Cannabis and cannabinoid use in autism spectrum disorder: a systematic review

Abstract Introduction Autism spectrum disorder (ASD) is a neurodevelopmental disorder characterized by persistent deficits in social communication and social interaction, associated with the presence of restricted and repetitive patterns of behavior, interests, or activities. Cannabis has been used to alleviate symptoms associated with ASD. Method We carried out a systematic review of studies that investigated the clinical effects of cannabis and cannabinoid use on ASD, according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA checklist). The search was carried out in four databases: MEDLINE/PubMed, Scientific Electronic Library Online (SciELO), Scopus, and Web of Science. No limits were established for language during the selection process. Nine studies were selected and analyzed. Results Some studies showed that cannabis products reduced the number and/or intensity of different symptoms, including hyperactivity, attacks of self-mutilation and anger, sleep problems, anxiety, restlessness, psychomotor agitation, irritability, aggressiveness perseverance, and depression. Moreover, they found an improvement in cognition, sensory sensitivity, attention, social interaction, and language. The most common adverse effects were sleep disorders, restlessness, nervousness and change in appetite. Conclusion Cannabis and cannabinoids may have promising effects in the treatment of symptoms related to ASD, and can be used as a therapeutic alternative in the relief of those symptoms. However, randomized, blind, placebo-controlled clinical trials are necessary to clarify findings on the effects of cannabis and its cannabinoids in individuals with ASD. Systematic review registration International Prospective Register of Systematic Reviews (PROSPERO), code 164161.


Introduction
Conventional medical treatment includes several psychotropic drugs such as atypical antipsychotics, selective serotonin reuptake inhibitors, stimulants and anxiolytics; they do not treat ASD, but aim to eliminate inappropriate behavior, such as psychomotor agitation, aggressiveness, and obsessive-compulsive symptoms. [5][6][7][8] They may lead to severe side effects such as nephropathy, hepatopathy, and metabolic syndromes, among others. 9 Unfortunately, 40% of children with autism and disruptive behaviors do not respond well to standard medical and behavioral treatment. 4 This carries a high cost for the individual and society, causing life expectancy to be reduced by 20 years in patients with autism compared to the population average. 10 Among the possible pharmacological treatments, researchers began to explore other therapeutic alternatives, such as the use of substances derived from Cannabis sativa. 11 Cannabidiol (CBD) represents one of the major components of the plant, having been studied in several disorders. At present, preliminary evidence suggests that CBD can relieve spasticity, 12 pain, sleep disorders, 13 improve mobility in multiple sclerosis, 14 in addition to relieving anxious symptoms and social phobia 15 ; however, further studies are needed to prove its effectiveness.
In autism, cannabis and cannabinoids have also been used to treat symptomatic conditions. 16,17 CBD, and some other compounds in the plant, interact with the endocannabinoid system and can modulate different aspects related to cognition, socioemotional responses, susceptibility to seizures, nociception and neuronal plasticity, which are often altered in autism. [18][19][20][21] In mammals, the endocannabinoid system is mainly composed of two receptors, CB1 and CB2, endocannabinoids (endogenous substances that activate CB1 and CB2 receptors) and the enzymes responsible for their synthesis and metabolism. 22 CB1 receptors are expressed in both the central and peripheral nervous systems, with their most abundant expression in basal ganglia nuclei and pre-synaptic GABAergic and glutamatergic neurons. 23 Considering that the endocannabinoid system modulates emotional responses, mood, behavioral reactions to the context and social interaction, investigators have started to formulate the hypothesis that changes in this system would be present in the autistic phenotype. 24 Aran et al. 25 26  to autism (autistic, autism, Asperger, and pervasive development disorder). During the selection process, no restrictions were applied in terms of language, e,g., any article found was included in the eligibility analysis.
Descriptors were included in quotation marks, and the search operators "AND" and "OR" were used. Cannabisrelated terms were grouped using the "OR" operator; terms related to autism were grouped similarly. Then, these two groups of related terms were added and joined by the "AND" operator ( Figure 1).
We included all articles published until October 2020, in any language, in the form of clinical trials or case studies involving human beings. Articles unrelated to the topic, i.e., those reporting on illicit or recreational use of cannabis, as well as abstracts, book chapters, animal studies, and research on other pathologies or changes that were associated with signs and symptoms similar to those observed in autism, were rejected.
The articles found in the databases were initially screened by reading their titles and abstracts.
Subsequently, those articles considered to meet the proposed topic were read in full. At the end of the screening phase, we browsed the references of the articles ultimately selected in search of other studies that met the eligibility criteria.
The search and screening of the selected articles were carried out simultaneously and independently by two authors. In the end, the disagreements found were sent to another author, to make the final decision about whether or not to include a certain study, but always checking the eligibility criteria.
The searches conducted in the MEDLINE/PubMed, SciELO, Scopus, and Web of Science databases yielded 64, 1, 242, and 125 articles, respectively. Of these, respectively, 58, 1, 237, and 121 articles were eliminated because they did not meet the inclusion criteria. Thus, 14 studies were found, which, after eliminating duplicates, resulted in six articles. From the browsing of references of these six studies, another paper was selected to be part of the review, making a total of seven selected articles. Finally, the search carried out on Google Scholar yielded two more studies, reaching a final total of nine articles included in this systematic review, in accordance with the inclusion and exclusion criteria adopted ( Figure 1).
The data extraction method of each study consisted in filling a standardized information sheet. One reviewer extracted the scientific data, and a second reviewer verified the acquired information. Disagreements were resolved by discussion and consensus among the authors-reviewers.  Five studies used cannabis extract, in the presentation of CBD-rich oil, 16,17,28,30,31 two studies used CBD in oral solution, 26,28 one study used dronabinol, which is a synthetic analogue of THC (tetrahydrocannabinol), dissolved in sesame oil, 27 and one study used cannabidivarin (CBDV) 32 (Table 1).
The studies using CBD-enriched cannabis oil showed a variation between the proportions of CBD and THC, ranging from 6 to 75% CBD combined with 1 to 1.5% THC. Those who used pure CBD used a dose of 600 mg (oral solution), dronabinol was used at a dose ranging between of 0.62 and 3.62 mg/day (dissolved in sesame oil), and cannabidivarin was used at a dose of 600 mg.
The samples were composed of: 1) children in three studies 16,17,27 ; 2) children and adolescents in one study, with ages ranging from 5 to 19 years 29 ; and 3) adults in three studies. 26,28,32 Two studies did not specify the age group. 30,31 Only three studies used any imaging exam, namely, magnetic resonance spectroscopic imaging after the intervention with CBD in two studies and with CBDV in one study to search for brain changes. 26,28,32 The other studies used questionnaires, forms and subjective reports of family members or caregivers. Of the nine Administration of oil with CBD and THC (20:1), orally, with telephone interviews conducted every two weeks with parents or caregivers, asking about changes in symptoms, the data obtained were analyzed independently by specialists in search of these changes in symptoms and safety of medicines. The improvement resulting from CBD was also compared with conventional treatment for ASD. Patients were allocated in a randomized order: about half in each group participated in the placebo before CBD (600 mg oral solution) and the other half participated in CBD before the placebo. After administration, placebo or CBD, a check was scheduled to coincide with the maximum plasma concentration (2 h). It was evaluated by magnetic resonance spectroscopic imaging.
It was seen that patients with ASD had a drop in IQ compared to neurotypical controls (F 1 = 5,781; p = 0.022), but the difference in IQ did not influence the results: ASD (r < -0.008; p > 0.698); neurotypical (r < 0.068; p > 0.235). The excitatory mechanisms of response to glutamate were comparable, regardless of diagnosis, however the inhibitory response by GABA + was altered in ASD. There was a difference in the results found in the images in relation to the placebo group.
CBD can change the levels of glutamate, glutamine and GABA +, regulators of excitatory and inhibitory neurotransmission. The autistic brain reacts differently to GABA+, which helps to understand the mechanisms and targets of treatment for ASD.  Randomized, doubleblind, crossover study using magnetic resonance spectroscopic imaging comparing glutamate and GABA levels after the use of placebo and 600 mg CBDV. Information was collected from the dorsomedial region of the prefrontal cortex and the left basal ganglia (areas related to ASD) after 2 h (plasma peak of the substance) of administration.
Tests performed at least 13 days after using the drug/placebo indicated that CBDV increased the levels of glutamate in the left basal ganglia in both groups, but in those with ASD despite this increase, the basal concentration of the substance decreased. CBDV did not alter the levels of glutamate or GABA in the medial dorsal region of the prefrontal cortex of either group. There was a difference in the results found in the images in relation to the placebo group.   Endocannabinoids are known to regulate the main brain functions that are altered in ASDs. 41 A well validated animal model of ASD based on prenatal exposure to valproic acid in rats has been used to evaluate behavioral alterations. 42,43 There is strong evidence suggesting that altered levels of AEA, which already manifest in childhood and persist in adolescence and adulthood, may be associated with autistic symptoms, thus providing preclinical justification for a potential role of AEA signaling as a new therapeutic target for ASD. These results have corroborated a series of preclinical data that suggest that AEA signaling seems to play a modulating role on rodent behaviors associated with symptoms of ASD. 21 A pioneering clinical study was able to identify low levels of AEA in plasma from children with ASD compared to plasma from children without ASD. These preliminary results corroborate the preclinical evidence that signs of AEA may be impaired in patients with ASD. 44 A study conducted in 2019 by Aran et al. 25 showed strong evidence that serum levels of certain

fMRI pattern after using CBD
A 600 mg CBD oral solution was used in individuals with ASD who underwent fMRI to assess the effects of this treatment on their central nervous system. 26,28,32 All those studies were carried out by the same team of researchers; 17 neurotypical adults and 17 adults with autism were administered a 600 mg CBD oral solution at one occasion, and a placebo substance at another occasion (randomized order); patients were then examined using fMRI. CBDV increased the levels of glutamate in the left basal ganglia, assessed with spectroscopy; however, in patients with ASD, despite the increase, the basal concentration of the substance decreased. 32 It was noticed that CBD and CBDV altered the GABAergic system in all participants. The excitatory mechanisms of response to glutamate did not differ between the two groups, however the inhibitory response mediated by GABA was different in people with ASD, indicating that the brain of an autistic individual has a distinct GABAergic system from that of neurotypical individuals. In other words, the autistic brain reacts differently to GABA, and this discovery may help understand the mechanisms and targets of treatment in autism. Pretzsch et al. 28  In those authors' study, the symptoms of drowsiness, irritability, diarrhea, increased appetite, conjunctival hyperemia, and increased body temperature were seen in some cases and considered mild and/or transient.
Few participants had to interrupt treatment before the end of the first month, due to adverse effects such as insomnia, irritability, rapid heartbeat, and worsening of the psychobehavioral crisis. The patients who had relevant side effects were all taking several medications, including at least one antipsychotic. A possible bias could be that the presence of epilepsy (38.9% of participants) may have interfered with the outcome, as studies that report improvement in epilepsy often also describe ASD-related symptoms.
It is important to highlight that all the randomized double-blind studies found on the use of cannabis and cannabinoids for autism assessed brain structures through magnetic resonance imaging, but did not have a focus on the efficacy and safety of cannabis for ASD.
All evaluations were observational, either in individuals who started the medication in the study and were observed prospectively, or in those who had already used the substance and were analyzed retrospectively.
As general limitations of the studies included in this systematic review, it possible to cite the absence of follow-up evaluations and the lack of laboratory tests to help confirm the safety of the substances used. Also, only six studies evaluated the patients clinically; the others were based on image examination only. Samples were small, and several participants were lost along the study period. Finally, endocannabinoids were not dosed.

Conclusion
Cannabis and cannabinoids have very promising effects in the treatment of autistic symptoms and can be used in the future as an important therapeutic alternative to relieve those symptoms, especially bouts of self-mutilation and anger, hyperactivity, sleep problems, anxiety, restlessness, psychomotor agitation, irritability, and aggressiveness; as well as improve sensory sensitivity, cognition, attention, social interaction, language, perseverance, and depression.
In addition, it is important to note that CBD can also change the levels of glutamate, glutamine and GABA, substances that contribute to the regulation of excitatory and inhibitory neurotransmission in both neurotypical and autistic individuals. However, randomized, doubleblind and placebo-controlled clinical trials, as well as longitudinal studies, are necessary to clarify the findings on the effects of cannabis and its cannabinoids in individuals with autism.
Cannabis has been prescribed on an individual basis only, with autism being the second largest disease with available use, surpassed only by epilepsy. Therefore, it is essential to analyze what we have so far in the scientific literature, as cannabis is already being used worldwide as a phytopharmaceutical or as a CBD-rich cannabis extract for the autism spectrum.

Disclosure
No conflicts of interest declared concerning the publication of this article.