Attenuation of the Apoptosis Stimulating Protein of TP53-1 (ASPP1) Is a Predictive Marker for Therapy Resistance and Overall Survival in Acute Myeloid Leukemia (AML)
Authors: Schittenhelm Marcus M. , Kaiser Max , Tsintari Vasileia , Blumenstock Gunnar , Györffy Balazs , Kampa-Schittenhelm Kerstin Maria 
Published: November 2021

ASPP1 (PPP1R13B) belongs to a family of p53-binding proteins and enhances apoptosis by stimulation of p53-transactivation of selected proapoptotic target genes. It is preferentially expressed in hematopoietic stem cells (HSC) and together with p53 preserves the genomic integrity of the HSC pool. We have previously demonstrated that ASPP1 is attenuated in AML, which is linked to methylation of the promoter region. ASPP1-interferference models suggest a functional role in leukemogenesis and therapy response. We now provide evidence that ASPP1 is highly altered in AML and attenuated expression levels associate with inferior outcomes:

mRNA expression patterns of ASPP1 were assessed in an unselected patient cohort with newly diagnosed AML (n=39) - and were found to be significantly lower compared to bone marrow aspirates of 12 healthy donors (p &lt; 0.0001, Mann-Whitney test) with a median relative expression level of 0.33 (ASPP1 AML) vs. 0.95 (ASPP1 donor). Subcohort analysis by genetic risk profile according to the European LeukemiaNet (ELN) 2017 genetic risk stratification revealed significantly lower expression levels of the intermediate/adverse risk group (n=27) compared to the favorable risk group (n=10, p=0,013) with a median relative expression level of 0.21 (ASPP1 int/adv) vs. 0.46 (ASPP1 good). In addition, analysis of patients undergoing induction chemotherapy demonstrated a significantly lower complete remission (CR) rate in patients with attenuated ASPP1 levels (p=0.0015) with a median relative expression level of 0.13 (ASPP1 nonCR) vs. 0.51 (ASPP1 CR). Of note, patients with a &gt;10x fold decrease of ASPP2 expression were exclusively found in the patient cohort failing induction therapy. To validate these observations in an independent patient set, we next performed a database RNAseq screen on a defined, unselected AML cohort (n=142, FAB M3 were excluded). Together, ASPP1 was independently confirmed as an adverse prognostic factor: Overall survival (OS) was longer in the ASPP1 high cohort vs. the ASPP1 low group with a hazard ratio for death of 0.63. Even though this analysis closely failed significance (0.082) due to lack of power, probability to be alive after 70 months was 45% (ASPP1 high) vs. 10% (ASPP1 low) with a median survival of 19.23 months (ASPP1 low) vs. 26.4 months (ASPP1 high). Most interestingly, looking at the prognostic good risk population: attenuation of ASPP1 resulted in a dramatic decrease of survival with a hazard ratio of 0.29 and a probability to be alive after 70 months of 25% (ASPP1 low) vs. 85% (ASPP1 high). Consequently, screening for ASPP1 expression may define a patient cohort likely to fail (induction) therapy - which might be a target population for hypomethylating agents (HMA) to reconstitute ASPP1 and sensitize towards (chemo)therapy.

To address this hypothesis, MOLM-14 acute leukemia cells and patient derived freshly-isolated AML samples were treated with decitabine - resulting in upregulation of ASPP1 mRNA expression levels in all assessed samples. As expected, cells were more chemo-susceptible when exposed to decitabine followed by daunorubicin and cytarabine in an ASPP1-dependent manner: MOLM-14.ASPP1i cell line strains did not reveal a significant change of proapoptotic efficacy when compared to mock strains which significantly benefitted from this approach with more than doubled proapoptotic rates (**** p &lt; 0.0001, t-test), again confirming ASPP1 target specificity.

Our results demonstrate that dysfunctional regulation of ASPP1 expression is frequently observed in AML, which associates with therapy responses and survival outcome. Prospective clinical studies are warranted to evaluate the role as a biomarker for risk stratification. Further, we provide a rationale to re-sensitize high-risk patients (defined as ASPP1 low-expressors) towards chemotherapy using HMA priming upfront - and this strategy should be followed in future trials.

Disclosures

Schittenhelm: BMS: Other: advisory board; Takeda: Other: advisory board; Astellas: Other: advisory board; University of Tuebingen: Patents & Royalties: patent for ASPP2k. Kampa-Schittenhelm: University of Tuebingen: Patents & Royalties: patent related to ASPP2k.