The SLC6 family is a diverse set of transporters that mediate solute translocation across cell plasma membranes by coupling solute transport to the cotransport of sodium and chloride down their electrochemical gradients. These transporters probably have 12 transmembrane domains, with cytoplasmic N- and C-terminal tails, and at least some may function as homo-oligomers. Family members include the transporters for the inhibitory neurotransmitters GABA and glycine, the aminergic transmitters norepinephrine, serotonin, and dopamine, the osmolytes betaine and taurine, the amino acid proline, and the metabolic compound creatine. In addition, this family includes a system B(0+) cationic and neutral amino acid transporter, and two transporters for which the solutes are unknown. In general, SLC6 transporters act to regulate the level of extracellular solute concentrations. In the central and the peripheral nervous system, these transporters can regulate signaling among neurons, are the sites of action of various drugs of abuse, and naturally occurring mutations in several of these proteins are associated with a variety of neurological disorders. For example, transgenic animals lacking specific aminergic transporters show profoundly disturbed behavioral phenotypes and probably represent excellent systems for investigating psychiatric disease. SLC6 transporters are also found in many non-neural tissues, including kidney, intestine, and testis, consistent with their diverse physiological roles. Transporters in this family represent attractive therapeutic targets because they are subject to multiple forms of regulation by many different signaling cascades, and because a number of pharmacological agents have been identified that act specifically on these proteins.

The present study investigated the effect of renal impairment and hemodialysis on the pharmacokinetics of lenalidomide following a single 25-mg oral dose in 30 subjects aged 39 to 76 years. A single 25-mg dose was well tolerated by renally impaired subjects. Renal impairment did not alter the oral absorption, protein binding, or nonrenal elimination of lenalidomide. Mean urinary recovery of unchanged lenalidomide was 84% of the dose in subjects with normal renal function (creatinine clearance [CL(Cr)] > 80 mL/min), and it declined to 69%, 38%, and 43% in subjects with mild (50 < or = CL(Cr) < or = 80 mL/min), moderate (30 < or = CL(Cr) < 50 mL/min), and severe (CL(Cr) < 30 mL/min) renal impairment, respectively. The differences in pharmacokinetic parameters between normal renal function and mild renal impairment were minor to modest (11%-32%). As renal impairment progressed to moderate, severe, or end-stage renal disease, total and renal lenalidomide clearance decreased drastically, area under the concentration-time curve increased by approximately 185% to 420%, and t((1/2)) was prolonged by approximately 6 to 12 hours. A 4-hour hemodialysis removed 31% of lenalidomide in the body. Therefore, lenalidomide dose adjustments should be considered for patients with CL(Cr) < 50 mL/min, and the recommendations are given for the starting doses.

The present study addressed the role of N-linked glycosylation of the human dopamine transporter (DAT) in its function with the help of mutants, in which canonical N-glycosylation sites have been removed (N181Q, N181Q,N188Q, and N181Q,N188Q,N205Q), expressed in human embryonic kidney-293 cells. Removal of canonical sites produced lower molecular weight species as did enzymatic deglycosylation or blockade of glycosylation, and all three canonical sites were found to carry sugars. Prevention of N-glycosylation reduced both surface and intracellular DAT. Although partially or non-glycosylated DAT was somewhat less represented at the surface, no evidence was found for preferential exclusion of such material from the plasma membrane, indicating that glycosylation is not essential for DAT expression. Non-glycosylated DAT was less stable at the surface as revealed by apparently enhanced endocytosis, consonant with weaker DAT immunofluorescence at the cell surface and stronger presence in cytosol in confocal analysis of the double and triple mutant. Non-glycosylated DAT did not transport dopamine as efficiently as wild-type DAT as judged from the sharp reduction in uptake V max , and prevention of N-glycosylation enhanced the potency of cocaine-like drugs in inhibiting dopamine uptake into intact cells without changing their affinity for DAT when measured in membrane preparations prepared from these cells. Thus, non-glycosylated DAT at the cell surface displays appreciably reduced catalytic activity and altered inhibitor sensitivity compared with wild type.Biogenic amine carriers include the dopamine transporter (DAT), 1 the norepinephrine transporter, and the serotonin transporter, all part of the larger family of Na ϩ ,Cl Ϫ -dependent neurotransmitter transporters (1), which also includes the nonbiogenic amine subfamily of ␥-aminobutyric acid-related transporters (2). In the brain, the DAT is responsible for the clearance of extraneuronal dopamine thought to be the crucial process for terminating dopamine action (3). The DAT is a heavily glycosylated protein with sugars attached to asparagine (4 -6). In general, carbohydrate units of glycoproteins can play several roles, such as controlling protein folding, stabilizing protein conformation, protecting against proteolysis, and regulating intracellular and surface trafficking (7). Early on, for the DAT, glycosylation has been proposed to alter uptake function (6); thus, the generally higher K m values for dopamine uptake in cell systems (micromolar) as compared with striatal synaptosomes (submicromolar) is speculated to be due to environment-specific differences in glycosylation, both in extent and type of sugars.So far, only limited information is available to answer the question as to whether or how glycosylation affects DAT function. In a preliminary report, we describe the generation of three N-glycosylation mutants in which the three putative N-linked glycosylation sites in human DAT are removed: the single mutant N181Q, the double mutant N181Q,N188Q, and the tripl...

The human dopamine (DA) transporter (hDAT) contains multiple tryptophans and acidic residues that are completely or highly conserved among Na 1 /Cl 2 -dependent transporters. We have explored the roles of these residues using nonconservative substitution. Four of 17 mutants (E117Q, W132L, W177L and W184L) lacked plasma membrane immunostaining and were not functional. Both DA uptake and cocaine analog (i.e. 2b-carbomethoxy-3b-(4-¯uorophenyl)tropane, CFT) binding were abolished in W63L and severely damaged in W311L. Four of ®ve aspartate mutations (D68N, D313N, D345N and D436N) shifted the relative selectivity of the hDAT for cocaine analogs and DA by 10±24-fold. In particular, mutation of D345 in the third intracellular loop still allowed considerable The data taken together indicate that mutation of conserved tryptophans or acidic residues in the hDAT greatly impacts ligand recognition and substrate transport. Additionally, binding of cocaine to the transporter may not be the only way by which cocaine analogs inhibit DA uptake.