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Chapter 11. Medical Biotechnology. Detecting and Diagnosing Human Disease Conditions. Models of Human Disease Identify diseases and test therapies before clinical trials in humans Clinical trials: three phases Phase I: safety studies- safe dose and how to administer the dose (ADME)
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Chapter 11 Medical Biotechnology
Detecting and Diagnosing Human Disease Conditions • Models of Human Disease • Identify diseases and test therapies before clinical trials in humans • Clinical trials: three phases • Phase I: safety studies- safe dose and how to administer the dose (ADME) • Phase II: few hundred patients for the purpose of testing effectiveness • Phase III: effectiveness compared to other drugs –involve thousands of patients often with different backgrounds and stages of illness throughout the country
Detecting and Diagnosing Human Disease Conditions • FDA Oversight of Drug Development Process • Clinical Trials Video: What is a Clinical Trial?
Detecting and Diagnosing Human Disease Conditions • Lab to Market Failure • Limited biomarkers to optimize treatment • Inadequate experimental approaches for patient selection • Limitations in animal models Interactive Map of Current US Clinical Trials
Detecting and Diagnosing Human Disease Conditions • Models of Human Disease • Rat gene, ob, also found in humans therefore homologous • Codes for a protein hormone called leptin if missing leads to obesity • Found that treating obese children defected in this gene with leptin decreases their weight
Detecting and Diagnosing Human Disease Conditions • Models of Human Disease • Organism has 959 cells, 131 of them are destined to go through apoptosis • Study programmed cell death in this organism • Programmed cell death important to correct development of the fetus and improper cell death is implicated in: Alzheimers, Lou Gehrig’s, Huntington’s, Parkinson’s
Detecting and Diagnosing Human Disease Conditions • Models of Human Disease • Heart attack mice • Defect in genes for cholesterol uptake • HIV small animal model HIV Clinical Trials
Detecting and Diagnosing Human Disease Conditions • Detecting Genetic Diseases
Detecting and Diagnosing Human Disease Conditions • Molecular Diagnostics • The use of DNA, RNA, and proteins to facilitate disease detection, diagnosis, subclassification, prognosis, and monitoring response to therapy
Detecting and Diagnosing Human Disease Conditions • Advantages of Molecular Diagnostics • Improvement in sensitivity • High specificity • Cost less • Faster analysis time
Detecting and Diagnosing Human Disease Conditions • Detecting Genetic Diseases • Fetal testing for chromosome abnormalities and defective genes • Amniocentesis (Test at 16 weeks - karyotype) • Chorionic villus sampling (Test at 8 to 10 weeks - karyotype)
Detecting and Diagnosing Human Disease Conditions • Detecting Genetic Diseases • Testing for chromosome abnormalities and defective genes • Fluorescence in situ hybridization (FISH) • Fluorescence probes that are specific for chromosomes and/or genes • Spectral karotype
Detecting and Diagnosing Human Disease Conditions • Detecting Genetic Diseases • Testing for chromosome abnormalities and defective genes • RFLP (restriction fragment length polymorphisms)
Detecting and Diagnosing Human Disease Conditions • Hybridization - single-stranded oligonucleotides are permitted to interact so that complexes, or hybrids, are formed by molecules with sufficiently similar, complementary sequences • Target - the nucleotide sequence the oligonucleotide is designed to hybridize with • Probe - the nucleic acid that carries a marker for detection Making SNPs Make Sense
Detecting and Diagnosing Human Disease Conditions • Dot Blots • Assay for detecting SNPs • Uses PCR amplified DNA blotted onto a membrane • Unbound ASO probe is washed off • Bound ASO probe is detected by radioactive or colorimetric assays Dot Blot animation
Detecting and Diagnosing Human Disease Conditions • Allele-Specific Oligonucleotide (ASO) Dot Blot to detect Sickle Cell Anemia
Detecting and Diagnosing Human Disease Conditions • Reverse Dot Bot • Instead of binding DNA to the membrane, an array of ASOs are bound to a membrane and hybridized to labeled target DNA Reverse Dot Blot video
Detecting and Diagnosing Human Disease Conditions • Detecting Genetic Diseases • Single Nucleotide Polymorphisms (SNPs) • One of the most common forms of genetic variation • Estimated that one SNP occurs approximately every 1,000-3,000 bp in the human genome • 99.9 percent of the DNA sequence will be exactly the same –> 80% of 0.1 percent variation will be SNPs • Most have no effect because they occur in non-protein coding regions (introns) • 10 pharmaceuticals donated millions in a collaborative partnership called the SNP Consortium
Detecting and Diagnosing Human Disease Conditions • Microarray • A chip containing thousands of pieces of single stranded DNA molecules • DNA is isolated from a patient, fluorescently labeled, and hybridized to the microarray • A laser scanner measures the intensity of the fluorescence to indicate the binding of the patients DNA to the SNP or gene on the microarray
Detecting and Diagnosing Human Disease Conditions • Detecting Genetic Diseases • Identifying sets of disease genes by microarray analysis • Microarray created with known diseased genes or SNPs • DNA from a patient is tagged with fluorescent dyes and then hybridized to the chip • Binding of a patient’s DNA to a gene sequence on the chip indicates that the person’s DNA has a particular mutation or SNP
Medical Products and Applications of Biotechnology • Pharmacogenomics – individualized medicine based on a person’s genetic information Pharmacogenomics Animation
Medical Products and Applications of Biotechnology • The search for new medicines and drugs • Oncogenes- genes that produce proteins that may function as transcription factors and receptors for hormones and growth factors, as well as serve as enzymes involved in a wide variety of ways to change growth properties of cells that cause cancer • Tumor Suppressor Genes – regulate oncogenes
Medical Products and Applications of Biotechnology • The search for new medicines and drugs • Personalized Medicine • BRCA1 or 2 – increases risk of developing breast cancer • But there are many other cases of breast cancer that do not exhibit this mode of inheritance • They SHOULD be treated differently (i.e. different chemotherapy!)
Medical Products and Applications of Biotechnology • Microarray technology • Can compare levels of gene expression in different tissues • Applications in cancer research Microarray animation
Medical Products and Applications of Biotechnology • The search for new medicines and drugs • Improving techniques for drug delivery • Factors that influence drug effectiveness • Drug solubility • Drug breakdown • Drug elimination
Medical Products and Applications of Biotechnology • Microparticle Drug Delivery • Microspheres – tiny particles that can be filled with drugs Drug Delivery Video
Medical Products and Applications of Biotechnology • Nanotechnology – area of science involved in designing, building, and manipulating structures at the nanometer (nm) scale Nanotechnology video
Medical Products and Applications of Biotechnology • Polymer based nanomedicine Targeted Medicine Video
Medical Products and Applications of Biotechnology • Nanobots – manufactured device with embedded sensors to facilitate target identification and drug delivery
Medical Products and Applications of Biotechnology • Vaccines and Therapeutic Antibodies • Vaccines stimulate immune response • Also hope that vaccination may be useful against conditions such as Alzheimer’s disease or drug addiction • Using antibodies in some types of therapies: Development of Monoclonal Antibodies
Gene Therapy • Gene Therapy – treating disease by inserting functional genes to replace defective ones
Gene Therapy • A vector delivers the therapeutic gene into a patient’s target cell • Functional proteins are created from the therapeutic gene Gene Therapy Animation
Gene Therapy • In vivo Gene Therapy • Ex vivo Gene Therapy Gene Therapy Video
Gene Therapy • Vectors for Therapeutic Gene Delivery • Viral • Non-Viral Animation: Gene Therapy Vectors
Gene Therapy • 1990 – 4 year old Ashanti DaSilva had a genetic disorder called severe combined immunodeficiency (SCID) • Defect in ADA gene results in an accumulation of dATP, which is toxic to certain types of T cells • Takes down the entire immune system
Gene Therapy • Case Study: Cystic Fibrosis • Defective cystic fibrosis transmembrane conductance regulator (CFTR) • Normally it serves as a pump at the cell membrane to move electrically charged chloride atoms out of the cells • If cells can’t move chloride out, they absorb water trying to dilute the chloride in the cell • This leads to the production of THICK sticky mucus Animation: Cystic Fibrosis Case Study
Gene Therapy • Gene Therapy in Clinical Trials
Gene Therapy • Challenges • Adverse effects of viral vectors • Targeting specific cells • Controlling expression of the therapeutic gene • Long lasting therapy Video: Challenges of Gene Therapy
Regenerative Medicine • Growing cells and tissues that can be used to replace or repair defective tissues and organs Regenerative Medicine Video
Regenerative Medicine • Tissue Engineering - replacement of tissues and organs by growing them in culture • Cell based • Scaffold guided Video: Tissue Engineering
Regenerative Medicine • Cells Based Tissue Regeneration • Fetal tissue grafts • Nanofibers
Regenerative Medicine • Scaffold Guided Tissue Regeneration • Creates framework onto which cells are seeded and bathed in growth factors
Regenerative Medicine • Application: Bone regeneration Bone regeneration animation
Regenerative Medicine • Application: Blood Vessel regeneration Animation of blood vessel regeneration
Regenerative Medicine • Cells and Tissue Transplantation • Organ transplantation • Autograft – transplanting a patient’s own tissue from one region of the body to another- ex. Vein from leg used in coronary bypass-organ transplants are between individuals and so must be checked for compatibility • Histocompatibility complex - >70 genes which produce tissue typing proteins (must match!) • There are many different types of MHC proteins (one group is called human leukocyte antigens or HLAs)- have been using immunosuppressive drugs but there are problems.
Regenerative Medicine • Cells and Tissue Transplantation • Organ transplantation • Xenotransplantation – transfer between species (pig to human) • University of Missouri scientists have produced cloned, knockout pigs that lack a gene called GGTA1 (or 1,3 galactosyltransferase) • The gene normally codes for a sugar that would be recognized as foreign by humans
Regenerative Medicine Video: Growing Body Parts
Regenerative Medicine • Bioprinting - The construction of a biological structure by computer-aided, automatic, layer-by-layer depositing of bioink onto biopaper. • Bioink – cells from patient • Biopaper – thin gel layers Bioprinting Video
Regenerative Medicine • Embryonic Stem Cells – cells derived from a blastocyst that can differentiate into any type of cell in the body Stem Cell Animation
Regenerative Medicine • Culturing embryonic stem cells