Abstract 651: Chemotherapy induces a transient senescence-like state capable of initiating AML recurrence
Authors: Li Meng , Teater Matt , Meydan Cem , Garrett-Bakelman Francine E. , Chin Christopher , Carroll Martin , Melnick Ari , Duy Cihangir 
Published: September 2020

In acute myeloid leukemia (AML), chemotherapy is frequently followed by disease relapse, yet the mechanism by which AML reemerges is not fully understood. We hypothesized that chemotherapy-induced senescence (CIS) facilitates survival to genotoxic exposure, allowing AML cells to endure treatment in a transiently dormant state while retaining potential for leukemic repopulation.

Here we show that, following treatment with cytarabine (AraC), primary AML cells exhibit hallmark senescence features, including morphological changes, senescence-associated transcriptomic alterations, and senescence-associated-beta-galactosidase (SA-β-gal) activity. To quantify SA-β-gal activity, we used a fluorogenic substrate that enabled sensitive cytometric quantification of the senescence marker in viable cells. Mechanistically, we demonstrate that genotoxic stress following AraC treatment activated the ATR kinase, a key regulator of DNA damage response. Inhibition of ATR abrogated CIS in leukemia cells and induced cell death, indicating that CIS mediates stress survival.

We next analyzed the gene expression profiles of the minor residual fraction of chemotherapy-surviving cells ex vivo and in patients after chemotherapy. We found that leukemia stem cell (LSC) gene signatures were largely depleted in chemotherapy-surviving cells. However, we did find that the CIS signature was positively enriched. To recapitulate relapse of disease, we established an AML relapse model using primary human specimens engrafted in NSG mice. Treatment of AML-engrafted NSG mice with a physiologically relevant regimen of AraC reduced leukemic burden to a small residual leukemia fraction 8 days after therapeutic initiation. This reduction in leukemic burden was reversed to full blown leukemia present at day 29. Gene expression analysis of purified human AML cells at nadir (day 8) showed significant enrichment for senescence signatures. This enrichment was reduced by day 29 post-treatment, as expression was partially reversed to untreated levels. While we did not observe an enrichment of LSC signatures at nadir, we found an enrichment of LSC gene signatures in 60% of xenografted AML cases after overt recurrence of disease, suggesting partial reversal of the senescence phenotype and recovery of the LSC phenotype. We observed a similar pattern, when we performed RNA-seq analysis in matched AML specimens from patients at diagnosis vs relapse. To examine if LSCs might be present in the CIS cell population, we performed scRNA-seq in primary AML before and after chemotherapy. Although chemotherapy killed a majority of AML cells, we did not find an enrichment of LSCs on single cell level compared to untreated control cells in most AML specimens. Instead, we found increased expression of the CIS gene signature. Next, to test if CIS cells possess self-renewal capability, we sorted for SA-β-gal+ cells after AraC exposure and seeded them in CFC assays. We found colony-forming potential of CIS cells, suggesting a capacity to repopulate leukemia. To confirm that leukemia can be initiated from CIS AML in vivo, we transplanted sorted CIS AML cells into NSG mice and found that these cells successfully repopulated leukemia.

Altogether, we propose a mechanism of AML relapse whereby AML cells survive chemotherapy via acquisition of a transient senescent-like state.

Citation Format: Meng Li, Matt Teater, Cem Meydan, Francine E. Garrett-Bakelman, Christopher Chin, Martin Carroll, Ari Melnick, Cihangir Duy. Chemotherapy induces a transient senescence-like state capable of initiating AML recurrence [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 651.