Evolutionary histories of breast cancer and related clones.

Recent studies have documented frequent evolution of clones carrying common cancer mutations in apparently normal tissues, which are implicated in cancer development1-3. However, our knowledge is still missing with regard to what additional driver events take place in what order, before one or more of these clones in normal tissues ultimately evolve to cancer. Here, using phylogenetic analyses of multiple microdissected samples from both cancer and non-cancer lesions, we show unique evolutionary histories of breast cancers harbouring der(1;16), a common driver alteration found in roughly 20% of breast cancers. The approximate timing of early evolutionary events was estimated from the mutation rate measured in normal epithelial cells. In der(1;16)(+) cancers, the derivative chromosome was acquired from early puberty to late adolescence, followed by the emergence of a common ancestor by the patient's early 30s, from which both cancer and non-cancer clones evolved. Replacing the pre-existing mammary epithelium in the following years, these clones occupied a large area within the premenopausal breast tissues by the time of cancer diagnosis. Evolution of multiple independent cancer founders from the non-cancer ancestors was common, contributing to intratumour heterogeneity. The number of driver events did not correlate with histology, suggesting the role of local microenvironments and/or epigenetic driver events. A similar evolutionary pattern was also observed in another case evolving from an AKT1-mutated founder. Taken together, our findings provide new insight into how breast cancer evolves.

Frequent mutations that converge on the NFKBIZ pathway in ulcerative colitis.

Chronic inflammation is accompanied by recurring cycles of tissue destruction and repair and is associated with an increased risk of cancer1-3. However, how such cycles affect the clonal composition of tissues, particularly in terms of cancer development, remains unknown. Here we show that in patients with ulcerative colitis, the inflamed intestine undergoes widespread remodelling by pervasive clones, many of which are positively selected by acquiring mutations that commonly involve the NFKBIZ, TRAF3IP2, ZC3H12A, PIGR and HNRNPF genes and are implicated in the downregulation of IL-17 and other pro-inflammatory signals. Mutational profiles vary substantially between colitis-associated cancer and non-dysplastic tissues in ulcerative colitis, which indicates that there are distinct mechanisms of positive selection in both tissues. In particular, mutations in NFKBIZ are highly prevalent in the epithelium of patients with ulcerative colitis but rarely found in both sporadic and colitis-associated cancer, indicating that NFKBIZ-mutant cells are selected against during colorectal carcinogenesis. In further support of this negative selection, we found that tumour formation was significantly attenuated in Nfkbiz-mutant mice and cell competition was compromised by disruption of NFKBIZ in human colorectal cancer cells. Our results highlight common and discrete mechanisms of clonal selection in inflammatory tissues, which reveal unexpected cancer vulnerabilities that could potentially be exploited for therapeutics in colorectal cancer.

Germ line DDX41 mutations define a unique subtype of myeloid neoplasms.

Germ line DDX41 variants have been implicated in late-onset myeloid neoplasms (MNs). Despite an increasing number of publications, many important features of DDX41-mutated MNs remain to be elucidated. Here we performed a comprehensive characterization of DDX41-mutated MNs, enrolling a total of 346 patients with DDX41 pathogenic/likely-pathogenic (P/LP) germ line variants and/or somatic mutations from 9082 MN patients, together with 525 first-degree relatives of DDX41-mutated and wild-type (WT) patients. P/LP DDX41 germ line variants explained ∼80% of known germ line predisposition to MNs in adults. These risk variants were 10-fold more enriched in Japanese MN cases (n = 4461) compared with the general population of Japan (n = 20 238). This enrichment of DDX41 risk alleles was much more prominent in male than female (20.7 vs 5.0). P/LP DDX41 variants conferred a large risk of developing MNs, which was negligible until 40 years of age but rapidly increased to 49% by 90 years of age. Patients with myelodysplastic syndromes (MDS) along with a DDX41-mutation rapidly progressed to acute myeloid leukemia (AML), which was however, confined to those having truncating variants. Comutation patterns at diagnosis and at progression to AML were substantially different between DDX41-mutated and WT cases, in which none of the comutations affected clinical outcomes. Even TP53 mutations made no exceptions and their dismal effect, including multihit allelic status, on survival was almost completely mitigated by the presence of DDX41 mutations. Finally, outcomes were not affected by the conventional risk stratifications including the revised/molecular International Prognostic Scoring System. Our findings establish that MDS with DDX41-mutation defines a unique subtype of MNs that is distinct from other MNs.

Age-related remodelling of oesophageal epithelia by mutated cancer drivers.

Clonal expansion in aged normal tissues has been implicated in the development of cancer. However, the chronology and risk dependence of the expansion are poorly understood. Here we intensively sequence 682 micro-scale oesophageal samples and show, in physiologically normal oesophageal epithelia, the progressive age-related expansion of clones that carry mutations in driver genes (predominantly NOTCH1), which is substantially accelerated by alcohol consumption and by smoking. Driver-mutated clones emerge multifocally from early childhood and increase their number and size with ageing, and ultimately replace almost the entire oesophageal epithelium in the extremely elderly. Compared with mutations in oesophageal cancer, there is a marked overrepresentation of NOTCH1 and PPM1D mutations in physiologically normal oesophageal epithelia; these mutations can be acquired before late adolescence (as early as early infancy) and significantly increase in number with heavy smoking and drinking. The remodelling of the oesophageal epithelium by driver-mutated clones is an inevitable consequence of normal ageing, which-depending on lifestyle risks-may affect cancer development.

TP53 and RB1 alterations characterize poor prognostic subgroups in pediatric acute myeloid leukemia.

Pediatric acute myeloid leukemia (AML) is a poor prognostic subtype of pediatric leukemia. However, the detailed characteristics of many genetic abnormalities are yet to be established in this disease. Although TP53 and RB1 are established as representative tumor suppressor genes in various cancers, alterations of these two genes, especially RB1, have not been characterized in pediatric AML. We performed next-generation sequencing in 328 pediatric AML patients from the Japanese AML-05 trial to ascertain TP53 and RB1 alterations, and their prognostic implications. We identified seven patients with TP53 alterations (2.1%) and six patients with RB1 alterations (1.8%). These alterations were found in only patients without RUNX1::RUNX1T1, CBFB::MYH11, or KMT2A rearrangements. TP53 and RB1 were frequently co-deleted with their neighboring genes PRPF8 and ELF1, respectively. Patients with TP53 alterations had significantly lower 5-year overall survival (OS; 14.3% vs. 71.4%, p < 0.001) and lower 5-year event-free survival (EFS; 0% vs. 56.3%, p < 0.001); similarly, patients with RB1 had significantly lower 5-year OS (0% vs. 71.8%, p < 0.001) and lower 5-year EFS (0% vs. 56.0%, p < 0.001) when compared to patients without these alterations. In gene expression analyses, oxidative phosphorylation, glycolysis, and protein secretion were upregulated in patients with TP53 and/or RB1 alterations. Additionally, Kaplan-Meier analysis revealed that high expressions of SLC2A5, KCNAB2, and CD300LF were related to poor OS of non-core-binding factor AML patients (p < 0.001, p = 0.001, and p = 0.021, respectively). This study will contribute to the development of risk-stratified therapy and precision medicine in pediatric AML.

Frequent genetic alterations in immune checkpoint-related genes in intravascular large B-cell lymphoma.

Intravascular large B-cell lymphoma (IVLBCL) is a unique type of extranodal lymphoma characterized by selective growth of tumor cells in small vessels without lymphadenopathy. Greater understanding of the molecular pathogenesis of IVLBCL is hampered by the paucity of lymphoma cells in biopsy specimens, creating a limitation in obtaining sufficient tumor materials. To uncover the genetic landscape of IVLBCL, we performed whole-exome sequencing (WES) of 21 patients with IVLBCL using plasma-derived cell-free DNA (cfDNA) (n = 18), patient-derived xenograft tumors (n = 4), and tumor DNA from bone marrow (BM) mononuclear cells (n = 2). The concentration of cfDNA in IVLBCL was significantly higher than that in diffuse large B-cell lymphoma (DLBCL) (P < .0001) and healthy donors (P = .0053), allowing us to perform WES; most mutations detected in BM tumor DNA were successfully captured in cfDNA and xenograft. IVLBCL showed a high frequency of genetic lesions characteristic of activated B-cell-type DLBCL, with the former showing conspicuously higher frequencies (compared with nodal DLBCL) of mutations in MYD88 (57%), CD79B (67%), SETD1B (57%), and HLA-B (57%). We also found that 8 IVLBCL (38%) harbored rearrangements of programmed cell death 1 ligand 1 and 2 (PD-L1/PD-L2) involving the 3' untranslated region; such rearrangements are implicated in immune evasion via PD-L1/PD-L2 overexpression. Our data demonstrate the utility of cfDNA and imply important roles for immune evasion in IVLBCL pathogenesis and PD-1/PD-L1/PD-L2 blockade in therapeutics for IVLBCL.

Immunotherapy as a potential treatment approach for currently incurable bone metastasis.

Once cancer metastasizes to the bone, the prognosis of cancer patients becomes extremely poor. Unfortunately, the current most successful treatment for bone metastasis can extend their survival by only a few months. Although recent studies have revealed promising impacts of cancer immunotherapies, their treatment efficacy on bone metastatic diseases remains controversial. Therefore, in this review, we discussed (i) preclinical and clinical evidence of the immunotherapeutic strategies for cancer bone metastasis, mainly focusing on cell-based immunotherapy, cytokine-based immunotherapy, and immune checkpoint blockade, and (ii) current shortcomings of immunotherapy for bone metastasis and their potential future directions. Although the evidence on treatment efficacy and safety, as well as long-term effects, is limited, immunotherapies could induce partial or complete remissions in a few cancer patients with bone metastasis. However, there are still hurdles, such as the immunosuppressive nature of the bone marrow microenvironment and poor distribution of cell-based immunotherapies to bone, that need to be overcome to enhance the treatment efficacy of immunotherapies on bone metastasis. While it is apparent that further investigation is needed regarding immunotherapeutic treatment efficacy in patients with bone metastasis, this therapy may prove to be clinically novel in this subset of cancer patients.

A PCR-amplified transgene fragment flanked by a single copy of a truncated inverted terminal repeat for recombinant adeno-associated virus production prevents unnecessary plasmid DNA packaging.

The application of recombinant adeno-associated viruses (rAAVs) for gene therapy faces certain challenges, including genome packaging of non-vector sequences. Inverted terminal repeats (ITRs) flanking the rAAV genome, comprising three inverted repeat regions (A, B, and C) and a non-inverted repeat region (D), contribute to non-vector genome packaging. We aimed to circumvent this issue by comparing the properties of rAAV containing DNA plasmids and PCR-amplified transgenes, including a single copy of the AD sequence (rAAV-pAD/L-AD, respectively), which is a truncated form of ITR, with those of wild-type ITR genome (single-stranded and self-complementary AAV; ssAAV and scAAV). The packaging efficiency of rAAV-pAD/L-AD was found to be comparable to that of scAAV, whereas the transduction efficiency of rAAV-pAD/L-AD was lower than that of ss/scAAV. Remarkably, rAAV-L-AD reduced the plasmid backbone packaging contamination compared to ss/scAAV. Furthermore, to confirm the functionality of this system, we generated a rAAV-L-AD harboring a short hairpin RNA targeting ATP5B (rAAV-L-AD-shATP5B) and found that it caused a significant decrease in ATP5B mRNA levels when transduced into HEK293EB cells, suggesting that it was functional. Thus, our system successfully packaged L-AD into capsids with minimal contamination of plasmid DNA, offering a novel functional packaging platform without causing plasmid backbone encapsidation.

The landscape of genetic aberrations in myxofibrosarcoma.

Myxofibrosarcoma (MFS) is a rare subtype of sarcoma, whose genetic basis is poorly understood. We analyzed 69 MFS cases using whole-genome (WGS), whole-exome (WES) and/or targeted-sequencing (TS). Newly sequenced genomic data were combined with additional deposited 116 MFS samples. WGS identified a high number of structural variations (SVs) per tumor most frequently affecting the TP53 and RB1 loci, 40% of tumors showed a BRCAness-associated mutation signature, and evidence of chromothripsis was found in all cases. Most frequently mutated/copy number altered genes affected known disease drivers such as TP53 (56.2%), CDKN2A/B (29.7%), RB1 (27.0%), ATRX (19.5%) and HDLBP (18.9%). Several previously unappreciated genetic aberrations including MUC17, FLG and ZNF780A were identified in more than 20% of patients. Longitudinal analysis of paired diagnosis and relapse time points revealed a 1.2-fold mutation number increase accompanied with substantial changes in clonal composition over time. Our study highlights the genetic complexity underlying sarcomagenesis of MFS.

De Novo Mutations Activating Germline TP53 in an Inherited Bone-Marrow-Failure Syndrome.

Inherited bone-marrow-failure syndromes (IBMFSs) include heterogeneous genetic disorders characterized by bone-marrow failure, congenital anomalies, and an increased risk of malignancy. Many lines of evidence have suggested that p53 activation might be central to the pathogenesis of IBMFSs, including Diamond-Blackfan anemia (DBA) and dyskeratosis congenita (DC). However, the exact role of p53 activation in each clinical feature remains unknown. Here, we report unique de novo TP53 germline variants found in two individuals with an IBMFS accompanied by hypogammaglobulinemia, growth retardation, and microcephaly mimicking DBA and DC. TP53 is a tumor-suppressor gene most frequently mutated in human cancers, and occasional germline variants occur in Li-Fraumeni cancer-predisposition syndrome. Most of these mutations affect the core DNA-binding domain, leading to compromised transcriptional activities. In contrast, the variants found in the two individuals studied here caused the same truncation of the protein, resulting in the loss of 32 residues from the C-terminal domain (CTD). Unexpectedly, the p53 mutant had augmented transcriptional activities, an observation not previously described in humans. When we expressed this mutant in zebrafish and human-induced pluripotent stem cells, we observed impaired erythrocyte production. These findings together with close similarities to published knock-in mouse models of TP53 lacking the CTD demonstrate that the CTD-truncation mutations of TP53 cause IBMFS, providing important insights into the previously postulated connection between p53 and IBMFSs.

Genomic landscape and clonal evolution of acute myeloid leukemia with t(8;21): an international study on 331 patients.

Acute myeloid leukemia with t(8;21)(q22;q22) is characterized by considerable clinical and biological heterogeneity leading to relapse in up to 40% of patients. We sequenced coding regions or hotspot areas of 66 recurrently mutated genes in a cohort of 331 t(8;21) patients. At least 1 mutation, in addition to t(8;21), was identified in 95%, with a mean of 2.2 driver mutations per patient. Recurrent mutations occurred in genes related to RAS/RTK signaling (63.4%), epigenetic regulators (45%), cohesin complex (13.6%), MYC signaling (10.3%), and the spliceosome (7.9%). Our study identified mutations in previously unappreciated genes: GIGYF2, DHX15, and G2E3 Based on high mutant levels, pairwise precedence, and stability at relapse, epigenetic regulator mutations were likely to occur before signaling mutations. In 34% of RAS/RTKmutated patients, we identified multiple mutations in the same pathway. Deep sequencing (∼42 000×) of 126 mutations in 62 complete remission samples from 56 patients identified 16 persisting mutations in 12 patients, of whom 5 lacked RUNX1-RUNX1T1 in quantitative polymerase chain reaction analysis. KIT high mutations defined by a mutant level ≥25% were associated with inferior relapse-free survival (hazard ratio, 1.96; 95% confidence interval, 1.22-3.15; P = .005). Together with age and white blood cell counts, JAK2, FLT3-internal tandem duplicationhigh, and KIT high mutations were identified as significant prognostic factors for overall survival in multivariate analysis. Whole-exome sequencing was performed on 19 paired diagnosis, remission, and relapse trios. Exome-wide analysis showed an average of 16 mutations with signs of substantial clonal evolution. Based on the resemblance of diagnosis and relapse pairs, genetically stable (n = 13) and unstable (n = 6) subgroups could be identified.

Progress in Targeted Alpha-Particle-Emitting Radiopharmaceuticals as Treatments for Prostate Cancer Patients with Bone Metastases.

Bone metastasis remains a major cause of death in cancer patients, and current therapies for bone metastatic disease are mainly palliative. Bone metastases arise after cancer cells have colonized the bone and co-opted the normal bone remodeling process. In addition to bone-targeted therapies (e.g., bisphosphonate and denosumab), hormone therapy, chemotherapy, external beam radiation therapy, and surgical intervention, attempts have been made to use systemic radiotherapy as a means of delivering cytocidal radiation to every bone metastatic lesion. Initially, several bone-seeking beta-minus-particle-emitting radiopharmaceuticals were incorporated into the treatment for bone metastases, but they failed to extend the overall survival in patients. However, recent clinical trials indicate that radium-223 dichloride (223RaCl2), an alpha-particle-emitting radiopharmaceutical, improves the overall survival of prostate cancer patients with bone metastases. This success has renewed interest in targeted alpha-particle therapy development for visceral and bone metastasis. This review will discuss (i) the biology of bone metastasis, especially focusing on the vicious cycle of bone metastasis, (ii) how bone remodeling has been exploited to administer systemic radiotherapies, and (iii) targeted radiotherapy development and progress in the development of targeted alpha-particle therapy for the treatment of prostate cancer bone metastasis.

Genomic analysis of multiple myeloma using targeted capture sequencing in the Japanese cohort.

Previous genomic studies have revealed the genomic landscape of myeloma cells. Although some of the genomic abnormalities shown are believed to be correlated to the molecular pathogenesis of multiple myeloma and/or clinical outcome, these correlations are not fully understood. The aim of this study is to elucidate the correlation between genomic abnormalities and clinical characteristics by targeted capture sequencing in the Japanese multiple myeloma cohort. We analysed 154 patients with newly diagnosed multiple myeloma. The analysis revealed that the study cohort consisted of a less frequent hyperdiploid subtype (37·0%) with relatively high frequencies of KRAS mutation (36·4%) and IGH-CCND1 translocation (26·6%) compared with previous reports. Moreover, our targeted capture sequencing strategy was able to detect rare IGH-associated chromosomal translocations, such as IGH-CCND2 and IGH-MAFA. Interestingly, all 10 patients harboured MAX mutations accompanied by 14q23 deletion. The patients with del(17p) exhibited an unfavourable clinical outcome, and the presence of KRAS mutation was associated with shorter survival in patients with multiple myeloma, harbouring IGH-CCND1. Thus, our study provides a detailed landscape of genomic abnormalities, which may have potential clinical application for patients with multiple myeloma.

Physiological Srsf2 P95H expression causes impaired hematopoietic stem cell functions and aberrant RNA splicing in mice.

Splicing factor mutations are characteristic of myelodysplastic syndromes (MDS) and related myeloid neoplasms and implicated in their pathogenesis, but their roles in the development of MDS have not been fully elucidated. In the present study, we investigated the consequence of mutant Srsf2 expression using newly generated Vav1-Cre-mediated conditional knockin mice. Mice carrying a heterozygous Srsf2 P95H mutation showed significantly reduced numbers of hematopoietic stem and progenitor cells (HSPCs) and differentiation defects both in the steady-state condition and transplantation settings. Srsf2-mutated hematopoietic stem cells (HSCs) showed impaired long-term reconstitution compared with control mice in competitive repopulation assays. Although the Srsf2 mutant mice did not develop MDS under the steady-state condition, when their stem cells were transplanted into lethally irradiated mice, the recipients developed anemia, leukopenia, and erythroid dysplasia, which suggests the role of replicative stress in the development of an MDS-like phenotype in Srsf2-mutated mice. RNA sequencing of the Srsf2-mutated HSPCs revealed a number of abnormal splicing events and differentially expressed genes, including several potential targets implicated in the pathogenesis of hematopoietic malignancies, such as Csf3r, Fyn, Gnas, Nsd1, Hnrnpa2b1, and Trp53bp1 Among the mutant Srsf2-associated splicing events, most commonly observed were the enhanced inclusion and/or exclusion of cassette exons, which were caused by the altered consensus motifs for the recognition of exonic splicing enhancers. Our findings suggest that the mutant Srsf2 leads to a compromised HSC function by causing abnormal RNA splicing and expression, contributing to the deregulated hematopoiesis that recapitulates the MDS phenotypes, possibly as a result of additional genetic and/or environmental insults.

Prognostic relevance of integrated genetic profiling in adult T-cell leukemia/lymphoma.

Adult T-cell leukemia/lymphoma (ATL) is a heterogeneous group of peripheral T-cell malignancies characterized by human T-cell leukemia virus type-1 infection, whose genetic profile has recently been fully investigated. However, it is still poorly understood how these alterations affect clinical features and prognosis. We investigated the effects of genetic alterations commonly found in ATL on disease phenotypes and clinical outcomes, based on genotyping data obtained from 414 and 463 ATL patients using targeted-capture sequencing and single nucleotide polymorphism array karyotyping, respectively. Aggressive (acute/lymphoma) subtypes were associated with an increased burden of genetic and epigenetic alterations, higher frequencies of TP53 and IRF4 mutations, and many copy number alterations (CNAs), including PD-L1 amplifications and CDKN2A deletions, compared with indolent (chronic/smoldering) subtypes. By contrast, STAT3 mutations were more characteristic of indolent ATL. Higher numbers of somatic mutations and CNAs significantly correlated with worse survival. In a multivariate analysis incorporating both clinical factors and genetic alterations, the Japan Clinical Oncology Group prognostic index high-risk, older age, PRKCB mutations, and PD-L1 amplifications were independent poor prognostic factors in aggressive ATL. In indolent ATL, IRF4 mutations, PD-L1 amplifications, and CDKN2A deletions were significantly associated with shorter survival, although the chronic subtype with unfavorable clinical factors was only marginally significant. Thus, somatic alterations characterizing aggressive diseases predict worse prognosis in indolent ATL, among which PD-L1 amplifications are a strong genetic predictor in both aggressive and indolent ATL. ATL subtypes are further classified into molecularly distinct subsets with different prognosis. Genetic profiling might contribute to improved prognostication and management of ATL patients.

Genome analysis of myelodysplastic syndromes among atomic bomb survivors in Nagasaki.

Ionizing radiation is a risk factor for myeloid neoplasms including myelodysplastic syndromes (MDS), and atomic bomb survivors have been shown to have a significantly higher risk of MDS. Our previous analyses demonstrated that MDS among these survivors had a significantly higher frequency of complex karyotypes and structural alterations of chromosomes 3, 8, and 11. However, there was no difference in the median survival time between MDS among survivors compared with those of de novo origin. This suggested that a different pathophysiology may underlie the causative genetic aberrations for those among survivors. In this study, we performed genome analyses of MDS among survivors and found that proximally exposed patients had significantly fewer mutations in genes such as TET2 along the DNA methylation pathways, and they had a significantly higher rate of 11q deletions. Among the genes located in the deleted portion of chromosome 11, alterations of ATM were significantly more frequent in proximally exposed group with mutations identified on the remaining allele in 2 out of 5 cases. TP53, which is frequently mutated in therapy-related myeloid neoplasms, was equally affected between proximally and distally exposed patients. These results suggested that the genetic aberration profiles in MDS among atomic bomb survivors differed from those in therapy-related and de novo origin. Considering the role of ATM in DNA damage response after radiation exposure, further studies are warranted to elucidate how 11q deletion and aberrations of ATM contribute to the pathogenesis of MDS after radiation exposure.

Neuromuscular ultrasound for taxane peripheral neuropathy in breast cancer.

BACKGROUND: Our study aim was to evaluate neuromuscular ultrasound (NMUS) for the assessment of taxane chemotherapy-induced peripheral neuropathy (CIPN), the dose-limiting toxicity of this agent. METHODS: This cross-sectional study of breast cancer patients with taxane CIPN measured nerve cross-sectional area (CSA) by NMUS and compared with healthy historical controls. Correlations were determined between CSA and symptom scale, nerve conduction studies, and intraepidermal nerve fiber density (IENFD). RESULTS: A total of 20 participants reported moderate CIPN symptoms at a median of 3.8 months following the last taxane dose. Sural nerve CSA was 1.2 mm2 smaller than healthy controls (P ≤ .01). Older age and time since taxane were associated with smaller sural nerve CSA. For each 1 mm2 decrease in sural nerve CSA, distal IENFD decreased by 2.1 nerve/mm (R2 0.30; P = .04). CONCLUSIONS: These data support a sensory predominant taxane neuropathy or neuronopathy and warrant future research on longitudinal NMUS assessment of CIPN.

The Roles of Bone Marrow-Resident Cells as a Microenvironment for Bone Metastasis.

It has been appreciated that the cross talk between bone metastatic cancer cells and bone marrow microenvironment influence one another to worsen bone metastatic disease progression. Bone marrow contains various cell types, including (1) cells of mesenchymal origin (e.g., osteoblasts, osteocytes, and adipocytes), (2) cells of hematopoietic origin (e.g., osteoclast and immune cells), and (3) others (e.g., endothelial cells and nerves). The recent studies have enabled us to discover many important cancer-derived factors responsible for the development of bone metastasis. However, many critical questions regarding the roles of bone microenvironment in bone metastatic progression remain elusive. To answer these questions, a deeper understanding of the cross talk between bone metastatic cancer and bone marrow microenvironment is clearly warranted.

Gene expression and risk of leukemic transformation in myelodysplasia.

Myelodysplastic syndromes (MDSs) are a heterogeneous group of clonal hematopoietic disorders with a highly variable prognosis. To identify a gene expression-based classification of myelodysplasia with biological and clinical relevance, we performed a comprehensive transcriptomic analysis of myeloid neoplasms with dysplasia using transcriptome sequencing. Unsupervised clustering of gene expression data of bone marrow CD34+ cells from 100 patients identified 2 subgroups. The first subtype was characterized by increased expression of genes related to erythroid/megakaryocytic (EMK) lineages, whereas the second subtype showed upregulation of genes related to immature progenitor (IMP) cells. Compared with the first so-called EMK subtype, the IMP subtype showed upregulation of many signaling pathways and downregulation of several pathways related to metabolism and DNA repair. The IMP subgroup was associated with a significantly shorter survival in both univariate (hazard ratio [HR], 5.0; 95% confidence interval [CI], 1.8-14; P = .002) and multivariate analysis (HR, 4.9; 95% CI, 1.3-19; P = .02). Leukemic transformation was limited to the IMP subgroup. The prognostic significance of our classification was validated in an independent cohort of 183 patients. We also constructed a model to predict the subgroups using gene expression profiles of unfractionated bone marrow mononuclear cells (BMMNCs). The model successfully predicted clinical outcomes in a test set of 114 patients with BMMNC samples. The addition of our classification to the clinical model improved prediction of patient outcomes. These results indicated biological and clinical relevance of our gene expression-based classification, which will improve risk prediction and treatment stratification of MDS.

Genetic abnormalities in myelodysplasia and secondary acute myeloid leukemia: impact on outcome of stem cell transplantation.

Genetic alterations, including mutations and copy-number alterations, are central to the pathogenesis of myelodysplastic syndromes and related diseases (myelodysplasia), but their roles in allogeneic stem cell transplantation have not fully been studied in a large cohort of patients. We enrolled 797 patients who had been diagnosed with myelodysplasia at initial presentation and received transplantation via the Japan Marrow Donor Program. Targeted-capture sequencing was performed to identify mutations in 69 genes, together with copy-number alterations, whose effects on transplantation outcomes were investigated. We identified 1776 mutations and 927 abnormal copy segments among 617 patients (77.4%). In multivariate modeling using Cox proportional-hazards regression, genetic factors explained 30% of the total hazards for overall survival; clinical characteristics accounted for 70% of risk. TP53 and RAS-pathway mutations, together with complex karyotype (CK) as detected by conventional cytogenetics and/or sequencing-based analysis, negatively affected posttransplant survival independently of clinical factors. Regardless of disease subtype, TP53-mutated patients with CK were characterized by unique genetic features and associated with an extremely poor survival with frequent early relapse, whereas outcomes were substantially better in TP53-mutated patients without CK. By contrast, the effects of RAS-pathway mutations depended on disease subtype and were confined to myelodysplastic/myeloproliferative neoplasms (MDS/MPNs). Our results suggest that TP53 and RAS-pathway mutations predicted a dismal prognosis, when associated with CK and MDS/MPNs, respectively. However, for patients with mutated TP53 or CK alone, long-term survival could be obtained with transplantation. Clinical sequencing provides vital information for accurate prognostication in transplantation.