# Presentation: 464547

##  Cytogenetic Effects of Methylphenidate Pediatric Advisory Committee

**June 30, 2005**
**David Jacobson-Kram, Ph.D. DABT**
**Office of New Drugs**
**Center for Drug Evaluation and Research**
**Food and Drug Administration**

## The Issue

## Study design

- Examined three endpoints in 12 children diagnosed with ADHD. Blood drawn before and after  3 month treatment with methylphenidate.
- Sister chromatid exchange
- Chromosomal aberrations
- Micronuclei
- Therapeutic doses were 20 to 54 mg/day

## What are sister chromatid exchanges (SCE)?

- Reciprocal exchanges of chromatid arms visualized in metaphase cells that have undergone two rounds of DNA replication in the presence of the nucleotide analogue bromodeoxyuridine.
- While the mechanism of SCE is poorly understood, increases in their frequencies are generally indicative of DNA damage.

## What are sister chromatid exchanges (SCE)?

## What are chromosomal aberrations?

- Chromosomal aberrations represent unrepaired or misrepaired chromosomal lesions that are visible under the light microscope.
- The same processes that give rise to these events are associated with chromosomal alterations resulting in cancer, e.g. Burkitt’s lymphoma.

## What are chromosomal aberrations?

## What are micronuclei?

- Micronuclei result from acentric chromosome fragments or whole chromosomes left behind in the cytoplasm after mitosis
- They are visualized in binucleated cells that have been blocked for cytokinesis
- Indicative of chromosome breakage or nondisjunction.

## What are micronuclei?

## What is the significance of these cytogenetic endpoints?

- Chromosome aberration frequency in peripheral blood lymphocytes is an independent risk factor for cancer.*
- If reproducible, data from the El-Zein paper suggest that patients taking methylphenidate may be at increased risk for cancer.

- *Chromosomal aberrations and risk of cancer in humans: an epidemiologic perspective. Bonassi et al. 2004

## What else is known about the mutagenicity/carcinogenicity of methylphenidate?

**No structural alerts**
**Metabolism qualitatively similar in humans and animals but quantitative differences exist**
**Negative in rat carc study and mouse p53 study**
**Positive for liver tumors in mouse 2-year study**
**Negative in Ames assay, mouse lymphoma gene mutation assay and in vivo micronucleus test; some positive or equivocal results for in vitro chromosomal aberrations and SCE**
**Review of pharmacy and medical records of 143,574 patients found fewer cancer cases than expected (Selby et al., Cancer Res. 1989).**

## Data summary from El-Zein et al.

## Data summary from El-Zein et al.

## Questions Regarding El-Zein Study

- Lack of placebo controls
- Use of unusual data presentation
- Aberrations/cell instead of % damaged cells
- Total SCE in 25 cells
**Presence of 6 subjects with 0 SCE/cell**
- Investigators agreed to a site visit to answer questions

## Site visit to Univ. of Texas

- Representatives from NIEHS, NICHD, FDA and EPA site visited U of T on May 23rd.
- Reviewed:
- Patient selection
- Methods
- Raw data
- Slide evaluation

## Observations at site visit

**Investigators cordial, cooperative and responded to all inquiries. **
**Good concordance between raw data sheets and data in publication.**
**The slides were evaluated in a “blinded” fashion but the same technician coded, evaluated and decoded slides. **
**A number of slides were chosen at random and were found to have low mitotic indices and poor differential staining for SCE.**

## Effect of preparation quality on SCE frequencies

- Good preparation

- Bad preparation

## Ongoing efforts to assess methylphenidate clastogenic potential organized under BPCA

**El-Zein et al., are seeking funding to perform larger (100 informative subjects) study**
**NICHD, NIEHS and Duke are collaborating to reproduce the El-Zein study**
**CDC has developed a protocol for a cross-sectional study that incorporates cytogenetic endpoints**
**NIMH will assess stable chromosomal rearrangements as part of an ongoing cross sectional study**

## Ongoing efforts to assess methylphenidate clastogenic potential organized under BPCA

**Division of Neuropharm drugs is asking IND holders to assess clastogenic potential**
**NCTR will perform experimental studies in non-human primates and transgenic mice**
**Other drugs used to treat ADD and ADHD will also be studied**
**First results will likely be available in about 1 year**

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