No full-text available
To read the full-text of this research,
you can request a copy directly from the authors.
Fusion of ETV6 to Neurotrophin-3 Receptor TRKC in Acute Myeloid Leukemia With t(12;15)(p13;q25)
Abstract
Chromosome translocations involving band 12p13 are known to be involved in a variety of hematologic malignancies, some of them resulting in rearrangement of the ETV6/TEL gene. Applying the fluorescence in situ hybridization (FISH) method, we found a cryptic translocation t(12;15)(p13;q25) in an adult acute myeloid leukemia (AML) patient. Hybridization with cosmid probes showed that the ETV6 gene was rearranged in this translocation. A patient-specific cDNA library was screened with ETV6 cDNA, and a novel fusion transcript was identified between the ETV6 and TRKC/NTRK3 gene located on 15q25. TRKC is a receptor tyrosine kinase that is activated by neurotrophin-3 (NT-3). It is known to be expressed broadly in neural tissues but not in hematologic cells, so far. ETV6-TRKC chimeric transcript encoded the pointed (PNT) domain of the ETV6 gene that fused to the protein-tyrosine kinase (PTK) domain of the TRKC gene. Two types of fusion transcript were determined, one that included the entire PTK domain of TRKC and the other in which the 3'-terminal 462 bp of TRKC was truncated within the PTK domain. Western blot analysis showed the expression of both chimeric proteins of 52 and 38 kD in size. Our results suggest that chimeric PTK expressed in the leukemic cells may contribute to cellular transformation by abnormally activating TRK signaling pathways. Moreover, this is the first report on truncated neurotrophin receptors associated in leukemia.
... Although other, non-NTRK ETV6-based fusions have been long reported to play a role in leukemogenesis [42,[48][49][50][51], an EN fusion was first reported in 1999 by Eguchi et al. [52,53] in a 59-year-old female with AML-M2 using fluorescence in situ hybridization (FISH). They identified two variants of the fusion with FISH (Fig. 2a). ...
... Moreover, one fusion included the entire PTK domain (encoding a 52 kD protein), while the other variant involved a truncated PTK domain (encoding a 38 kD protein). However, both of these chimeric proteins lack a 42-base-pair exon near the C-terminus of the NTRK3 protein that was reported thereafter in the EN oncofusion found in cases of congenital fibrosarcoma [52]. In addition to the 42-base-pair stretch, the fusion found in congenital fibrosarcoma contains ETV6 exons 1-5 versus 1-4 that are seen in EN fusions found in acute myeloid leukemia (AML). ...
... Analysis of this variant suggested that the first five exons of ETV6 fused with the kinase domain of NTRK3 (Fig. 2a). This was in contrast to the oncofusion described by Eguchi et al. [52], where only the first four exons of ETV6 were fused to NTRK3. ...
NTRK fusions are dominant oncogenic drivers found in rare solid tumors. These fusions have also been identified in more common cancers, such as lung and colorectal carcinomas, albeit at low frequencies. Patients harboring these fusions demonstrate significant clinical response to inhibitors such as entrectinib and larotrectinib. Although current trials have focused entirely on solid tumors, there is evidence supporting the use of these drugs for patients with leukemia. To assess the broader applicability for Trk inhibitors in hematological malignancies, this review describes the current state of knowledge about alterations in the NTRK family in these disorders. We present these findings in relation to the discovery and therapeutic targeting of BCR–ABL1 in chronic myeloid leukemia. The advent of deep sequencing technologies has shown that NTRK fusions and somatic mutations are present in a variety of hematologic malignancies. Efficacy of Trk inhibitors has been demonstrated in NTRK-fusion positive human leukemia cell lines and patient-derived xenograft studies, highlighting the potential clinical utility of these inhibitors for a subset of leukemia patients.
... The NTRK family consists of NTRK1, NTRK2, and NTRK3, which encode tropomyosin receptor kinase (TRK) A, B, and C, respectively. Oncogenic TRK activation is mainly caused by the fusion of NTRK genes and is involved in the pathogenesis of many tumors, including infantile fibrosarcoma [19], mesoblastic nephroma [20], acute myeloid and chronic eosinophilic leukemia [21,22], secretory breast carcinoma [23], mammary analog secretory carcinoma of the salivary gland [24], radiation-induced thyroid carcinoma [25], myofibroblast tumors [26] and WT GISTs [12][13][14][15]27]. TRK inhibitors, such as larotrectinib and entrectinib, have been approved to treat multiple cancers with NTRK fusions [28][29][30][31]. ...
... The NTRK genes encode TRK proteins that exert oncogenic effects on tumors, and the activation of most TRK proteins is caused by NTRK fusions [43]. NTRK fusions occur in a variety of cancers with different incidences [19][20][21][22][23][24][25][26]44]. TRK inhibitors, such as larotrectinib and entrectinib, showed encouraging antitumor efficacies in tumors with NTRK rearrangements and have been approved as treatments for multiple cancers harboring NTRK fusions [29][30][31]34,[45][46][47]. ...
Simple Summary
Wild-type GISTs are generally not sensitive to tyrosine kinase inhibitors. Tropomyosin receptor kinase inhibitors have been approved to be effective in multiple cancers with neurotrophic tyrosine receptor kinase (NTRK) fusions. Although NTRK fusions are rare in wild-type GISTs, the unambiguous diagnosis can bring clinical benefits to the patients. The immunohistochemistry staining of Pan-TRK, next-generation sequencing or fluorescence in situ hybridization have been used to screen NTRK fusions in a few cases of wild-type GIST, and each technique has its advantages and drawbacks. This study aimed to identify NTRK fusions in wild-type GISTs with the above three methods and explore the clinicopathological and genetic features of GISTs with NTRK functions based on our patients and the literature. The findings from this study provide new evidence to establish a clinical protocol for screening GISTs with NTRK fusions and an overall view of the clinicopathological characteristics of GISTs with NTRK fusions.
Abstract
The most common mutations in gastrointestinal stromal tumors (GISTs) are KIT or PDGFRA mutations. Recently, neurotrophic tyrosine receptor kinase (NTRK) fusions have been reported in WT GISTs, which increased interest in introducing tropomyosin receptor kinase (TRK) inhibitors as treatments for GISTs with NTRK fusions. Hence, we aimed to screen NTRK fusions in WT GISTs; we used fluorescence in situ hybridization (FISH), next-generation sequencing (NGS), and immunohistochemistry (IHC) to screen NTRK fusions in 46 WT GISTs and evaluate each method. We further reviewed NTRK fusion-positive GISTs from the literature and performed clinical and pathological analyses; two GISTs with an ETV6-NTRK3 fusion (5%) were identified, while only one (50%) was positive for Pan-TRK expression. On the other hand, among the six GISTs with Pan-TRK-positive expression, only one (17%) harbored NTRK fusion. The literature review revealed the strong consistency between FISH and NGS and the limited value of Pan-TRK IHC in screening NTRK fusions in GISTs. In addition, the clinical and pathological analysis showed that GISTs with NTRK rearrangement occurred less frequently in the stomach, were more frequently larger in size, and the epithelioid type presented with a higher risk of recurrence. The NTRK3 fusion has been more common than the NTRK1 fusion in GISTs to date; our study identified two ETV6-NTRK3 fusions in 46 WT GISTs. Compared with FISH and IHC, NGS is preferred for screening WT GISTs, including NTRK rearrangements. However, since GISTs with NTRK fusions are rare, further studies including more samples and mechanistic investigations should be conducted in the future.
... The pathway enrichment analysis (blue bars) revealed that 25 pathways were highly enriched for the target genes of the upregulated miRNAs studied. Among them, the highly enriched pathways related to hematopoiesis were MAPK [21], insulin signaling pathway [2,40], neurotrophin signaling pathway [18], Ras [27], FOXO [38], and mTOR signaling pathways [50], in addition to pathways involved in cancer especially AML and CML as shown in Fig. 3. ...
... Of them, insulin growth factors (IGF-1 and IGF-2) and insulin signaling pathway have a major function in hematopoiesis. IGF-1 stimulates erythropoiesis in vitro and in vivo [2,40] as well as the neurotrophin signaling pathway [18]. Moreover, Ras is one of the signaling molecules activated during erythropoiesis and appear important for the balance of proliferation/differentiation/apoptosis of erythroid cells [27]. ...
Background
Fetal hemoglobin (HbF) induction has shown promise for the treatment of β-hemoglobinopathies. HbF induction in β-thalassemia could overcome ineffective hematopoiesis and thus terminate transfusion dependency for formerly transfusion dependant patients. Several miRNAs have been found to reactivate γ-globin expression and increase HbF. In this study, we aimed to investigate the expression of 4 miRNAs (miR-15a, miR-16-1, miR-96, and miR-486-3p) in high HbF thalassemia patients and correlate their levels with the patients’ HbF levels then, in order to predict the exact role of the studied miRNAs in hematopoiesis, a bioinformatic analysis was carried out. We went through this bioinformatic analysis to determine the network of genes regulated by miRNAs and further investigate the interaction between all of them through their involvement in hematopoiesis. In this study, the differential expression was measured by qRT-PCR for 40 patients with high HbF and compared to 20 healthy controls. Bioinformatics was conducted involving functional annotation and pathway enrichment analyses.
Results
The studied microRNAs were significantly deregulated in thalassemia patients in correlation with HbF. Functional annotation and pathway enrichment analyses revealed a major role of miR-486-3p and miR-15a in HbF induction.
Conclusion
MiR-486-3p and miR-15a are crucial for HbF induction. Further validating studies are needed.
... 21 The tolerability and effectiveness of entrectinib and larotrectinib for Trk-positive solid tumors have sparked our interest in assessing the role of NTRKs in hematological malignancies. Studies thus far have primarily focused on ETV6-NTRK3 fusions in patients with acute myeloid leukemia (AML), 22,23 chronic eosinophilic leukemia, 24 and Philadelphia chromosome-like acute lymphoblastic leukemia (ALL). 25,26 In animal models of AML, entrectinib treatment resulted in tumor regression and elimination of residual ETV6-NTRK3 1 cancer cells from the marrow. ...
... **P , .01; ***P , .001.NTRK POINT MUTATIONS IN HEMATOLOGIC NEOPLASMSblood® 11 JUNE 2020 | VOLUME 135, NUMBER 24 2165 activated by much broader mechanisms, including oncogenic fusions,22,25,28 deletion/truncation events, 51,52 transcriptional upregulation,40,53,54 and point mutations32,33 in leukemia. ...
Much of what is known about the neurotrophic receptor tyrosine kinase (NTRK) genes in cancer is through the identification and characterization of activating Trk fusions across many tumor types. A resurgence of interest in these receptors has emerged owing to the realization that they are promising therapeutic targets. The remarkable efficacy of the pan-Trk inhibitors, larotrectinib and entrectinib, in clinical trials led to their accelerated, tissue agnostic FDA approval for adult and pediatric patients with Trk-driven solid tumors. Despite our enhanced understanding of Trk biology in solid tumors, the importance of Trk signaling in hematological malignancies is underexplored and warrants further investigation. Herein, we describe mutations in NTRK2 and NTRK3 that were identified via deep sequencing of 185 patients with hematological malignancies. Ten patients contained a point mutation in NTRK2 or NTRK3. Among these patients, we identified nine unique point mutations. Of these nine mutations, four were oncogenic-NTRK2A203T, NTRK2R458G, NTRK3E176D, and NTRK3L449F-as determined via cytokine-independent cellular assays. Our data demonstrate that these mutations have transformative potential to promote downstream survival signaling and leukemogenesis. Specifically, the three mutations located within the extracellular (i.e., NTRK2A203T and NTRK3E176D) and transmembrane (i.e., NTRK3L449F) domains increased receptor dimerization and cell-surface abundance. The fourth mutation, NTRK2R458G, residing in the juxtamembrane domain, activates TrkB via non-canonical mechanisms that may involve altered interactions between the mutant receptor and lipids in the surrounding environment. Importantly, these four activating mutations can be clinically targeted using entrectinib. Our findings contribute to ongoing efforts focused on defining the mutational landscape that drives hematological malignancies and underscore the utility of FDA-approved Trk inhibitors for patients with aggressive Trk-driven leukemias.
... TRK expression occurs in myeloid leukemia cell lines and in primary leukemic cells from patients with AML (66). Subsequent studies have shown that activation of TRK receptors in hematologic malignancies could result from chromosomal rearrangements (67), point mutations (68), truncations (69), and transcriptional changes (70). Although the prevalence of NTRK-mediated cancers is <1%, fusions involving NTRK3 were observed in various hematologic malignancies including AML (67). ...
... Subsequent studies have shown that activation of TRK receptors in hematologic malignancies could result from chromosomal rearrangements (67), point mutations (68), truncations (69), and transcriptional changes (70). Although the prevalence of NTRK-mediated cancers is <1%, fusions involving NTRK3 were observed in various hematologic malignancies including AML (67). The efficacy of TRK inhibitors has been demonstrated in NTRK fusion-positive human leukemia cell lines and patient-derived xenograft studies, highlighting the potential clinical utility of these inhibitors for a subset of patients with leukemia (71,72). ...
To study mechanisms underlying resistance to the BCL2 inhibitor venetoclax in acute myeloid leukemia (AML), we used a genome-wide CRISPR/Cas9 screen to identify gene knockouts resulting in drug resistance. We validated TP53, BAX, and PMAIP1 as genes whose inactivation results in venetoclax resistance in AML cell lines. Resistance to venetoclax resulted from an inability to execute apoptosis driven by BAX loss, decreased expression of BCL2, and/or reliance on alternative BCL2 family members such as BCL2L1. The resistance was accompanied by changes in mitochondrial homeostasis and cellular metabolism. Evaluation of TP53 knockout cells for sensitivities to a panel of small-molecule inhibitors revealed a gain of sensitivity to TRK inhibitors. We relate these observations to patient drug responses and gene expression in the Beat AML dataset. Our results implicate TP53, the apoptotic network, and mitochondrial functionality as drivers of venetoclax response in AML and suggest strategies to overcome resistance.
Significance
AML is challenging to treat due to its heterogeneity, and single-agent therapies have universally failed, prompting a need for innovative drug combinations. We used a genetic approach to identify genes whose inactivation contributes to drug resistance as a means of forming preferred drug combinations to improve AML treatment.
See related commentary by Savona and Rathmell, p. 831.
This article is highlighted in the In This Issue feature, p. 813
... Furthermore, they found that 12p aberrations were associated with better prognosis in myelodysplasia and less conversion to AML. 17 The most important genes that is carried on 12p and was identified as one of the genes involved in the pathogenesis of AML is ETV6. 18,19 This gene is a member of transcription regulators and responsible for DNA binding activities. 18 In our study, we found that the overall survival and leukemia free survival was less in 12p AML than 12p negative AML. ...
... 18,19 This gene is a member of transcription regulators and responsible for DNA binding activities. 18 In our study, we found that the overall survival and leukemia free survival was less in 12p AML than 12p negative AML. This consistent with literature that found that 12p aberrations were associated with worse prognosis. ...
Objectives
To investigate the correlation of homeobox (HOX) transcript antisense RNA expression with clinicopathological features and the clinical prognosis of the patients with chromosome 12p abnormalities associated acute myeloid leukemia (AML). We also investigate the association of 12p chromosomal on the expression of HOTAIR, miRNA‐193a, and c‐kit gene as targeting genes for HOTAIR in AML.
Methods
AML patients with 12p chromosomal abnormalities were recruited and compared to AML with other chromosomal abnormalities rather than 12p. The long noncoding RNA (lncRNA) “HOTAIR,” miR‐193a, and c‐Kit genes expression were measured in bone marrow samples using Syber green based real‐time polymerase chain reaction.
Results
We found a significant difference for the expression levels of HOTAIR, c‐kit, and miR‐193a between 12p abnormalities associated AML and those without. The survival analysis revealed that patient's with low expression levels of HOTAIR and c‐kit had significantly better survival and leukemia free survival. In contrast, miR‐193a was associated with better overall survival but not leukemia free survival.
Conclusion
12p abnormalities associated AML were associated with worse prognosis. Our results proved that HOTAIR, miR‐193a, and c‐kit genes are independent prognostic predictors in 12p chromosomal associated AML; therefore it may represent a novel therapeutic application in AML in the future.
... Despite the promise of TRK inhibition in solid tumors, the frequency and characteristics of TRK fusions in hematologic malignancies has not been systematically evaluated nor is there knowledge of the clinical efficacy of TRK inhibition in any hematologic malignancy. Given that individual patients with TRK fusions have been described in acute myeloid leukemia (AML), (12)(13)(14) Philadelphia chromosome-like B-cell acute lymphoblastic leukemia (Ph-like B-ALL), (15) and histiocytosis,(16) TRK fusions may have clinical importance in hematologic malignancies. Here we describe the frequency and characteristics of NTRK1-3 fusions across >7,000 pediatric and adult patients with hematologic malignancies and report the efficacy of TRK inhibition in a refractory AML patient with a novel ETV6-NTRK2 fusion. ...
... We herein describe that TRK fusions occur across patients with a wide variety of hematologic malignancies and are amenable to TRK inhibition. In addition to prior data identifying TRK fusions in patients with AML, (12)(13)(14) Ph-like B-ALL, (15) and histiocytosis, (16) we also document TRK fusions in multiple myeloma and dendritic cell neoplasms. Although the frequency of TRK fusions appears to be rare across a very wide spectrum of leukemias and lymphoid malignancies studied here, the antitumor activity of larotrectinib seen here calls for systematic identification of TRK fusions in the clinical evaluation of hematologic malignancy patients with refractory disease. ...
Rearrangements involving the neurotrophic receptor kinase genes (NTRK1, NTRK2, and NTRK3; hereafter referred to as TRK) produce oncogenic fusions in a wide variety of cancers in adults and children. Although TRK fusions occur in <1% of all solid tumors, inhibition of TRK results in profound therapeutic responses resulting in breakthrough FDA-approval of the TRK inhibitor larotrectinib for adult and pediatric solid tumor patients regardless of histology. In contrast to solid tumors, the frequency of TRK fusions and clinical effects of targeting TRK in hematologic malignancies is unknown. Here, through an evaluation for TRK fusions across > 7,000 patients with hematologic malignancies, we identified TRK fusions in acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), histiocytosis, multiple myeloma and dendritic cell neoplasms. Although TRK fusions occurred in only 0.1% of patients (8 out of 7,311 patients), they conferred responsiveness to TRK inhibition in vitro and in vivo in a patient-derived xenograft and a corresponding AML patient with ETV6-NTRK2 fusion. These data identify that despite their individual rarity, collectively TRK fusions are present in a wide variety of hematologic malignancies and predict clinically significant therapeutic responses to TRK inhibition.
... However, due to the fusion of NTRK3 and ETS variant transcription factor 6 (ETV6), ETV6-NTRK3 (EN) can self-phosphorylate (13). The EN fusion gene has been identified in congenital fibrosarcoma (14), secretory breast carcinoma (15), AML (16,17), mammary analog secretory carcinoma of the salivary gland (18), chronic eosinophilic leukemia (19) congenital mesoblastic nephroma (20), and thyroid cancer associated with 131 I radiation exposure (21). Due to the already low prevalence of NTRK fusions in most tumours (<1%) EN cases are even more uncommon (22). ...
ETS variant transcription factor 6 (ETV6)-neurotrophic receptor tyrosine kinase 3 (NTRK3) (EN) fusions are typically found in rare diseases, such as primary renal fibrosarcoma (only six cases have been reported), secretory carcinoma of the breast and salivary gland (1 case), and AML (4 cases). Few cases have been reported, and expression of the EN gene fusion requires additional clinical data and fundamental research to be supported. The aim of the present study was to determine the inhibitory effect of Andrographis paniculata methanol extract (MeAP) on EN-related cell lines, IMS-M2 and BaF3/EN, as well as evaluate the mechanism of action. Vero cells were used as control cells. Trypan blue staining and MTT were used to evaluate the inhibitory effect of MeAP on tested cells. Western blotting and immunoprecipitation were used to detect the activation of EN after MeAP treatment. The IC50 values of MeAP were found to be 12.38±0.57 µg/ml (IMS-M2) and 13.06±0.49 µg/ml (BaF3/EN). MeAP was observed to inhibit cell proliferation in a time, dose, and cell density-dependent manner. The IC50 value for MeAP in Vero cells was markedly higher, at 109.97±4.24 (µg/ml), indicating a much less sensitive effect. Furthermore, MeAP treatment inhibited EN phosphorylation and induced apoptosis in these cells. Collectively, the present study demonstrated that MeAP has an oncogenic effect on EN fusion-positive cell lines, in particular.
... JAK2-activating alterations may predict sensitivity to HDAC inhibitors [156][157][158]; a phase I/II study of givinostat for patients with JAK2 V617F-mutated polycythemia vera reported ORRs from 72.7% (8/11, 1 CR) to 80.7% (25/31, 3 CRs) across trial arms [159]. Other alterations that activate JAK2, such as fusions [160][161][162][163][164][165] or amplifications [166], may also confer sensitivity to JAK2 inhibitors on the basis of clinical data in myeloid neoplasms as well as preclinical data. ...
Comprehensive genomic profiling examinations (CGPs) have recently been developed, and a variety of tumor-agnostic mutations have been detected, leading to the development of new molecular-targetable therapies across solid tumors. In addition, the elucidation of hereditary tumors, such as breast and ovarian cancer, has pioneered a new age marked by the development of new treatments and lifetime management strategies required for patients with potential or presented hereditary cancers. In acute myeloid leukemia (AML), however, few tumor-agnostic or hereditary mutations have been the focus of investigation, with associated molecular-targeted therapies remaining poorly developed. We focused on representative tumor-agnostic mutations such as the TP53, KIT, KRAS, BRCA1, ATM, JAK2, NTRK3, FGFR3 and EGFR genes, referring to a CGP study conducted in Japan, and we considered the possibility of developing molecular-targeted therapies for AML with tumor-agnostic mutations. We summarized the frequency, the prognosis, the structure and the function of these mutations as well as the current treatment strategies in solid tumors, revealed the genetical relationships between solid tumors and AML and developed tumor-agnostic molecular-targeted therapies and lifetime management strategies in AML.
... For example, NTRK2 forms fusions with partners such as AGBL4, NACC2, QKI, and VCL, which were identified in non-brainstem high-grade glioma, soft tissue liposarcoma, head and neck squamous cell carcinoma, pilocytic astrocytoma, ganglioglioma, and diffuse intrinsic pontine glioma. NTRK3 fusion partners include ETV6, BTBD1, and EML4, which were identified in diffuse intrinsic pontine glioma, congenital fibrosarcoma, papillary thyroid carcinoma, mammary analog secretory carcinoma (MASC) of the thyroid gland, secretory breast cancer, and inflammatory myofibroblastic tumor [48,[57][58][59][60] , as well as in hematological malignancies such as acute myeloid leukemia, Philadelphia-like acute lymphoblastic leukemia, and chronic eosinophilic leukemia [61][62][63] . ...
The rapid advances in the understanding of oncogenic process and the maturation of affordable precision diagnostic tools have enabled the development of targeted therapeutic agents, such as those targeting BCR-ABL, epithelial growth factor receptor L858R, EML4-anaplastic lymphoma kinase, and BRAF V600E, to treat cancers that harbor specific molecular alterations. Traditionally, each targeted drug has been developed for a particular tumor type where such alteration is most frequently found. Recently, the widespread adoption of next generation sequencing has led to an increase in the identification of rare and ultra-rare alterations, and, in some cases, the same rare alterations are found across multiple tumor types. The rarity of these alterations makes clinical trials traditionally designed for specific tumor types infeasible. As a result, tissue-agnostic trials have been developed to study the efficacy of these treatments and increase patient access. This review summarizes current successful cases of tissue-agnostic development, such as drugs targeting tropomyosin receptor kinase fusions, and proposes the next wave of potential tissue-agnostic targets, including fusions of ROS1, anaplastic lymphoma kinase, fibroblast growth factor receptor, and rearranged during transfection. In addition, the advantages and the challenges of such approach are discussed in the context of clinical development and approval.
... Mammakarzinom (Eguchi et al., 1999;Tognon et al., 2002) TEL/JAK2 t(9;12)(p24;p13) Prä-B-Zell-ALL, T-ALL Peeters et al., 1997) ...
Background: Dendritic cell (DC) therapy offers a novel therapeutic approach in the treatment of different forms of cancer. In the case of leukemia the aim of the DC therapy would be to differentiate leukemic blasts through cytokine stimulation into dendritic cells. Thus known and unknown tumor-associated antigens are presented to the immune system (Blair et al., 2001; B. A. Choudhury et al., 1999; Cignetti et al., 1999). Further an ideal target as a tumor-associated antigen could be the oncoprotein TEL/AML1, which is a result of the chromosomal translocation (12;21)(p13;q22) and represents the most common fusion gene in childhood B-cell precursor acute lymphoblastic leukemia (BCP-ALL) (Shurtleff et al., 1995). Yotnda et al. has shown that this epitope created through the fusion gene of TEL and AML1 leads to a cytotoxic T-cell response (Yotnda et al., 1998). In 2009 Schmidt et al. successfully developed cytokin-cocktails, which differentiated TEL/AML1-positive leukemic blasts into dendritic cells (Schmidt et al., 2009). Due to these findings, it is assumed that the addition of the TEL/AML1 peptide would enhance the cytokine- stimulated differentiation of TEL/AML1 positive leukemic blasts into leukemic DCs. This could lead to a specific DC-therapy against the TEL/AML1 positive ALL. Methods: We used a peptide sequence of the TEL/AML1 oncoprotein representing the fusion region. As this peptide is not synthesized naturally by healthy cells, it would have the highest probability of triggering an immunogenic reaction. Via flow cytometry several concentration levels of the TEL/AML1 peptide were examined on the differentiation of the cell lines into APCs in vitro. As a source for leukemic blasts we used the human TEL/AML1 positive acute lymphoblastic leukemic (ALL) cell line of REH cells comparing them with the human TEL/AML1 negative chronic myeloid leukemic (CML) cell line of K562 and human monocytes. Results: It was possible to moderately induce monocytes into mature antigen presenting cells. Unexpectedly though the TEL/AML1 peptide decreased the differentiation of the TEL/AML1 positive ALL cell line (REH) into DCs. In comparison the peptide didn’t have any effect on the TEL/AML1 negative CML cell line (K562). Conclusion: The TEL/AML1 fusion sequence seems to inhibit the differentiation into leukemic DCs in spite of cytokine stimulation. Due to this, the peptide is probably not suitable for a dendritic cell therapy. This effect seems to be specific for the TEL/AML1 positive leukemic cell line, as no effect was observed on the TEL/AML1 negative CML cell line. The results show that the TEL/AML1 peptide could be responsible in conserving a leukemic state on TEL/AML1 positive leukemic blasts. A possible cause could be an intracellular binding or interaction with another protein. This could lead to an inhibition of differentiation. Due to these discoveries further research is needed to evaluate the function of the TEL/AML1 fusion peptide in the pathomechanism of the TEL/AML1 positive ALL.
... 15 In rare instances, the same translocation and gene fusion have been demonstrated in 2 or more unrelated neoplasms, for example, ALK-CLTC in inflammatory myofibroblastic tumor and anaplastic large cell lymphoma and ETV6-NTRK3 in infantile fibrosarcoma, acute myeloid leukemia, secretory carcinoma of the breast, and the mammary analogue secretory carcinoma of the salivary glands. [16][17][18][19][20][21] These occurrences suggest that the type of cell the fusion oncogene is targeting may determine whether the neoplastic transformation is of mesenchymal, hematologic, or epithelial lineage. tumors whose fusion oncogenes encode for chimeric tyrosine kinases that elicit deregulation of kinase signaling pathways. ...
Context.—Soft tissue pathology encompasses a remarkably diverse assortment of benign and malignant soft tissue tumors. Rendering a definitive diagnosis is complicated not only by the large volume of existing histologic subtypes (>100) but also frequently by the presence of overlapping clinical, histologic, immunohistochemical, and/or radiographic features. During the past 3 decades, mesenchymal tumor–specific, cytogenetic and molecular genetic abnormalities have demonstrated an increasingly important, ancillary role in mesenchymal tumor diagnostics.
Objectives.—To review molecular diagnostic tools available to the pathologist to further classify specific soft tissue tumor types and recurrent aberrations frequently examined. Advantages and limitations of individual approaches will also be highlighted.
Data Sources.—Previously published review articles, peer-reviewed research publications, and the extensive cytogenetic and molecular diagnostic experience of the authors to include case files of The University of Nebraska Medical Center.
Conclusions.—Cytogenetic and molecular genetic assays are used routinely for diagnostic purposes in soft tissue pathology and represent a powerful adjunct to complement conventional microscopy and clinicoradiographic evaluation in the formulation of an accurate diagnosis. Care should be taken, however, to recognize the limitations of these approaches. Ideally, more than one technical approach should be available to a diagnostic laboratory to compensate for the shortcomings of each approach in the assessment of individual specimens.
... The chromosomal translocation t(12;15)(p13;q25), through which the N-terminal helix-loop-helix (HLH) dimerization domain of ETV6 fuses to the C-terminal protein tyrosine kinase (PTK) domain of NTRK3, has been implicated in both solid tumors and leukemias. (4)(5)(6)(7)(8)(9)(10) Oligomerization of the ETV6 HLH domain and activation of NTRK3 PTK domain, which leads to constitutive stimulation of Ras-Erk1/2, PI3K-Akt, and IGF1R signaling pathways (10,11), are essential for ETV6-NTRK3-induced cellular transformation. (12,13) Transduction of murine BM cells with ETV6-NTRK3expressing retroviruses followed by transplantation results in a rapid and fatal myeloproliferative disease resembling AML. ...
Background: The ETV6-NTRK3 fusion transcript has been found to recurrently identified in both solid tumors and leukemias. It has attracted a lot of interest for the clinical targeted therapy and genetic features in ETV6-NTRK3 positive solid tumors. While the t(12;15)(p13;q25)/ETV6-NTRK3 is a rare genetic aberration in hematologic malignancies at a low frequency of ≤1%. An accumulation of reported cases would be needed to discuss clinical and cytogenetic characteristics of the important entity. Therefore, it is useful to report every case for clinical implication of prognosis or therapy.
Case presentation: We report the case of a previously healthy 30-year-old female, who was diagnosed as acute myeloid leukemia (AML) and presented with chemoresistance and short survival. The patient was treated with four cycles of chemotherapy but failed to achieve remission. Then the patient underwent a salvage haploidentical stem cell transplantation. Unfortunately, she worsened within 1 month and died of the refractory leukemia 35 days after transplantation. ETV6-NTRK3 rearrangement was revealed by RNA sequencing but the chromosomal translocation t(12;15)(p13;q25) was cryptic by conventional karyotype analysis. We review the literature and find that the ETV6-NTRK3 fusion transcript is associated with cryptic karyotype, trisomy 8, aggressive and poor prognosis in hematologic malignancy. The clinical and laboratory characteristics of ETV6-NTRK3 positive hematologic malignancies are different from those of solid tumors. Nevertheless, tropomyosin receptor kinase (TRK) inhibitor has powerful anti-tumor activity in patients with TRK fusion–driven cancers, regardless of the tumor type.
Conclusions: We demonstrated that TRK inhibitor larotrectinib is an effective treatment on the primary bone marrow (BM) cells derived from the patient described here with ETV6-NTRK3 positive AML. Our report stresses the importance of screening for ETV6-NTRK3 fusion transcript in newly diagnosed leukemias and clinical treatment of TRK inhibitor in hematologic malignancies.
... Over the last years, new tumor entities, especially in the already very heterogenous group of soft tissue tumors, were described harboring NTRK fusions [6,8,[19][20][21]. In addition, NTRK fusions were detected in other more common cancer types, such as papillary thyroid carcinoma, gastrointestinal stromal tumor (GIST), gliomas, non-small cell lung cancer (NSCLC), colorectal carcinoma (most frequently found in MSI-high carcinoma associated with MLH1 promoter hypermethylation), as well as malignant melanomas, uterine sarcomas, and pancreatic adenocarcinomas, however in less than 1% of all solid tumors overall [3,6,[22][23][24][25][26][27][28][29][30][31][32][33][34][35]. ...
Fusions involving NTRK1, NTRK2, and NTRK3 are oncogenic drivers occurring in a spectrum of mesenchymal neoplasms ranging from benign to highly malignant tumors. To gain further insights into the staining profile with the pan-TRK assay, we analyzed a large number of soft tissue sarcomas and correlated our findings with molecular testing. Additionally, we expand the spectrum of NTRK-fusion tumors by reporting a mesenchymal lesion in the lung as well as a mesenchymal skin lesion in the spectrum of benign fibrous histiocytoma with NTRK—fusion. We retrospectively reviewed soft tissue sarcomas diagnosed at the Diagnostic and Research Institute of Pathology, Medical University of Graz, between 1999 and 2019, and cases from the consultation files of one of the authors (BLA). In total, 494 cases were analyzed immunohistochemically with pan-TRK antibody (clone EPR17341, RTU, Roche/Ventana) and positive cases (defined as any cytoplasmic/nuclear staining in more than 1% of tumor cells) underwent next-generation sequencing (NGS). Immunohistochemical staining was observed in 16 (3.2%) cases. Eleven cases with focal weak and moderate cytoplasmic/membranous or focal moderate to strong nuclear staining did not harbor an NTRK-fusion (three synovial sarcomas, three leiomyosarcomas, two extraskeletal myxoid chondrosarcomas, and one each: dedifferentiated liposarcoma, pleomorphic liposarcoma, and myxofibrosarcoma). Four cases showed strong diffuse nuclear and/or cytoplasmatic staining, and one case showed diffuse, but weak cytoplasmic staining. All these cases demonstrated an NTRK-fusion (LMNA-NTRK1, IRF2BP2-NTRK1, TMB3-NTRK1, ETV6-NTRK3, RBPMS-NTRK3). Pan-TRK assay (clone EPR17341, RTU, Roche, Ventana) immunohistochemistry serves as a reliable diagnostic marker that can also be expressed in non-NTRK-rearranged mesenchymal neoplasms. It can be used as a surrogate marker for identification of NTRK fusion, nevertheless, an RNA-based NGS for detection of the specific fusion should be performed to confirm the rearrangement, if patients are undergoing targeted therapy. Additionally, we identified NTRK-fusion-positive, primary mesenchymal tumors of the lung and the skin.
... Secretory carcinomas of all anatomic sites harbor the t(12;15)(p13;q25) balanced translocation fusing ETV6 on chromosome 12 with NTRK3 on chromosome 15 [74,86]. Not unique to SC, ETV6-NTRK3 fusions have been identified in a range of other malignancies including PTC and infantile fibrosarcoma [87][88][89][90][91][92]. According to a study of over 38,000 malignancies of different types and sites, NTRK1-3 fusions are most common in inflammatory myofibroblastic tumor (17.7%), salivary gland carcinomas (5.1%), and thyroid carcinomas (2.3%) and are infrequent in other tissues (< 1.0%) [86]. ...
Thyroid carcinomas represent 3.2% of all new cases of cancer in the United States. Whereas most thyroid tumors arise from follicular cells or, less commonly, parafollicular cells, the derivation of some rare primary thyroid carcinoma subtypes is less clear and represents an area of evolving knowledge. Primary thyroid carcinomas that resemble neoplasms characteristic of the salivary glands (“salivary-like” primary thyroid carcinomas) arguably represent some of the most unusual primary thyroid tumors. Herein, we have undertaken a review of the literature in order to present a comprehensive overview of salivary-like primary thyroid carcinomas including: mucoepidermoid carcinoma, sclerosing mucoepidermoid carcinoma with eosinophilia, and secretory carcinoma. Awareness of these unusual, distinct primary tumors is important for timely diagnosis and optimal patient management. This review highlights these three salivary-like carcinomas, with special emphasis on developments since publication of the World Health Organization (WHO) 2017 Classification of Tumours of Endocrine Organs.
... The ETV6-NTRK3 gene fusion was originally identified in congenital fibrosarcomas with t(12;15)(p13;q25) translocations (Table 1) [57]. Subsequent studies demonstrated that the ETV6-NTRK3 fusion, in contrast to many other gene fusions, may occur in a variety of neoplasms in different anatomical locations, including secretory carcinomas of the breast and salivary glands [58,59], mesoblastic nephromas [60], acute leukemias [61], pediatric non-brainstem high-grade gliomas [62], papillary thyroid carcinomas [63], colorectal carcinomas [64], and inflammatory myofibroblastic tumors (Table 1) [65]. The fusion links the dimerization domain of the ETV6 transcription factor to the tyrosine kinase domain of NTRK3 (neurotrophin-3 receptor) leading to constitutive activation of the NTRK3 kinase [66]. ...
Chromosome rearrangements resulting in pathogenetically important gene fusions are a common feature of many cancers. They are often potent oncogenic drivers and have key functions in central cellular processes and pathways and encode transcription factors, transcriptional co-regulators, growth factor receptors, tyrosine kinases, and chromatin modifiers. In addition to being useful diagnostic biomarkers, they are also targets for development of new molecularly targeted therapies. Studies in recent decades have shown that several oncogenic gene fusions interact with the insulin-like growth factor (IGF) signaling pathway. For example, the MYB–NFIB fusion in adenoid cystic carcinoma is regulated by IGF1R through an autocrine loop, and IGF1R is a downstream target of the EWSR1–WT1 and PAX3–FKHR fusions in desmoplastic small round cell tumors and alveolar rhabdomyosarcoma, respectively. Here, we will discuss the mechanisms behind the interactions between oncogenic gene fusions and the IGF signaling pathway. We will also discuss the role of therapeutic inhibition of IGF1R in fusion gene driven malignancies.
... The ETV6/NTRK3 oncogene fusion was first clinically described in 1998 in congenital fibrosarcoma [6]. Since then, this novel oncogene has been implicated in carcinogenesis of secretory breast carcinoma, congenital mesoblastic nephroma, acute myeloid leukemia (AML), salivary gland tumors, and PTC [7][8][9][10][11][12]. ...
Patient: Female, 32
Final Diagnosis: Papillary thyroid carcinoma
Symptoms: Cervical node metastasis
Medication: —
Clinical Procedure: Radioactive iodine treatment
Specialty: Oncology
Objective
Rare co-existance of disease or pathology
Background
Theranostics is a combined diagnostic and treatment approach to individualized patient care. Kostmann syndrome, or severe congenital neutropenia, is an autosomal recessive disease that affects the production of neutrophils. Papillary thyroid carcinoma (PTC) is the most common type of thyroid malignancy associated with gene alterations, including in the mitogen-activated protein kinase (MAPK) signaling pathway gene. Translocation of the ETS variant 6/neurotrophic receptor tyrosine kinase 3 (ETV6/NTRK3) gene has been implicated in radiation-induced and pediatric forms of thyroid carcinoma but has rarely been described in sporadic PTC. This report is of a case of PTC in a patient with Kostmann syndrome associated with ETV6/NTRK3 gene translocation.
Case Report
A 32-year-old woman with a history of Kostmann syndrome, acute myeloid leukemia (AML), and chronic graft versus host disease (GVHD) was diagnosed with PTC with cervical lymph node metastases and soft tissue invasion following total thyroidectomy and bilateral modified radical neck dissection. Her postoperative radioactive iodine (RAI) scan confirmed lymph node metastasis. Gene expression studies identified increased expression of iodine-handling genes and ETV6/NTRK3 gene fusion. Because of the bone marrow compromise due to Kostmann syndrome and AML, a careful genomic and molecular analysis was performed to guide therapy.
Conclusions
This is the first reported case of the association between PTC, Kostmann syndrome, and ETV6/NTRK3 gene translocation in which multimodality treatment planning was optimized by genomic profiling.
... The ETV6-NTRK3 fusion gene leads to constitutive activation of the TRKC kinase domain, with downstream activation of the PI3K/AKT and MAPK pathways [18,26]. The ETV6-NTRK3 fusion gene is also found in a small subset of acute myeloid leukaemia [27,28], but the breakpoints are distinct from those found in solid malignancies. NTRK1/2/3 fusion genes have also been detected in small subsets of common tumour types, and, in this context, the fusions typically occur in a mutually exclusive fashion with other strong mitogenic drivers, i.e. genetic alterations affecting the most common driver genes belonging to the MAPK signalling pathway (e.g. ...
Background:
NTRK1, NTRK2 and NTRK3 fusions are present in a plethora of malignancies across different histologies. These fusions represent the most frequent mechanism of oncogenic activation of these receptor tyrosine kinases, and biomarkers for the use of TRK small molecule inhibitors. Given the varying frequency of NTRK1/2/3 fusions, crucial to the administration of NTRK inhibitors is the development of optimal approaches for the detection of human cancers harbouring activating NTRK1/2/3 fusion genes.
Materials and methods:
Experts from several Institutions were recruited by the European Society for Medical Oncology (ESMO) Translational Research and Precision Medicine Working Group (TR and PM WG) to review the available methods for the detection of NTRK gene fusions, their potential applications, and strategies for the implementation of a rational approach for the detection of NTRK1/2/3 fusion genes in human malignancies. A consensus on the most reasonable strategy to adopt when screening for NTRK fusions in oncologic patients was sought, and further reviewed and approved by the ESMO TR and PM WG and the ESMO leadership.
Results:
The main techniques employed for NTRK fusion gene detection include immunohistochemistry, fluorescence in situ hybridization (FISH), RT-PCR, and both RNA-based and DNA-based next generation sequencing (NGS). Each technique has advantages and limitations, and the choice of assays for screening and final diagnosis should also take into account the resources and clinical context.
Conclusion:
In tumours where NTRK fusions are highly recurrent, FISH, RT-PCR or RNA-based sequencing panels can be used as confirmatory techniques, whereas in the scenario of testing an unselected population where NTRK1/2/3 fusions are uncommon, either front-line sequencing (preferentially RNA-sequencing) or screening by immunohistochemistry followed by sequencing of positive cases should be pursued.
... SC of the salivary gland was originally defined by the t(12;15)(p13;q25) chromosomal rearrangement and resulting ETV6-NTRK3 gene product identical to that found in SC of the breast [5]. This balanced translocation and fusion gene have also been described in congenital fibrosarcoma, the cellular variant of congenital mesoblastic nephroma, and rare cases of acute myeloid leukemia [33][34][35][36][37]. Our case and all of the nine other cases of SC of thyroid reported to date that were tested have all harbored the characteristic translocation ( Table 2). ...
After being described in the salivary glands as a malignancy with features essentially identical to those of the breast, secretory carcinoma (SC) (formerly mammary analogue SC) has now been identified in other sites including the skin, lung, and thyroid gland. In the breast, SC has a relatively favorable prognosis. Likewise when arising in the salivary glands, it is generally considered to be a low to intermediate grade carcinoma; however, there is a range of clinical behavior with occasional patients dying of progressive disease. SCs of the thyroid gland are rare, and reports suggest a relatively aggressive behavior, at least relative to well differentiated carcinomas such as papillary carcinoma and minimally invasive follicular carcinoma. We present a patient with a highly aggressive thyroid gland SC that mimicked undifferentiated carcinoma clinically. The patient had widespread metastatic disease and died rapidly from airway compromise. We also review the literature for reported cases of thyroid gland SC in order to better establish the clinical features and expected clinical course of such tumors occurring at this site. © 2018, Springer Science+Business Media, LLC, part of Springer Nature.
... Über alternatives Spleißen werden zwei verschiedene Isoformen gebildet: die katalytisch aktive Volllänge Isoform und eine verkürzte, inaktive Variante. In AML und kindlicher B-ALL wurde über einzelne Fälle berichtetet, in denen eine ETV6-NTRK3-Fusion mit konstitutiv aktivierter Kinasedomäne vorliegt(Eguchi et al., 1999;Roberts et al., 2014). Dieses Fusionsprotein bewirkt über die Die Expressionsanalyse ergab, dass NTRK3 spezifisch in der T-Linien ALL stark erhöht war, durchschnittlich um das 63-fache und teilweise noch sehr viel höher, während gesunde Probanden dieses Gen kaum exprimierten. ...
Tyrosinkinasen (TK) sind Schlüsselregulatoren der zellulären Signaltransduktion und beeinflussen Zellzyklus, Zellüberleben, Apoptose, Proliferation und Differenzierung. Die Dysregulation der TK-Aktivität trägt zur Entwicklung von Leukämie und anderen malignen Erkrankungen bei. So sind 25% der akuten lymphatischen Leukämien bei Erwachsenen (ALL) durch die BCR-ABL1-Translokation bedingt. Trotz intensiver Therapie beträgt das 5-Jahres-Überleben von erwachsenen Patienten mit ALL nur etwa 50 %. Als Alternative zu herkömmlichen Chemotherapeutika bietet der Einsatz von spezifisch wirkenden TK-Inhibitoren einen individualisierten Therapieansatz mit idealerweise weniger Nebenwirkungen und einem dadurch verbesserten Outcome. Um mögliche neue therapeutische Ziele zu identifizieren, wurde eine systematische Untersuchung der Expressionsveränderungen des gesamten Tyrosinkinoms durchgeführt. Eine Vielzahl verschiedener Tyrosinkinasen zeigte starke Veränderungen im Expressionsprofil von ALL-Zellen. Ein Teil dieser Expressionsänderungen kam durch das veränderten Methylierungsprofil der ALL-Zellen zustande. EPHA7 und PTK2 sind potentielle Marker für B-Linien ALL und NTRK3, ERBB4 und ZAP70 für T-Linien ALL. Die interindividuell variierende Expression der Tyrosinkinasen EPHA3, EPHB3, KIT, ZAP70 und PDGFRB könnte eine genauere Risikoeinstufung ermöglichen. Insbesondere sind die Tyrosinkinasen ABL1, DDR1, EPHA7, FGFR1, ERBB4, FLT1, FLT3, FLT4, LCK, LTK, PTK2, PTK2B, PTK7, SRC, TEC und TYK2 vielversprechende therapeutische Ziele, die im hämatopoetischen System die Proliferation fördern und / oder die Apoptose hemmen. Eine proliferationsfördernde Wirkung von überexprimiertem FLT4 konnte erstmals gezeigt werden. Die Vielfalt der Veränderungen in der Tyrosinkinase-Expression scheint eine wichtige Rolle bei der Entwicklung von ALL zu spielen und TK könnte vielversprechende neue therapeutische Ziele sein.
... This discovery sparked the identification of a specific inhibitor of the fusion protein, which has dramatically improved the prognosis for this patient group (127). Since then, numerous types of recurrent fusion genes have been identified across a wide spectrum of different tumor types, a large proportion of which are tumor-type specific, whereas others are more promiscuously involved in various malignancies (128)(129)(130)(131). Initially, fusion genes were thought to be predominantly found in hematological malignancies, but they have since been shown to be frequent in sarcomas and carcinomas (123,128). ...
Adenoid cystic carcinoma (ACC) is a rare malignancy most often affecting the salivary glands. Despite its slow growth, it has a grave prognosis characterized by frequent local recurrences, distant metastases, and tumor-related mortality often several years after a seemingly uncomplicated clinical course. The same features characterize ACC in the lacrimal gland, whereas ACC in the breast is an indolent disease. The molecular mechanisms involved in the metastatic process of ACC are poorly-understood despite being a major cause of ACC-related mortality. Also, molecular markers with prognostic value in salivary gland ACC are highly warranted in order to identify patients at high risk of recurrent disease. Increased understanding of the molecular characteristics involved in ACC has the potential to aid in individualized follow-up programs and ultimately in novel treatments.In order to investigate the molecular background of this, we conducted a series of studies evaluating i) the clinical, genetic, and microRNA (miRNA) expressional differences between ACC of the salivary gland, lacrimal gland, and breast, ii) the genetic and miRNA expressional differences between paired primary and metastatic salivary gland ACC, and iii) the value of miRNA as independent prognostics factors in a large material of salivary gland ACC.We found that ACC from the salivary gland, lacrimal gland, and breast were very similar in all parameters except for miRNA expression. Here, breast ACC was different from ACC of the two other sites and were more similar to normal breast tissue. In contrast, paired primary and metastatic salivary gland ACC were similar in their patterns of chromosomal aberrations, point mutations were few and heterogeneous, and miRNA expression did not differ significantly. Different subgroups of salivary gland ACC separated according to miRNA expression, and several miRNAs were found to be independent prognostic markers for overall and recurrence-free survival.In conclusion, salivary gland and lacrimal gland ACC are highly similar entities with breast ACC having a normal-like pattern of miRNA expression. The development of metastatic disease in salivary gland ACC is a heterogeneous biological process in which the involvement of miRNA is not clear, whereas several, well-characterized miRNAs function as independent prognostic markers in salivary gland ACC.
... This fusion gene has been discovered in various tumors such as congenital infantile fibrosarcoma, acute myeloblastic leukemia, secretory breast cancer, mammary analog secretory carcinoma of the salivary gland, and papillary thyroid carcinoma. [12][13][14][15][16] Tumors with ETV6-NTRK3 expression demonstrated downstream hyperactivation of phosphoinositide 3-kinase and RAS signaling pathways, 17 and only show a limited response to cytotoxic anticancer agents; 18 however, acute lymphoblastic leukemia cell lines with ETV6-NTRK3 fusions were sensitive to crizotinib. 19 Although crizotinib may be a promising therapeutic agent for IMT with the ETV6-NTRK3 fusion gene, the effect of crizotinib on this tumor is unknown. ...
Inflammatory myofibroblastic tumors (IMTs) are neoplasms with low malignant potential, and the most common tumor in the lung and orbit. Their occurrence in the uterus is rare. Approximately 50% of IMT patients have anaplastic lymphoma kinase gene (ALK) rearrangements. Recent studies described novel fusions involving ROS1, platelet-derived growth factor receptor beta (PDGFR-β), and ETS translocation variant (ETV6) genes in a subset of ALK-negative patients. We report a 44-year-old woman with anemia and uterine IMT. Ultrasonography and magnetic resonance imaging revealed a myxoid degenerative myoma-like mass, 7.4 cm in maximum diameter, on the left uterine side wall. Hysterectomy was performed as a definitive treatment. Microscopic examination revealed spindle cell proliferation with numerous lymphocytes and plasma cells. Immunohistochemically, the spindle cells were negative for ALK-1, desmin, and smooth muscle actin. The pathological diagnosis was IMT arising from the uterus. Fluorescence in situ hybridization demonstrated an ETV6–neurotrophic tyrosine kinase, receptor, type 3 gene (NTRK3) translocation but no ALK, ROS1, or PDGFR-β translocations. Lung and abdomen computed tomography at 31 months postoperatively revealed no disease recurrence. This association of an ETV6–NTRK3 fusion oncogene with an ALK-negative uterine IMT increases our understanding of this neoplasm, which may help the development of specific therapies.
... Previous work indicated that fusion products containing ETV6 involve in numerous regulated errant mechanisms and can influence the fusion partner's activity through the HLH domain [12]. The ETV6 gene rearrangements were found in a translocation t (12;15)(p13;q25) via Hybridization with cosmid probes which was found in an adult acute myeloid leukaemia (AML) patient with the use of fluorescence in situ hybridization (FISH) method in 1999 [13]. In 2012, Ding L and his union performed whole-genome sequencing of primary tumor and relapse genomes from eight AML patients and certified hundreds of somatic mutations. ...
Objectives: E26 transformation-specific variant 6 gene (ETV6) is one of the most consistently rearranged genes in acute leukaemia. It encodes a principal hematopoietic transcription factor.
Methods: We performed a systematic review focusing on the mechanisms responsible for etv6 acquisition, and its effect on the development of AML. We also review the Characteristics of ETV6 mutations and its fusion genes. Finally, for using ETV6 as a molecular target, we discuss future therapeutic approaches available to mitigate the associated disease.
Results: ETV6 rearrangements often accompany other molecular mutations. Thirty-three distinct partner bands of ETV6 that contain various fusion genes were detected which plays a vital role in obtaining information about leukaemia genesis. RXDX-101 and PKC412 were reported to be inhibitors of ETV6-NTRK3.
Discussion: Future researches are needed to explain how ETV6 mutations act within the microenvironment of leukemic cells and how it affects the progression of leukaemia.
... The TRK pathway has been implicated in the pathogenesis of many cancer types, with TRK fusions being the best characterized. 7 ETV6-NTRK3 has been identified in a range of malignancies, including secretory breast carcinoma, infantile sarcoma, acute myeloid leukemia, and more recently, pediatric glioma [8][9][10][11] ; however, the oncogenic role of ETV6-NTRK3 in ALL has not been investigated. Furthermore, although we and others have shown moderate sensitivity of ETV6-NTRK3 to the ALK inhibitor, crizotinib, 12,13 the identification of potent and specific TRK inhibitors is required for clinical efficacy. ...
... It is in the form of an oncogenic fusion protein that TrkA has been identified [86][87][88]. Such rearrangements have been found for TrkC in particular in congenital fibrosarcoma and acute myeloid leukaemia [89,90]. Because of their kinase activity, these receptors have been shown to play an important role in the biology of cancers, especially in the ones of neuronal or neuroendocrine origin. ...
Transmembrane receptors are usually seen as on and off switch: when the specific ligand is bound, the receptor is on and transduces a downstream signal, while when the ligand is absent, the receptor is off. Over the last two decades several reports have argued from an alternative view where some receptors, depending on the context, will be active both in the presence and in the absence of ligand, being sort of onA and onB switch rather than on and off. These receptors have been named Dependence Receptors (DR) and they share the ability to actively trigger cell death when unbound by their respective ligands. DRs have been shown to be important guardians of tissue homeostasis. In pathological settings such as cancer, DRs are seen as tumor suppressors and a clinical trial is ongoing to assay whether these DRs can be used to provide clinical benefit by triggering cancer cell death. In this review we are reviewing this functional family of receptors and underlying their promising potential for targeted therapy against cancer. This article is protected by copyright. All rights reserved.
... A temporal lobe oligoastrocytoma (NOS) with ETV6-NTRK3 fusion was detected by NanoString assay (4%, 1/26). ETV6-NTRK3 fusion was originally identified in congenital fibrosarcoma (CFS), a pediatric spindle cell malignancy of the soft tissues [32] and subsequently in secretory breast cancer, cellular mesoblastic nephroma (CMN) which is a related pediatric tumor, and a case report of adult acute myeloid leukemia (AML) [33][34][35]. The ETV6-NTRK3 (EN) protein could function as an active protein tyrosine kinase (PTK) with potent transforming activity [36,37]. ...
Although oligodendrogliomas appear histologically similar in adult and pediatric patients, the latter have only been rarely studied and most of those studies did not have long follow-up. We examined 55 oligodendroglial tumors from pediatric and teenage patients for their biomarkers with formalin-fixed paraffin-embedded tissues and studied their survival status. None of the tumors harbored 1p/19q codeletion or IDH mutation. Mutations in TERTp (4%), BRAF (11%), FGFR1 (3%) and H3F3A (5%), fusions of BRAF (8%) and FGFR1 (8%) were found sparingly and almost all in a mutually exclusive manner. Molecular events were exclusively found in tumors with classic oligodendroglial histology. Survival analysis showed remarkably excellent prognosis compared to the adult counterparts. 5-year overall survival was 95% in our cohort with median follow-up of 8.1 years and in nine patients with follow-up more than 10 years, the 10-year overall survival was 100%. The 5-year and 10-year progression-free survivals of our cohort were 89 and 77%, respectively. FGFR1 fusion seemed to confer a poor prognosis in pediatric oligodendrogliomas. Patients receiving adjuvant chemotherapy (p = 0.046) or harboring Grade II histology (p < 0.001) had longer interval to recurrence. Our study demonstrated the distinct indolent clinical course of pediatric and teenage oligodendrogliomas compared to the adult tumors. Molecular markers commonly seen in adult oligodendrogliomas and other pediatric low-grade gliomas were only rarely seen. Since there is no clinical or molecular evidence suggesting that pediatric “oligodendrogliomas” are the same as adult oligodendrogliomas albeit histologic similarity, a case can be made for their separation from adult oligodendrogliomas in the next WHO classification.
... This holds especially true for the ETV6-RET fusion, as this fusion gene has now been reported only in ETV6-rearranged low-grade SNAC and SC, in contrast to ETV6-NTRK3, which is found in various malignancies including congenital fibrosarcoma, acute myeloid leukemia, and mesoblastic nephroma. [9][10][11] The significance of separating the morphologically different, but genetically identical ETV6-rearranged low-grade SNAC and sinonasal SC, awaits reports of clinical outcomes in larger series of both tumor types. Conflicts of Interest and Source of Funding: The authors have disclosed that they have no significant relationships with, or financial interest in, any commercial companies pertaining to this article. ...
Artemisia vulgaris L. has a long history of use in traditional medicine for the treatment of a wide range of ailments. Advancements in science and technology established scientific evidence for this medicinal plant. Recent studies have shown that A. vulgaris inhibits the growth of numerous cancer cell lines, including MCF-7, HepG2, Hela, and K-562. To access the potential anti-leukemia activity of A. vulgaris crude methanol extract (MetAV) on the ETV6-NTRK3-carrying cells, the IMS-M2, MO-91, and BaF3-CFS cell lines were co-cultured with MetAV for 48 h before being stained with Trypan Blue to calculate the percentage of viable cells. With IC50 values of 26.98 ± 2.25; 21.85 ± 0.92; and 18.70 ± 1.70 µg/ml for IMS-M2, MO-91, and BaF3-CFS, respectively, the results indicated that MetAV had a significant effect on the examined cells.
NTRK fusions are validated oncogenic drivers of various adult and pediatric tumor types, including thyroid cancer, and serve as a therapeutic target. Recently, tropomyosin receptor kinase (TRK) inhibitors, such as entrectinib and larotrectinib, display promising therapeutic efficacy in NTRK -positive solid tumors. Although some NTRK fusion partners have been identified in thyroid cancer, the spectrum of NTRK fusion is not fully characterized. In this study, a dual NTRK3 fusion was identified by targeted RNA-Seq in a 47-year-old female patient with papillary thyroid carcinoma. The patient harbors a novel in-frame fusion between NTRK3 exon 13 and AJUBA exon 2, co-existing with a known in-frame fusion between ETV6 exon 4 and NTRK3 exon 14. The dual NTRK3 fusion was validated by Sanger sequencing and fluorescence in situ hybridization (FISH) but lack TRK protein expression as defined by pan-TRK immunohistochemistry (IHC). We supposed the pan-TRK IHC result to be falsely negative. In conclusion, we present the first case of a novel NTRK3-AJUBA fusion co-existing with a known ETV6-NTRK3 fusion in thyroid cancer. These findings extend the spectrum of translocation partners in NTRK3 fusion, and the effect of dual NTRK3 fusion on TRK inhibitor therapy and prognosis needs long-term follow-up.
Introduction:
: Glioblastoma (GBM) is the most common primary brain tumor in adults. GBM treatment options have been the same for the past 30 years and have only modestly extended survival, despite aggressive multimodal treatments. The progressively better knowledge of GBM biology and a comprehensive analysis of its genomic profile have elucidated GBM heterogeneity, contributing to a more effective molecular classification and to the development of innovative targeted therapeutic approaches.
Areas covered:
: In this article, the report all the noteworthy innovations for immunotherapy and targeted therapy, providing insights into the current advances in trial designs, including combination therapies with immuno-oncology agents and target combinations.
Expert opinion:
: GBM molecular heterogeneity and brain anatomical characteristics critically restrain drug effectiveness. Nevertheless, stimulating insights for future research and drug development come from innovative treatment strategies for GBM, such as multi-specific "off-the-shelf" CAR-T therapy, oncolytic viral therapy and autologous dendritic cell vaccination. Disappointing results from targeted therapies-clinical trials are mainly due to complex interferences between signaling pathways and biological processes leading to drug resistance: hence, it is imperative in the future to develop combinatorial approaches and multimodal therapies, such as dual block of PI3K and MAPK signaling or PI3K/MTOR inhibition, to improve therapeutic benefit and survival.
Secretory carcinoma is a salivary gland neoplasm first described as a mammary analogue secretory carcinoma by Skalova et al. in 2010 and redesignated as a secretory carcinoma in the 2017 World Health Organization Classification of Head and Neck Tumors. Secretory carcinoma diagnosis is reliant on specific cytological and histological findings and the detection of an ETV6-NTRK3 fusion gene. Here, we examined the clinical and cytopathological features of four cases of secretory carcinoma occurring in three males and a female, aged between 39 and 74 years. All four tumors involved the parotid gland, and were found to have the ETV6-NTRK3 fusion gene. Fine-needle aspiration-based cytology smears of all tumors displayed papillary and/or dendritic pattern clusters, some of which were associated with blood vessels. The neoplastic cells displayed enlarged nuclei with fine chromatin and small, distinct, single nucleoli. Furthermore, several neoplastic cells with a characteristic vacuolated cytoplasm were identified in each specimen. Giemsa staining revealed cytoplasmic vacuolation, intracytoplasmic metachromatic secretions and/or various sized metachromatic granules, and a background of metachromatic mucin in all four specimens. Given this, we conclude that these cytological findings, especially those of the Giemsa staining, might be helpful in the diagnosis of secretory carcinoma.
Clear cell sarcoma of the kidney (CCSK) comprises 3% of all childhood renal cancers. Accurate diagnosis is vital for appropriate therapy, which results in a 70% to 90% overall survival rate in this previously lethal tumor. Renowned for its ability to mimic and be mimicked by every other pediatric renal tumor, and even some extrarenal retroperitoneal tumors, CCSK has a unique metastatic pattern and molecular aberrations, as well as a generally consistent clinical presentation which is of great utility in the differential. A case of CCSK in a 4-month-old boy is presented, only the fifth case reported in a patient younger than 6 months.
The NTRK gene family is composed of NTRK1, NTRK2, and NTRK3, which encode three tropomyosin-receptor kinases, belonging to a class of tyrosine kinase receptors. These proteins are known to play roles in cell proliferation, differentiation, apoptosis, and survival. Fusions involving the NTRK genes are long known as drivers in many tumors. Although they occur in less than 5% of all malignancies, their occurrence in a great diversity of tumors has been documented. Several rare tumors including infantile fibrosarcoma, secretory breast carcinoma, and mammary analogue secretory carcinoma are accompanied by NTRK fusions in more than 90% of cases, demonstrating a diagnostic value for the NTRK fusion testing in these tumors. More recently, the development of effective targeted therapies has created a demand for their detection in all malignancies. A variety of approaches are available for testing including immunohistochemistry, fluorescence in situ hybridization (FISH), reverse transcription polymerase chain reaction (RT-PCR), and DNA- and RNA-based next-generation sequencing (NGS). This article reviews the molecular biology and tumorigenesis of NTRK fusions, their prevalence and clinical significance with a focus on available methods for fusion detection. The advantages and limitations of different technologies, the best practice algorithms for NTRK fusion detection, and the future direction of NTRK testing are also discussed.
Biliary tract cancer (BTC) is a leading cause of cancer-related death, due to the limited benefits of current systematic therapies and the heterogeneity of the tumor itself. High heterogeneity means that the clinical and molecular features vary between different subtypes of BTC, while the underlying molecular mechanisms remain unclear. Targeted therapy, where inhibitors are developed to selectively combine with targeted molecules in order to block abnormal signaling pathways in BTC, has shown promise as an emerging form of treatment for various types of cancer. In this article, a comprehensive review is conducted to examine potential molecular targets for BTC targeted therapy and their mechanisms. Furthermore, preliminary data published from clinical trials is utilized to analyze the main drugs used to combat BTC. The collective information presented in this article has provided useful insights into the current understanding of BTC.
The ETV6-CBFA2 (TEL-AML1) fusion gene arises from the chromosomal translocation, t(12;21)(p13;q22) which occurs in approximately 30% of childhood B-cell acute lymphoblastic leukaemia. The role of the fusion gene in leukaemogenesis was investigated by using a DNA construct containing the murine Etv6 exon 5 fused in frame to human CBFA2 (exon 2-8) to mimic the ETV6-CBFA2 fusion identified in human acute lymphoblastic leukaemia. This DNA-targeting construct was introduced by homologous recombination into mouse embryonic stem cells and the fusion gene shown to be transcribed under the control of endogenous Etv6 elements. High level chimeras were generated by blastocyst injection of targeted embryonic stem cells. The chimeras were subsequently crossed to generate offspring heterozygous for the Etv6-CBFA2 mutation. To date, neither the chimeric animals (at 14.5 months) nor the heterozygous offspring (at 10 months) have demonstrated any overt phenotypic evidence of leukaemia. However, mice homozygous for the Etv6-CBFA2 mutation died between dE10.5-11.5. The phenotype of these embryos was similar to that described in the Etv6 knock-out model. Our study demonstrates that the Etv6-CBFA2 fusion gene resulting from the t(12;21) translocation is not, by itself, oncogenic and we postulate that further genetic events are required for the development of leukaemia. Furthermore, the lethal phenotype observed in the mice carrying the homozygous disruption of the Etv6 gene is likely to be due to a lack of the Etv6 DNA binding domain.
In this issue of Blood, Joshi et al report and characterize novel and recurrent NTRK2/3 point mutations in ∼5% of patients with different hematologic neoplasms, including acute myeloid leukemia and lymphoblastic leukemia, as well as myeloproliferative disorders.
Tropomyosin receptor kinase (Trk) C contributes to the clinicopathology of a variety of human cancers, and new chimeric oncoproteins containing the tyrosine kinase domain of TrkC occur after fusion to the partner genes. Overexpression of TrkC and TrkC fusion proteins was observed in patients with a variety of cancers, including mesenchymal, hematopoietic, and those of epithelial cell lineage. Both microRNAs (miRNAs) and long non-coding RNAs (lncRNAs) were involved in the regulation of TrkC expression through transcriptional and posttranscriptional alteration. Aberrant activation of TrkC and TrkC fusion proteins markedly induces the epithelial-mesenchymal transition (EMT) program, growth rate, tumorigenic capacity via constitutive activation of Ras-MAP kinase (MAPK), PI3K-AKT, and the JAK2-STAT3 pathway. The clinical trial of TrkC or TrkC fusion-positive cancers with newly developed Trk inhibitors demonstrated that Trk inhibitors were highly effective in inducing tumor regression in patients who do not harbor mutations in the kinase domain. Recently, there has been a progressive accumulation of mutations in TrkC or the TrkC fusion protein detected in the clinic and its related cancer cell lines caused by high-throughput DNA sequencing. Despite given the high overall response rate against Trk or Trk fusion proteins-positive solid tumors, acquired drug resistance was observed in patients with various cancers caused by mutations in the Trk kinase domain. To overcome acquired resistance caused by kinase domain mutation, next-generation Trk inhibitors have been developed, and these inhibitors are currently under investigation in clinical trials.
Philadelphia chromosome (Ph)-like acute lymphoblastic leukemia (ALL) is a subtype of Ph-negative ALL that molecularly resembles Ph-positive ALL. It shares the adverse prognosis of Ph-positive ALL, but lacks the BCR-ABL1 fusion oncogene. Instead, Ph-like ALL is associated with alternative mutations in signaling pathways. We describe a case of Ph-like ALL that harbored 2 genomic alterations, which activated signaling, an NRASGly12Asp mutation, and an ETV6-NTRK3 rearrangement. Initially, the NRAS mutation was detected at high frequency, whereas the gene fusion was only detectable with a targeted next-generation sequencing-based fusion assay, but not by fluorescence in situ hybridization analysis. The disease failed to respond to multiagent chemotherapy but investigational CD19-directed chimeric antigen receptor T-cell therapy resulted in a complete remission. However, the leukemia relapsed after 6 weeks. Intriguingly, the NRAS mutation was extinguished during the chimeric antigen receptor T-cell therapy and did not contribute to the relapse, which was instead associated with a rise in ETV6-NTRK3. The relapsed leukemia progressed with further chemo- and immunotherapy but was controlled for 6 weeks with substantial leukemic cytoreduction using the TRK inhibitor larotrectinib. Unfortunately, recovery of normal hematopoiesis was only marginal and the patient eventually succumbed to infections. These results demonstrate that larotrectinib has clinical activity in ETV6-NTRK3-associated Ph-like ALL.
The tropomyosin receptor kinase (TRK) family of receptor tyrosine kinases are encoded by NTRK genes and have a role in the development and normal functioning of the nervous system. Since the discovery of an oncogenic NTRK gene fusion in colorectal cancer in 1986, over 80 different fusion partner genes have been identified in a wide array of adult and paediatric tumours, providing actionable targets for targeted therapy. This review describes the normal function and physiology of TRK receptors and the biology behind NTRK gene fusions and how they act as oncogenic drivers in cancer. Finally, an overview of the incidence and prevalence of NTRK gene fusions in various types of cancers is discussed.
The Ets variant gene 6 (ETV6/TEL) gene is rearranged in the majority of patients with 12p13 translocations fused to a number of different partners. We present here a case of acute myeloid leukemia M4 with eosinophilia (AML-M4Eo) positive for the CBFb/MYH11 rearrangement and carrying a t(1;12)(q25;p13) that involves the ETV6 gene at 12p13. By 3′rapid amplification of cDNA ends-polymerase chain reaction (3′RACE-PCR), a novel fusion transcript was identified between the ETV6 and the Abelson-related gene (ARG) at 1q25, resulting in a chimeric protein consisting of the HLH oligomerization domain of ETV6 and the SH2, SH3, and protein tyrosine kinase (PTK) domains of ARG. The reciprocal transcript ARG-ETV6 was also detected in the patient RNA by reverse transcriptase-polymerase chain reaction (RT-PCR), although at a lower expression level. The ARG gene encodes for a nonreceptor tyrosine kinase characterized by high homology with c-Abl in the TK, SH2, and SH3 domains. This is the first report on ARGinvolvement in a human malignancy.
NTRK gene fusions affecting the tropomyosin receptor kinase (TRK) protein family have been found to be oncogenic drivers in a broad range of cancers. Small molecule inhibitors targeting TRK activity, such as the recently Food and Drug Administration-approved agent larotrectinib (Vitrakvi®), have shown promising efficacy and safety data in the treatment of patients with TRK fusion cancers. NTRK gene fusions can be detected using several different approaches, including fluorescent in situ hybridization, reverse transcription polymerase chain reaction, immunohistochemistry, next-generation sequencing, and ribonucleic acid-based multiplexed assays. Identifying patients with cancers that harbor NTRK gene fusions will optimize treatment outcomes by providing targeted precision therapy.
Gene rearrangements or fusions as a tumorigenic genomic driver event have been identified as a common recurrent occurrence in a variety of human malignancies. The neurotrophic tyrosine receptor kinase gene family contains NTRK1, NTRK2, and NTRK3, which encode the proteins tropomyosin receptor kinase A, B, and C (TRKA, TRKB, TRKC), respectively. TRKA, TRKB, and TRKC can be activated by the specific ligands, such as nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin-3 (NT3). Interestingly, although NTRK gene fusions occur relatively rarely in human cancers overall, they have been found to be present broadly in many different tumor types, including both pediatric and adult malignancies. The recognition of NTRK fusions as driver genomic event in recent years have prompted impactful clinical therapeutic development which demonstrated the efficacy and safety of TRK inhibitors, with a recent approval of larotrectinib by the US Food and Drug Administration in a cancer-agnostic manner for NTRK fusion-positive cancers. Here, we reviewed the biology of NTRK gene fusions, antitumor activity of TRK inhibitors, clinical trials development, and challenges and future perspectives of NTRK-targeted therapies in human cancer with a special focus on lung cancer.
The neurotrophic tyrosine receptor kinase ( NTRK ) gene family encodes three tropomyosin receptor kinases (TRKA, TRKB, TRKC) that contribute to central and peripheral nervous system development and function. NTRK gene fusions are oncogenic drivers of various adult and paediatric tumours. Several methods have been used to detect NTRK gene fusions including immunohistochemistry, fluorescence in situ hybridisation, reverse transcriptase polymerase chain reaction, and DNA- or RNA-based next-generation sequencing. For patients with TRK fusion cancer, TRK inhibition is an important therapeutic target. Following the FDA approval of the selective TRK inhibitor, larotrectinib, as well as the ongoing development of multi-kinase inhibitors with activity in TRK fusion cancer, testing for NTRK gene fusions should become part of the standard diagnostic process. In this review we discuss the biology of NTRK gene fusions, and we present a testing algorithm to aid detection of these gene fusions in clinical practice and guide treatment decisions.
Chromosomal rearrangements involving the NTRK1, NTRK2, and NTRK3 genes (NTRK genes), which encode the high-affinity nerve growth factor receptor (TRKA), brain-derived neurotrophic factor/neurotrophin-3 (BDNF/NT-3) growth factor receptor (TRKB), and neurotrophin-3 (NT-3) growth factor receptor (TRKC) tyrosine kinases (TRK proteins), act as oncogenic drivers in a broad range of pediatric and adult tumor types. NTRK gene fusions have been shown to be actionable genomic events that are predictive of response to TRK kinase inhibitors, making their routine detection an evolving clinical priority. In certain exceedingly rare tumor types, NTRK gene fusions may be seen in the overwhelming majority of cases, whereas in a range of common cancers, reported incidences are in the range of 0.1% to 2%. Herein, we review the structure of the three NTRK genes and the nature and incidence of NTRK gene fusions in different solid tumor types, and we summarize the clinical data showing the importance of identifying tumors harboring such genomic events. We also outline the laboratory techniques that can be used to diagnose NTRK gene fusions in clinical samples. Finally, we propose a diagnostic algorithm for solid tumors to facilitate the identification of patients with TRK fusion cancer. This algorithm accounts for the widely varying frequencies by tumor histology and the underlying prevalence of TRK expression in the absence of NTRK gene fusions and is based on a combination of fluorescence in situ hybridization, next-generation sequencing, and immunohistochemistry assays.
Pediatric glioblastoma (pGBM) is a lethal cancer with no effective therapies. To understand the mechanisms of tumor evolution in this cancer, we performed whole-genome sequencing with linked reads on longitudinally resected pGBM samples. Our analyses showed that all diagnostic and recurrent samples were collections of genetically diverse subclones. Clonal composition rapidly evolved at recurrence, with less than 8% of nonsynonymous single-nucleotide variants being shared in diagnostic-recurrent pairs. To track the origins of the mutational events observed in pGBM, we generated whole-genome datasets for two patients and their parents. These trios showed that genetic variants could be (i) somatic, (ii) inherited from a healthy parent, or (iii) de novo in the germlines of pGBM patients. Analysis of variant allele frequencies supported a model of tumor growth involving slow-cycling cancer stem cells that give rise to fast-proliferating progenitor-like cells and to nondividing cells. Interestingly, radiation and antimitotic chemotherapeutics did not increase overall tumor burden upon recurrence. These findings support an important role for slow-cycling stem cell populations in contributing to recurrences, because slow-cycling cell populations are expected to be less prone to genotoxic stress induced by these treatments and therefore would accumulate few mutations. Our results highlight the need for new targeted treatments that account for the complex functional hierarchies and genomic heterogeneity of pGBM.
Significance
This work challenges several assumptions regarding the genetic organization of pediatric GBM and highlights mutagenic programs that start during early prenatal development.
Pediatric glioblastoma (pGBM) is a lethal cancer with no effective therapies. Intratumoral genetic heterogeneity and mode of tumor evolution have not been systematically addressed for this cancer. Whole-genome sequencing of germline-tumor pairs showed that pGBM is characterized by intratumoral genetic heterogeneity and consequent subclonal architecture. We found that pGBM undergoes extreme evolutionary trajectories, with primary and recurrent tumors having different subclonal compositions. Analysis of variant allele frequencies supported a model of tumor growth involving slow-cycling cancer stem cells that give rise to fast-proliferating progenitor-like cells and to non-dividing cells. pGBM patient germlines had subclonal structural variants, some of which underwent dynamic frequency fluctuations during tumor evolution. By sequencing germlines of mother-father-patient trios, we found that inheritance of deleterious germline variants from healthy parents cooperate with de novo germline and somatic events to the tumorigenic process. Our studies therefore challenge the current notion that pGBM is a relatively homogeneous molecular entity.
Context.—:
Biphenotypic sinonasal sarcoma (BSNS) is a rare, slow-growing soft tissue sarcoma of the sinonasal tract, typically presenting with nonspecific obstructive nasal symptoms. Although recurrences are common, no metastases have been reported, and only 1 patient has died of disease thus far. It characteristically demonstrates rearrangements of PAX3 with multiple fusion partners, the most common of which is MAML3.
Objectives.—:
To highlight the most important diagnostic features, including morphologic, immunohistochemical, and molecular findings, and to provide comparisons to other entities in the differential diagnosis. We also aim to provide a summary of the clinical features and outcomes in cases reported to date.
Data sources.—:
Recently published literature encompassing BSNS and its synonym, low-grade sinonasal sarcoma with neural and myogenic differentiation.
Conclusions.—:
BSNS is a sinonasal tumor that is important to recognize because its biologic behavior differs from most of the entities in the differential diagnosis. The diagnosis can typically be rendered through a combination of morphology, immunohistochemical stains, and ancillary testing for characteristic PAX3 rearrangements.
Oncogenic somatic chromosomal rearrangements involving the NTRK1, NTRK2 or NTRK3 genes (NTRK gene fusions) occur in up to 1% of all solid tumors, and have been reported across a wide range of tumor types. The fusion proteins encoded by such rearranged sequences have constitutively activated TRK tyrosine kinase domains, providing novel therapeutic anticancer targets. The potential clinical effectiveness of TRK inhibition in patients with tumors harboring NTRK gene fusions is being assessed in phase I and II trials of TRK inhibitors, such as larotrectinib and entrectinib. Clinical trial results have demonstrated that larotrectinib is generally well tolerated and has shown high response rates that are durable across tumor types. These data validate NTRK gene fusions as actionable genomic alterations. In this review, we present the clinical data, discuss the different approaches that might be used to routinely screen tumors to indicate the presence of NTRK gene fusions, explore the issue of acquired resistance to TRK inhibition, and reflect on the wider regulatory considerations for tumor site agnostic TRK inhibitor drug development.
Tumors of the kidney comprise approximately 7% of all childhood malignancies. The vast majority are Wilms tumors, which have an overall incidence of approximately 500 cases per year. In recent years, the systematic study of pediatric renal tumors has allowed for more accurate characterization of these different entities, including their histologic appearance and molecular features that aid in risk stratification. In this chapter, we discuss the pathologic and molecular features of the most common pediatric renal tumors, including genetic alterations with diagnostic and prognostic implications.
Gene rearrangements resulting in the aberrant activity of tyrosine kinases have been identified as drivers of oncogenesis in a variety of cancers. The tropomyosin receptor kinase (TRK) family of tyrosine receptor kinases is emerging as an important target for cancer therapeutics. The TRK family contains three members, TRKA, TRKB, and TRKC, and these proteins are encoded by the genes NTRK1, NTRK2, and NTRK3, respectively. To activate TRK receptors, neurotrophins bind to the extracellular region stimulating dimerization, phosphorylation, and activation of downstream signaling pathways. Major known downstream pathways include RAS/MAPK/ERK, PLCγ, and PI3K/Akt. While being rare in most cancers, TRK fusions with other proteins have been well-established as oncogenic events in specific malignancies, including glioblastoma, papillary thyroid carcinoma, and secretory breast carcinomas. TRK protein amplification as well as alternative splicing events have also been described as contributors to cancer pathogenesis. For patients harboring alterations in TRK expression or activity, TRK inhibition emerges as an important therapeutic target. To date, multiple trials testing TRK-inhibiting compounds in various cancers are underway. In this review, we will summarize the current therapeutic trials for neoplasms involving NTKR gene alterations, as well as the promises and setbacks that are associated with targeting gene fusions.
TEL is a novel member of the ETS family of transcriptional regulators which is frequently involved in human leukemias as the result of specific chromosomal translocations. We show here by co-immunoprecipitation and GST chromatography analyses that TEL and TEL-derived fusion proteins form homotypic oligomers in vitro and in vivo. Deletion mutagenesis identifies the TEL oligomerization domain as a 65 amino acid region which is conserved in a subset of the ETS proteins including ETS-1, ETS-2, FLI-1, ERG-2 and GABP alpha in vertebrates and PNTP2, YAN and ELG in Drosophila. TEL-induced oligomerization is shown to be essential for the constitutive activation of the protein kinase activity and mitogenic properties of TEL-platelet derived growth factor receptor beta (PDGFR beta), a fusion oncoprotein characteristic of the leukemic cells of chronic myelomonocytic leukemia harboring a t(5;12) chromosomal translocation. Swapping experiments in which the TEL oligomerization domain was exchanged by the homologous domains of representative vertebrate ETS proteins including ETS-1, ERG-2 and GABP alpha show that oligomerization is a specific property of the TEL amino-terminal conserved domain. These results indicate that the amino-terminal domain conserved in a subset of the ETS proteins has evolved to generate a specialized protein-protein interaction interface which is likely to be an important determinant of their specificity as transcriptional regulators.
Ewing's sarcoma and related subtypes of primitive neuroectodermal tumours share a recurrent and specific t(11;22) (q24;q12) chromosome translocation, the breakpoints of which have recently been cloned. Phylogenetically conserved restriction fragments in the vicinity of EWSR1 and EWSR2, the genomic regions where the breakpoints of chromosome 22 and chromosome 11 are, respectively, have allowed identification of transcribed sequences from these regions and has indicated that a hybrid transcript might be generated by the translocation. Here we use these fragments to screen human complementary DNA libraries to show that the translocation alters the open reading frame of an expressed gene on chromosome 22 gene by substituting a sequence encoding a putative RNA-binding domain for that of the DNA-binding domain of the human homologue of murine Fli-1.
In myeloid and lymphoid leukemias recurrent chromosomal aberrations can be detected in chromosome region 12p13. We characterized the genes involved in t(12;22) (p13;q11) in two patients with myeloid leukemia and one with myelodysplastic syndrome (MDS). MN1, a gene on chromosome 22q11 was shown to be fused to TEL, a member of the family of ETS transcription factors on chromosome 12p13. The translocation results in transcription of the reciprocal fusion mRNAs, MN1-TEL and TEL-MN1, of which MN1-TEL is likely to encode an aberrant transcription factor containing the ETS DNA-binding domain of TEL. In addition to fusion of TEL to the PDGF beta receptor in t(5;12) in chronic myelomonocytic leukemia (CMML), our data suggest that the involvement of this protein in myeloid leukemogenesis could be dual; its isolated protein-protein dimerization and DNA-binding domains may be crucial for the oncogenic activation of functionally different fusion proteins.
Translocations involving chromosome band 11q23, found in 5-10% of human acute leukemias, disrupt the ALL-1 gene. This gene is fused by reciprocal translocation with a variety of other genes in acute lymphoblastic and myelogenous leukemias, and it undergoes self-fusion in acute myeloid leukemias with normal karyotype or trisomy 11. Here we report an alteration of the ALL-1 gene in a gastric carcinoma cell line (Mgc80-3). Characterization of this rearrangement revealed a three-way complex translocation, involving chromosomes 1 and 11, resulting in a partial duplication of the ALL-1 gene. Sequencing of reverse transcription-PCR products and Northern blot analysis showed that only the partially duplicated ALL-1 gene was transcribed, producing an mRNA with exon 8 fused to exon 2. This report of ALL-1 gene rearrangement in a solid tumor suggests that ALL-1 plays a role in the pathogenesis of some solid malignancies. The absence of the normal transcript in this cell line, in association with the loss-of-heterozygosity studies on chromosome 11q23 seen in solid tumors, suggests that ALL-1 is involved in tumorigenesis by a loss-of-function mechanism.
Chromosomal rearrangements involving band 12p13 are found in a wide variety of human leukemias but are particularly common in childhood acute lymphoblastic leukemia. The genes involved in these rearrangements, however, have not been identified. We now report the cloning of a t(12;21) translocation breakpoint involving 12p13 and 21q22 in two cases of childhood pre-B acute lymphoblastic leukemia, in which t(12;21) rearrangements were not initially apparent. The consequence of the translocation is fusion of the helix-loop-helix domain of TEL, an ETS-like putative transcription factor, to the DNA-binding and transactivation domains of the transcription factor AML1. These data show that TEL, previously shown to be fused to the platelet-derived growth factor receptor beta in chronic myelomonocytic leukemia, can be implicated in the pathogenesis of leukemia through its fusion to either a receptor tyrosine kinase or a transcription factor. The TEL-AML1 fusion also indicates that translocations affecting the AML1 gene can be associated with lymphoid, as well as myeloid, malignancy.
Analysis of a growing number of chromosomal translocations in human tumors have shown that they frequently result in gene fusions encoding chimeric proteins. We have characterized the recurrent t(12;21)(p12;q22) translocation present in human B-lineage acute leukemias. This translocation fused two genes, tel and AML1, that have previously been described in chromosomal translocations specific for myeloid malignancies. These two genes therefore belong to an increasing number of human genes that are involved in a variety of hematopoietic malignant disorders and can be rearranged with numerous partners. Interestingly, in these acute leukemias, deletion of the other tel allele from the normal chromosome 12 was associated with the tel rearrangement, whereas both tel alleles were present in the chronic leukemias bearing a t(5;12) that we have tested.
Balanced translocations involving band q12 of human chromosome 22 are the most frequent recurrent translocations observed in human solid tumours. It has been shown recently that this region encodes EWS, a protein with an RNA binding homologous domain. In Ewing's sarcoma and malignant melanoma of soft parts, translocations of band 22q12 to chromosome 11 and 12 result in the fusion of EWS with the transcription factors FLI-1 and ATF1, respectively. The present analysis of 89 Ewing's sarcomas and related tumours show that in addition to the expected EWS-FLI-1 fusion, the EWS gene can be fused to ERG, a transcription factor closely related to FLI-1 but located on chromosome 21. The position of the chromosome translocation breakpoints are shown to be restricted to introns 7-10 of the EWS gene and widely dispersed within introns 3-9 of the Ets-related genes. This heterogeneity generates a variety of chimeric proteins that can be detected by immuno-precipitation. On rare occasions, they may be associated with a truncated EWS protein arising from alternate splicing. All 13 different fusion proteins that were evidenced contained the N-terminal domain of EWS and the Ets domain of FLI-1 or ERG suggesting that oncogenic conversion is achieved by the linking of the two domains with no marked constraint on the connecting peptide.
The trkC gene product gp145trkC is a high affinity signaling receptor for neurotrophin-3 (NT-3), a member of the NGF family of neurotrophic factors. We now report that trkC encodes at least two additional tyrosine protein kinase receptors. These receptors, designated TrkC K2 and TrkC K3, have the same amino acid sequences as gp145trkC (now designated TrkC K1) except for the presence of 14 and 25 additional amino acid residues between kinase subdomains VII and VIII, just downstream from the TDYYR motif which encompasses the putative autophosphorylation site of the Trk receptor family. Upon interaction with their cognate ligand, NT-3, all three TrkC receptor isoforms become rapidly phosphorylated on tyrosine residues and induce DNA synthesis in quiescent cells. However, only TrkC K1 has mitogenic activity in NIH3T3 cells and induces neuronal differentiation of PC12 cells. The different biological properties of these TrkC receptor isoforms probably result from their engagement with different signaling pathways. Whereas TrkC K1 phosphorylates phospholipase C gamma 1 and phosphatidylinositol-3 kinase, TrkC K2 and TrkC K3 do not. TrkC K2 and transcripts encoding TrkC K3 have been identified in various structures of the adult murine brain. These observations suggest that the trophic activities of NT-3 in the mammalian nervous system might be mediated by different TrkC receptor isoforms.
When used at concentrations allowing interactions only with its high-affinity receptors, neurotrophin-3 (NT-3) promotes the survival of sensory neurons isolated from embryonic day 8 (E8) chicks, but not the survival of E11 sympathetic neurons. These sympathetic neurons (which can be rescued by the addition of NGF) display high-affinity receptors for NT-3 (Kd of 1.6 x 10(-11) M) that cannot be distinguished from the high-affinity NT-3 receptors on sensory neurons using equilibrium binding or kinetic criteria. This represents the first example of embryonic neurons that cannot be rescued by the in vitro addition of a neurotrophin in spite of the presence of corresponding neurotrophin high-affinity receptors. At elevated concentrations, beyond the saturation of its high-affinity receptors, NT-3 supports the survival of some E11 sympathetic neurons, an effect that might be mediated by the high-affinity NGF receptor. Using E7 sympathetic neurons, about 40% of the cells initially plated can be rescued in vitro by the addition of low concentrations of NT-3 (but not of NGF) and produce profuse neurites. This indicates that NT-3 may play a role in the early development of some sympathetic neurons.
The genetic locus for the TrkC/neurotrophin 3 (NT-3) receptor tyrosine kinase encodes multiple isoforms including receptors
with inserts in the catalytic domain. This study examines the signaling capabilities of TrkC and related kinase insert isoforms
TrkC14 and TrkC25. We show that in PC12 cells expressing both TrkC and TrkA/nerve growth factor (NGF) receptors, different
morphological changes occur upon addition of NGF or NT-3. NT-3-treated cells exhibit longer neurites and larger cell bodies
as compared to NGF-treated cells. Both TrkC and TrkA mediate qualitatively similar increases in the tyrosine phosphorylation
of phospholipase C (PLC)-1, Shc, SNT, and MAPK and the transcription of the c-fos, c-jun, NGFI-A, and NGFI-B immediate early genes. However, the TrkC kinase insert forms fail to stimulate these events. Furthermore, TrkC14 and TrkC25
have only a low intrinsic tyrosine kinase activity, and insertion of the TrkC14 kinase insert into TrkA at an equivalent position
results in a dramatic reduction of the kinase activity and signaling capabilities of TrkA. The TrkC14 and −25 isoforms may
fail to transmit signals due to their low intrinsic kinase activity and failure to activate and/or tyrosine phosphorylate
targets shown to be involved in neurotrophin signal transduction pathways.
TEL is a member of the Ets family of transcription factors which are frequently rearranged in human leukemia. The mechanism
of TEL-mediated transformation, however, is unknown. We report the cloning and characterization of a chromosomal translocation
associated with acute myeloid leukemia which fuses TEL to the ABL tyrosine kinase. The TEL-ABL fusion confers growth factor-independent
growth to the marine hematopoietic cell line Ba/F3 and transforms Rat-1 fibroblasts and primary murine bone marrow cells.
TEL-ABL is constitutively tyrosine phosphorylated and localizes to the cytoskeleton. A TEL-ABL mutant containing an ABL kinase-inactivating
mutation is not constitutively phosphorylated and is nontransforming but retains cytoskeletal localization. However, constitutive
phosphorylation, cytoskeletal localization, and transformation are all dependent upon a highly conserved region of TEL termed
the helix-loop-helix (HLH) domain. TEL-ABL formed HLH-dependent homo-oligomers in vitro, a process critical for tyrosine kinase
activation. These experiments suggest that oligomerization of TEL-ABL mediated by the TEL HLH domain is required for tyrosine
kinase activation, cytoskeletal localization, and transformation. These data also suggest that oligomerization of Ets proteins
through the highly conserved HLH domain may represent a previously unrecognized phenomenon.
The TEL/PDGF beta R fusion protein is the product of the t(5;12) translocation in patients with chronic myelomonocytic leukemia. The TEL/PDGF beta R is an unusual fusion of a putative transcription factor, TEL, to a receptor tyrosine kinase. The translocation fuses the amino terminus of TEL, containing the helix-loop-helix (HLH) domain, to the transmembrane and cytoplasmic domain of the PDGF beta R. We hypothesized that TEL/PDGF beta R self-association, mediated by the HLH domain of TEL, would lead to constitutive activation of the PDGF beta R tyrosine kinase domain and cellular transformation. Analysis of in vitro-translated TEL/ PDGF beta R confirmed that the protein self-associated and that self-association was abrogated by deletion of 51 aa within the TEL HLH domain. In vivo, TEL/PDGF beta R was detected as a 100-kDa protein that was constitutively phosphorylated on tyrosine and transformed the murine hematopoietic cell line Ba/F3 to interleukin 3 growth factor independence. Transformation of Ba/F3 cells required the HLH domain of TEL and the kinase activity of the PDGF beta R portion of the fusion protein. Immunoblotting demonstrated that TEL/PDGF beta R associated with multiple signaling molecules known to associate with the activated PDGF beta R, including phospholipase C gamma 1, SHP2, and phosphoinositol-3-kinase. TEL/PDGF beta R is a novel transforming protein that self-associates and activates PDGF beta R-dependent signaling pathways. Oligomerization of TEL/PDGF beta R that is dependent on the TEL HLH domain provides further evidence that the HLH domain, highly conserved among ETS family members, is a self-association motif.
The human TEL gene is involved in several 12p13 chromosomal abnormalities present in various human hematological malignancies, the most frequent being the t(12;21)(p13;q22), specific for childhood acute lymphoblastic leukemia. The predicted product of TEL harbours an amino acid region similar to the ETS DNA binding domain. We now report the isolation of the murine TEL cDNA and the characterization of the human TEL proteins. Human and murine TEL proteins are particularly homologous within their aminoterminal regions and their ETS domains. TEL proteins are nuclear and display specific DNA binding activity toward classical ETS binding sites. In addition, we show that TEL mRNAs initiate translation at either of the two first inframe ATGs (codon 1 and 43) to encode 50 kDa and 57 kDa TEL proteins. In vivo, each of these primary translational products is modified by multiple phosphorylation events.
We identified a fusion between ETV6 on 12p13 and MDS1/EVI1 on 3q26 in a t(3;12)(q26;p13) found in two cases of myeloproliferative disorder. The resulting chimeric transcript consists of the first two exons of ETV6 fused to MDS1 sequences, which in turn is fused to the second exon of the EVI1 gene. It has recently been reported that MDS1 can be expressed in normal tissues both as a single gene and fused to EVI1. ETV6 does not contribute any known functional domain to the predicted fusion protein. Association with blast crisis and myelodysplastic syndrome-derived leukemia, bad prognosis, and relative complex karyotype are in agreement with observations made in other cases of t(3;12)(q26;p13). Furthermore, a comparison can be made with the formation of an AML1/MDS1/EVI1 fusion gene in translocations (3;21)(q26;q22).
Translocations in hematologic disease of myeloid or lymphoid origin with breakpoints at chromosome band 12p13 frequently result in rearrangements of the Ets variant gene 6 (ETV6). As a consequence either the ETS DNA-binding domain or the Helix-Loop-Helix (HLH) oligomerization domain of ETV6 is fused to different partner genes. We show here that a t(9;12)(p24;p13) in a case of early pre-B acute lymphoid leukemia and a t(9;15;12)(p24;q15;p13) in atypical chronic myelogenous leukemia in transformation involve the ETV6 gene at 12p13 and the JAK2 gene at 9p24. In each case different fusion mRNAs were found, with only one resulting in an open reading frame for a chimeric protein consisting of the HLH oligomerization domain of ETV6 and the protein tyrosine kinase (PTK) domain of JAK2. The cloning of the complete human JAK2 coding and genomic sequences and of the genomic junction fragments of the translocations allowed a characterization of the different splice events leading to the various mRNAs. JAK2 plays a central role in non-protein tyrosine kinase receptor signaling pathways, which could explain its involvement in malignancies of different hematologic lineages. Besides hop in Drosophila no member of the JAK family has yet been implicated in tumorigenesis.
Using fluorescence in situ hybridization (FISH) and probes located on 12p12.1 to 13.3, we studied the breakpoints of 23 patients who had various hematologic malignant diseases and who had 12p13-balanced translocations (21 patients), inversion (1 patient), or insertion (1 patient). Among them, 14 patients had breakpoints within YAC964c10, which contains the TEL (ETV6 ) gene and in 12 of these with balanced translocations or insertion, the FISH results suggested that TEL was involved. Two of the 14 patients, patients no. 13 and 14, had breakpoints in YAC 964C10 that were centromeric to TEL but telomeric to KIP1. In the other 9 patients whose breakpoints did not fall within the YAC, the breakpoints were found telomeric to the YAC in at least three different locations on distal 12p. These results indicated that TEL was involved in only half (12 of 23) of the patients with balanced 12p13 rearrangements and that there probably were several other breakpoint cluster regions on 12p13, suggesting that genes other than TEL were involved in these rearrangements.
The NTRK1 gene in the q arm of chromosome I encodes one of the receptors for the nerve growth factor and is frequently activated as an oncogene in papillary thyroid carcinomas. The activation is due to chromosomal rearrangements juxtaposing the NTRK1 tyrosine kinase domain to 5′‐end sequences from different genes. The thyroid TRK oncogenes are activated by recombination with at least three different genes: the gene coding for tropomyosin and TPR, both on chromosome I, and TFG on chromosome 3. In a previous study, we showed that two tumors carrying the TPR/NTRK1 rearrangement contained structurally different oncogenes named TRK‐T1 and TRK‐T2. In this paper, we report (1) the cDNA structure of TRK‐T2. (2) evidence that TRK‐T2 is generated by different rearrangements in two thyroid tumors, and (3) a detailed analysis of the three different TPR/NTRK1 rearrangements. With molecular studies based on Southern blot hybridization, cloning, and sequencing, we show that all the rearrangements are nearly balanced, involving deletion, insertion, or duplication of only few nucleotides. In one case, an additional rearrangement involving sequences derived from chromosome 17 was detected. Genes Chromosom. Cancer 19:112–123, 1997. © 1997 Wiley‐Liss, Inc.
We report the isolation and molecular characterization of trkC, a new member of the trk family of tyrosine protein kinase genes. trkC is preferentially expressed in the brain. In situ hybridization studies revealed trkC transcripts in the hippocampus, cerebral cortex, and the granular cell layer of the cerebellum. The product of the trkC gene has been identified as a glycoprotein of 145,000 daltons, gp145trkC, which is equally related to the previously characterized gp140trk and gp145trkB tyrosine kinases. gp145trkC is a functional receptor for neurotrophin-3 (NT-3). However, gp145trkC does not bind the highly related neurotrophic factors NGF or BDNF. In proliferating cells, the interaction between gp145trkC and NT-3 elicits a more efficient biological response than when NT-3 binds to its other receptors gp140trk and gp145trkB. These results indicate that gp145trkC may play an important role in mediating the neurotrophic effects of NT-3.
Oncogenic activation of the proto-oncogene c-abl in human leukemias occurs as a result of the addition of exons from the gene
bcr and truncation of the first abl exon. Analysis of tyrosine kinase activity and quantitative measurement of transformation
potency in a single-step assay indicate that variation in bcr exon contribution results in a functional difference between
p210bcr-abl and p185bcr-abl proteins. Thus, foreign upstream sequences are important in the deregulation of the kinase activity
of the abl product, and the extent of deregulation correlates with the pathological effects of the bcr-abl proteins.
A biologically active complementary DNA clone of a transforming gene present in a human colon carcinoma contains gene sequences of both tropomyosin and a previously unknown protein tyrosine kinase. The predicted protein (641 amino acids) encoded by this oncogene seems to have been formed by a somatic rearrangement that replaced the extracellular domain of a putative transmembrane receptor by the first 221 amino acids of a non-muscle tropomyosin molecule.
Most Ewing's sarcomas or related primitive neuroectodermal tumors have the (11;22)(q24;q12) or less frequently the (21;22)(q22;q12) translocation. These rearrangements fuse the EWS gene on chromosome 22q12 to either the FLI1 or ERG genes, both members of the ETS family of transcription factors. Simple variant chromosomal translocations have been occasionally described in these tumors. We have identified a third Ewing's sarcoma translocation, the t(7;22)(p22;q12), that fuses EWS to the human homologue of the murine ETS gene ER81. This gene, designated ETV1 (for ETS Translocation Variant), is located on chromosome band 7p22. Identical EWS nucleotide sequences found in the majority of EWS-FLI1 and EWS-ERG chimeric transcripts are fused to a portion of ETV1 encoding an ETS domain with sequence specific DNA-binding activity. These findings confirm that the fusion of EWS to different ETS family members can result in a similar tumor phenotype.
In human leukemia, activation of the ABL proto-oncogene locus on chromosome 9 most commonly occurs as a result of its fusion to the BCR locus on chromosome 22. The resulting chimeric protein displays an elevated tyrosine kinase activity. We have identified a novel activation of ABL which involves a gene located on chromosome 12, designated TEL. Like BCR, TEL is fused in-frame with ABL and produces a fusion protein with an elevated tyrosine kinase activity when assayed in an immune complex. The amino-terminal sequences of TEL encode a helix-loop-helix motif which may mediate dimerization.
TrkC is a receptor tyrosine kinase that is activated by neurotrophin-3, a factor important in the development of certain areas of the central nervous system. We have cloned and sequenced the human trkC cDNA and found that the predicted amino acid sequence is 97 to 98% homologous to the rat and porcine trkC sequences, respectively. The rat trkC has several isoforms due to alternative splicing in the tyrosine kinase domain. We cloned one human splice variant that has a nucleic acid sequence identical to the rat isoform with an insert of 14 amino acids. The human trkC cDNA also has a (CGG)n repeat in the 5'-untranslated region. This sequence was not highly polymorphic in that 79 of 80 chromosomes examined had eight repeats, while 1 chromosome had four repeats. By PCR analysis of a somatic cell hybrid panel and fluorescence in situ hybridization with the cDNA clone, human trkC was mapped to chromosome 15q24-q25.
The expression of the low-affinity NGF receptor (p75) and the trkA proto-oncogene product was analyzed in a series of human hematopoietic cell lines at protein and RNA levels. We did not detect any form of NGF receptor in cell lines displaying a myelomonocytic phenotype (HL60 and U937). In contrast, cells displaying a more immature erythroleukemic phenotype (TF1 and K562) expressed TrkA in the absence of detectable p75. Scatchard analysis showed a single high-affinity site for NGF (kd = 10(-10) mol/L), with a copy number ranging from 300 to 3,000 sites per cell depending on the studied cell line. In addition, NGF induced autophosphorylation of TrkA and could substitute for granulocyte-monocyte colony-stimulating factor to trigger the proliferation of the TF1 cell line, with a half-maximal signal observed at 50 pmol/L, indicating that p75 is not required for DNA synthesis in this cell line. The physiologic relevance of NGF in early hematopoiesis was confirmed by showing that 12% to 15% of progenitor blood cells from mice treated with 5-fluorouracil expressed TrkA and that these cells could be induced to proliferate and differentiate in response to NGF in association with macrophage colony-stimulating factor. Our study demonstrates for the first time that trkA proto-oncogene expression and activation is not restricted to the nervous system, but is also an important element in early hematopoiesis.
Chronic myelomonocytic leukemia (CMML) is a myelodysplastic syndrome characterized by abnormal clonal myeloid proliferation and by progression to acute myelogenous leukemia (AML). CMML thus offers an opportunity to study early genetic events in the transition to AML. A recently recognized subgroup of CMML has a t(5;12)(q33;p13) balanced translocation. We report that the consequence of the t(5;12) translocation is expression of a fusion transcript in which the tyrosine kinase domain of the platelet-derived growth factor receptor beta (PDGFR beta) on chromosome 5 is coupled to a novel ets-like gene, tel, on chromosome 12. The tel-PDGFR beta fusion demonstrates the oncogenic potential of PDGFR beta and may provide a paradigm for early events in the pathogenesis of AML.
The t(16;21)(p11;q22) translocation is a recurrent chromosomal abnormality found in several types of myeloid leukemia. We have previously demonstrated that the breakpoints of this translocation are clustered in a specific intron of the ERG gene on chromosome 21, which has recently been reported to be involved in Ewing's sarcoma. We show here that the TLS/FUS gene on chromosome 16 is fused with the ERG gene to produce the TLS/FUS-ERG chimeric transcript by this translocation. The TLS/FUS gene has been identified as a translocated gene in myxoid liposarcoma by the t(12;16)(q13;p11) translocation and encodes an RNA-binding protein that is highly homologous to the product of the EWS gene involved in Ewing's sarcoma. Thus, the TLS/FUS-ERG gene fusion in t(16;21) leukemia is predicted to produce a protein that is very similar to the EWS-ERG chimeric protein responsible for Ewing's sarcoma.
We have developed a method that allows reliable construction of high quality FISH probes from yeast artificial chromosomes (YACs) based on the separation of YACs by pulse-field gel electrophoresis and a rapid sequence-independent amplification procedure (SIA). These probes can be used to localize YACs on metaphase chromosomes and also with high efficiency, in interphase nuclei.
Human neuroblastomas have been found to express the neurotrophin receptors TrkA and TrkB. Expression of TrkA correlates with favorable outcome, while expression of full-length TrkB is associated with unfavorable, more aggressive, N-myc amplified tumors. In this study we have determined the expression of TrkC in neuroblastoma primary tumors and cell lines. Using probes for the extracellular domain and the tyrosine kinase domain of human TrkC, we found by Northern analysis that TrkC mRNA is expressed in 14 of 55 (25%) tumors from a representative panel of neuroblastomas. A 14 kb transcript was detected by both probes, indicating that it would encode the full-length TrkC protein. A significant association was found between TrkC mRNA expression detected by Northern analysis and lower stage tumors [stage 1, 2, 4S, 11 of 30 (37%); vs stage 3, 4, 3 of 25 (12%), chi2 = 4.4, P < 0.04]. Only one of eight primary tumors with N-myc amplification had detectable TrkC mRNA expression and none of the eight neuroblastoma cell lines expressed TrkC by Northern analysis. Our results suggest that TrkC is involved in the biology of favorable neuroblastomas.
We have constructed a detailed map of the genomic region containing the ETS-variant gene 6 (ETV6), involved in translocations and deletions associated with hematologic malignancies. Thirty-eight cosmids were characterized belonging to two contigs spanning 340 kb, and an EcoRl restriction map was developed. The gap between the two contigs, 2 kb in size, was closed by PCR. The contigs contain the complete coding sequence and the 5' and 3' UTRs of ETV6. Eight exons accounting for the ETV6 cDNA sequence were identified. The helix-loop-helix (HLH) motif is coded by exons 3 and 4, whereas exons 6-8 code for the ETS DNA-binding domain. All introns show consensus 5' donor and 3' acceptor splice sites. Introns 1 and 2 span 100 and 82 kb, respectively, and introns 3-7 range from 15 to 1.3 kb. An alternative exon 1 (exon 1B) is localized in intron 2. The 5' end of the ETV6 gene is associated with a CpG island characterized by the presence of four Notl, four Sacll, and three BssHll recognition sites and several SP1- and AP2-binding motifs. Alternative polyadenylation at the 3' end of the ETV6 gene generates the three transcripts of 6200, 4300, and 2400 nucleotides, respectively. The ETV6 gene spans 240 kb and is flanked at its 5' and 3' end by D12S1697 and D12S98, respectively. The markers D12S1095 and D12S89 are located in the first intron. Two new DNA polymorphisms were identified in the ETV6 gene, which will be useful for the analysis of loss of heterozygosity reported for the ETV6 gene in leukemia.
The NTRK1 gene in the q arm of chromosome 1 encodes one of the receptors for the nerve growth factor and is frequently activated as an oncogene in papillary thyroid carcinomas. The activation is due to chromosomal rearrangements juxtaposing the NTRK1 tyrosine kinase domain to 5'-end sequences from different genes. The thyroid TRK oncogenes are activated by recombination with at least three different genes: the gene coding for tropomyosin and TPR, both on chromosome 1,and TFG on chromosome 3. In a previous study, we showed that two tumors carrying the TPR/NTRK1 rearrangement contained structurally different oncogenes named TRK-T1 and TRK-T2. In this paper, we report (1) the cDNA structure of TRK-T2, (2) evidence that TRK-T2 is generated by different rearrangements in two thyroid tumors, and (3) a detailed analysis of the three different TPR/NTRK1 rearrangements. With molecular studies based on Southern blot hybridization, cloning, and sequencing, we show that all the rearrangements are nearly balanced, involving deletion, insertion, or duplication of only few nucleotides. In one case, an additional rearrangement involving sequences derived from chromosome 17 was detected.
Thirty-eight sex-mismatched bone marrow transplantation patients with various hematological diseases were followed-up using fluorescence in situ hybridization. Probes specific for various translocations, the X chromosome (DXZ1) and the whole Y chromosome (WCP Y), were used to assess successful engraftment and residual host cells. The combination of translocation and WCP Y probes enabled the identification of host and donor cells in addition to the identification of malignant vs. normal cells in the transplant recipient. Fifteen patients were sequentially followed up. The results obtained using the combination of translocation plus WCP Y probes were more reliable than those with DXZ1 plus WCP Y probes, or the translocation probe alone, especially when the percentage of residual leukemic cells detected by the translocation probe alone was around the cut-off level.
The Janus family of tyrosine kinases (JAK) plays an essential role in development and in coupling cytokine receptors to downstream
intracellular signaling events. A t(9;12)(p24;p13) chromosomal translocation in a T cell childhood acute lymphoblastic leukemia
patient was characterized and shown to fuse the 3′ portion ofJAK2 to the 5′ region of TEL, a gene encoding a member of the ETS transcription factor family. The TEL-JAK2 fusion protein includes the catalytic domain
of JAK2 and the TEL-specific oligomerization domain. TEL-induced oligomerization of TEL-JAK2 resulted in the constitutive
activation of its tyrosine kinase activity and conferred cytokine-independent proliferation to the interleukin-3–dependent
Ba/F3 hematopoietic cell line.
Congenital (or infantile) fibrosarcoma (CFS) is a malignant tumour of fibroblasts that occurs in patients aged two years or younger. CFS is unique among human sarcomas in that it has an excellent prognosis and very low metastatic rate. CFS is histologically identical to adult-type fibrosarcoma (ATFS); however, ATFS is an aggressive malignancy of adults and older children that has a poor prognosis. We report a novel recurrent t(12;15)(p13;q25) rearrangement in CFS that may underlie the distinctive biological properties of this tumour. By cloning the chromosome breakpoints, we show that the rearrangement fuses the ETV6 (also known as TEL) gene from 12p13 with the 15q25 NTRK3 neurotrophin-3 receptor gene (also known as TRKC). Analysis of mRNA revealed the expression of ETV6-NTRK3 chimaeric transcripts in all three CFS tumours analysed. These were not detected in ATFS or infantile fibromatosis (IFB), a histologically similar but benign fibroblastic proliferation occurring in the same age-group as CFS. ETV6-NTRK3 fusion transcripts encode the helix-loop-helix (HLH) protein dimerization domain of ETV6 fused to the protein tyrosine kinase (PTK) domain of NTRK3. Our studies indicate that a chimaeric PTK is expressed in CFS and this may contribute to oncogenesis by dysregulation of NTRK3 signal transduction pathways. Moreover, ETV6-NTRK3 gene fusions provide a potential diagnostic marker for CFS.
A t(6;12)(q23;p13) results in the fusion of ETV6 to a novel gene, STL, in a B-cell ALL cell line.
- Suto
Expression of the neurotrophin receptor TrkC is linked to a favorable outcome in medulloblastoma.
- Segal