AMPK activation induces immunogenic cell death in AML.

Survival of patients with acute myeloid leukemia (AML) can be improved by allogeneic hematopoietic stem cell transplantation (allo-HSCT) because of the antileukemic activity of T and natural killer cells from the donor. However, the use of allo-HSCT is limited by donor availability, recipient age, and potential severe side effects. Similarly, the efficacy of immunotherapies directing autologous T cells against tumor cells, including T-cell recruiting antibodies, chimeric antigen receptor T-cell therapy, and immune checkpoint inhibitors are limited in AML because of multiple mechanisms of leukemia immune escape. This has prompted a search for novel immunostimulatory approaches. Here, we show that activation of adenosine 5'-monophosphate-activated protein kinase (AMPK), a master regulator of cellular energy balance, by the small molecule GSK621 induces calreticulin (CALR) membrane exposure in murine and human AML cells. When CALR is exposed on the cell surface, it serves as a damage-associated molecular pattern that stimulates immune responses. We found that GSK621-treated murine leukemia cells promote the activation and maturation of bone marrow-derived dendritic cells. Moreover, vaccination with GSK621-treated leukemia cells had a protective effect in syngeneic immunocompetent recipients bearing transplanted AMLs. This effect was lost in recipients depleted of CD4/CD8 T cells. Together, these results demonstrate that AMPK activation by GSK621 elicits traits of immunogenic cell death and promotes a robust immune response against leukemia. Pharmacologic AMPK activation thus represents a new potential target for improving the activity of immunotherapy in AML.

Rac1 mediates cadherin-11 induced cellular pathogenic processes in aortic valve calcification.

BACKGROUND: Calcific aortic valve disease (CAVD), a major cause for surgical aortic valve replacement, currently lacks available pharmacological treatments. Cadherin-11 (Cad11), a promising therapeutic target, promotes aortic valve calcification in vivo, but direct Cad11 inhibition in clinical trials has been unsuccessful. Targeting of downstream Cad11 effectors instead may be clinically useful; however, the downstream effectors that mediate Cad11-induced aortic valve cellular pathogenesis have not been investigated. APPROACH AND RESULTS: Immunofluorescence of calcified human aortic valves revealed that GTP-Rac1 is highly upregulated in calcified leaflets and is 2.15 times more co-localized with Cad11 in calcified valves than GTP-RhoA. Using dominant negative mutants in porcine aortic valve interstitial cells (PAVICs), we show that Cad11 predominantly regulates Runx2 nuclear localization via Rac1. Rac1-GEF inhibition via NSC23766 effectively reduces calcification in ex vivo porcine aortic valve leaflets treated with osteogenic media by 2.8-fold and also prevents Cad11-induced cell migration, compaction, and calcification in PAVICs. GTP-Rac1 and Trio, a known Cad11 binding partner and Rac1-GEF, are significantly upregulated in Nfatc1Cre; R26-Cad11Tg/Tg (Cad11 OX) mice that conditionally overexpress Cad11 in the heart valves by 3.1-fold and 6.3-fold, respectively. Finally, we found that the Trio-specific Rac1-GEF inhibitor, ITX3, effectively prevents Cad11-induced calcification and Runx2 induction in osteogenic conditions. CONCLUSION: Here we show that Cad11 induces many cellular pathogenic processes via Rac1 and that Rac1 inhibition effectively prevents many Cad11-induced aortic disease phenotypes. These findings highlight the therapeutic potential of blocking Rac1-GEFs in CAVD.