Optimal initial antibiotic regimen for the treatment of acute appendicitis: a systematic review and network meta-analysis with surgical intervention as the common comparator.

BACKGROUND: The optimal antibiotic regimen for the medical management of acute appendicitis remains unknown due to a lack of head-to-head comparisons between different antibiotic regimens. METHODS: We systematically searched the PubMed, EMBASE, Scopus and Cochrane Central Register of Controlled Trials databases from their inception through to August 2020. We selected randomized controlled trials (RCTs) or observational studies comparing antibiotic therapy and appendectomy as the initial treatment for adult or paediatric patients with acute appendicitis. We performed a Bayesian network meta-analysis (NMA) to obtain the indirect comparison results between different antibiotic regimens by employing the group managed by surgery as a common comparator. Antibiotic regimens were classified into three categories: those including a carbapenem; those including a cephalosporin; and those including a ß-lactam/ß-lactamase inhibitor combination. RESULTS: A total of 9 RCTs (adults, n = 8; paediatrics, n = 1) and 12 observational studies (adults, n = 3; paediatrics, n = 9) were included in the NMA, with a total of 4551 patients. The most commonly administered regimen was a ß-lactam/ß-lactamase inhibitor combination (9/21; 43%), followed by a cephalosporin (7/21; 33%) or a carbapenem (5/21; 24%). The NMA indicated that surgery significantly increased 1 year treatment success, compared with cephalosporins [OR: 16.79; 95% credible interval: 3.8-127.64] or ß-lactam/ß-lactamase inhibitor combinations (OR: 19.99; 95% credible interval: 4.87-187.57), but not carbapenems (OR: 3.50, 95% credible interval: 0.55-38.63). In contrast, carbapenems were associated with fewer treatment-related complications compared with surgery (OR: 0.12; 95% credible interval: 0.01-0.85). CONCLUSIONS: Carbapenems might be recommended as the initial antibiotic regimen for the non-operative management of adult patients with acute appendicitis. Nevertheless, due to the imprecise estimates in our NMA, additional RCTs are needed to corroborate these findings, especially for paediatric patients.

Mrc1/Claspin: a new role for regulation of origin firing.

Mrc1 and its vertebrate homologue Claspin serve as a mediator for replication stress checkpoint signaling, receiving the signal from Mec1/Rad3/ATR sensor kinase and transmitting it to the effector Rad53/Cds1/Chk1 kinase. They are likely to be a part of the replisome and facilitate the S-phase progression by promoting replication fork progression. Recent reports on Mrc1/Claspin indicate their new role in regulating the replication initiation through interaction with Cdc7, a key conserved serine-threonine kinase that triggers firing at each replication origin. Mrc1/Claspin has a specific domain that specifically interacts with Cdc7, and this domain is involved also in intramolecular interaction with its N-terminal segment. Mechanisms for novel regulation of origin firing and its timing through recruitment of Cdc7 to Mrc1/Claspin will be discussed.

Claspin is Required for Growth Recovery from Serum Starvation through Regulating the PI3K-PDK1-mTOR Pathway in Mammalian Cells.

Claspin plays multiple important roles in regulation of DNA replication as a mediator for the cellular response to replication stress, an integral replication fork factor that facilitates replication fork progression and a factor that promotes initiation by recruiting Cdc7 kinase. Here, we report a novel role of Claspin in growth recovery from serum starvation, which requires the activation of PI3 kinase (PI3K)-PDK1-Akt-mTOR pathways. In the absence of Claspin, cells do not proceed into S phase and eventually die partially in a ROS- and p53-dependent manner. Claspin directly interacts with PI3K and mTOR, and is required for activation of PI3K-PDK1-mTOR and for that of mTOR downstream factors, p70S6K and 4EBP1, but not for p38 MAPK cascade during the recovery from serum starvation. PDK1 physically interacts with Claspin, notably with CKBD, in a manner dependent on phosphorylation of the latter protein, and is required for interaction of mTOR with Claspin. Thus, Claspin plays a novel role as a key regulator for nutrition-induced proliferation/survival signaling by activating the mTOR pathway. The results also suggest a possibility that Claspin may serve as a common mediator that receives signals from different PI3K-related kinases and transmit them to specific downstream kinases.

Claspin-Dependent and -Independent Chk1 Activation by a Panel of Biological Stresses.

Replication stress has been suggested to be an ultimate trigger of carcinogenesis. Oncogenic signal, such as overexpression of CyclinE, has been shown to induce replication stress. Here, we show that various biological stresses, including heat, oxidative stress, osmotic stress, LPS, hypoxia, and arsenate induce activation of Chk1, a key effector kinase for replication checkpoint. Some of these stresses indeed reduce the fork rate, inhibiting DNA replication. Analyses of Chk1 activation in the cell population with Western analyses showed that Chk1 activation by these stresses is largely dependent on Claspin. On the other hand, single cell analyses with Fucci cells indicated that while Chk1 activation during S phase is dependent on Claspin, that in G1 is mostly independent of Claspin. We propose that various biological stresses activate Chk1 either directly by stalling DNA replication fork or by some other mechanism that does not involve replication inhibition. The former pathway predominantly occurs in S phase and depends on Claspin, while the latter pathway, which may occur throughout the cell cycle, is largely independent of Claspin. Our findings provide evidence for novel links between replication stress checkpoint and other biological stresses and point to the presence of replication-independent mechanisms of Chk1 activation in mammalian cells.

Correction: Hsiao et al. Claspin-Dependent and -Independent Chk1 Activation by a Panel of Biological Stresses. Biomolecules 2023, 13, 125.

In the original publication [...].

Cdc7 activates replication checkpoint by phosphorylating the Chk1-binding domain of Claspin in human cells.

Replication checkpoint is essential for maintaining genome integrity in response to various replication stresses as well as during the normal growth. The evolutionally conserved ATR-Claspin-Chk1 pathway is induced during replication checkpoint activation. Cdc7 kinase, required for initiation of DNA replication at replication origins, has been implicated in checkpoint activation but how it is involved in this pathway has not been known. Here, we show that Cdc7 is required for Claspin-Chk1 interaction in human cancer cells by phosphorylating CKBD (Chk1-binding-domain) of Claspin. The residual Chk1 activation in Cdc7-depleted cells is lost upon further depletion of casein kinase1 (CK1γ1), previously reported to phosphorylate CKBD. Thus, Cdc7, in conjunction with CK1γ1, facilitates the interaction between Claspin and Chk1 through phosphorylating CKBD. We also show that, whereas Cdc7 is predominantly responsible for CKBD phosphorylation in cancer cells, CK1γ1 plays a major role in non-cancer cells, providing rationale for targeting Cdc7 for cancer cell-specific cell killing.

Claspin recruits Cdc7 kinase for initiation of DNA replication in human cells.

Claspin transmits replication stress signal from ATR to Chk1 effector kinase as a mediator. It also plays a role in efficient replication fork progression during normal growth. Here we have generated conditional knockout of Claspin and show that Claspin knockout mice are dead by E12.5 and Claspin knockout mouse embryonic fibroblast (MEF) cells show defect in S phase. Using the mutant cell lines, we report the crucial roles of the acidic patch (AP) near the C terminus of Claspin in initiation of DNA replication. Cdc7 kinase binds to AP and this binding is required for phosphorylation of Mcm. AP is involved also in intramolecular interaction with a N-terminal segment, masking the DNA-binding domain and a newly identified PIP motif, and Cdc7-mediated phosphorylation reduces the intramolecular interaction. Our results suggest a new role of Claspin in initiation of DNA replication during normal S phase through the recruitment of Cdc7 that facilitates phosphorylation of Mcm proteins.