Methamphetamine (Meth) is among the most frequently abused medicines worldwide. for Meth misuse. Methamphetamine (Meth) misuse leads to cognitive dysfunction, neurodegeneration, infections, and several complications1,2,3, and has become a general public health and Rabbit Polyclonal to GPR113. social concern around the world. Currently, there is no effective pharmacological therapy for Meth misuse in individuals4,5. Selective antibodies against Meth have been examined for the treatment of Meth addiction through LY2140023 active or passive immunization6,7, an approach that was initially developed for the treatment of snake venom or digoxin8. In active immunization, Meth-like small molecules, linked to immunogenic carriers, were used to stimulate the production of Meth antibodies in hosts7,9,10. For passive immunization, hosts were infused intravenously with specific Meth antibodies derived from vaccinated animals or the synthetic antibody libraries11,12,13. These high-affinity Meth antibodies capture the Meth molecule in the circulatory system, reduce its access to the activation sites in the brain, and attenuate Meth-mediated behavioral changes6,11,14,15. The effectiveness of Meth antibodies is generally limited by several factors. For example, Meth is a weak immunogen; a regular and well-scheduled course of vaccination is required to maintain Meth antibody titer in the blood9,16. The antibodies induced by active immunization may cross-react with other molecules or endogenous ligands structurally similar to Meth. Furthermore, HIV or other immune-compromised conditions, commonly seen in LY2140023 drug addicts, may yield insufficient antibodies after active immunization17,18. These limitations may be partially overcome by passive immunization. However, repeated administration of purified Meth antibodies would generally be required to reduce Meth responses over an extended period of time, as would be necessary for chronic users. The high cost of passive immunization would also be a complicating factor likely to reduce compliance in Meth addicts. We hereby propose an alternative approach to producing Meth antibodies in the host through adeno-associated virus (AAV) infection. A recombinant AAV serotype-8 vector (AAV-MethAb) carrying the Meth monoclonal antibody gene was used to express a LY2140023 monoclonal antibody specific for Meth (Fig. 2b). Mice (n?=?10) were treated with AAV-MethAb (109 VGC/animal, i.p.). Sera were collected at 9 weeks post-injection. Four molecules (bupropion, L-DOPA, methadone, and Meth) were used to compete with (+)-Meth-HRP for the binding to serum-derived anti-Meth antibodies. The IC50 for Meth (~10?4?mM) was much lower (>10,000-fold) than that for bupropion (~1?mM), L-DOPA (~10?mM), and methadone (>1?mM), suggesting that the antibodies derived from the AAV-MethAb infection were specific to (+)-Meth expression of a functional anti-Meth monoclonal antibody following gene delivery through AAV vectors. This finding is consistent with previous reports that combination of 2A LY2140023 self-processing system and AAV-mediated gene transfer can achieve long-term stable expression of full-length antigen-specific antibodies in the periphery for the treatment of cancers and Alzheimers disease in rodent models23,24,25. The MethAb, generated by hybridoma cell lines, has high selectivity to Meth without cross-reactivity with a broad range of small molecules26. Similar high selectivity to Meth was found in the current study. We demonstrated that L-DOPA (structurally similar to Meth), bupropion, and methadone did not compete with Meth for the binding to MethAb in the sera collected from the animals receiving AAV-MethAb. These data suggest that AAV-MethAb infection produced a LY2140023 specific antibody for (+)-Meth DH5 by the anion-exchange-based endotoxin-free plasmid purification kit (Cat. No. 12362, Qiagen). At six h post-transfection, the culture medium was changed with fresh DMEM plus 2% FBS, and the ethnicities were additional incubated for 48?h. The tradition press had been centrifuged and gathered at 2,500??g in 4?C for 30?min to eliminate the cell.