We explored whether breast cancer outcomes are associated with endoxifen and other metabolites of tamoxifen, and to examine potential correlates of endoxifen concentrations including CYP2D6 metabolizer phenotype and body mass index (BMI). Tamoxifen, endoxifen, 4-hydroxytamoxifen and N-desmethyltamoxifen concentrations were measured from 1370 estrogen receptor positive breast cancer patients participating in the Women's Healthy Eating and Living (WHEL) Study, and tested for associations with breast cancer outcomes. Breast cancer outcomes were not associated with tamoxifen, 4-hydroxytamoxifen or N-desmethyltamoxifen concentrations. For endoxifen, a threshold was identified suggesting that women in the upper four quintiles of endoxifen had a 26% lower recurrence rate than women in the bottom quintile. (HR=0.74; 95% CI, [0.55, 1.00]). Predictors of membership in this higher risk bottom quintile were poor/ intermediate metabolizer genotype, higher BMI, and low tamoxifen concentrations. This study suggests a minimal threshold at which endoxifen is effective against breast cancer recurrence, which 80% of tamoxifen-takers achieve.

CYP2D6 genotype and concomitant potent CYP2D6 inhibitors are highly associated with endoxifen plasma concentration and may have an impact on the response to tamoxifen therapy. These iterative approaches may be valuable in the study of other complex genotype-phenotype relationships.

Bone marrow progenitor cells differentiate into mononuclear phagocytes in the presence of colony stimulating factor‐1 (CSF‐1). Characterization of the human CSF‐1 gene shows that it contains 10 exons and 9 introns, which span 20 kb. Analysis of multiple CSF‐1 transcripts demonstrates that alternate use of exon 6 splice acceptor sites and 3′ noncoding sequence exons occurs. These alternatively spliced transcripts can encode either a 224 or a 522 amino acid CSF‐1. Implications of differential splicing for the production and function of CSF‐1 are discussed.

In the yeast Saccharomyces cerevisiae, biochemical and genetic evidence have established that a number of phospholipid biosynthetic enzymes are coordinately regulated in response to the soluble precursors inositol and choline and a common set of regulatory factors (1). The ino2 and ino4 mutants show pleiotropic defects in phospholipid metabolism. Recessive mutations at the IN02 locus lead to reduced phosphatidylcholine synthesis and inositol auxotrophy due to an inability to derepress expression of the INO] structural gene (which encodes inositol-1-phosphate synthase) (2), (3). Ino2 mutant extracts also lack a specific DNAprotein complex that is present in wildtype extracts (4). Thus, the IN02 locus encodes a positive regulatory factor required for derepression of the coregulated phospholipid biosynthetic enzymes.The wildtype IN02 gene was isolated by functional complementation of the inositol auxotrophy in an ino2 mutant. Upon transformation with a partial Sau3A genomic library, one plasmid harboring a 1.8 kilobase SmaI-Xbal insert restored inositol prototrophy. Integrative transformation established linkage to the ino2-21 mutation. In a cross between an integrant and wildtype strain 48 tetrads showed 4:0 segregation for the Ino+ phenotype, confirming that the cloned DNA represents the authentic IN02 locus. The nucleotide sequence of the IN02 gene was determined on both strands by the Sanger dideoxy chain termination method (5). Computer-assisted sequence analysis revealed 912 base pair open reading frame, capable of encoding a 304 amino acid protein with a predicted molecular mass of 34,234 daltons. The Ino2 protein (Ino2p) is largely hydrophilic and acidic (pl = 5.76). Proline residues comprise 8.5% of the protein. A potential structural similarity between the carboxy-terminus of Ino2p and the helix-loop-helix (HLH) domain of the proto-oncogene c-myc mapped to residues 253-291 of Ino2p (see Figure 1) (6). Basic residues precede the HLH domain of Ino2p. Interestingly, the Ino4 protein, which encodes a known transcriptional activator of phospholipid synthesis shares sequence similarity to Ino2p in a region that is restricted the the 68 amino acid HLH structural domain (9). Extracts prepared from ino4 mutants lack the same DNA-protein complex that is missing in extracts prepared from ino2 mutants (4). Thus, it is tempting to speculate that Ino2p and Ino4p may be dimerization partners that associate in a DNA-protein complex to regulate the expression of phospholipid structural genes. ACKNOWLEDGEMENTSWe thank Dr John E.Hill for expertise in the computer sequence analysis.

Introduction: An important technological advance in genetic testing is the DNA microarray, which allows for the simultaneous testing of thousands of DNA sequences. The AmpliChip CYP450 Test employs this microarray technology for cytochrome P450 (CYP) 2D6 and CYP2C19 genotyping. Isoenzymes encoded by these genes are responsible for the metabolism of many widely prescribed drugs. The objectives of this study were to identify CYP2D6 and CYP2C19 alleles and phenotypes in a psychiatric patient population in Kentucky, and to describe practical issues associated with DNA microarray technology.Methods: A total of 4,532 psychiatric patients were recruited from three state hospitals in Kentucky. Whole blood, buccal swabs, or saliva samples were genotyped with the AmpliChip CYP450 Test to derive a predicted phenotype.Results: In this cohort, the overall prevalence of CYP2D6 poor metabolizers was 7.6% (95% CI 7%, 8.3%), 8.2% in the Caucasians (95% CI 7.4%, 9.1%) and 1.8% in the African Americans (95% CI 0.9%, 3.5%). The overall prevalence of CYP2D6 ultrarapid metabolizers was 1.5% (95% CI 1.2%, 1.9%), 1.5% in the Caucasians (95% CI 1.1%, 1.9%) and 2.0% in the African Americans (95% CI 1.1%, 3.7%). The overall prevalence of CYP2C19 poor metabolizers was 2.0% (95% CI 1.8%, 2.7%), 2.2% in Caucasians (95% CI 1.6%, 2.5%) and 4.0% in African Americans (95% CI 2.6%, 6.1%).Conclusion: We also propose a numeric system for expression of CYP2D6 and CYP2C19 enzyme activity to aid clinicians in determining treatment strategy for patients receiving therapeutics that are metabolized by the CYP2D6 or CYP2C19 gene products.