Below is the uncorrected machine-read text of this chapter, intended to provide our own search engines and external engines with highly rich, chapter-representative searchable text of each book. Because it is UNCORRECTED material, please consider the following text as a useful but insufficient proxy for the authoritative book pages.
4 Translating Research from Nonhuman Primates to Humans Highlights â¢ Translating discoveries from nonhuman primates to humans may be possible because of the similar size and complexity of their brains (Smith). â¢ Gene-edited monkeys provide an approach to understanding the different phenotypes of autism (Amaral). â¢ Sophisticated nonhuman primate models of diseases such as au- tism will be useful to identify therapeutic targets and biomarkers, and possibly develop new treatments (Parker). â¢ Behavioral assays in monkeys that replicate the kinds of assays used in humans have been developed to demonstrate that the mon- key model accurately reflects features of human autism (Parker, Poo, Roberts). â¢ Tools that demonstrate similar cognitive processes and neurobio- logical correlates in humans and nonhuman primates enable both forward and back translation of discoveries (Roberts). â¢ Marmosets are useful for longitudinal imaging studies of brain de- velopment because they reach maturity in only 2 years (Roberts). â¢ The marmoset brain is ideal for mapping brain circuits involved in cognition and other behaviors because although small, it is struc- turally similar to humans (Okano). â¢ Calcium imaging in deep layers of the marmoset cortex is possible in freely moving animals, enabling the monitoring and decoding of complex behaviors (Okano). NOTE: These points were made by the individual speakers identified above; they are not intended to reflect a consensus among workshop participants 27 PREPUBLICATION COPY: UNCORRECTED PROOFS
28 TRANSGENIC NEUROSCIENCE RESEARCH The hope embedded in the nonhuman primate research taking place around the world is that discoveries made in these animals will translate to humans in ways that research in other animal models has not. Yoland Smith said that making the leap from nonhuman primates to humans will be neither trivial nor easy, but the size and complexity of the primate brain will make it easier to target a sufficient number of neurons to facilitate the diffusion of vectors or achieve other forms of genetic modification. A TRANSLATIONAL RESEARCH ROADMAP IN AUTISM David G. Amaral, distinguished professor of psychiatry and neurosci- ence and Beneto Foundation Chair of the MIND [Medical Investigations of Neurodevelopmental Disorders] Institute at the University of Califor- nia, Davis, has been studying nonhuman primates for many years, and in the past two decades has also been studying human children with autism. He believes that gene-edited monkeys offer a way to understand the differ- ent phenotypes of autism and parse this heterogeneous disorder into mean- ingful subtypes that have clinical and possibly therapeutic implications. For example, Amaral was involved in a study published in 2014 that investigated the role of CHD8 [chromodomain helicase DNA binding pro- tein 8] mutations in children with autism (Bernier et al., 2014). What they found, according to Amaral, was that of the 15 children in the study that had loss-of-function truncations in the CHD8 gene, 13 had autism. Most of these children also had macrocephaly, or large brains. Amaral said that about 15 percent of boys with autism have large brains, and that this sub- group also has more severe symptoms and a worse prognosis (Amaral et al., 2017). To investigate the neurobiology of this large-brain form of au- tism, an animal model was needed, but according to Amaral, disrupting CHD8 in rodents produces macroencephaly, but not the behavioral char- acteristics typical of autism. He suggested that manipulating the gene in nonhuman primates could provide not only a model for understanding the neurobiology of this form of autism, but also for testing new therapeutic interventions. Karen Parker described what she called a translational research roadmap for autism that begins by developing sophisticated animal models to identify biomarkers and targets; translates those monkey findings to au- tism patients; and then uses that model to test novel treatments. PREPUBLICATION COPY: UNCORRECTED PROOFS
TRANSLATING RESEARCH FROM NONHUMAN PRIMATES TO HUMANS 29 Earlier work in twins had demonstrated that autistic traits and the pol- ygenic burden or risk underlying these traits is distributed across popula- tions (Constantino and Todd, 2003), suggesting it would be valuable to model autism polygenically in monkeys, said Parker. She started with an extremes group approach in a population of 5,000 rhesus monkeys at the California National Primate Research Center. These animals are bred to preserve genetic heterogeneity and housed in ecologically valid environ- ments (outdoors in mixed male and female groups with access to play gyms and swinging perches). Parker said that like humans, these animals exhibit stable, individual differences in social behavior. At the extremes are individuals who can be identified as âlow socialâ or âhigh social.â Low-social animals, like humans with autism, initiate and receive fewer affiliative interactions, spend less time grooming and playing, display more inappropriate social behaviors, and show diminished interest and less social competence than their typical peers (Sclafani et al., 2016). Parker asked a group of clinicians to identify the kinds of behavioral assays that would provide face validity for her monkey model, that is, that would convince them that the monkeys were accurately modeling human autism. She and her colleagues then used this information to develop a monkey social behavior test battery that would allow them to interrogate the core symptoms seen in patients. Criteria for these tests included eco- logical relevance, prior successful use in macaques or related macaque species, the ability to generate variation in performance, and no need for extensive training prior to test administration. For example, Parker said there is evidence that children with autism have face recognition but not object recognition problems, so her team created a monkey version to test these two domains. Other tests included in the battery test social competence, joint attention, ability to read social cues, social motivation, and intellectual disabilities, all using species- specific behaviors, such as lip smacking and vocalizations. To validate this model from a biological perspective, Parker and col- leagues turned to biomarkers. Because reproducibility problems have plagued much of the work done in humans on blood biomarkers, Parker thought it might be better to look at cerebrospinal fluid (CSF), particularly at some of the signaling pathways that have been implicated in regulating prosocial behavior in mammals and those dysregulated in syndromic forms of autism. They discovered that CSF vasopressin levels could, with high accuracy, predict whether an animal was high or low social (Parker et al., 2018). To see if this is replicated in human children, they piggy- PREPUBLICATION COPY: UNCORRECTED PROOFS
30 TRANSGENIC NEUROSCIENCE RESEARCH backed on clinical indications where children were getting lumbar punc- ture for standard of care reasons. In their first study, they found that 13 of 14 children could be classified with high accuracy based on their CSF vas- opressin level alone. They have since replicated the study and shown that CSF vasopressin levels correlate with social deficits (Oztan et al., 2018). Based on these results, Parker and colleagues have also conducted a small clinical trial of vasopressin as a possible treatment for autism. ASSESSMENT TOOLS FOR NONHUMAN PRIMATES In addition to developing these new disease models, tools will be needed to assess behavior and cognition in nonhuman primates in a man- ner that translates to human conditions, said Angela Roberts. For example, more than 20 years ago, Roberts and colleagues developed a battery of tests for use in nonhuman primates to assess various aspects of cognitive function, including working memory, visual recognition memory, visual spatial processing, planning, and behavioral flexibility. Their lab also demonstrated that damage to two distinct regions of the prefrontal cortex affected different aspects of behavioral flexibility. They developed a test to assess these two forms of behavioral flexibility in mon- keys and were able to demonstrate that damage to the orbito-frontal cortex impaired a monkeyâs ability, having learned that one of two stimuli was associated with reward, to switch responding to the other stimulus as the association between stimuli and rewards changed (reversal learning). On the other hand, damage to the ventrolateral prefrontal cortex disrupted higher-order attentional set shifting, the ability to switch attention from one aspect of a stimulus (e.g., shape) to another (e.g., color) (Dias et al., 1996). Roberts said this was subsequently forward translated into humans preclinically by showing a similar dissociation using functional imaging, and clinically by showing that a deficit in shifting attention may be a vul- nerability marker for obsessive-compulsive disorder (Chamberlain et al., 2006) and possibly an early marker of Huntingtonâs disease. Roberts and colleagues have also developed a test to measure threat- driven behaviors in the marmoset that relate to fear and anxiety. They used this test to fractionate out the different forms of anxiety produced by le- sions to different areas of the prefrontal cortex (Agustin-Pavon et al., 2012; Shiba et al., 2017). They showed that animals with lesions in the ven- trolateral prefrontal cortex are anxious because they have problems shift- ing attention, while those with lesions in the anterior orbitofrontal cortex PREPUBLICATION COPY: UNCORRECTED PROOFS
TRANSLATING RESEARCH FROM NONHUMAN PRIMATES TO HUMANS 31 are anxious because they have problems tracking punishment and reward in their environment. The human analog, said Roberts, would be two pa- tients with a similar anxiety phenotype but different underlying causes, who thus should receive different treatment. Another human characteristic that demonstrates high cognitive func- tion is self-recognition. Mu-Ming Poo and colleagues have developed a test of mirror self-recognition in macaques that replicates the self- recognition test used as an indicator of self-consciousness and body awareness in humans and apes. Because some people with psychiatric con- ditions or autism demonstrate impairments in this test, Poo reasoned that if macaques could be trained to perform mirror self-recognition, their brains could be examined with brain imaging to study the neural processes underlying this ability. Although macaques do not spontaneously demon- strate mirror self-recognition, Poo and colleagues were able to train them to recognize face marks through the use visual-somatosensory training where face marks seen in the mirror were simultaneously associated with laser- light stimulation-induced sensation of the corresponding area of the face (Chang et al., 2015). Subsequently, they also trained the monkeys to learn precise visual-proprioceptive association for mirror images, and that this ability could be generalized to novel situations where the monkeys demon- strated self-directed behaviors after seeing mirror images of themselves (Chang et al., 2017). BRAIN MAPPING AND IMAGING IN GENETICALLY MODIFIED NONHUMAN PRIMATES Marmosets are an ideal model for longitudinal imaging studies of brain development because they are fully developed by 2 years of age, said Roberts. She and her colleagues imaged a group of marmosets every 3 months between the ages of 3 months and 2 years to measure growth tra- jectories within 53 cortical regions. Thus, they were able to document sub- stantial variability in the prefrontal cortex during development, which may relate to different symptoms that emerge during childhood and adoles- cence, she said (Sawiak et al., 2018). She suggested that transgenic models may help to show how stress interacts temporally with the genome in order to affect prefrontal circuits. The marmoset brain is also ideal for mapping brain circuits of cogni- tive behaviors because it is small but with primate structures, said Hideyuki PREPUBLICATION COPY: UNCORRECTED PROOFS
32 TRANSGENIC NEUROSCIENCE RESEARCH Okano (see Figure 4-1). He and his colleagues recently performed FIGURE 4-1 Functional brain imaging. Technologies such as electrocorticogra- phy, local field potential, intracellular recording, and calcium imaging have made it possible to functionally map deep-brain microcircuits in both humans and nonhuman primates. SOURCE: Presented by Hideyuki Okano, October 4, 2018. microscope mounted on the skull to enable deep imaging of freely behav- ing animals (Ghosh et al., 2011) combined with adeno-associated virus (AAV)-delivered GCaMP (Park et al., 2016) to measure calcium influx into cells as a marker of neuronal excitation (Broussard et al., 2014). GCaMP is a molecule created by fusing a calcium-binding protein with green fluorescent protein (Akerboom et al., 2012). This approach enabled them to monitor and decode the activity of 100 to 1,000 neurons in the motor cortex during a complex behavior such as pellet reaching (Kondo et al., 2018). Okano said this technology using free-moving marmosets will make it possible to dissect large-scale neural circuits during human- PREPUBLICATION COPY: UNCORRECTED PROOFS
TRANSLATING RESEARCH FROM NONHUMAN PRIMATES TO HUMANS 33 relevant behaviors under natural conditions, enabling the study of com- plex behaviors such as social interactions, fear, anxiety, and complex mo- tor tasks. In combination with transgenic technologies, this has the potential to transform our understanding, diagnosis, and treatment of hu- man diseases, he said. Okano is now developing techniques to monitor the activity of neu- rons in control and disease model marmosets. He predicted that the micro- endoscope will also make possible other imaging advances, including imaging of deeper brain areas such as the basal ganglia, pathway-specific and cell-type-specific imaging using Cre lines and in vivo genome editing, and imaging multiple sites simultaneously. It may also be used in combi- nation with optogenetics, Okano added. PREPUBLICATION COPY: UNCORRECTED PROOFS