contains two highly conserved domains: an ∼140-amino-acid VEFS domain and a Cys2-His2 zinc finger (ZnF). Analysis of recombinant PRC2 bearing VEFS domain alterations, including some modeled after leukemia mutations, identifies distinct elements needed for SU(Z)12 assembly with E(Z) and stimulation of histone methyltransferase. The results define an extensive VEFS subdomain that organizes the SU(Z)12-E(Z) interface. Although the SU(Z)12 ZnF is not needed for methyltransferase in vitro, genetic rescue assays show that the ZnF is required in vivo. Chromatin immunoprecipitations reveal that this ZnF facilitates PRC2 binding to a genomic target. This study defines functionally critical SU(Z)12 elements, including key determinants of SU(Z)12-E(Z) communication. Together with recent findings, this illuminates PRC2 modulation by conserved inputs from its noncatalytic subunits.

Subunit contributions to histone methyltransferase activities of fly and worm polycomb group complexes.

The ESC-E(Z) complex of Drosophila melanogaster Polycomb group (PcG) repressors is a histone H3 methyltransferase (HMTase). This complex silences fly Hox genes, and related HMTases control germ line development in worms, flowering in plants, and X inactivation in mammals. The fly complex contains a catalytic SET domain subunit, E(Z), plus three noncatalytic subunits, SU(Z)12, ESC, and NURF-55. The four-subunit complex is >1,000-fold more active than E(Z) alone. Here we show that ESC and SU(Z)12 play key roles in potentiating E(Z) HMTase activity. We also show that loss of ESC disrupts global methylation of histone H3-lysine 27 in fly embryos. Subunit mutations identify domains required for catalytic activity and/or binding to specific partners. We describe missense mutations in surface loops of ESC, in the CXC domain of E(Z), and in the conserved VEFS domain of SU(Z)12, which each disrupt HMTase activity but preserve complex assembly. Thus, the E(Z) SET domain requires multiple partner inputs to produce active HMTase. We also find that a recombinant worm complex containing the E(Z) homolog, MES-2, has robust HMTase activity, which depends upon both MES-6, an ESC homolog, and MES-3, a pioneer protein. Thus, although the fly and mammalian PcG complexes absolutely require SU(Z)12, the worm complex generates HMTase activity from a distinct partner set.