Ma

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University of Maryland Baltimore
Experimental Therapeutics Symposium 2009

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Ma

  1. 1. Develop in vivoExperimental Therapeutic Model for Anti-Cancer Research <br />Mark (Zhi-Qing) Ma, Ph.D.<br />
  2. 2. New Preclinical Models to Improve Predictive Value<br /><ul><li>Heterotransplant model of human tumor xenograft
  3. 3. Transgenic mouse mode (Over-expression of oncogene)
  4. 4. Gene Knockout mouse model (deletion of tumor suppressor) </li></li></ul><li>Concept of Heterotransplants<br />
  5. 5. Tumor Genome Analysis <br />Pre- and Post Treatment<br />intact tumor <br />microenvironment<br />GeneChip Human Exon 1.0 ST Array<br />DNA Methylation Analysis<br />other genetic and epigenetic assays<br />Genome-Wide Human SNP Array 6.0<br />
  6. 6. matched<br />matched<br />50 mouse models<br />Repeatable<br />Systematic Preclinical Efficacy Evaluation of Drugs (SPEED)<br />CTRL Arm : 50 Pts<br />Tx Arm : 50 Pts <br />Pathak et al., Am J Trans Res, 2009<br />
  7. 7. Transgenic Mouse Lines Over-expressing DNMT3B<br />SPC/Δ3B4/IGFP (Δ3B4G)<br />DrdI<br />DrdI<br />NdeI<br />NotI<br />NotI<br />IRES-GFP<br />pA+<br />SPC promoter<br />ΔDNMT3B4<br />Geno#1<br />Forward<br />Geno#1<br />Reverse<br />Geno#2<br />Forward<br />Geno#2<br />Reverse<br />SPC/Δ3B4 del/IGFP (Δ3B4DG)<br />DrdI<br />DrdI<br />NotI<br />NdeI<br />NotI<br />IRES-GFP<br />pA+<br />SPC promoter<br />ΔDNMT3B4 del<br />Geno#1<br />Forward<br />Geno#1<br />Reverse<br />Geno#2<br />Forward<br />Geno#2<br />Reverse<br />
  8. 8. Purification of Transgene fragments for Microinjectin<br />SPC/Δ3B4Del/IGFP<br />(Δ3B4G)<br />SPC/Δ3B4/IGFP<br />(Δ3B4G)<br />Transgene<br />10 kb<br />8<br />6<br />4<br />3<br />Vector <br />
  9. 9. Genotyping Strategy of Transgenic Mice <br />Tail DNA + Δ3B4DG<br />Tail DNA + Δ3B4DG<br />Tail DNA + Δ3B4G<br />Tail DNA + Δ3B4G<br />100 bp Marker<br />1 kB Marker<br />Tail DNA<br />Tail DNA<br />Geno#1<br />Geno#2<br />
  10. 10. Generation of Conditional Mutant<br />Cre<br />floxed<br />breeding<br />ATG<br />Promoter<br />Cre<br />Exon3<br />Exon2<br />Tissue Specific<br />Conditional KO mice<br />
  11. 11. Generation of FloxedHuR Mouse Line<br />5 ARM <br />(5.1KB)<br />3 ARM<br />(1.7 KB)<br />5 ARM probe<br />3 ARM probe<br />E<br />E<br />A<br />A<br />A<br />A<br />A<br />A<br />E<br />E<br />E<br />E2<br />E1<br />E3<br />Neo<br />PGK-TK<br />Targeting Vector<br />LoxP<br />LoxP<br />LoxP<br />WT Mutant<br />5ARM (Apl I) 10.0kb 11.3kb<br />3ARM (EcoRI) 5.5kb 7.5kb<br />

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