Azacitidine Switch to Lenalidomide Eradicated the TP53/ CDKN2A Co-Mutated Clone and Induced Long-Term Erythroid Response in Del(5q) MDS

Abstract

Management of progressing Myelodysplastic Syndrome (MDS) represents a very difficult task for clinicians. While targeted therapy with lenalidomide is administered at low-risk phase and is primarily effective in patients with deletion of chromosome 5q, it is largely ineffective upon progression at high-risk MDS phase characterized by accumulation of adverse somatic mutations involving the tumor suppressor protein TP53. We herein report a 68-year old male MDS del(5q) patient that progressed to higher risk MDS EB1 with red blood cell transfusion dependency. Administration of 17 cycles of azacitidine inhibited further progression by stabilizing disease with Complete Marrow Response (mCR) without hematology improvement. The patient despite of reaching mCR on AZA further increased the allelic burden of gene mutations in the Cyclin- Dependent Kinase Inhibitor 2A, CDKN2A at residue D74A, Tumor Suppressor TP53 (K373R+T377P), and the splicing factor ZRSR2 (P13T). Based on the fact that the patient reached lower blast count on AZA but was accumulating adverse mutations we decided to switch the therapy into lenalidomide. The lenalidomide therapy repelled the progression-prone subclones characterized by the somatic mutations, fully normalized blood counts, and produced a longlasting remission. Our data suggest that the del(5q) patient progressing to high risk MDS could be treated by azacitidine to block MDS progression, however, only additional therapeutic line of lenalidomide was capable to suppress the progression-prone clones characterized by unfavorable mutations involving also TP53.

Keywords: Myelodysplastic syndrome; Azacitidine; Lenalidomide; NGS; ddPCR

Abbreviations

EPO: Erythropoietin; AZA: Azacitidine; LEN: Lenalidomide; TU: Transfusion Unit; HB: Hemoglobin; MCV: Mean Corpuscular Volume; DG: Diagnosis; MDS: Myelodysplastic Syndrome

Case Presentation

Management of progressing Myelodysplastic Syndrome (MDS) represents a very difficult task for clinicians. While targeted therapy with Lenalidomide (LEN) is administered at low-risk phase and is primarily effective in patients with deletion of chromosome 5q, it is largely ineffective upon progression at High-Risk (HR) MDS phase characterized by accumulation of adverse somatic mutations involving the tumor suppressor protein TP53. LEN therapy is not administered to MDS-progressing patients. However, it may be of clinical benefit if LEN could prolong the Azacitidine (AZA)-mediated response that is often complicated by AZA-resistance. We herein present longitudinal data from MDS del(5q) patient that progressed to HR-MDS EB1 with red blood cell transfusion dependency. While administration AZA induced the Complete marrow Response (mCR) the switch to LEN therapy repelled the progression-prone sub clones characterized by adverse somatic mutations and restored hematology parameters.

MDS progression is often associated with the accumulation of genetic aberrations that allow high survival properties for myeloblast outgrowth. In MDS subtype characterized by chromosome 5q deletion, the sensitivity to LEN treatment may lead to red blood cell recovery of transfusion-dependent anemia. LEN therapy induces tumor-specific cell lethality via Cereblon-dependent degradation of haplodeficient proteins encoded within the commonly deleted region at chromosome 5q. In non-del(5q) MDS, LEN sensitivity was documented to enhance the Erythropoietin (EPO) receptor-initiated transcriptional response. Monitoring of cytogenetic aberrations by FISH and nucleotide variants by NGS provides useful data for assessment of clinical outcomes example includes a study showing that non-del(5q) ancestral clones containing a distinct pattern of mutations may expanded over time on LEN [1]. Inversely, loss of LEN sensitivity may lead to re-expansion of del(5q) clone with transient sensitivity to DNA-methylation inhibitor AZA [1]. Despite the promising potential of AZA and LEN therapy to block progression in HR-MDS and to induce complete remission, the AZA-based combinations had similar response rate (ORR) to AZA alone (Sekeres, Othus et al. 2017). The del(5q) MDS patients may progress to HR-MDS, which is indicated for AZA therapy. Since this process has been in some patients associated with mutations of tumor suppressor TP53, the response to AZA may be limited. Allele burden of the TP53 mutant clone in del(5q) MDS is inversely proportional to Overall Survival (OS), confirming the importance of p53 as a negative prognostic variable of therapy response [2]. However, recently we and also others have observed that some but not all MDS patients with TP53 mutations treated by AZA have relatively longer survival, which creates a possibility that AZA can partly control growth of these clones [3]. To ensure that the mutation-bearing clone was a target of therapy it is essential to monitor the mutation pattern in respect to therapeutic lines.

We herein present complex clinical, laboratory and molecular data of 68-year-old male MDS patient that was initially diagnosed with MDS with isolated cytogenetically detected del(5q) in 46% of cells. Risk of progression was calculated as low (IPSS-R was determined to be 3=intermediate). However, after 14 months the patient became transfusion-dependent as he progressed to EB1 (excess of blasts 1). 4-months EPO administration achieved no response. Besides cytogenetic del(5q) aberration (detected by standard and advanced FISH technology), his Bone Marrow (BM) contained 7% Myeloblasts (MB). BM was analyzed by TruSight Myeloid Sequencing Panel (Illumina, San Diego, USA) that determines integrity of 54 target gene regions and is a set of 568 amplicons and ~141 kb designed to detect somatic variants previously associated with myeloid malignancies.

Mutations of CDKN2A (D74A) and TP53 (K373R) with pathogenic FATHMM score 0.71 were noted in BM CD3-depleted myeloid cells. The TP53 K373R mutation has been previously described in AML [4] and reported in the COSMIC (https://cancer.sanger.ac.uk/cosmic) database. Additional two less-pathogenic variants in TP53 (T377P) and ZRSR2 (P13T) also exceeded a 5% cutoff. Next, the patient was treated with AZA 75mg/m2 (5-2-2 regimen). After 4 cycles and 13 cycles the restaging analyses indicated the marrow CR. Nevertheless, the transfusion dependency (>4 transfusion units/6 weeks) stayed unimproved with AZA as the patient collectively received 72 blood units, which aggravated his iron overload characterized by increasing Ferritin levels. Importantly, tumor burden measured as a function of abundance of del(5q) + the somatic mutations was not affected by AZA (Figure 1).

    

    

    Figure 1: Timeline of patients’ disease course. Shortly after diagnosis of del(5q) MDS there was a development of transfusion dependency, patient started the EPO
therapy, however without any effect. Control BM examinations indicated progression to EB1 and the patient started the AZA treatment lasting for 17 cycles, which
led to a decrease of bone marrow blasts (mCR) but concurrently increased the allelic burden of somatic mutations. The patient remained transfusion-dependent and
therefore the therapy was switched to LEN that led to a complete response (Hemoglobin levels and Mean corpuscular volumes are shown at the top). Middle panel
shows dynamics of bone marrow blast counts and variant allele frequencies (in %) of somatic mutations during the therapies. Besides persisting del(5q) aberration
the patient contained also mutations in CDKN2A D74A, TP53 K373R+T377P and ZRSR2 P13T; the clone/s was eradicated by LEN therapy.