1. Ukraine Clinical Project CIFASD Meeting
Christina Chambers, PhD, MPH
Austin Texas
March 8-9, 2016

2. AIMS
Determine if beneficial effects of a prenatal MVM supplementation intervention in alcohol exposed children persist at preschool age
Determine if prenatal alcohol exposure in the context of the prenatal nutritional nutritional environment affects child nutritional status and determine the extent to which nutritional status in children prenatally exposed to alcohol affects growth and performance
Determine if a miRNA biomarker for alcohol exposure previously identified in a sheep model translates to the human as a marker of recent or distant exposure to various levels of alcohol
In collaboration with CIFASD investigators, explore selected genetic and epigenetic risk factors for FASD, the effect of nutrition on 3D facial images, and telemedicine approaches to early diagnosis

3. Aim 1

4. Progress toward completion of Aim
Preliminary analyses will be presented by Claire Coles
This aim was powered to require the n of 240
The remaining pool of eligible subjects is adequate to achieve the sample size
We will complete sample recruitment within the next four months

5. Relevant publications

6. MDI at 6 months by alcohol group and micronutrient supplement
Alcohol dose p <0.001
Coles et al, Matern Child Health J, 19(12), , 2015

7. Information Processing COR
Choline supplementation resulted in greater change in heart rate in visual habituation trials in both groups (p<0.001)
Choline supplementation resulted in greater change in heart rate in visual dishabituation trials in the low/no alcohol-exposed group (p<0.013)
Latency of response was reduced in both groups and both conditions with choline supplementation
Kable et al, Alcohol 49 (7):647-56, 2015

8. Information Processing COR
Change in maternal choline blood level and maternal dimethylglycine blood level from enrollment to third trimester was positively correlated with change in heart rate, and negatively correlated with latency in visual habituation trials
There was no evidence of an effect of multivitamin mineral supplementation in any analysis, or any effect of supplementation or alcohol on responses to auditory stimuli
Kable et al, Alcohol 49 (7):647-56, 2015

9. 2 additional manuscripts related to neurobehavioral outcomes under review
Kable et al, Cardiac Orienting Responses Differentiate the Impact of Prenatal Alcohol Exposure in Ukrainian Toddlers – submitted to ACER
Bandoli et al, Assessing the independent and joint effects of unmedicated prenatal depressive symptoms and alcohol consumption in pregnancy and infant neurodevelopmental outcomes – submitted to ACER

10. Maternal depression and alcohol
Depressive symptoms reported in pregnancy (BDI in third trimester) were not independent predictors of Bayley score in the overall sample (n=344), but interacted with alcohol exposure
Larger deficits in Bayley scores among those with prenatal alcohol exposure
Joint effects in female infants were stronger than in males

11. Aim 2
Collection of venous blood samples or, alternatively, blood spots continues
A total of 103 child blood samples have been collected in recent months and shipped to Davis, of varying sample quantities

12. Aim 2
Initial analysis of 46 samples from preschoolers conducted by a graduate student at Davis -
Alcohol exposed with FASD n = 7
Alcohol exposed without FASD n = 8
Low or unexposed n = 31
No differences between groups on transferrin receptor, C- reactive protein, homocysteine, or cortisol levels
Group 1 FASD had significantly lower plasma ferritin levels compared to Group 3 but were similar to Group 2
Student submitted abstract to Teratology Society meeting 2016

13. Aim 2 Additional work on nutrition and growth focused on:
Does supplemental vitamin use in pregnancy, accounting for other factors, relate to postnatal growth measures at 2 and 4 years of age, and does this vary by alcohol exposure status?
Does prenatal micronutrient status, accounting for other factors, relate to postnatal growth measures at 2 and 4 years of age, and does this vary by alcohol exposure status?
Does child’s eating style at 2 or 4 years of age relate to growth trajectory over the first 4-5 years of life, and does this vary by alcohol exposure status?

14. 2 year old growth by prenatal vitamin status
Alcohol Exposed
N = 68
Alcohol Unexposed
N = 64
Weight
Kg (SE)
Height
Cm (SE)
OFC
No multivitamins at all
12.82
(0.31)
93.76
(1.41)
48.03
(0.45)
13.07
(0.28)
94.64
(1.18)
48.68
(0.27)
Multivitamins at enrollment OR assigned to MVM by study
12.86
(0.37)
91.63
48.44
(0.46)
13.30
(0.25)
91.28
(1.09)
48.72
(0.26)
Both multivitamins at enrollment and assigned to MVM by study
13.53
(0.67)
90.47
(2.91)
47.45
(0.92)
12.94
(0.42)
92.26
(1.94)
48.70
(0.47)
Marginal means adjusted for birth size and child’s age at the postnatal growth measurement

15. 4 year old growth by prenatal vitamin status
Alcohol Exposed
N = 58
Alcohol Unexposed
N = 97
Weight
Kg (SE)
Height
Cm (SE)
OFC
No multivitamins at all
16.01
(0.62)
109.33
(2.08)
50.07
(0.60)
15.37
(0.48)
100.51
(1.30)
50.03
(0.40)
Multivitamins at enrollment OR assigned to MVM by study
16.33
(0.34)
105.80
(1.15)
49.89
(0.33)
16.50
(0.30)
104.42
(0.80)
50.04
(0.25)
Both multivitamins at enrollment and assigned to MVM by study
16.47
(0.63)
102.32
49.54
15.99
(0.43)
102.44
(1.18)
50.17
(0.36)
Marginal means adjusted for birth size and child’s age at the postnatal growth measurement

16. 2 year old growth by maternal micronutrient status at enrollment
Growth Outcome
Overall
N = 43
Alcohol Exposed
N = 18
Weight
Height
OFC Cu + Zn
Folate
B12
Homocysteine
Choline
Betaine
DMG Fe

17. 4 year old growth by maternal micronutrient status at enrollment
Growth Outcome
Overall
N = 127
Alcohol Exposed
N = 51
Weight
Height
OFC Cu - Zn
Folate
B12 +
Homocysteine
Choline
Betaine
DMG Fe

18. Overweight/obesity in children with FASD?
Prevalence of overweight or obesity in 445 children with an FASD diagnosis and 171 without compared to national and state prevalence
Increased rates of overweight or obesity seen in children with partial FAS diagnosis, and with any FASD diagnosis in adolescents, particularly among females
Fugelstad et al, Alc Clin Exp Res, 2014

19. Obesity and alcohol: common links to addictive behaviors, cognitive dysfunction
Volkaw N et al, Overlapping neuronal circuits in addiction and obesity: evidence of systems pathology, 2008
Volkow N et al, Food and drug reward: overlapping circuits in human obesity and addiction, 2011
Volkow N et al, NOW vs LATER brain circuits: implications for obesity and addiction, 2015
Kanoski SE et al, Western diet consumption and cognitive impairment links to hippocampal dysfunction and obesity, 2011

21. CEBQ and association with body mass index in children
Food Approach constructs - all associated with higher BMI-Z in 6-7 year old children
Food responsiveness
Emotional overeating
Enjoyment of food
Food Avoidance constructs – all associated with lower BMI-Z
Satiety responsiveness
Slowness in eating
Emotional undereating
Food fussiness
Sleddens et al, Child Eating Behavior Questionnaire: factorial validity and associationwith BMI in Dutch children Age 6-7 Intl Jl Behavior Nutrition and Physical Activity 2008

22. 2 year old children CEBQ Unexposed N = 81 Exposed N = 47 P-value
Food Resp
2.1 (07)
2.1 (0.8)
0.697
Enjoy Food
3.4 (9.7)
3.1 (0.9)
0.065
Satiety
3.0 (0.6)
2.9 (0.7)
0.688
Slow Eating
2.4 (0.7)
2.7 (1.0)
0.190
Emotional Over Eating
2.6 (1.0)
2.4 (1.0)
0.292
Fussy Eating
2.8 (0.8)
2.7 (0.8)
0.785

23. 4 year old children CEBQ Unexposed N = 50 Exposed N = 33 P-value
Food Resp
2.0 (0.6)
2.3 (0.5)
0.002
Enjoy Food
3.3 (0.7)
0.996
Satiety
3.2 (0.6)
3.2 (0.5)
0.855
Slow Eating
2.6 (1.0)
3.0 (0.9)
0.052
Emotional Over Eating
2.3 (1.0)
2.8 (1.2)
0.100
Fussy Eating
3.0 (0.8)
2.9 (0.9)
0.627

24. 4 year old females CEBQ Unexposed N = 19 Exposed N = 15 P-value
Food Resp
1.8 (0.6)
2.6 (0.6)
0.002
Enjoy Food
Satiety
Slow Eating
Emotional Over Eating
2.1 (0.9)
3.1 (1.1)
0.008
Fussy Eating

25. Growth trajectory
Postnatal BMI Z scores available for 2-5 time points from 1 month to 4 years of age
Correlations between CEBQ constructs and growth trajectory compared

29. Description of the sample
Exposed
N = 75
Unexposed
N = 126
P-value
Mean (SE)
Birth Weight
3159 (63.75)
3379 (39.51)
<0.01
Mean Z-score (SE)
-1.14 (0.13)
-0.74 (0.09)

32. Interaction with others in CIFASD
This project has interacted with others in the consortium to:
Provide the samples, clinical data, and statistical analysis staff support for the miRNA project
Provide the samples, clinical data, and statistical analysis staff support for Weinberg’s inflammatory markers Developmental Project
Provide the samples and clinical data for Sarkar’s Developmental Project
Provide 3D images for Hammond/Foroud’s project
This project depends on the dysmorphology core

33. Interaction with others in CIFASD
An example of how this project has gained from the consortium is that similar biological samples and clinical data with some overlap have been used for four different types of hypotheses being pursued by other consortium investigators; comparison and contrast, as well as combined analyses utilizing these various domains are very promising
Separate unconnected R01’s using this clinical project as the source of the clinical data, biological samples, and interpretation would not have the synergy that the consortium provides

34. Future directions Earlier diagnosis Interventions
Longer term outcomes e.g. growth and adult diseases of fetal origin
Prediction of resiliency/risk in children prenatally exposed

35. Aims to consider for a future direction: 1. Microbiome
Microbiome is a hot topic as a contributor to early and late developmental effects (and is potentially modifiable)
We have a collaboration with Rob Knight at UCSD who is the leading researcher in this field
In the current cohort, obtain new skin swabs and fecal samples
Utilize existing samples that have been banked from mothers and children to evaluate microbiome (urine, saliva, cheek swabs; protocol being developed to measure microbiome in blood)
Continue to enroll a small number of new pregnant women at the two sites in Ukraine
Obtain skin swabs and fecal samples during pregnancy, infant samples at delivery, and breastmilk samples from those who breastfeed
Core analytical costs $50 per sample and coming down

36. Aims to consider for future direction: 2
Aims to consider for future direction: 2. Pre and early adolescent growth and other DOHAD outcomes
Continue to follow the ~300 children previously retained in the cohort into later development to evaluate
Growth trajectory
Onset of adult disease

37. Aims to consider for future direction: 3
Aims to consider for future direction: 3. Validate previous findings and expand on miRNA and inflammatory profile
Continue to follow the ~300 children previously retained in the cohort into later development and validate and expand on previous pilot work by Miranda and Weinberg
Supplement with additional new pregnant women and capture placental samples as well as a broader range of measures of stress

38. Aims to consider for future direction: 4
Aims to consider for future direction: 4. Create a resiliency/risk prediction model across CIFASD
Using accumulated data on exposures and modifying or mediating factors within CIFASD, for a given age child create a model that can help with understanding the likely trajectory of that child to help better direct resources for services and interventions

39. Biorepository Samples UC UBC Texas A&M
Original maternal blood sample aliquots: 23 per draw
13,152
Maternal plasma aliquots: 1 per draw
567
177
Maternal buffy coats
592
Child plasma aliquots: 10 per draw
452
Child blood spots 92
Maternal cheek swab
268
Child cheek swab
301
Child urine sample –
5 per collection
905