In this n-of-1 trial, we observed that leptin was associated with an improved rate of development in various aspects of neurocognitive functioning. As evidence that leptin replacement was beneficial to the patient, we documented resolution of hypertension, dyslipidemia and hyperinsulinemia, in the context of decrease in food intake and weight loss. To the best of our knowledge, this is the first longitudinal study that shows that leptin has a cognitive enhancing role in the developing CNS of humans. To achieve that result, we employed the only possible human model: a leptin-deficient child under leptin replacement therapy. Treatment of leptin-sufficient children poses many ethical issues, and results are contaminated by the fact that these children are already under the effects of their endogenous leptin. Due to the leptin-naïve status of the patient, we were able to evaluate the effects of leptin on the developing CNS.
To compute rates of development at different time points, we inferred a steady rate from birth to 61 months (pre-leptin treatment) because we did not have two pretreatment baselines. The inferred rate of development was much slower from birth to 61 months (T1), suggesting developmental age below expectations for chronological age, than the rate of development observed during the treatment phase (T1 to T3) on most of the neurocognitive measures administered. By T3 (two years after initiation of leptin therapy), developmental relative chronological age ratios increased on all but one subtest compared to the baseline evaluation, and in many cases, approached or exceeded chronological age expectations. Because the patient had not experienced any environmental changes that would affect his neurocognitive abilities (i.e., starting school, receiving tutoring) between T1 and T2, we cautiously attribute the increased rate of development to leptin treatment and its subsequent effects on the CNS. However, the patient had completed the first year of school in his home country by the time of the third evaluation, which may partly have contributed to the apparent increase in rate of development between T2-T3. Nonetheless, these improvements are consistent with the finding that leptin replacement tends to normalize certain brain structures in leptin deficient mice. It would be instructive to compute correlations between rates of change in cognitive scores and rates of change in brain volume pre- and post-treatment to further support this assertion. Nonetheless, initial results suggest that in this leptin deficient patient, leptin may have had a cognitive enhancing role.
After the initiation of r-metHuLeptin, we observed a slower rate of development at T2 relative to T1 on a few subtests (“Recall of Objects,” “Memory for Faces,” and “Blocks”). However, scores at T2 were no more than 20% lower than scores predicted for age and in most cases, not clinically different. The few exceptions to the general pattern of increase in developmental rate may in part be within the error variance of the instrument. These normal variations in performance are difficult to characterize given that there is only subject in the study.
Leptin has a role in the central nervous system beyond its regulatory function through the hypothalamus on food intake and energy balance. Leptin is expressed in the brain, including the cerebellum, pyriform cortex, cerebral cortex, thalamus, hippocampus, amygdala, olfactory tract and substantia nigra 
. It may improve cognition by the selective enhancement of N-methyl D-aspartate (NMDA) receptors, facilitating long-term potentiation (LTP) 
. In low frequency stimulation, leptin inhibits hippocampal neurons, regulating hippocampal excitability 
. In adult mice, leptin increases hippocampal neurogenesis, both in vitro and in vivo 
. Hippocampal synaptic plasticity is also regulated by leptin. Animals with leptin receptor mutations (db/db
mice or fa/fa
rats) present deficits in hippocampal-specific memory tasks 
, impairments in hippocampal LTP, and long-term depression 
. Leptin is also an important hormonal signal in the developing CNS. In adult leptin deficient (ob/ob
) or leptin resistant (db/db
) mice, for example, leptin deficiency leads to lower brain weight and protein content 
. Many of these effects normalize after postnatal administration of leptin 
. In addition, brain myelin 
, neuronal soma size 
, and several synaptic proteins are reduced 
. Growth-associated proteins in the neocortex, striatum and hippocampus are elevated, and dendritic orientation is altered 
In humans, normal neonates have two to three times higher serum leptin levels, when compared to adults 
. Elevated circulating leptin levels are originated from the maternal and the neonatal adipose tissue, as well as from the placenta. Moreover, the increased adipose leptin mRNA expression in fetal adipose tissue, and the delayed onset of synthesis of leptin prior to the maturation of its receptor also explain the high levels of leptin 
. Several maternal, placental and neonatal factors are potentially associated with production and functions of leptin in early life, and have been reviewed by Alexe et al. 
. Leptin does not appear to regulate food intake and body weight during neonatal life 
, and the metabolically irrelevant surge of leptin may act as a developmental signal 
, which coincides with the development of major hypothalamic feeding circuits 
. This neurodevelopmental activity of leptin is limited to a neonatal window of maximum sensitivity, corresponding to a period of elevated leptin secretion 
. However, in adults, leptin can still affect brain plasticity by causing synaptic rearrangement of excitatory and inhibitory inputs on arcuate neurons 
, suggesting that these circuits remain relatively plastic throughout life.
In abnormal conditions, low levels of leptin at birth are seen in small for gestational age infants 
, but the neurodevelopmental consequences of this alteration have not been evaluated. The neurodevelopmental actions of leptin are not restricted to the hypothalamus: it has been demonstrated that cortical and hippocampal development can also be influenced by leptin 
. It is known that, besides its effects on the components of feeding circuits, leptin also acts as a cognitive enhancer in the hippocampus and on excitatory synaptic strength, which may regulate the processes involved in learning and memory 17
. These processes would be explained, at the cellular level, by the leptin-mediated modulation of hippocampal synaptic plasticity, leading to the conversion of hippocampal short-term potentiation (STP) into LTP, via enhancing NMDA receptor function 
. In the hippocampus, leptin also regulates neuronal excitability via its ability to activate the large conductance Ca2+
(BK) channels. Leptin can also evoke a novel form of NMDA receptor-dependent LTD in the CA1 region of the hippocampus, but this is only apparent under conditions of enhanced excitability 
. The signaling processes underlying these effects involve activation of a phosphatidylinositol 3-kinase-dependent process 
. Activation of mitogen-activated protein kinases and Src tyrosine kinases has also been implicated in this pathway.
Morphological studies in adult humans with genetic leptin deficiency suggest that leptin effect on the CNS extends beyond the hypothalamic nuclei. Six months of leptin replacement treatment for these three genetically leptin-deficient adults produced increased gray matter concentration in the anterior cingulate gyrus, inferior parietal lobule, and the cerebellum, findings that were maintained for 18 months following treatment 
The most marked effect of leptin replacement is weight loss, which is primarily achieved through reducing caloric intake. Leptin also leads to weight loss by increasing resting energy expenditure (REE) in animals 
, but not in humans 
. In this study, we did not evaluate the effects of leptin on REE, which prevents us from stating that weight loss was entirely attributed to the decrease in energy intake. Following weight loss, we observed a mild deceleration in growth. Height changed from the 50th
percentile to the 25th
percentile between the first and the second year under treatment. Growth will be monitored and adequate hormonal parameters will be assessed in the next follow-up visit. Subsequent to losing weight, hypertension, dyslipidemia and hyperinsulinemia were reversed. We believe these changes are attributable to weight loss, but we cannot exclude a direct effect of leptin on these parameters, independent of weight loss. In particular, it is known that leptin directly affects the pancreatic beta cells to decrease insulin secretion 
, but it is problematic to evaluate whether these changes are caused by leptin alone, by weight loss or both.
We acknowledge that our study has some limitations. First, the significance of our findings is hard to judge, due to the fact that we report the findings on only one patient. To our knowledge, eleven leptin-deficient children have been identified in the world thus far (including our patient). Neurocognitive evaluations of these children are warranted, preferably at early ages, and while leptin-naïve. Second, we did not have an age-, sex-, weight- and cultural background-matched control group. To compensate for this limitation, we gave careful consideration to our choice of measurement instruments so as to select tests and subtests that are less culturally biased and measure general skills. Further, we considered a strength of the study to the fact that we have longitudinal evaluation, which in essence allowed us to use the patient's baseline performance as his own control for subsequent evaluations. Although this is an n-of-1 trial, we chose not to conduct additional leptin-free evaluations (other than those performed at T1) because we believe that the absence of leptin would be harmful to the child's development.
In conclusion, treatment of a leptin-deficient child with r-metHuLeptin was beneficial, leading to improvements in neurocognition, substantial weight loss, decrease in caloric intake and normalization of the parameters of metabolic syndrome. Further studies need to confirm these neurocognitive findings additional in leptin-deficient patients across the life span. Additional findings may support a rationale for the development of further studies to test the hypothesis that the effects of leptin replacement may improve cognition in leptin-sufficient patients with delay or decline of cognitive function.