Dissecting the molecular mechanisms of breast cancer bone metastasis for therapeutic targeting

Breast cancer that has metastasized to bone is currently an incurable disease, causing significant morbidity and mortality. The aim of this thesis work was to elucidate molecular mechanisms of bone metastasis and thereby gain insights into novel therapeutic approaches. First, we found that L‐serine biosynthesis genes, phosphoglycerate dehydrogenase (PHGDH), phosphoserine aminotransferase 1 (PSAT1) and phosphoserine phosphatase (PSPH), were up‐regulated in highly bone metastatic MDA‐MB‐231(SA) cells as compared with the parental breast cancer cell line. Knockdown of serine biosynthesis inhibited proliferation of MDA‐MB‐231(SA) cells, and L‐serine was essential for the formation of bone resorbing osteoclasts. Clinical data demonstrated that high expression of PHGDH and PSAT1 was associated with decreased relapse‐free and overall survival and with features typical of poor outcome in breast cancer. Second, RNA interference screening pointed out heparan sulfate 6‐O‐sulfotransferase 2 (HS6ST2) as a critical gene for transforming growth factor β (TGF‐β)‐induced interleukin 11 (IL‐11) production in MDA‐MB‐231(SA) cells. Exogenous heparan sulfate glycosaminoglycans heparin and K5‐NSOS also inhibited TGF‐β‐induced IL‐11 production in MDA‐MB‐231(SA) cells. Furthermore, K5‐NSOS decreased osteolytic lesion area and tumor burden in bone in mice. Third, we discovered that the microRNAs miR‐204, ‐211 and ‐379 inhibited IL‐11 expression in MDA‐MB‐231(SA) cells through direct targeting of the IL‐11 mRNA. MiR‐379 also inhibited Smad‐mediated signaling. Gene expression profiling of miR‐204 and ‐379 transfected cells indicated that these microRNAs down‐regulate several bone metastasis‐relevant genes, including prostaglandin‐endoperoxide synthase 2 (PTGS2). Taken together, this study identified three potential treatment strategies for bone metastatic breast cancer: inhibition of serine biosynthesis, heparan sulfate glycosaminoglycans and restoration of miR‐204/‐211/‐379.

Intracranial Nimodipine Implant: Feasibility and Implications for the Treatment of Subarachnoid Hemorrhage – A Pre-Clinical Study

Intracranial aneurysmal subarachnoid hemorrhage (aSAH) is a life-threatening condition requiring immediate neurocritical care. A ruptured aneurysm must be isolated from arterial circulation to prevent rebleeding. Open surgical clipping of the neck of the aneurysm or intra-arterial filling of the aneurysm sack with platinum coils are major treatment strategies in an acute phase. About 40% of the patients suffering from aSAH die within a year of the bleeding despite the intensive treatment. After aSAH, the patient may develop a serious complication called vasospasm. Major risk for the vasospasm takes place at days 5–14 after the primary bleeding. In vasospasm, cerebral arteries contract uncontrollably causing brain ischemia that may lead to death. Nimodipine (NDP) is used to treat of vasospasm and it is administrated intravenously or orally every four hours for 21 days. NDP treatment has been scientifically proven to improve patients’ clinical outcome. The therapeutic effect of L-type calcium channel blocker NDP is due to the ability to dilate cerebral arteries. In addition to vasodilatation, recent research has shown the pleiotropic effect of NDP such as inhibition of neuronal apoptosis and inhibition of microthrombi formation. Indeed, NDP inhibits cortical spreading ischemia. Knowledge of the pathophysiology of the vasospasm has evolved in recent years to a complex entity of early brain injury, secondary injuries and cortical spreading ischemia, instead of being pure intracranial vessel spasm. High NDP levels are beneficial since they protect neurons and inhibit the cortical spreading ischemia. One of the drawbacks of the intravenous or oral administration of NPD is systemic hypotension, which is harmful particularly when the brain is injured. Maximizing the beneficial effects and avoiding systemic hypotension of NDP, we developed a sustained release biodegradable NDP implant that was surgically positioned in the basal cistern of animal models (dog and pig). Higher concentrations were achieved locally and lower concentrations systemically. Using this treatment approach in humans, it may be possible to reduce incidence of harmful hypotension and potentiate beneficial effects of NDP on neurons. Intracellular calcium regulation has a pivotal role in brain plasticity. NDP blocks L-type calcium channels in neurons, substantially decreasing intracellular calcium levels. Thus, we were interested in how NDP affects brain plasticity and tested the hypothesis in a mouse model. We found that NDP activates Brain-derived neurotrophic factor (BDNF) receptor TrkB and its downstream signaling in a reminiscent of antidepressant drugs. In contrast to antidepressant drugs, NDP activates Akt, a major survival-promoting factor. Our group’s previous findings demonstrate that long-term antidepressant treatment reactivates developmental-type of plasticity mechanisms in the adult brain, which allows the remodeling of neuronal networks if combined with appropriate rehabilitation. It seems that NDP has antidepressant-like properties and it is able to induce neuronal plasticity. In general, drug induced neuronal plasticity has a huge potential in neurorehabilitation and more studies are warranted.