HDAC inhibitors enhance the antidepressant and

anxiolytic effects of fluoxetine.Vanessa Lim

The Bronx High School of Science

Mentor: Dr. Claudia Schmauss M.D

Columbia University

Mood Disorders

• Mood disorders such depression and anxiety

are the most prevalent mental illnesses

diagnosed today.

• Despite of increasing sales of antidepressant

medications in the past decade, there is little of

evidence of greater efficacy.

Histone Lysine 4k12

Adaptive Epigenetic Mechanisms

• Epigenetic mechanisms act at the interface between genes and

the environment.

• Previous studies showed that histone H4 acetylation is an

adaptive epigenetic mark that reduces the severity of the

emotional psychopathology.

Ac

Histone Lysine 4k12

Histone Lysine 4k12

Ac

Ac

Solution to the Problem…Acetylation has also been shown to be responsible for the

enhanced responsiveness of IMS Balb/mice to adolescent

fluoxetine treatment.

• This finding led to the hypothesis that the efficacy of

fluoxetine is enhanced when histone deacetylase (HDAC)

activity is low.

To increase

acetylation….

HDACi

Add an inhibitor!

Animal Care

Animals

• All studies were performed on adult male and female

Balb/c mice. All procedures involving the animals were

approved by the Animal Care and Use Committee of

Columbia University

Behavioral Tests

Elevated Plus Maze Light/Dark Exploration

Behavioral Test

Elevated plus Maze (EPM): A test for anxiety.

Light/Dark Exploration Test (L/D) A test for anxiety.

Forced Swim Test (FST): A test for depression.

Improvement Improvement

Drug Treatment

Drug Treatment

Starting at postnatal age P60 mice received fluoxetine 16 mg/kg/day. Additional groups received the same dose of fluoxetine and either entinostat (MS-275) or trichostatin A (TSA).

Chromatin Immunoprecipitation (ChIP)

I used real-time PCR to analyze the levels of acetylated H4K12 (acH4K12) and RNA Polymerase II (Pol II) at the bdnf

promotor using promotor-specific specific DNA primers.

Emotional Phenotype• Mice treated with fluoxetine alone showed

significant improvement on the FST but not in the EPM and L/D test. Only mice co-treated with the TSA and fluoxetine showed significant improvement in EPM and L/D test.

Molecular Phenotype

Previous studies have shown that P4 and P5 do not play a role in

the epigenetic modifications of antidepressants

Significant Change between control and treated mice.

The highest levels of acetylated histone 4K12 and Pol II are located at bdnf promoter 3.

Significant Change between control and treated mice.

Co-treatment with fluoxetine and HDAC inhibitor TSA led to the highest expression

of bdnf transcript variant 3 mRNA

Discussion• Studies on fluoxetine-treated adolescent SFR mice have

shown that class I HDAC inhibitors, targeting HDAC 1 and 3, are efficaciously improving depression-like behavior but are ineffective in tests of anxiety-like behaviors.

• Class II HDAC inhibitors, effectively exert both antidepressant and anxiolytic effects in adolescent mice co-treated with fluoxetine.

• Also in adult mice treated with fluoxetine, class II HDAC inhibitors enhance anxiolytic effects. (*Fluoxetine alone is a potent antidepressant drug in adulthood.)

• Importantly, the HDAC-dependent mechanism underlying the potentiating effects in adulthood are similarly to those identified in adolescents.

Discussion (continued)

• In both treatment regimens, the most effective enhancement of fluoxetine’s effect was detected when bdnf transcript variant 3 levels were the highest.

• This is of importance because bdnf expression levels seem to decrease as people deal with long-term stress.

Discussion (continued)

• Class I HDACs inhibits HDAC 1, 2, 3, and 8.

• Class II HDACs inhibits HDACs 5, 6, 7, 9, and 10.

• Previous studies have shown that HDACs 5, 6, 7, 9 do not play a major role in antidepressant-triggered epigenetic mechanisms

• This suggests that reduced activity of HDAC 10 plays a significant role in enhancing the anxiolytic effects of fluoxetine treatment.

Conclusion

• Contrary to adolescent fluoxetine, adult fluoxetine alone has potent antidepressant effects. However, only the co-administration of fluoxetine and a class II HDAC inhibitor also triggers anxiolytic effects in adult SFR mice.

• This decrease in anxiety-like behaviors measured in the EPM and L/D test is accompanied by increased enrichment of acH4K12 and RNA polymerase II at promotor 3 of the bdnf gene, ultimately resulting in increased bdnf transcript variant 3 mRNA expression.

Future Experiments

• Future clinical studies can be conducted on human subjects who suffer from depression and are resistant to antidepressant treatment with fluoxetine alone.

• Identify other genes (in addition to bdnf) whose transcriptions are also epigenetically modified by fluoxetine/HDACi interaction (done possibly by ChIP-seq)

• Focus studies on HDAC 10 activity to clarify its role on mediating anxiolytic effects.

Acknowledgement

I would like to thank Dr. Claudia Schmauss , MD from Columbia University for her supervision and mentorship. I would also like to thank Dr.

Gensert, for being my research teacher in school.

Citations

• Castren, E., Rantamaki, Tomi. (2010). The Role of BDNF and Its Receptors in Depression and Antidepressant Drug Action: Reactivation of Developmental Plasticity. Developmental Neurobiology, 70, 289-294.

•Hoyert, Donna L., Xu Jiaquan. (2012). Deaths: Preliminary data for 2011. National Vital Statistics Reports, 61,4.

•Kovacs, D., Gonda, X., Petschner, P., Edes, Andrea., Esziari, Nora., Bagdy, G,. Juhasz, G. (2014). Antidepressant treatment response is modulated by genetic and environmental factors and their interactions. Annals of General Psychiatry, 13, 1-9.

•Lee, S., Jeong, J., Kwak, Y., Park, S.K. (2010). Depression research: where are we now?. Molecular Brain, 3, 1.

•Levine, A., Worell, T.R., Zimnisky, R., & Schmauss, C. (2012). Early life stress triggers sustained changes in histone deacetylase expression and histone H4 modifications that alter responsiveness to adolescent antidepressant treatment. Neurobiology of Disease, 45, 488-498.