Targeting sclerostin loop3 to promote bone formation with low cardiovascular risk in animal models

Sclerostin is a novel target for bone anabolic therapy. Antibody against sclerostin (romosozumab) for postmenopausal osteoporosis has been approved by the Advisory Committee of US FDA, with a black box warning for cardiovascular risk. Due to this risk, romosozumab was rejected by the European Medicines Agency. Importantly, there is a growing cardiovascular concern for osteoporotic patients with cardiovascular diseases history after therapeutic antibody treatment. Therefore, developing a next generation of sclerostin inhibitors with low cardiovascular risk is inevitable.

Sclerostin has a protective role in cardiovascular system through inhibiting the proinflammatory cytokines to prevent both aortic aneurysm and atherosclerosis development in a typical cardiovascular model using ApoE-/- mice induced by angiotensin II. Indeed, using the same mouse model, we found that anti-sclerostin antibody promoted both aortic aneurysm and atherosclerosis development, and enhanced the serum levels of proinflammatory cytokines such as IL-6, MCP-1 and TNF-α. Sclerostin has three loops within its central region. Interestingly, our in vitro studies indicated that sclerostin participated its function in bone and cardiovascular system through different loops. Both loop2 and loop3 played important roles in inhibiting osteogenesis and bone formation through Wnt signaling, whereas only loop2 played the role in inhibiting the proinflammatory cytokines. Based on these findings, we hypothesized that specifically targeting sclerostin loop3 could promote bone formation and keep its function in cardiovascular protection intact. Aptamers are ssDNA/RNA which bind to their targets through 3D complementarity. Aptamers can be tailored selected by introducing positive and negative targets. Thus, we designed aptamer selectively targets the sclerostin loop3 (aptamer aptscl56). Micro-CT and bone histomorphometry analysis revealed that aptscl56 could promote bone formation, increase bone mass and improve bone microarchitecture integrity, in ovariectomized osteoporosis rats. Importantly, aptscl56 has no effects on aortic aneurysm and atherosclerosis development, as well as the proinflammatory cytokines in the cardiovascular model using ApoE-/- mice. In toxicity evaluation in rats, the serum levels of the liver, kidney function indexes and hematologic parameters were not notably altered after a single or multiple administration(s) of aptscl56. Consistently, our in vitro data showed that aptscl56 could promote osteogenesis and bone formation through Wnt signaling, but not affect the levels of proinflammatory cytokines, which fully supported our in vivo data.

To further evaluate the effects of aptscl56 on bone anabolic and cardiovascular safety in vivo, we have the following three specific aims in this proposal:

Aim 1: To evaluate how the doses of aptscl56 affects the bone anabolic efficacy in ovariectomized-rats with established osteoporosis;
Aim 2: To determine how aptscl56 exerts the bone anabolic effect in ovariectomized-rats with established osteoporosis;
Aim3: To evaluate whether aptscl56 affects the myocardiac ischemic development in the typical cardiovascular model using ApoE-/- mice.

Taken together, this study could facilitate the development of the next generation sclerostin inhibitor specifically targeting sclerostin loop3 for bone anabolic treatment in established osteoporosis, with a low safety concern of cardiovascular risk.