History of Medicine and clinical research

Evidence of clinical research has been found as far back as 600 BC. In the biblical book of Daniel, Daniel describes how he and three companions were captured in their native land of Judah and transported to Persia where they were forced to serve under the King of Babylon. Daniel wrote that he objected to the king’s food and described a comparative protocol for diet and health. He showed that a diet of legumes and water made for healthier individuals than the King of Babylon’s diet of meat, fish, eggs, other rich foods, and wine. Two thousand years before Roger Bacon originated the scientific method, Daniel described his experiment:

Ancient Chinese herbalists such as Shen Nung (2700 BC) described clinical studies. Nung tasted and tested plants for their medicinal properties, gaining practical experience that allowed him to categorize medicines and record their toxicity and lethal dosage.

Evidence suggests that surgical techniques were being performed in ancient India as early as 800 BC. Sushruta studied anatomy and described plastic surgery techniques that he developed. He specialized in cataract extraction and rhinoplasty (restoration of a mutilated nose). The details of his surgeries were recorded in Sushruta Samahita (“Sushruta’s Compendium”). The steps Sushruta followed are remarkably similar to those used during advanced surgery today.

The practice of medicine in Europe dates to the 5th century BC, around the time of the birth of the Greek physician Hippocrates in 460 BC. Hippocrates believed that there was a rational explanation for all physical illnesses, rejecting the then-popular belief that the cause of disease was the disfavor of the gods or evil spirits. He founded a medical school and traveled throughout Greece practicing medicine.

Hippocrates accurately described disease symptoms of malaria, respiratory infections, diarrhea, and epilepsy. He was the first physician to use a careful, systematic approach while examining a patient’s condition. His observations included facial appearance, pulse, respiration, temperature, localized pains, appearance of bodily fluids (sputum, feces, and urine), and movements of the body. Hippocrates described the importance of cleanliness in managing patient wounds. The Hippocratic Oath taken by all physicians today before they begin medical practice is a modern version of a medical ethics doctrine authored by Hippocrates.

Aelius Galenus often anglicized as Galen (AD 129–217), who is considered the father of experimental physiology, challenged the teachings of Hippocrates. Galen was a Greek physician who traveled extensively throughout his life while practicing medicine, teaching, and studying. He was a prolific writer, authoring more than 70 books on anatomy, physiology, pharmacy, pathology, and temperaments, and was the first physician to perform animal experiments with cats, monkeys, pigs, and oxen. Through his animal experiments, Galen demonstrated that an excised heart would beat outside of the body, showing that the heartbeat is not dependent on the nervous system.

Medicine continued to improve throughout the Middle Ages (476–1453) and Renaissance (1453–1600). Hospitals were built in England, Scotland, France, and Germany starting around the 1100s, and the first medical textbooks were published in the 1470s. Andreas Vesalius (1514–1564), a Belgian physician who may have been inspired by the anatomical drawings of Leonardo da Vinci (FIGURE 17-2), changed the history of the study of anatomy by dissecting the human body.

Vesalius and his students resorted to grave-robbing and other activities to secure cadavers for their studies. In 1543, Vesalius published two anatomy manuals that corrected the works of Galen. His research on human anatomy provided a strong foundation for all future medical research (FIGURE 17-3). During the 17th century, emphasis was placed on studying a patient’s blood and vital statistics. Extraordinary advances in medical research and microbiology occurred in the 18th century. Microbiology pioneer Antoni von Leeuwenhoek (1632–1723) invented the light microscope. James Lind conducted clinical trials in 1747 involving the treatment of scurvy with citrus fruits. Edward Jenner’s 1796 vaccination trials used cowpox scabs taken from the hands of milkmaids to prevent smallpox.

Ignaz Semmelweis (1818–1865) led the most sophisticated preventative clinical trial of the 19th century by requiring physicians to wash their hands with chloride of lime to prevent the spread of “childbed,” or puerperal, fever in the maternity ward of a Vienna General Hospital in 1846. Mortality rates of women in childbirth dying from puerperal fever were as high as 50% from 1841 to 1846. Semmelweis studied the cadavers of fever victims, including a fellow physician, Jakob Kolletschka, who died of a similar infection sustained from a small cut on his finger during an autopsy of a deceased mother. Semmelweis reasoned that the disease was caused by an infectious living organism and insisted on handwashing practices. After doctors complied with the handwashing practices, the mortality rate of pregnant women at the Vienna General Hospital dropped to 1.27% in 1848.

Medicine was a male-dominated field, with the exception of women’s significant role in assisting childbirth. The first woman to graduate from an American medical school was Elizabeth Blackwell (1821–1910; FIGURE 17-4A). In her first career, Blackwell was a teacher, which was considered a suitable career for a woman. After a close friend died, her interests shifted toward caring for the sick. After applying to and being rejected by more than 20 medical schools, Blackwell gained admittance, despite the reluctance of most students and faculty at Geneva Medical College in upstate New York. In fact, the admissions committee accepted her application as a “joke.” Two years later, in 1849, she earned her degree.

After graduation, Blackwell worked in clinics in London and Paris for 2 years. Blackwell lost her sight in one eye after contracting a severe, purulent eye infection from a patient while working and studying midwifery at La Maternite in Paris, which ended her dreams of becoming a surgeon. She returned to New York in 1851 but was refused work as a physician. In 1853, with the help of her friends, she opened her own dispensary in a single rented room. Her sister also became a doctor, and together they opened the New York Infirmary for Women and Children in 1857, where they provided training and experience for women doctors and medical care for the poor. The hospital was staffed by women.

Another pioneer in medicine was Canadian Emily Jennings Stowe (1831–1903), the first woman to practice medicine in Canada (FIGURE 17-4B). Like Blackwell, Jennings became a teacher at the age of 15. She married John Stowe in 1856, and for the first several years of their marriage she stayed at home while raising their children. When her husband contracted tuberculosis and was committed to a sanatorium, Stowe returned to teaching for financial reasons. Her husband’s illness inspired her interest in medicine, though, and she decided to pursue a medical career.

Stowe faced the same adversities as Elizabeth Blackwell, and she was denied entrance into medical schools in Canada. She persisted and was later accepted at the New York Medical College for Women, from which she graduated with a specialty in diseases of women and children. Stowe returned to Canada in 1867 and established a private practice in Toronto. She saw patients on a regular basis, but the College of Physicians and Surgeons in Ontario initially denied Stowe a medical license because the college admissions committee did not accept women into their program. Eventually, Stowe was admitted and earned her license to practice medicine in 1880. Other medical pioneers in the 19th century were Louis Pasteur, Robert Koch, Emil von Behring, and Elie Metchnikoff.

Insulin and Penicillin: great Medical breakthroughs in the 20th century

The discovery of penicillin and insulin early in the 20th century was monumental in driving clinical research forward. The collaborative efforts of a team of Canadian doctors at the University of Toronto led to the discovery in 1921 of insulin, a lifesaving treatment for individuals suffering from diabetes. Diabetics cannot metabolize sugars due to the lack of insulin production, which results in a high concentration of sugar in the blood and urine. Before the discovery of insulin, diabetics were put on special restrictive diets. Patients lost weight, and many died of poor nutrition.

The insulin project began when Fredrick Banting hypothesized that the pancreas secretes a hormone responsible for the metabolism of sugar. In 1921, he discussed his hypothesis with the researcher John James Rickard Macleod, an expert on carbohydrate metabolism. Macleod cautiously agreed to provide Banting with laboratory space and funds to begin testing his hypothesis. In addition, he was offered the assistance of graduate student Charles Best during the summer of 1921, and together they perfected a surgical procedure that would create diabetic dogs and invented a technique to measure the dogs’ blood sugar levels. They were able to keep the diabetic dogs alive with a crude extract from the pancreas of dogs.

James Bertram Collip, a biochemist with protein purification expertise, joined Banting’s team in the fall of 1921. His contribution to the project was a purified extract of the pancreas that contained “antidiabetic” properties. In January 1922, Collip’s extract—insulin— was tested on 14-year-old Leonard Thompson, who had been a diabetic for 3 years. At the time, Thompson weighed a mere 65 pounds and was near coma and death. After receiving the extract, his blood sugar returned to normal and he regained his health. In 1923, Banting and Macleod shared the Nobel Prize in Physiology or Medicine with Best and Collip.

Penicillin may have saved the lives of your parents and grandparents (FIGURE 17-5). In contrast to the discovery of insulin by hypothesis-driven medical research, penicillin was an accidental discovery. Alexander Fleming earned a medical degree from St. Mary’s Medical School in London in 1906. After doing research and serving in World War I as a captain of the Army Medical Corps, Fleming returned to lecturing and research at St. Mary’s. His research focus was antiseptics produced by human tissues and secretions. In 1921, he discovered an enzyme secreted in tears called lysozyme. Lysozyme breaks down the cell walls of certain bacteria and can act as a mild antiseptic.

Like most scientists, Fleming was not in the habit of discarding old experimental bacterial culture plates. Many scientists keep their experimental reagents as long as possible in case they need to return to the experiment. One day in 1928 Fleming glanced at a stack of old Petri plates containing staphylococci colonies. He examined a greenish mold contaminating the surface of the solid medium in the plate. None of the staphylococci bacteria were growing near the mold, which appeared to be producing a substance that caused the staphylococci to lyse or rupture.

Fleming began experiments to identify the laboratory mold, which he determined was Penicillium, and to screen other molds for their antibacterial properties. In 1929, his results were published in the British Journal of Experimental Pathology as a research report titled, “On the Anti-bacterial Action of Cultures of a Penicillium, with Special Reference to Their Use in the Isolation of B. influenzae.” Fleming demonstrated that the mold inhibited Streptococcus, Staphylococcus, and Corynebacterium bacterial species, but that it was not toxic to many gram-negative bacteria. He named the active substance penicillin and suggested that it could be used to isolate bacteria in the laboratory and as an antiseptic to remove penicillin-sensitive bacteria on patient dressings.

Fleming did not follow up on the application of penicillin in treating bacterial infections. Instead, a decade later, Howard Florey and Ernst Chain followed through on the medical applications of penicillin. Chain extracted penicillin from the Penicillium mold and with the aid of Florey began testing it on mice that had been infected with Streptococcus. All mice infected with Streptococcus survived after penicillin injections, whereas mice that were not given penicillin injections died. The experiments were expanded to include human trials, which required the mass production of penicillin. Fleming, Florey, and Chain shared the Nobel Prize in Physiology or Medicine in 1945 “for their discovery of penicillin and its curative effect in various infectious diseases.”

Before the 20th century, the infant mortality rate in the United States was as high as 20%, and many children died before reaching 1 year of age. The rate decreased to 5.98% by 2015. The reduction has been attributed to vaccine-preventable diseases, the introduction of antibiotics, handwashing, and other methods used to prevent and control infections. In the 1950s the average life expectancy in the United States was 62 years; in 2012, it was 79.

The past 50 years have seen tremendous advances in biomedical research. Therapies being developed today will gradually become commonplace in treating future health problems. Each new application will require all of us—researchers, policymakers, and the public—to consider the benefits, risks, and implications of new and innovative treatments.

Gene Therapy

Gene therapy is an experimental treatment that involves the introduction of genes into a person’s cells to replace or compensate for defective genes in a person’s body that are responsible for a disease or medical problem. But what does gene therapy have to do with viruses?

With gene therapy, the “good” genes must be delivered or find their way to the location in the body in which their product is normally active. The “good” genes can be carried and delivered to cells by customized vectors such as viruses. The engineered viruses are manipulated using genetic engineering techniques to escape the body’s immunosurveillance system, ensuring that the patient is not harmed. The safety of gene therapy was intensely reevaluated after Jesse Gelsinger died of complications during a 1999 clinical trial on OTC, as described in Section 17.1.

Gelsinger was one of two individuals in the study who were administered 300 times the normal gene therapy virus vector dose as the initial patients. The vector was a modified adenovirus. Gelsinger died as a result of the accumulation of a cytokine known as interleukin 6 (IL-6). High levels of IL-6 cause adult respiratory distress syndrome (ARDS). Normally, another cytokine, interleukin 10 (IL-10), suppresses IL-6 production, as does another cytokine, tumor necrosis factor alpha (TNF-α). TNF-α is a cellular protein involved in the body’s inflammation response to an infection. Unfortunately, Gelsinger’s IL-10 and TNF-α levels did not effectively reduce the increased IL-6 levels, resulting in an immunological complication and ultimately his death.

After Gelsinger’s death, gene therapy trials at the University of Pennsylvania were stopped and the FDA performed an investigation. The FDA determined that the IHGT had violated 18 specific FDA rules and regulations during the OTC study, and the field of gene therapy in general was scrutinized and questioned. The FDA and NIH created new regulations and policies to police experiments and new protocols. Rules issuing the disclosure of information regarding gene therapy trials are available to the general public through Web-based databases.

In January 2003, gene therapy had another major setback. The FDA temporarily halted 27 gene therapy trials in which retrovirus vectors were being used. The halt was enforced after the FDA learned that a second child had developed leukemia after being treated for X-linked severe combined immunodeficiency disease (X-SCID). The syndrome only affects boys. Boys with X-SCID have a defective copy of a gene that encodes the cytokine interleukin 2 (IL-2), which modulates T helper (T_H) cell production, located on their X chromo-some. T_H cells play a vital role in cell-mediated immunity to fight infections. Without gene therapy treatment, X-SCID children usually die before their first birthday.

The second child who developed leukemia was participating in a trial led by Alain Fischer at Necker Hospital in Paris, France. A retroviral vector was used to genetically modify the boy’s own bone marrow stem hematopoietic cells. Unfortunately, the modified retrovirus integrated its genome into a cellular gene involved in normal cellular growth. The retroviral integration event disrupted the gene’s function, resulting in uncontrolled white blood cell division (leukemia). The theoretical risk of retroviral integration was known; however, this was the second time the retrovirus gene therapy vector integrated into the same gene. Both patients got leukemia during gene therapy treatment. The risk of retroviral integration into the chromosome of a host cell appeared to be much greater than researchers thought it would be.

The role of gene therapy in the death of Jolee Mohr was determined to be unrelated to gene therapy. Mohr died on July 24, 2007, at the University of Chicago Medical Center after her second gene therapy treatment. The 36-year-old woman received two injections of a gene therapy vector into her right knee to treat rheumatoid arthritis, a chronic inflammatory disorder. The injections contained a genetically engineered adeno-associated virus (AAV) that contained a gene that encodes the receptor for TNF-α. TNF-α plays a pivotal role in regulating the body’s inflammation response in rheumatoid arthritis. The leading cause of her death was a massive Histoplasma capsulatum fungal infection. Mohr was taking immunosuppressive drugs to treat rheumatoid arthritis. The drugs may have played a role in weakening her body’s ability to fend off the fungal infection. Initial tests determined that only a trace amount of AAV was found in tissues outside of her knee joint.

Despite these rare, highly publicized events, gene therapy trials continue to move forward. Gene therapy holds promise for treating genetic disorders caused by single-gene defects such as Huntington’s chorea, Duchenne muscular dystrophy, polycystic kidney disease, familial hypercholesterolemia, sickle cell anemia, hemophilia A and B, phenylketonuria, and cystic fibrosis. Other gene therapy applications are used to treat other health problems, such as cancer, diabetes, high blood pressure, and heart disease. The hope is that gene therapy will provide cures for diseases in which the good gene must function throughout one’s life, such as hemophilia. In other instances, gene therapy may be used to restore eyesight (including color blindness) or hearing, repair a wound, or grow new blood vessels. In the latter examples, the treatment would be a temporary solution.

At the time of this writing, the majority of gene therapy clinical trials are taking place in the United States (1,386 trials; 65%), Europe (524 trials; 25%), Asia (109 trials; 5.1%), and Australasia (34 trials; 1.6%). About 83 (4%) clinical trials are multicountry collaborations. Of these, 3.5% are in Phase III (76 trials) and 0.1% are in Phase IV (2 trials). As guidelines have become more stringent, fewer trials are being approved. The number of clinical trials approved decreased from 2009 to 2012. In 2013, the number of trials approved worldwide was 120—the same number approved in 2008 (FIGURE 17-6).

Gene Delivery by Viruses: The Key to Gene Therapy

One of the major challenges of gene therapy is delivering genes to the correct cells or tissues where they must function. For example, if the good gene must function in the liver, the good genes must be targeted to and function correctly in the liver cells. How do scientists ensure that the good gene is targeted to the liver and not the big toe?

Viruses can act as gene delivery vehicles, or vectors. They make good vectors because they can be genetically engineered using recombinant DNA techniques and polymerase chain reaction (PCR) to target and enter specific types of host cells. To safeguard the patient, viruses are engineered so that they cannot replicate and destroy or harm the cells of the patient. Some drawbacks remain regarding viral vectors, though. For example, they are limited in the size of gene that they can carry. Some genes may be too large for certain viral vectors. Sometimes the patient may get sick (as in the case of Jesse Gelsinger) and the patient may mount an immune response toward the viral vector, preventing further treatments from working. The hallmarks of a good gene delivery system are its ability to:

• 
  

  Target the appropriate host cells.

  
  
• 
  

  Integrate the correct gene into the cell’s chromosomal DNA.

  
  
• 
  

  Transcribe and translate the gene of interest so that its gene product can function properly.

  
  
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  Cause no toxic or harmful effects.

Two methods are used to target the genes of interest into the patient’s cells using viral vectors:

1. 
   

   In vivo gene therapy: The patient’s body is directly injected or infused with the modified gene therapy viral vector.

   
   
2. 
   

   Ex vivo gene therapy: The patient’s cells, such as cells from bone marrow, are removed, grown in culture dishes in the laboratory, infected with the viral vector to introduce the gene of interest, and subsequently infused back into the patient.

Ex vivo gene therapy requires highly specialized facilities in contrast to in vivo gene therapy. In vivo gene therapy is a scalable method; ex vivo gene therapy is not (FIGURE 17-7). The following are the most popular viral vectors currently being used in gene therapy trials:

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  Retroviruses

  
  
• 
  

  Adenoviruses

  
  
• 
  

  Adeno-associated virus (AAV)

  
  
• 
  

  Herpes simplex virus 1

  
  
• 
  

  Vaccinia or modified vaccinia Ankara (MVA)

As detailed in TABLE 17-1, each type of gene therapy viral vector has advantages and limitations.

Gene Delivery Without Viruses

Plasmid DNA, or “naked” DNA, is also used to deliver genes into patients’ cells. Plasmids have fewer limitations regarding the size of the DNA of the corrected gene that can be inserted. Plasmid DNA does not induce an immune response by the body, making plasmids a safer alternative than viral vectors. Plasmid DNA delivery is less efficient than viral delivery, however. Research in the past 5 years has revolutionized the efficiency of nonviral gene transfer. To aid plasmid DNA cell entry, the DNA is either complexed with liposomes or other chemical polymers or physical energy is applied. Physical energy methods include electroporation, pressure-mediated delivery, ultrasound, laser, magnetic fields, and ballistic delivery.

Triple helix–forming nucleotides, antisense technology, ribozymes, and RNA interference systems (RNAi) are all being used as gene therapy approaches when inserting a good copy of a gene will not correct the cell’s problem; for example, a defective gene may code for a gene product that prevents the normal gene product from functioning correctly in the cell. In this case, a good copy or normal gene will not help. Instead, the approach to repairing the problem is to remove or inactivate the defective gene. The majority of the types of genes engineered into vectors used in today’s clinical trials function as:

• 
  

  Specific antigens (involved in immune modulation)

  
  
• 
  

  Cytokines

  
  
• 
  

  Tumor suppressors

  
  
• 
  

  Receptors

  
  
• 
  

  DNA replication inhibitors

  
  
• 
  

  Cell protection/drug resistance

  
  
• 
  

  Proteins that compensate for defective gene products

  
  
• 
  

  Growth factors

  
  
• 
  

  Proteins that induce apoptosis (programmed cell death)

A smaller percentage of clinical trials involve transferring other types of genes that function as hormones, adhesion molecules, porins, ion channels, transporters, ribozymes, gene silencers (siRNA), and transcription factors.

Why Pigs?

Pig-to-human xenotransplantation is a possible solution to the world shortage of organ donors. Why would transgenic pig donors be able to save or improve the quality of human lives? Why not choose to use closer genetic relatives, such as apes or monkeys? Using pigs for donor organs for human transplants has a number of advantages (TABLE 17-2). Pig organs are similar in size to human organs, and pig physiology is similar to that of humans. A pig kidney will act similarly to a human kidney, and the circulatory system is similar. The size of a pig heart would meet the cardiac output of a human.

Another advantage is that pigs are readily available and can be bred quickly. Pigs have a short gestation period and can carry up to 18 piglets in a litter. They can be raised in special pathogen-free environments following cesarean protocols to exclude serious human pathogens at minimal cost. In addition, pigs have been genetically modified to decrease immunological rejection and were successfully cloned in 2000. Overall, the genetic modifications result in a more “human” pig.

Organizations working on small-scale pig cloning projects include the University of Missouri–Columbia; PPL Therapeutics PLC of Scotland (the company that helped clone Dolly the sheep); and the Department of Bio-technology, Institute of Farm Animal Genetics, Friedrich-Loeffler-Institute, Mariensee, Neustadt, Germany.

Xenotransplantation: Molecular roadblocks and Immunological Hurdles

Hyperacute rejection (HAR) occurs when a transplanted organ becomes inflamed, turns dark/blackish and cyanotic, and loses its ability to contract. The organ tissue never becomes vascularized. The organ is dead within minutes to hours of a transplant and the recipient dies quickly (FIGURE 17-10). This occurs in xenotransplantation because the blood vessels of pigs contain α-1,3-galactose, a carbohydrate epitope unique to pigs. The human immune system recognizes the α-1,3-galactose as foreign and rejects the pig carbohydrates by quickly destroying the newly transplanted organ. Naturally occurring antibodies and complement activation are directed against the transplanted organ tissues.

	Pigs	Apes	Monkeys
Physiology compared to humans	Similar	Nearly identical	Similar
Organ size compared to humans	Similar	Identical	Too small
Gestation	100 days	251–289 days	170–193 days
Progeny	10–18	1–2	1–2
Protection of species	No	Yes, very strong	Yes
Availability	Unlimited	None	Low
Transgenic animals	Done	Not yet	Done
Cloning	Done	Not yet	Done
Costs	Low	Very high	High
Microbiological risks	Low (can be grown in specified pathogen-free environments)	High	High
Information from Denner, J. 2003. “Porcine endogenous retroviruses (PERVs) and xenotransplantation: Screening for transmission in several clinical trials and in experimental models using non-human primates.” Ann Transplant 8(3):39–48. Only gold members can continue reading. Log In or Register to continue Share this: Click to share on Twitter (Opens in new window) Click to share on Facebook (Opens in new window) Like this: Like Loading... Related Related posts: Poxviruses Herpesviruses Epidemiology Host Resistance to Viral Infections Stay updated, free articles. Join our Telegram channel Join Tags: Understanding Viruses_gate Jul 29, 2017 | Posted by admin in MICROBIOLOGY | Comments Off on The History of Medicine, Clinical Trials, Gene Therapy, and Xenotransplantation Full access? Get Clinical Tree Get Clinical Tree app for offline access Get Clinical Tree app for offline access %d