Genomic evolution shapes prostate cancer disease type.

The development of cancer is an evolutionary process involving the sequential acquisition of genetic alterations that disrupt normal biological processes, enabling tumor cells to rapidly proliferate and eventually invade and metastasize to other tissues. We investigated the genomic evolution of prostate cancer through the application of three separate classification methods, each designed to investigate a different aspect of tumor evolution. Integrating the results revealed the existence of two distinct types of prostate cancer that arise from divergent evolutionary trajectories, designated as the Canonical and Alternative evolutionary disease types. We therefore propose the evotype model for prostate cancer evolution wherein Alternative-evotype tumors diverge from those of the Canonical-evotype through the stochastic accumulation of genetic alterations associated with disruptions to androgen receptor DNA binding. Our model unifies many previous molecular observations, providing a powerful new framework to investigate prostate cancer disease progression.

Whole-Genome Sequencing Analysis of Male Breast Cancer Unveils Novel Structural Events and Potential Therapeutic Targets.

The molecular characterization of male breast cancer (MaBC) has received limited attention in research, mostly because of its low incidence rate, accounting for only 0.5% to 1% of all reported cases of breast cancer each year. Managing MaBC presents significant challenges, with most treatment protocols being adapted from those developed for female breast cancer. Utilizing whole-genome sequencing (WGS) and state-of-the-art analyses, the genomic features of 10 MaBC cases (n = 10) were delineated and correlated with clinical and histopathologic characteristics. Using fluorescence in situ hybridization, an additional cohort of 18 patients was interrogated to supplement WGS findings. The genomic landscape of MaBC uncovered significant genetic alterations that could influence diagnosis and treatment. We found common somatic mutations in key driver genes, such as FAT1, GATA3, SMARCA4, and ARID2. Our study also mapped out structural variants that impact cancer-associated genes, such as ARID1A, ESR1, GATA3, NTRK1, and NF1. Using a WGS-based classifier, homologous recombination deficiency (HRD) was identified in 2 cases, both presenting with deleterious variants in BRCA2. Noteworthy was the observation of FGFR1 amplification in 21% of cases. Altogether, we identified at least 1 potential therapeutic target in 8 of the 10 cases, including high tumor mutational burden, FGFR1 amplification, and HRD. Our study is the first WGS characterization of MaBC, which uncovered potentially relevant variants, including structural events in cancer genes, HRD signatures, and germline pathogenic mutations. Our results demonstrate unique genetic markers and potential treatment targets in MaBC, thereby underlining the necessity of tailoring treatment strategies for this understudied patient population. These WGS-based findings add to the growing knowledge of MaBC genomics and highlight the need to expand research on this type of cancer.

NIPBL::NACC1 Fusion Hepatic Carcinoma.

Several reports describing a rare primary liver tumor with histologic features reminiscent of follicular thyroid neoplasms have been published under a variety of descriptive terms including thyroid-like, solid tubulocystic, and cholangioblastic cholangiocarcinoma. Although these tumors are considered to represent histologic variants, they lack classic features of cholangiocarcinoma and have unique characteristics, namely immunoreactivity for inhibin and NIPBL::NACC1 fusions. The purpose of this study is to present clinicopathologic and molecular data for a large series of these tumors to better understand their pathogenesis. We identified 11 hepatic tumors with these features. Immunohistochemical and NACC1 and NIPBL fluorescence in situ hybridization assays were performed on all cases. Four cases had available material for whole-genome sequencing (WGS) analysis. Most patients were adult women (mean age: 42 y) who presented with abdominal pain and large hepatic masses (mean size: 14 cm). Ten patients had no known liver disease. Of the patients with follow-up information, 3/9 (33%) pursued aggressive behavior. All tumors were composed of bland cuboidal cells with follicular and solid/trabecular growth patterns in various combinations, were immunoreactive for inhibin, showed albumin mRNA by in situ hybridization, and harbored the NIPBL::NACC1 fusion by fluorescence in situ hybridization. WGS corroborated the presence of the fusion in all 4 tested cases, high tumor mutational burden in 2 cases, and over 30 structural variants per case in 3 sequenced tumors. The cases lacked mutations typical of conventional intrahepatic cholangiocarcinoma. In this report, we describe the largest series of primary inhibin-positive hepatic neoplasms harboring a NIPBL::NACC1 fusion and the first WGS analysis of these tumors. We propose to name this neoplasm NIPBL:NACC1 fusion hepatic carcinoma.

Subclonal Somatic Copy-Number Alterations Emerge and Dominate in Recurrent Osteosarcoma.

Multiple large-scale genomic profiling efforts have been undertaken in osteosarcoma to define the genomic drivers of tumorigenesis, therapeutic response, and disease recurrence. The spatial and temporal intratumor heterogeneity could also play a role in promoting tumor growth and treatment resistance. We conducted longitudinal whole-genome sequencing of 37 tumor samples from 8 patients with relapsed or refractory osteosarcoma. Each patient had at least one sample from a primary site and a metastatic or relapse site. Subclonal copy-number alterations were identified in all patients except one. In 5 patients, subclones from the primary tumor emerged and dominated at subsequent relapses. MYC gain/amplification was enriched in the treatment-resistant clones in 6 of 7 patients with multiple clones. Amplifications in other potential driver genes, such as CCNE1, RAD21, VEGFA, and IGF1R, were also observed in the resistant copy-number clones. A chromosomal duplication timing analysis revealed that complex genomic rearrangements typically occurred prior to diagnosis, supporting a macroevolutionary model of evolution, where a large number of genomic aberrations are acquired over a short period of time followed by clonal selection, as opposed to ongoing evolution. A mutational signature analysis of recurrent tumors revealed that homologous repair deficiency (HRD)-related SBS3 increases at each time point in patients with recurrent disease, suggesting that HRD continues to be an active mutagenic process after diagnosis. Overall, by examining the clonal relationships between temporally and spatially separated samples from patients with relapsed/refractory osteosarcoma, this study sheds light on the intratumor heterogeneity and potential drivers of treatment resistance in this disease. SIGNIFICANCE: The chemoresistant population in recurrent osteosarcoma is subclonal at diagnosis, emerges at the time of primary resection due to selective pressure from neoadjuvant chemotherapy, and is characterized by unique oncogenic amplifications.

The importance of escalating molecular diagnostics in patients with low-grade pediatric brain cancer.

Pilocytic astrocytomas are the most common pediatric brain tumors, typically presenting as low-grade neoplasms. We report two cases of pilocytic astrocytoma with atypical tumor progression. Case 1 involves a 12-yr-old boy with an unresectable suprasellar tumor, negative for BRAF rearrangement but harboring a BRAF p.V600E mutation. He experienced tumor size reduction and stable disease following dabrafenib treatment. Case 2 describes a 6-yr-old boy with a thalamic tumor that underwent multiple resections, with no actionable driver detected using targeted next-generation sequencing. Whole-genome and RNA-seq analysis identified an internal tandem duplication in FGFR1 and RAS pathway activation. Future management options include FGFR1 inhibitors. These cases demonstrate the importance of escalating molecular diagnostics for pediatric brain cancer, advocating for early reflexing to integrative whole-genome sequencing and transcriptomic profiling when targeted panels are uninformative. Identifying molecular drivers can significantly impact treatment decisions and improve patient outcomes.

Clonal evolution during metastatic spread in high-risk neuroblastoma.

Patients with high-risk neuroblastoma generally present with widely metastatic disease and often relapse despite intensive therapy. As most studies to date focused on diagnosis-relapse pairs, our understanding of the genetic and clonal dynamics of metastatic spread and disease progression remain limited. Here, using genomic profiling of 470 sequential and spatially separated samples from 283 patients, we characterize subtype-specific genetic evolutionary trajectories from diagnosis through progression and end-stage metastatic disease. Clonal tracing timed disease initiation to embryogenesis. Continuous acquisition of structural variants at disease-defining loci (MYCN, TERT, MDM2-CDK4) followed by convergent evolution of mutations targeting shared pathways emerged as the predominant feature of progression. At diagnosis metastatic clones were already established at distant sites where they could stay dormant, only to cause relapses years later and spread via metastasis-to-metastasis and polyclonal seeding after therapy.

Diagnostic utility of whole genome sequencing in adults with B-other acute lymphoblastic leukemia.

Genomic profiling during the diagnosis of B-cell precursor acute lymphoblastic leukemia (BCP-ALL) in adults is used to guide disease classification, risk stratification, and treatment decisions. Patients for whom diagnostic screening fails to identify disease-defining or risk-stratifying lesions are classified as having B-other ALL. We screened a cohort of 652 BCP-ALL cases enrolled in UKALL14 to identify and perform whole genome sequencing (WGS) of paired tumor-normal samples. For 52 patients with B-other, we compared the WGS findings with data from clinical and research cytogenetics. WGS identified a cancer-associated event in 51 of 52 patients, including an established subtype defining genetic alterations that were previously missed with standard-of-care (SoC) genetics in 5 of them. Of the 47 true B-other ALL, we identified a recurrent driver in 87% (41). A complex karyotype via cytogenetics emerges as a heterogeneous group, including distinct genetic alterations associated with either favorable (DUX4-r) or poor outcomes (MEF2D-r and IGK::BCL2). For a subset of 31 cases, we integrated the findings from RNA sequencing (RNA-seq) analysis to include fusion gene detection and classification based on gene expression. Compared with RNA-seq, WGS was sufficient to detect and resolve recurrent genetic subtypes; however, RNA-seq can provide orthogonal validation of findings. In conclusion, we demonstrated that WGS can identify clinically relevant genetic abnormalities missed with SoC testing as well as identify leukemia driver events in virtually all cases of B-other ALL.

Subclonal somatic copy number alterations emerge and dominate in recurrent osteosarcoma.

Multiple large-scale tumor genomic profiling efforts have been undertaken in osteosarcoma, however, little is known about the spatial and temporal intratumor heterogeneity and how it may drive treatment resistance. We performed whole-genome sequencing of 37 tumor samples from eight patients with relapsed or refractory osteosarcoma. Each patient had at least one sample from a primary site and a metastatic or relapse site. We identified subclonal copy number alterations in all but one patient. We observed that in five patients, a subclonal copy number clone from the primary tumor emerged and dominated at subsequent relapses. MYC gain/amplification was enriched in the treatment-resistant clone in 6 out of 7 patients with more than one clone. Amplifications in other potential driver genes, such as CCNE1, RAD21, VEGFA, and IGF1R, were also observed in the resistant copy number clones. Our study sheds light on intratumor heterogeneity and the potential drivers of treatment resistance in osteosarcoma.

Bone Marrow Surveillance of Pediatric Cancer Survivors Identifies Clones that Predict Therapy-Related Leukemia.

PURPOSE: Therapy-related myelodysplastic syndrome and acute leukemias (t-MDS/AL) are a major cause of nonrelapse mortality among pediatric cancer survivors. Although the presence of clonal hematopoiesis (CH) in adult patients at cancer diagnosis has been implicated in t-MDS/AL, there is limited published literature describing t-MDS/AL development in children. EXPERIMENTAL DESIGN: We performed molecular characterization of 199 serial bone marrow samples from 52 patients treated for high-risk neuroblastoma, including 17 with t-MDS/AL (transformation), 14 with transient cytogenetic abnormalities (transient), and 21 without t-MDS/AL or cytogenetic alterations (neuroblastoma-treated control). We also evaluated for CH in a cohort of 657 pediatric patients with solid tumor. RESULTS: We detected at least one disease-defining alteration in all cases at t-MDS/AL diagnosis, most commonly TP53 mutations and KMT2A rearrangements, including involving two novel partner genes (PRDM10 and DDX6). Backtracking studies identified at least one t-MDS/AL-associated mutation in 13 of 17 patients at a median of 15 months before t-MDS/AL diagnosis (range, 1.3-32.4). In comparison, acquired mutations were infrequent in the transient and control groups (4/14 and 1/21, respectively). The relative risk for development of t-MDS/AL in the presence of an oncogenic mutation was 8.8 for transformation patients compared with transient. Unlike CH in adult oncology patients, TP53 mutations were only detectable after initiation of cancer therapy. Last, only 1% of pediatric patients with solid tumor evaluated had CH involving myeloid genes. CONCLUSIONS: These findings demonstrate the clinical relevance of identifying molecular abnormalities in predicting development of t-MDS/AL and should guide the formation of intervention protocols to prevent this complication in high-risk pediatric patients.

Skeletal muscle metastases in neuroblastoma share common progenitors with primary tumor and biologically resemble stage MS disease.

Introduction: While subcutaneous metastases are often observed with stage MS neuroblastoma, an entity that usually resolves spontaneously, skeletal muscle metastases (SMM) have been rarely described. The purpose of this retrospective study was to investigate the significance of SMM in neuroblastoma. Patients and methods: Seventeen patients with neuroblastoma SMM were diagnosed at a median age of 4.3 (0.1-15.6) months. All had SMM at diagnosis and metastases at other sites. Fifteen (88%) had ≥ 2 SMM in disparate muscle groups. One, 14, and 2 patients had low, intermediate, and high-risk disease respectively. Fifteen tumors had favorable histology without MYCN amplification, and 2 were MYCN-amplified. Most SMM (80%; n=12/15 evaluated) were MIBG-avid. Results: Only 1 patient (with MYCN-non-amplified neuroblastoma) had disease progression. All survive at median follow-up of 47.9 (16.9-318.9) months post-diagnosis. Biological markers (histology, chromosomal and genetic aberrations) were not prognostic. Whole genome sequencing of 3 matched primary and SMM lesions suggested that both primary and metastatic tumors arose from the same progenitor. SMM completely resolved in 10 patients by 12 months post-diagnosis. Of 4 patients managed with watchful observation alone without any cytotoxic therapy, 3 maintain complete remission with SMM resolving by 5, 13, and 21 months post-diagnosis respectively. Conclusions: Children with neuroblastoma SMM have an excellent prognosis, with a clinical course suggestive of stage MS disease. Based on these results, the initial management of infants with non-MYCN-amplified NB with SMM could be watchful observation, which could eliminate or reduce exposure to genotoxic therapy.

Molecular International Prognostic Scoring System for Myelodysplastic Syndromes.

BACKGROUND: Risk stratification and therapeutic decision-making for myelodysplastic syndromes (MDS) are based on the International Prognostic Scoring System­Revised (IPSS-R), which considers hematologic parameters and cytogenetic abnormalities. Somatic gene mutations are not yet used in the risk stratification of patients with MDS. METHODS: To develop a clinical-molecular prognostic model (IPSS-Molecular [IPSS-M]), pretreatment diagnostic or peridiagnostic samples from 2957 patients with MDS were profiled for mutations in 152 genes. Clinical and molecular variables were evaluated for associations with leukemia-free survival, leukemic transformation, and overall survival. Feature selection was applied to determine the set of independent IPSS-M prognostic variables. The relative weights of the selected variables were estimated using a robust Cox multivariable model adjusted for confounders. The IPSS-M was validated in an external cohort of 754 Japanese patients with MDS. RESULTS: We mapped at least one oncogenic genomic alteration in 94% of patients with MDS. Multivariable analysis identified TP53multihit, FLT3 mutations, and MLLPTD as top genetic predictors of adverse outcomes. Conversely, SF3B1 mutations were associated with favorable outcomes, but this was modulated by patterns of comutation. Using hematologic parameters, cytogenetic abnormalities, and somatic mutations of 31 genes, the IPSS-M resulted in a unique risk score for individual patients. We further derived six IPSS-M risk categories with prognostic differences. Compared with the IPSS-R, the IPSS-M improved prognostic discrimination across all clinical end points and restratified 46% of patients. The IPSS-M was applicable in primary and secondary/therapy-related MDS. To simplify clinical use of the IPSS-M, we developed an open-access Web calculator that accounts for missing values. CONCLUSIONS: Combining genomic profiling with hematologic and cytogenetic parameters, the IPSS-M improves the risk stratification of patients with MDS and represents a valuable tool for clinical decision-making. (Funded by Celgene Corporation through the MDS Foundation, the Josie Robertson Investigators Program, the Edward P. Evans Foundation, the Projects of National Relevance of the Italian Ministry of University and Research, Associazione Italiana per la Ricerca sul Cancro, the Japan Agency for Medical Research and Development, Cancer Research UK, the Austrian Science Fund, the MEXT [Japanese Ministry of Education, Culture, Sports, Science and Technology] Program for Promoting Research on the Supercomputer Fugaku, the Japan Society for the Promotion of Science, the Taiwan Department of Health, and Celgene Corporation through the MDS Foundation.)

Association of BRAF V600E mutations with vasoactive intestinal peptide syndrome in MYCN-amplified neuroblastoma.

Very rarely, vasoactive intestinal peptide-related diarrhea (VIP-D) is observed in patients with high-risk neuroblastoma (HR-NB) where the associated fluid and electrolyte abnormalities can pose a major clinical challenge for administering the required aggressive multimodality treatment. Two patients with HR-NB developed VIP-D during induction and were found to have a somatic BRAF V600E mutation. Serum VIP levels and diarrhea promptly resolved in both patients after initiating treatment with BRAF and MEK inhibitors. This illustrates an association of VIP-D with BRAF V600E mutations and demonstrates a therapeutic strategy in the specific context of VIP-D and BRAF V600E mutations in HR-NB patients. The addition of BRAF and MEK inhibitors allows continued conventional tumor-directed treatment by decreasing the severity of symptoms caused by this life-threatening complication.

Comprehensive Molecular Profiling of Desmoplastic Small Round Cell Tumor.

Desmoplastic small round cell tumor (DSRCT) is characterized by the EWSR1-WT1 t(11;22) (p13:q12) translocation. Few additional putative drivers have been identified, and research has suffered from a lack of model systems. Next-generation sequencing (NGS) data from 68 matched tumor-normal samples, whole-genome sequencing data from 10 samples, transcriptomic and affymetrix array data, and a bank of DSRCT patient-derived xenograft (PDX) are presented. EWSR1-WT1 fusions were noted to be simple, balanced events. Recurrent mutations were uncommon, but were noted in TERT (3%), ARID1A (6%), HRAS (5%), and TP53 (3%), and recurrent loss of heterozygosity (LOH) at 11p, 11q, and 16q was identified in 18%, 22%, and 34% of samples, respectively. Comparison of tumor-normal matched versus unmatched analysis suggests overcalling of somatic mutations in prior publications of DSRCT NGS data. Alterations in fibroblast growth factor receptor 4 (FGFR4) were identified in 5 of 68 (7%) of tumor samples, whereas differential overexpression of FGFR4 was confirmed orthogonally using 2 platforms. PDX models harbored the pathognomic EWSR1-WT1 fusion and were highly representative of corresponding tumors. Our analyses confirm DSRCT as a genomically quiet cancer defined by the balanced translocation, t(11;22)(p13:q12), characterized by a paucity of secondary mutations but a significant number of copy number alterations. Against this genomically quiet background, recurrent activating alterations of FGFR4 stood out, and suggest that this receptor tyrosine kinase, also noted to be highly expressed in DSRCT, should be further investigated. Future studies of DSRCT biology and preclinical therapeutic strategies should benefit from the PDX models characterized in this study. IMPLICATIONS: These data describe the general quiescence of the desmoplastic small round cell tumor (DSRCT) genome, present the first available bank of DSRCT model systems, and nominate FGFR4 as a key receptor tyrosine kinase in DSRCT, based on high expression, recurrent amplification, and recurrent activating mutations.

Isabl Platform, a digital biobank for processing multimodal patient data.

BACKGROUND: The widespread adoption of high throughput technologies has democratized data generation. However, data processing in accordance with best practices remains challenging and the data capital often becomes siloed. This presents an opportunity to consolidate data assets into digital biobanks-ecosystems of readily accessible, structured, and annotated datasets that can be dynamically queried and analysed. RESULTS: We present Isabl, a customizable plug-and-play platform for the processing of multimodal patient-centric data. Isabl's architecture consists of a relational database (Isabl DB), a command line client (Isabl CLI), a RESTful API (Isabl API) and a frontend web application (Isabl Web). Isabl supports automated deployment of user-validated pipelines across the entire data capital. A full audit trail is maintained to secure data provenance, governance and ensuring reproducibility of findings. CONCLUSIONS: As a digital biobank, Isabl supports continuous data utilization and automated meta analyses at scale, and serves as a catalyst for research innovation, new discoveries, and clinical translation.

Cancer therapy shapes the fitness landscape of clonal hematopoiesis.

Acquired mutations are pervasive across normal tissues. However, understanding of the processes that drive transformation of certain clones to cancer is limited. Here we study this phenomenon in the context of clonal hematopoiesis (CH) and the development of therapy-related myeloid neoplasms (tMNs). We find that mutations are selected differentially based on exposures. Mutations in ASXL1 are enriched in current or former smokers, whereas cancer therapy with radiation, platinum and topoisomerase II inhibitors preferentially selects for mutations in DNA damage response genes (TP53, PPM1D, CHEK2). Sequential sampling provides definitive evidence that DNA damage response clones outcompete other clones when exposed to certain therapies. Among cases in which CH was previously detected, the CH mutation was present at tMN diagnosis. We identify the molecular characteristics of CH that increase risk of tMN. The increasing implementation of clinical sequencing at diagnosis provides an opportunity to identify patients at risk of tMN for prevention strategies.

Implications of TP53 allelic state for genome stability, clinical presentation and outcomes in myelodysplastic syndromes.

Tumor protein p53 (TP53) is the most frequently mutated gene in cancer1,2. In patients with myelodysplastic syndromes (MDS), TP53 mutations are associated with high-risk disease3,4, rapid transformation to acute myeloid leukemia (AML)5, resistance to conventional therapies6-8 and dismal outcomes9. Consistent with the tumor-suppressive role of TP53, patients harbor both mono- and biallelic mutations10. However, the biological and clinical implications of TP53 allelic state have not been fully investigated in MDS or any other cancer type. We analyzed 3,324 patients with MDS for TP53 mutations and allelic imbalances and delineated two subsets of patients with distinct phenotypes and outcomes. One-third of TP53-mutated patients had monoallelic mutations whereas two-thirds had multiple hits (multi-hit) consistent with biallelic targeting. Established associations with complex karyotype, few co-occurring mutations, high-risk presentation and poor outcomes were specific to multi-hit patients only. TP53 multi-hit state predicted risk of death and leukemic transformation independently of the Revised International Prognostic Scoring System (IPSS-R)11. Surprisingly, monoallelic patients did not differ from TP53 wild-type patients in outcomes and response to therapy. This study shows that consideration of TP53 allelic state is critical for diagnostic and prognostic precision in MDS as well as in future correlative studies of treatment response.

Author Correction: The evolutionary history of lethal metastatic prostate cancer.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.