Gene Expression Changes in Bone Marrow of MDS Patients with Del(5q) During Lenalidomide Treatment.

Genetic alterations reported in myelodysplastic syndromes (MDS) are not disease-specific and the underlying molecular causes of the disease remain poorly understood. It has been suggested that one or more of the genes mapping within the commonly deleted region of the 5q syndrome, together with other distant genes, may be critical to the development of the 5q syndrome. Potential candidate genes have been identified including the tumor suppressor gene SPARC, and the ribosomial protein gene, RPS14. Haploinsufficiency of RPS14 has been demonstrated and recent evidence indicates RPS14 as a causal gene for the 5q syndrome. Lenalidomide has proven efficacy in MDS patients with del(5q). Rapid and durable responses include transfusion-independence, with a rise in Hb, suppression of the 5q-deletion clone and improvement in bone marrow morphologic features.

In a multicenter Italian phase II trial to evaluate safety, changes in quality of life and efficacy of lenalidomide in primary MDS patients with del(5q) and low or Int-1 risk IPSS, we investigate changes in bone marrow cytogenetics and gene expression patterns during treatment. The starting dose of lenalidomide is 10 mg p.o once daily on a continuous daily schedule for a maximum of 12 months. Dosing is based upon clinical and laboratory findings. Bone marrow cytogenetics and gene expression profiling are performed on study entry and every 12 weeks up to end of study (week 52). Gene expression assays of 51 candidate genes from the published literature and genomic databases have been selected and are carried out with TaqMan® Low Density Array Fluidic card (TaqMan® Human Array, Applied Biosystems, Foster City, CA, USA) based on Applied Biosystems PRISM® 7900HT comparative dd CT method, according to manufacturer's instructions. Using an 18S mRNA gene pre-designed assay from Applied Biosystems to detect the expression of the housekeeping gene 18S in each sample, target gene expression is normalized with 18S gene expression derived from a bone marrow pool of normal healthy subjects and for each sample the ratio between the target and 18S are expressed.

Baseline values for 23 patients (mean age 73 ± 10 years) are available and 16 have been re-evaluated after 12 weeks. Mean Hb was 8.6 ± 0.9 g/dL and 20 patients were transfusion-dependent. Seven patients had additional cytogenetic abnormalities. At baseline, RPS14 was under-expressed in 19 out of 21 patients evaluated. After 12 weeks RPS14 was re-evaluated in 13 patients: all had erythroid responses and RPS14 increased significantly from 0.07 (IQ Range 0.03–0.13) to 76.1 (0.73– 304.0, p=0.002). SPARC expression was under-expressed in 15/23 patients and variations during treatment were not significant. Baseline FAS gene was under-expressed in all patients and increased above reference values (p=0,006) after 12 weeks in 7/14 cases. IL7R was over-expressed in all patients at baseline (median 3263.3, IQ range 1998.3–5027.1) and was significantly reduced after 12 weeks (median 0.17, IQ range 0.05–2.20, p<0.0001). TINAGL1 gene expression was very high at baseline (mean 5214 ± 3661) and significantly reduced below normal values during treatment (0.63±0.87, p<0.0001). WTI was over-expressed in almost all patients (median 3246, IQ range 1590–7785) and, though reduced during treatment, still remained highly over reference values at 12 weeks (p=0.148). The expression of the other genes did not vary significantly during treatment.

A preliminary study evaluating short-term effects of lenalidomide in low and Int-1 IPSS risk MDS and del(5q) demonstrates that gene expression is abnormal and varies during treatment. RPS14 is down-expressed and a significant increase in its expression is observed during treatment with lenalidomide in responsive patients. There is also an increase in FAS expression; the protein encoded by FAS gene is a member of the TNF-receptor superfamily and its interaction with its ligand leads to apoptosis.The IL7R gene on chromosome 5 (5p13) codifies for the IL7 receptor, which blocks apoptosis during differentiation and activation of T lymphocytes. TINAGL1 gene on chromosome 1 codifies for a Cathepsin B-like protein implicated in a number of human diseases such as cancer. During lenalidomide treatment, the expression of IL7R and TINAGL1 are reduced. Further results and repeated testing during this trial may develop new insights in the pathogenesis of MDS with del(5q) and in long-term effects of lenalidomide on bone marrow changes.

Oliva:Celgene: Consultancy. Balleari:Celgene: Consultancy. Finelli:Celgene: Consultancy.