Non-homologous end joining (NHEJ) is a major pathway to repair DNA double-strand breaks (DSBs), which can display different types of broken ends. However, it is unclear how NHEJ factors organize to repair diverse types of DNA breaks. Here, through systematic analysis of the human NHEJ factor interactome, we identify PAXX as a direct interactor of Ku. The crystal structure of PAXX is similar to those of XRCC4 and XLF. Importantly, PAXX-deficient cells are sensitive to DSB-causing agents. Moreover, epistasis analysis demonstrates that PAXX functions together with XLF in response to ionizing radiation-induced complex DSBs, whereas they function redundantly in response to Topo2 inhibitor-induced simple DSBs. Consistently, PAXX and XLF coordinately promote the ligation of complex but not simple DNA ends in vitro. Altogether, our data identify PAXX as a new NHEJ factor and provide insight regarding the organization of NHEJ factors responding to diverse types of DSB ends.

Relapse is the leading cause of mortality in children with acute lymphoblastic leukemia (ALL). Among chemotherapeutics, thiopurines are key drugs in the backbone of ALL combination therapy. Using whole-exome sequencing, we identified relapse-specific mutations in phosphoribosyl pyrophosphate synthetase 1 (PRPS1), a rate-limiting purine biosynthesis enzyme, in 24/358 (6.7%) relapse B-ALL cases. All individuals who harbored PRPS1 mutations relapsed early on-treatment, and mutated ALL clones expanded exponentially prior to clinical relapse. Our functional analyses of PRPS1 mutants uncovered a new chemotherapy resistance mechanism involving reduced feedback inhibition of de novo purine biosynthesis and competitive inhibition of thiopurine activation. Notably, the de novo purine synthesis inhibitor lometrexol can effectively abrogate PRPS1 mutant-driven drug resistance. Overall these results highlight the importance of constitutive activation of de novo purine pathway in thiopurine resistance, and offer therapeutic strategies for the treatment of relapsed and resistant ALL.

Drug molecules not only interact with specific targets, but also alter the state and function of the associated biological network. How to design drugs and evaluate their functions at the systems level becomes a key issue in highly efficient and low–side-effect drug design. The arachidonic acid metabolic network is the network that produces inflammatory mediators, in which several enzymes, including cyclooxygenase-2 (COX-2), have been used as targets for anti-inflammatory drugs. However, neither the century-old nonsteriodal anti-inflammatory drugs nor the recently revocatory Vioxx have provided completely successful anti-inflammatory treatment. To gain more insights into the anti-inflammatory drug design, the authors have studied the dynamic properties of arachidonic acid (AA) metabolic network in human polymorphous leukocytes. Metabolic flux, exogenous AA effects, and drug efficacy have been analyzed using ordinary differential equations. The flux balance in the AA network was found to be important for efficient and safe drug design. When only the 5-lipoxygenase (5-LOX) inhibitor was used, the flux of the COX-2 pathway was increased significantly, showing that a single functional inhibitor cannot effectively control the production of inflammatory mediators. When both COX-2 and 5-LOX were blocked, the production of inflammatory mediators could be completely shut off. The authors have also investigated the differences between a dual-functional COX-2 and 5-LOX inhibitor and a mixture of these two types of inhibitors. Their work provides an example for the integration of systems biology and drug discovery.