In 1974, VandenBerghe first reported five cases of 5q- syndrome. The clinical features include macrocytic anemia, normal or elevated platelets, and poorly lobulated megakaryocytes. 5q- syndrome has a low incidence of conversion to acute myeloid leukemia, less than 10%. The World Health Organization (WHO) classification of tumors of the hematolymphatic system (2001, 2008) has classified 5q-syndrome as a separate type of myelodysplastic syndrome (MDS). 5q-syndrome has less than 5% myeloid primitive cells and is associated solely with 5q-chromosomal abnormalities[i] and has been treated with thalidomide with good results in recent years. The pathogenesis and therapeutic progress of this disease are reviewed.
1, Pathogenesis.
5q- syndrome is a hematopoietic stem cell disorder in which more than 90% of hematopoietic stem cells are derived from malignant clones. Studies have shown that 5q- syndrome patients also have 5q-chromosomal abnormalities in B and NK cells [ii]. Their bone marrow is usually proliferatively active or markedly active. The massive proliferation of malignant clones impairs erythroid development and eventually leads to anemia.
The 5q-syndrome generalized deletion region (CDR) contains 40 genes expressed in hematopoietic stem progenitors. Numerous studies have found reduced expression of these genes, but all sequencing results confirm that none of these genes are mutated [iii]-[iv]. This suggests that hemiploid insufficiency (loss of function of one allele) may be the main cause of 5q- syndrome pathogenesis.
1.1 Ribosomal gene RPS14 hemiploidy insufficiency leads to impaired erythropoiesis
In 2008, Ebert et al [v] performed a series of RNA interference (RNAi) experiments on 40 genes within the CDR of 5q-32-33. By studying alterations in the bone marrow and peripheral blood of different RNAi mice, it was found that reduced expression of RPS14 resulted in poor erythroid lineage development and led to increased erythroid apoptosis. enhanced RPS14 expression in patients with 5q-syndrome significantly improved erythropoiesis.
Ribosomal proteins are essential for the regulation of ribosomal RNA precursor (pre-RNA) processing, assembly, stabilization, intracellular transport and translation, but are not involved in the translation process.RPS14 is one of the component units of ribosomal subunit 40S and contains 151 amino acids (5.9 Kb).RPS14 hemiploidy insufficiency leads to ribosome-associated and translation-associated gene regulation abnormalities [vi]. Mutations in other ribosomal protein genes, such as RPS19 and RPS24 mutations, can lead to congenital bone marrow failure syndromes, such as Diamond-Blackfan anemia. These congenital bone marrow failure syndromes possess similar tissue characteristics and clinical features to MDS. Animal models have demonstrated that ribosomal gene defects result in ineffective erythropoiesis and predisposition to tumorigenesis. It is now believed that ribosomal gene hemiploidy insufficiency leads to insufficient ribosomal subunit formation, which results in altered translation genes and activation of differentiation and apoptosis-related proteins [vii].
Oliva et al [viii] dynamically monitored RPS14 expression levels in 17 patients with low- and intermediate-risk 1MDS with chromosome 5q deletion. Before the application of ralidomide treatment, RPS14 expression was significantly reduced in 14 patients compared to normal controls. After 14 weeks of treatment with ralidomide, the hemoglobin increased by an average of 27 g/L and the expression level of RPS14 increased by an average of about 205-fold. The findings suggest that RPS14 hemiploidy insufficiency is associated with the pathogenesis of 5q- syndrome.
1.2. p53-mediated apoptosis is involved in the pathogenesis of 5q-syndrome
p53 is a key transcription factor that regulates cell growth and apoptosis. Under normal conditions, the expression level of p53 is very low. Upon the occurrence of various cellular stresses, p53 is rapidly activated. underfunctioning RPS14 hemiploidy leads to abnormal ribosomal protein synthesis, which impairs ribosome biosynthesis, resulting in cellular stress. barlow et al [ix] recently constructed a mouse model of 5q- syndrome. The genes within the human 5q-CDR region are located on chromosomes 11 and 18, respectively, in mice. Using genetic engineering, the regions where the different target genes are located were removed. Only the removal of the Cd74CNid67 (containing the RPS14 gene) fragment resulted in mice with significant hematological abnormalities, such as macrocytic anemia and impaired hematopoietic stem progenitor cell production. These mice had increased levels of bone marrow apoptosis and elevated p53 expression. In contrast, in mice with complete knockout of p53 followed by removal of the RPS14 gene fragment, hematopoietic stem cells were normal and erythropoiesis was normal. Thus, p53-mediated apoptosis may be an important mechanism of RPS14-mediated anemia.
All Barlow experimental designs of deletion mice did not show platelet elevation. And elevated platelets are one of the main clinical manifestations in the early stages of 5q- syndrome. Whether these deletions favor the proliferation of malignant clones remains to be further investigated.
Pellagatti et al. showed the presence of p53 activation and upregulation of the p53 pathway in patients with 5q- syndrome, and that RPS14 hemiploidy insufficiency may be responsible for p53 activation. 10 p53 pathway genes were significantly abnormally expressed in CD34+ cells of 5q- syndrome patient origin. These genes included FAS, CD82, WIG1, CASP3, SESN3, TNFRSF10B, BAX, DDB2, BID, and MDM4. p53 expression was significantly elevated in all genes except for the p53 repressor regulator MDM4, which was depressed. The majority of these genes were target genes downstream of p53. Immunohistochemical results of bone marrow biopsy specimens showed enhanced p53 expression in patients with 5q- syndrome, while normal bone marrow was largely inactive [x].
1.3. Micro-RNA deletion leads to platelet proliferation and clonal proliferation.
By studying micro-RNA in the CDR region of 5q- syndrome, Starczynowski et al. found that deletion of miR-145 and miR-146a was an important factor in the pathogenesis of 5q- syndrome. miR-145 and miR-146a were abundantly expressed in CD34+ bone marrow cells, but significantly reduced in 5q- syndrome bone marrow cells. Knockdown of these two micro-RNAs in mice resulted in the development of clinical features typical of 5q- syndrome: poorly lobulated megakaryocytes and elevated peripheral blood platelets. two genes of Toll-like receptor signaling pathway, TIRAP (miR-145) and TRAF6 (miR-146a), were identified as key target genes leading to the development of the disease. TIRAP interacts with TRAF6 to activate nuclear factor k-B (NFkB). TRAF6 overexpression mice are prone to bone marrow failure or AML [xi].
1.4 Other gene hemiploidy malfunctions of CDR may be an important cofactor in the pathogenesis of 5q- syndrome
Although reduced expression of RPS14, miR-145 and miR-145a can lead to many of the key clinical features of 5q- syndrome, it is not certain that these alterations are sufficient to cause the development of 5q- syndrome. Insufficient hemiploid function of other 5q- syndrome CDR genes may also be an important factor in disease development, such as SPARC, a candidate tumor suppressor gene that regulates cell adhesion and surrounding stroma, induces apoptosis, and inhibits angiogenesis. It is currently believed that hematopoietic stem cells expressing reduced SPARC are more likely to adhere to their supporting niches, leading to clonal dominance. SPARC expression was significantly elevated in erythroid primitive cells of the 5q-syndrome treated with ralidomide, thereby removing the clonal advantage. Since SPARC-deficient mice lack the clinical hematological features typical of 5q-syndrome, SPARC is not the key gene responsible for the 5q-syndrome phenotype, but is potentially an important cofactor [xii].
The 1.5 Mbp CDR region of chromosome 5q contains a number of genes associated with high-risk MDS and AML, such as EGR1 and CTNNA1. this region is exactly what is missing in typical 5q-syndrome. egr1 is an important regulator of stem cell silencing. egr1 genes are more likely to develop myeloid tumors after exposure to toxic substances in hemiploid-deficient mice [xiii]. CTNNA1 is a tumor suppressor gene and its reduced expression increases the chance of leukemic transformation.
1.45q-Red lineage progenitor cells have reduced proliferative capacity and roughly normal differentiation
Gardaret [xiv] conducted a study on the proliferation and differentiation of 5q-syndrome erythroid cells. The results showed that the proliferation of 5q-syndrome erythroid cells was significantly reduced compared to normal, and the differentiation was approximately normal. An 18-day single CD34+ progenitor cell proliferation assay showed that 5q-syndrome-derived CD34+ progenitor cells were significantly less capable of producing erythrocytes than normal CD34+ cells, averaging only about 1/88th of normal. 5q-cell culture cells had a significant defect in RPS14, especially during the first 11 days of culture. This defect was significantly associated with diminished proliferation. The study confirms that the 5q- syndrome may be primarily an impaired proliferation of erythrocytes rather than a differentiation disorder. rPS14 may be the main factor contributing to this alteration.
1.55q- syndrome bone marrow stromal alterations
Ximeri et al[xv] demonstrated altered bone marrow stroma in patients with 5q-syndrome. 5q-syndrome patients derived from bone marrow stromal cell layer, after irradiation, could not support the growth of CD34+ stem progenitor cells of normal origin. However, after achieving complete remission with the application of ralidomide treatment, the bone marrow stromal cells of 5q-syndrome patient origin were well able to support the growth of CD34+ stem progenitor cells of normal origin. The ability of their bone marrow stromal cells to support the growth of normal stem progenitor cells was significantly increased to the level of support of normal bone marrow stroma. Bone marrow cytokine levels were also significantly altered by the application of ralidomide, with marked increases in stromal cell-derived factor 1 (SDF-1) and intercellular adhesion molecule 1 (ICAM-1). SDF-1 binds to CXCR4 on hematopoietic stem cells and is an important regulator of stem cell homing. After thalidomide treatment, the altered bone marrow stroma can be normalized by reducing the number of malignant clones.
2. 5q-syndrome and ralidomide.
2.1 Clinical efficacy of relidomide
2.1.1 Ralidomide treatment of 5q-syndrome can achieve cytogenetic remission List et al[xvi]-[xvii] applied immunosuppressive drug-relidomide to treat 5q-syndrome with good efficacy. The clinical phase II trial (MDS003) included 148 patients with transfusion-dependent 5q-syndrome and low- or intermediate-risk1 MDS (IPSS). The mean onset of action was 4.6 weeks. 112 patients (67%) were discharged from transfusion. 38 (45%) of the 85 evaluable cytogenetic efficacy achieved complete cytogenetic remission. The major toxicities were severe neutropenia and thrombocytopenia. The odds of occurrence were 55 and 44%, respectively, requiring discontinuation of the drug and the application of granulocyte colony-stimulating factor support therapy. The median duration of therapy was 2 years. In some patients, remission lasted longer. However, even in these patients in complete remission, the presence of 5q-cells could still be detected with the application of more sensitive assays.
2.1.2 Ralidomide has shown efficacy in the treatment of non-5q-syndrome patients with low- and intermediate-risk-1MDS The phase II clinical trial of ralidomide in the treatment of transfusion-dependent low- and intermediate-risk-1MDS with non-5q-syndrome has shown that ralidomide is also efficacious in transfusion-dependent low- and intermediate-risk-1MDS patients with non-5q-syndrome. A total of 214 cases were included in the trial, and 56 (26%) patients were discharged from transfusion. The mean time to onset of efficacy was 4.8 weeks. The average duration of maintenance of efficacy was 41 weeks. In addition, 37 patients had a 50% or more reduction in transfusion volume. The overall efficiency was 43%. The major toxicities were neutropenia (30%) and thrombocytopenia (25%) [xviii].
2.1.3 Ralidomide is effective in the treatment of intermediate-risk-2 and high-risk MDS with 5q-chromosome abnormalities alone
The phase II clinical trial of ralidomide for intermediate-risk-2 and high-risk MDS with 5q- showed that ralidomide was also effective in this group of patients. 60 % of the 47 patients were at high risk and 40 % were at intermediate-risk-2. 13 patients (27 %) were hematologically effective, of which 7 had complete hematological remission, 4 had complete cytogenetic remission, and 3 had partial cytogenetic remission. The majority of patients who achieved efficacy were hemodynamically effective. The vast majority of patients who achieved efficacy were patients with simple 5q-abnormalities. Of the 11 patients with an additional chromosomal abnormality, only one was effective. None of the 27 patients with two or more additional chromosomal abnormalities was effective. This suggests that only intermediate-risk-2 and high-risk patients with 5q-chromosomal abnormalities alone are treated effectively with ralidomide [xix].
2.1.4 The efficacy of relidomide treatment may correlate with the degree of neutropenia and thrombocytopenia
Sekeres et al [xx] analyzed data from 2 phase II clinical trials of ralidomide (MDS003 and MDS002) showing that thrombocytopenia and neutropenia in patients with 5q-syndrome may be associated with detachment from transfusion. Seventy percent of 5q-syndrome patients with a 50% or greater reduction in platelets were off transfusion, while only 42% of 5q-syndrome patients with no or less than 50% reduction in platelets were off transfusion. Among 5q-syndrome patients with normal basal neutrophils, 82% of those with a neutrophil decline of more than 75% were off transfusion, while only 51% of those with no or less than 75% neutrophil decline were off transfusion. In patients with non-5q-syndrome, no association was found between platelet and neutropenia and disengagement from transfusion.
2.1.5 Relapse of 5q-syndrome after remission from ralidomide treatment is associated with proliferation of its residual drug-resistant malignant clone stem cells Tehranchi et al [xxi] examined changes in malignant clones during treatment with ralidomide in seven patients with 5q-syndrome. They found that the bone marrow of patients with 5q- syndrome before treatment was overwhelmingly 5q- malignant clones in both CD34+CD38-CD90+ hematopoietic stem cells and CD34+CD38+ hematopoietic progenitor cells. Ralidomide mainly acts on CD34+CD38+ 5q-hematopoietic progenitor cells, while the majority of CD34+CD38-/lowCD90+ 5q-malignant clone hematopoietic stem cells are in quiescent phase, and ralidomide cannot effectively eradicate this part of malignant clone hematopoietic stem cells. And this fraction of cells is the root cause of eventual relapse in patients with 5q- syndrome.
2.2 Mechanism of action of ralidomide
The mechanism of action of ralidomide is complex, and its specific mechanism of action in the treatment of 5q-syndrome is still not elucidated. Its effects include inhibition of tumor necrosis factor а, interleukin-6 and IL-12, induction of caspase-8-mediated apoptosis, inhibition of cell adhesion and angiogenesis, stimulation of T and NK cell proliferation through induction of interleukin-2 and interferonг, and inhibition of Akt phosphorylation. It also has the potential to inhibit the proliferation of malignant clonal cells and promote their apoptosis [xxii]-[xxiii].
Wei et al [xxiv] showed that ralidomide inhibits two phosphatases that regulate the cell cycle: Cdc25c and PP2Aca. The genes encoding both phosphatases are located at 5q, and both genes are absent in most patients with 5q- syndrome. Studies have shown that 5q- cells are more sensitive to ralidomide and that hemiploidy of Cdc25c and PP2Aca may be responsible for the increased sensitivity.
SPARC gene hemiploidy insufficiency is the main reason for the proliferation of MDS clones. Thalidomide normalized the SPARC expression level in 5q-stem progenitor cells. Thus, the ability of malignant clones to adhere to the stroma is diminished, eliminating their clonal advantage. It is also possible that the effect of ralidomide on NK cells and cell stroma is one of the mechanisms of action [xxv].
2.3 Problems with relidomide treatment
Although the US FDA approved the marketing of MDS003 in 2005 based on the data from the MDS003 trial, which showed very significant efficacy of ralidomide in the treatment of 5q- syndrome, the European EMEA has not approved the drug for marketing in Europe so far based on concerns about the safety of its application.
Long-term follow-up of MDS003 showed that cytogenetic remission was significantly associated with AML progression. Patients in the cytogenetic partial and complete remission group had a 15% chance of AML transformation; while patients in the cytogenetic no remission group had a 67% AML transformation rate. Previous data showed that patients with 5q-syndrome not treated with relidomide had only a 10% chance of converting to AML. Considering the complexity of the cases included in the group treated with ralidomide, such as some non-primary patients, some with chromosomal abnormalities other than 5q-, and some patients with slightly higher primitive cells, it cannot be simply concluded that ralidomide does increase the chance of AML conversion.
Gohring et al[xxvi] performed an in-depth analysis of 42 German cases from the MDS003 trial. 19 of the 42 patients met the diagnostic criteria for 5q- syndrome. 15 of the 42 patients (36%) converted to AML. 7 of the 19 patients (37%) who met the criteria for 5q- syndrome converted to AML. 17 of the 42 patients developed chromosomal changes other than 5q-. chromosomal alterations other than 5q- occurred in 17 of the 42 patients, 11 of which had complex karyotypes. This high rate of AML conversion greatly exceeds previous data.
The prognosis of classical 5q- syndrome is heterogeneous. j?dersten et al[xxvii] reported a case of 5q- syndrome in which a small number of p53 mutant cells were detected before the application of ralidomide in a patient with 5q- syndrome. Although this patient had a complete normalization of erythrocytes and achieved a partial cytogenetic remission after the application of ralidomide, the p53 mutant clone was progressively elevated during ralidomide treatment and eventually progressed to AML.
Tehranchi et al21 found in seven patients with 5q-syndrome treated with ralidomide at long-term follow-up that although all patients were off transfusion and achieved partial or complete cytogenetic remission, all patients developed drug resistance at about 2 years and all patients developed a new clonal evolution. four cases had a significant increase in primitive cells, three of which progressed to MDS-RAEB-2 The four cases had a significant increase in primary cells, three of which progressed to MDS-RAEB-2 and one to AML.
The results of the ongoing study of ralidomide treatment for AML transformation will help explain whether ralidomide does indeed improve AML transformation or whether it is just a matter of patient selection.
In conclusion, RPS14, miR-145 and miR146a hemiploidy insufficiency is an important link in the pathogenesis of 5q- syndrome. The stressful state resulting from hemiploid insufficiency of these key genes initiates the p53-mediated apoptotic pathway, leading to increased apoptosis in myeloid progenitors. In-depth studies of 5q- pathogenesis could contribute to new targeted therapies. Ralidomide has achieved good efficacy in the treatment of 5q- syndrome, but its safety should also be given sufficient attention.