SETD2 Heterozygous Loss in Leukemia Leads to Chemotherapy Resistance through Attenuation of the DNA Damage Response

Relapsed leukemia is chemotherapy resistant and survival is poor. We recently found that mutations in epigenetic regulators are enriched in relapsed pediatric B-cell acute lymphoblastic leukemia (B-ALL), suggesting that they are associated with clonal survival and chemotherapy resistance. The mutations included loss of function mutations in the epigenetic regulator SETD2, which had not before been described in B-ALL. SETD2 is the only known mammalian Histone 3 Lysine 36 (H3K36) trimethyltransferase and one or two copy loss leads to a global decrease or elimination, respectively, of its chromatin mark, H3K36me3.

Using CRISPR/Cas9 to engineer isogenic human and murine leukemia cells, we have demonstrated that SETD2 loss leads to resistance to DNA damaging chemotherapy used in standard ALL therapy, consistent with the gain of SETD2 mutations at relapse.  Leukemia cells with SETD2 heterozygous deletion demonstrated a significant increase in IC50 to the DNA damaging anti-metabolites 6-thioguanine (6TG) and cytarabine (AraC), but not the protein synthesis inhibitor L-Asparaginase (Asp). To model the enrichment of SETD2 mutations at relapse, we competed a small population of SETD2 heterozygous cells with isogenic control leukemia cells.  Within 3 weeks, treatment with 6TG and AraC, but not Asp or vehicle, led to a dramatic selection of SETD2 heterozygous cells, mimicking the selective pressure of SETD2 clones in patients.

Recent reports have shown that H3K36me3 is important in the DNA damage response (DDR).  Consistent with this, we find that heterozygous SETD2 loss impairs the DDR signaling pathway with attenuated phosphorylation of multiple DDR components, including Chk1 and Chk2, in response to chemotherapy.  Consequently, SETD2 heterozygous cells showed an abrogation of the apoptotic response to the DNA damaging chemotherapy agents 6TG and AraC, despite a normal apoptotic response to Asp.

In summary, we have demonstrated that SETD2 loss leads to resistance to DNA damaging chemotherapy. The resistance is caused by a failure to trigger DDR signaling and apoptosis after chemotherapy induced DNA damage. These findings identify heterozygous SETD2 loss as a novel mechanism of chemotherapy resistance in leukemia and have implications for therapy selection and novel therapeutic interventions.