Leukemia is a malignant clonal disease originating from hematopoietic stem and progenitor cells. Leukemia cells with proliferation and survival advantages proliferate and accumulate uncontrollably in the body, gradually replacing normal hematopoiesis and invading other organs and systems, resulting in a series of symptoms such as anemia, hemorrhage, infection and immersion. According to the degree of differentiation and maturation of leukemia cells and the natural course of disease, leukemia can be roughly divided into two categories: acute leukemia and chronic leukemia, and then divided into myelogenic/myeloid and lymphocytic/lymphoblastic according to the cell of origin.

SET-CAN/NUP214 fusion gene is formed by del(9)(q34.11;q34.13) or t(9;9)(q34;q34) and has been identified in the LOUCY cell line of T -ALL and the MEGAL cell line of AML(1, 2). In 1992, Von Lindern et al. first identified the SET-CAN/NUP214 fusion gene in a case of acute undifferentiated leukemia (AUL). Since then, with the development of detection technology and the deepening understanding of leukemia, subsequent cases of AML, MS, AUL, CML, and B-ALL have also been found (3–6). Overall, the disease experienced by most patients carrying SET-CAN/NUP214 is T-ALL.

The NUP214 protein, also known as CAN, is a nucleoporin with FG repeats rich in phenylalanine-glycine. The NUP214 gene is located on band 9q34.1 and it has a total of 36 exons numerically labeled from 1 to 36 ( Figure 1 ). Chromosome abnormality involving NUP214 occur repeatedly in leukemia, in addition to the SET-CAN/NUP214 reviewed here, other chromosome abnormalities were found such as DEK-NUP214, SQSTM1-NUP214 and NUP214-ABL1. DEK-NUP214 [t(6;9)(p22;q34)] was associated with AML, NUP214-ABL1 was identified in T-ALL patients, the rarest leukemia NUP214 fusion protein is SQSTM1-NUP214: to date, only two cases have been reported, one in ALL and the other in AML. The structure of the SQSTM1-NUP214 fusion gene consists of five exons located at the N-terminus of the SQSTM1 gene fused to a portion of the C-terminus of NUP214, including its last 14 FG repeats (7). In eukaryotic cells, nucleo-cytoplasmic transport plays an important role in maintaining the normal function and integrity of cells (8). Molecules with a molecular mass greater than 40kDa cannot move across the nuclear membrane by simple diffusion, but require to be facilitated by nuclear transporter receptors (NTRs) with the help of nuclear pore complexes (NPCs) embedded within the nuclear membrane (9–11). NUP214 interacts with NTRs via the FG repeat region in the cytoplasmic filaments of the nuclear pore complexes (NPCs) to control macromolecule trafficking (12). NUP214 has been shown to interact with exportin-1 (XPO1) and nuclear RNA export factor 1 (NXF1) of NTRs, which are highly mobile in cells (13) and play an important role in the response to NUP214 by nuclear export sequences (NES) protein; Furthermore, NUP214 fusion proteins such as SET-CAN/NUP214 and DEK-NUP214, reduce the mobility of XPO1 and lead to the accumulation of XPO1 cargo within the nucleus, impair nuclear output by sequestering XPO1 in the nucleus, interfere with nuclear-cytoplasmic transport of macromolecules, and potentially affect the transcriptional regulatory function of the NF-κB pathway (14), leading to various blood diseases (15). Moreover, genomic knockout of NUP214 led to embryonic lethality in mice (1).

SET, also referred to as TATA box binding protein-associated factor 1 (TAF1). SET is a component of the histone acetyltransferase inhibitor (Inhat), which has been reported to be a putative oncogene involved in transcription by regulating chromatin organization (16). SET encodes a protein which can exert an inhibitory effect on apoptosis induced by cytotoxic T lymphocytes (4). In eukaryotic cells, the occurrence of selective splicing in the first two exons of the TAF1 gene results in the formation of two forms of SET expression: the two heterodimeric forms, TAF1-α and TAF1-β ( 1). Whereas in SET-CAN/NUP214, only the TAF1-β isoform is present (17). The structure of SET/TAF1-β consists of three parts: an N-terminal dimerization domain, a central “Earmuff” domain named for its headphone-like structure, and an acidic and negatively charged C-terminal domain ( Figure 1 ). SET/TAF-Iβ has a variety of different activities, such as inhibiting phosphatase 2A activity, inducing cell transformation and differentiation, and transferring histones to naked DNA. The structural and negative regulatory functions may be related to glucocorticoid resistance (16, 18, 19).

SET-CAN/NUP214 fusion gene encodes a protein containing an almost complete portion of SET fused to the carboxy-terminal two-thirds of CAN, which is a rare gene rearrangement occurs primarily in hematological malignancies (3). The appearance of the fusion gene may be the result of prior cancer therapy, but it may also occur de novo.

SET-CAN/NUP214 positive patients often show resistance to chemotherapy including glucocorticoids, but the mechanism is not completely clear. The optional treatment has not been determined, previous studies have adopted different treatment options with varying prognoses for patients. Some previous studies have shown that SET-CAN/NUP214 fusion gene positive patients have a worse prognosis (3, 7, 20), while clinical studies have shown that there is no significant difference in 3-year event-free survival (EFS) and overall survival (OS) between patients with SET-CAN/NUP214 fusion gene positive and SET-CAN/NUP214 negative patients (21, 22). Conventional techniques such as chromosomal karyotype analysis may have limitations in detecting patients with SET-CAN/NUP214. Due to the emergence of more advanced detection techniques such as fluorescence in situ hybridization (FISH), previously challenging fusion genes like SET-CAN/NUP214 can now be detected with increasing frequency. This necessitates more precise disease classification and optimization of therapeutic regimens. Research shows that HSCT can improve the prognosis, the level of SET-CAN/NUP214 after transplantation can predict recurrence to a certain extent (23), new methods such as CAR-T may be effective for patients and further research is needed (24).

In this review, we summarized the general features and clinical advances of SET-CAN/NUP214 fusion gene in leukemia.

In the 2022 international consensus classification of acute lymphoblastic leukemia/lymphoma, SET-CAN/NUP214 fusion gene positive has been listed as a subtype of the HOXA gene family in the latest eight temporary entities (25). SET-CAN/NUP214 fusion gene is rare in leukemia patients, and there is no prospective clinical study for such patients. Relevant articles focus on case reports and mechanism studies. This section provides an overview of the general features of patients.

According to a statistic in 2016, a total of 42 SET-CAN/NUP214 positive patients were reported up to that year, including T-ALL(38/42,90.5%), AUL(2/42,4.8%), AML(1/42;2.4%) and B-ALL(1/42;2.4%) (4), another study involving 59 T-ALL patients showed that about 10.3% of T-ALL patients carried SET-CAN/NUP214 fusion gene (20), Ben Abdelli et al. reported that the positive rate of SET-CAN/NUP214 fusion gene in 196 patients with T-ALL was about 5.6% (21), in 2022, Yan C and others first reported two CML patients with positive SET-CAN/NUP214 fusion gene (7). The data revealed that although SET-CAN/NUP214 fusion gene occurs in various types of leukemia, it mainly occurs in T-ALL. This review compiled relevant literature containing more complete patient characteristics published since the emergence of the first SET-CAN/NUP214 fusion gene positive case to date, some articles were not included due to lack of patient information, a total of 81 patients’ information was collected, the overall statistical characteristics of the patients are listed in Table 1 , and detailed information on the individual characteristics of the patients are listed in Table 2 . Among the 81 patients in Table 2 , there are 57(57/81, 70.4%) patients with T-ALL, which is much higher than other types, consistent with the conclusion that the fusion gene is more likely to occur in T-ALL.

Among the fusion gene positive patients counted in this review, there are 59 male and 22 female patients, respectively, with the proportion of male patients reaching more than 70%, suggesting that the SET-CAN/NUP214 fusion gene is more likely to occur in male patients. The number of fusion gene positive T-ALL patients included 40 males and 17 females, with the proportion of males reaching 70.2%. Although there were fewer cases of other types of leukemia, there were still significantly more males than females, which suggests that the type of leukemia in fusion gene positive patients may not be an influencing factor in the proportion of males and females in the disease (4, 7, 20, 21).

There is a large difference in the age of patients at initial diagnosis, the youngest patient is only 8 years old (T-ALL), the oldest patient is 58 years old (T-ALL), the average age is 30.2 years old and the patients are distributed in all age groups (6, 17, 24, 26, 35, 41). Relatively speaking, the probability of fusion gene positive in adult leukemia patients is higher (40). Two CML patients with SET-CAN/NUP214 fusion gene positive were 37 and 42 years old, far from the average age of fusion gene positive patients. However, due to the small number of cases and the older age of CML patients, the relationship between age and fusion gene could not be established.

In previous cases, the patients with fusion gene positive leukemia did not show symptoms different from those with fusion gene negative leukemia, and most remained symptomatic with classic anemia, fever, and lower sternal segment tenderness. However, liver and spleen enlargement, lymph node enlargement, mediastinal involvement, as well as tumor bulk and rapid growth were more common than in fusion gene negative patients (5, 29, 34, 42). Some patients came to see doctors because of liver and spleen enlargement and related symptoms caused by mediastinal mass. Sang-Guk Lee et al. described a 28-year-old patient who complained of dyspnea and chest pain. Physical examination found that multiple lymph nodes in the neck were swollen. Chest CT showed that mediastinal mass compressed the main pulmonary artery with pleural effusion and splenomegaly. Finally, the patient was diagnosed as SET-CAN/NUP214 positive T-ALL (34). Song Y et al. (43) also confirmed that patients often have extramedullary infiltration at the onset of the disease, including areas such as the skin, liver and breast. According to the statistics, the median WBC count of the patients was 18.0×10⁹/L. Based on the collected patient information, the highest WBC count was 604.4 × 10⁹/L (T-ALL) and this patient died 5 months after diagnosis. The median percentage of leukemic blasts in the bone marrow was high (82.0-97.0%), probably reflecting the high proliferation status of fusion gene positive patients (4, 24).

Patients with fusion gene positive may have normal chromosome karyotype or complex karyotype, the existence of a complex karyotype may mask the presence of the fusion gene (34, 44). As a molecular abnormality with low frequency, this is also the reason why SET-CAN/NUP214 patients were not widely concerned at first.

In terms of immunophenotype, the fusion gene positive leukemia cells showed characteristics of extreme immaturity. Flow cytometry showed that their most frequent immunophenotype was CD7, except for the two CML cases mentioned previously (7), only one T-ALL patient and one AML patient reported by Zhang H (6)and Rosati R (29) did not detect CD7+. CD7 was highly frequent in SET-CAN/NUP214 fusion gene positive leukemia, and the other immunophenotypes with higher frequency were cCD3, CD34, CD33 and CD13. The immunophenotypic results suggest that the transformation of fusion gene positive leukemia may occur in the early stage of myeloid or T-lymphocyte differentiation, and it may be related to the inhibition of differentiation of primitive progenitor cells by the fusion gene (6, 7, 35, 45, 46).

Generally, myeloid markers such as CD13 and CD33 are only expressed in about 19% of T-ALL cases. The reason why the fusion gene induces myeloid marker expression remains to be further investigated.

SET-CAN/NUP214 fusion gene impairs the process of hematopoietic differentiation, but it alone is not sufficient to induce leukemia. Additional chromosomal aberrations and molecular events are required to mediate the development of leukemia. Understanding the process is greatly helpful for understanding the disease.

SET-CAN/NUP214 fusion gene may contribute to leukemia through direct and indirect effects. Saito S et al. (47) developed transgenic mice expressing SET-CAN/NUP214, which is active in different groups of hematopoietic cell groups, and the transgenic mice carrying SET-CAN/NUP214 gradually developed symptoms such as anemia, thrombocytopenia and splenomegaly, so that within 6 months, a considerable number of transgenic mice died successively, the course and characteristics of the lesions are more similar to those of leukemias, and the characterization of bone marrow cells in mice during the course of the disease showed that the SET-CAN/NUP214 fusion gene increased the number of immature cells and impaired the hematopoietic differentiation of erythroid, granulocytic, and megakaryocytic lineages (47).

Previous studies have shown that the fusion gene impairs the process of hematopoietic differentiation, but cannot induce the occurrence of leukemia alone. HOXA upregulation may be the key mechanism and play an intermediary role. HOX genes is a kind of gene that specially regulates biological form in organisms. The expression of HOX gene in various organisms is similar, and its sequence is related to its action sequence and action position. Human HOX gene can be divided into four gene clusters: HOXA, HOXB, HOXC, and HOXD, which are located on different chromosomes respectively. The DNA sequence of these gene family members is similar to the protein sequence transcribed. Quantitative and comparative analysis of bone marrow samples from SET-CAN/NUP214 positive patients during initial diagnosis and morphological remission using RT-PCR and other detection methods showed that the expression level of HOXA9 and HOXA10 at initial diagnosis was 3.53 and 4.15 times higher than that during morphological remission, while the expression level of HOXA5 was similar (34). Sang-Guk Lee and others found that the up-regulation of HOXA gene was caused by the interaction of SET-CAN/NUP214 fusion gene with XPO1, hDOT1L and HOXA promoters. The fusion genes with similar mechanism include CALM-AF10 and MLL-AF10 ( 33), which can lead to the H3K79 hypermethylation of HOXA genes and mediate the occurrence of leukemia. Hypermethylation and subsequent upregulation of HOXA genes play an important role in the pathogenesis of leukemia with positive fusion gene (48). Gorello P et al. detected the overexpression of HOXA7, HOXA9 and HOXA10 in the fusion gene positive patients selected from 256 ALL patients (7, 42). 17 fusion gene positive patients were detected by FISH analysis, and all patients(17/17, 100%) had overexpression of HOXA gene. There were also studies that summarized the up-regulation of HOXA gene with the positive expression of several NUP214 fusion gene subtypes. SET-NUP214, DEK-NUP214 and SQSTM1-NUP214 have the same characteristics, which can lead to the up-regulation of HOXA3, HOXA5, HOXA7, HOXA9, HOXA10 and HOXB in the HOX family (2, 33, 49, 50). These studies also confirmed the relationship between HOXA and SET-CAN/NUP214 fusion gene.

Na Lin et al. (40) evaluated common recurrent mutations in SET-CAN/NUP214 positive T-ALL patients through next-generation sequencing. The results showed that mutations were more common in NOTCH1(23/31,74.2%), PHF6(11/21,52.38%), KRAS(6/14,42.86%), JAK3(4/12,33.33%), CCND3(3/12,25%), JAK1(3/15,20%), STAT5B(2/10,20%), DNM2(2/10,20%) and EED(2/10,20%), these are common recurrent mutations in SET-CAN/NUP214 positive patients in T-ALL and ETP-ALL. The patients with fusion gene positive are accompanied by more molecular events than those with fusion gene negative. These complex molecular events may promote adverse reactions to induction therapy, and may also be one of the factors of poor prognosis (51). As the total number of cases remains low, these issues remain to be explored.

The protein encoded by NOTCH gene is a highly conserved cell surface receptor, which can regulate the development of a variety of biological cells. NOTCH signaling can affect a series of normal life processes of cells, including the differentiation of pluripotent progenitor cells, cell apoptosis, proliferation and cell boundary formation. The abnormality of NOTCH signaling is related to esophageal cancer, gastric cancer, leukemia and other diseases, Among them, abnormal NOTCH1 is most often detected in tumor diseases.

The activation mutation of NOTCH1 or the inactivation mutation of NOTCH1 negative regulatory factor(FBXW7) can be found in about 60% of T-ALL cases. However, the proportion of NOTCH1 mutation seems to be higher in SET-CAN/NUP214 positive leukemia patients. A gene sequencing of 6 SET-CAN/NUP214 positive T-ALL patients by Dai HP et al.(Jiangsu Institute of Hematology, China) showed most T-ALL patients with positive fusion gene have NOTCH1 mutations(5/6,83.3%) and PHF6 mutations(4/6,66.7%) (20). The next-generation sequencing of patients by Na Lin et al. (40) showed that the proportion of NOTCH1 mutations in 31 patients reached 74.2%, similarly, the results of the test performed by Wang Q et al. (52) on the association between 96 fusion gene positive patients and mutations such as NOTCH, JAK1 and others demonstrated a possible positive correlation between NOTCH1 mutations and fusion gene positivity.

The mutations of NOTCH1, PHF6 and JAK1 are closely linked in the process of leukemia, which may be the secondary genetic alterations of SET-CAN/NUP214 fusion gene. PHF6 is a tumor suppressor gene with transcriptional regulation linked to the X sex chromosome in the nucleus. Tumorigenic mutations have a higher incidence rate in T-ALL and can also be seen in AML, most of them occur in male patients. JAK1 plays a key role in initiating reactions related to a variety of major cytokine receptor families. It appears in about 20% of adult T-ALL patients, generally indicating poor prognosis. If the patients with positive fusion gene have co-mutation of NOTCH1 and PHF6, they are more likely to have symptoms such as splenomegaly and lymph node enlargement (2, 22, 52–54). In addition, the existence of SET-CAN/NUP214 fusion gene is related to the up-regulation of the expression level of lymphoblastic leukemia-associated hematopoietic regulator 1(LYL1) and myocyte enhancer 2C(MEF2C) genes (22). Contrary to the common mutations such as NOTCH1, PHF6 and JAK1, the overexpression of CALM-AF10, SIL-TAL, TLX1 or TLX3 is mutually exclusive with the existence of SET-CAN/NUP214 fusion gene. A gene test of 11 fusion gene positive T-ALL patients by Ben et al (21). showed that none of the 11 patients expressed CALM-AF10, SIL-TAL, TLX1 or TLX3(0/11,0%).

In the process of leukemogenesis mediated by SET-CAN/NUP214 fusion gene, it is generally accepted that additional chromosomal aberrations also play a role. Chae H et al. (35) reported del (12)(p13)/ETV6 in 3 of 4 patients, while Ben et al. (21) found this aberration numerous times in their cases. Similarly, the patients in the reports also presented del (6) (q21q23) and del (11) (q22q23) chromosomal aberrations (51, 55). The recurrent chromosomal aberrations in the rare fusion gene positive patients are intriguing and worth pondering.

In summary, SET-CAN/NUP214 fusion gene is relatively rare in leukemia and mainly occurs in adult male T-ALL patients. It has also been reported in AUL, MS, MPAL, AML, CML and B-ALL. Patients are generally resistant to chemotherapy, and the prognosis in different diseases may be different. The clinical symptoms of positive and negative fusion gene patients are relatively similar, and the common immunephenotypes are CD7, cCD3, CD34, CD33 and CD13. The karyotypes may be normal or complex, the concomitant molecular events can become the influencing factors of disease progression and prognosis. HSCT can significantly improve the survival rate of patients, CAR-T is also a potential treatment method. RQ-PCR is an effective monitoring method, and the monitoring of fusion gene may be more sensitive than FCM. Prognosis prediction and recurrence intervention based on the expression level of SET-CAN/NUP214 fusion gene can improve the treatment effect. Further research is needed to evaluate the role of SET-CAN/NUP214 fusion gene in leukemia.

JS: Data curation, Investigation, Methodology, Software, Visualization, Writing – original draft. HL: Conceptualization, Funding acquisition, Project administration, Supervision, Writing – review & editing. SF: Project administration, Supervision, Writing – review & editing.