Presentation at DPharm 2014

 

This past week, Dr. Jules T. Mitchel presented the current status of the eSource program at Target Health for the Disruptive Innovations meeting in Boston. This meeting is amust for all who think or want to think “outside of the box.“ Mark next year’s meeting on your calendars (Boston, September 9-11, 2015).

 

As of 14 September 2014, we have 18 ongoing or completed studies under 9 INDs and 2 IDEs. If all goes well with the trial results, a PMA submission will take place in Q4 2014 and an NDA submission in Q3 2015. As a full service eCRO, for all of these programs, we are providing data management and EDC services. For 2 of the programs we are also providing study management and monitoring, biostatistics and medical writing services. And for one program we are also providing full regulatory affairs services.

 

Five major studies include:

 

1. Men’s Health (Phase 3; 22 sites; 160 subjects)

2. Migraine (Phase 2; 40 sites; 400 subjects)

3. Autism (Phase 3; 25 sites; 300 subjects)

4. Alzheimer’s disease (Phase 3; 100 sites; 800 subjects)

5. Oncology (Pivotal Trial; 4 sites; 110 subjects)

 

Photos by Our Colleague, James Farley

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Pollinating Bee – ©JFarleyPhotography.com

 

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Pollinating Bee – ©JFarleyPhotography.com

 

ON TARGET is the newsletter of Target Health Inc., a NYC-based, full-service, contract research organization (eCRO), providing strategic planning, regulatory affairs, clinical research, data management, biostatistics, medical writing and software services to the pharmaceutical and device industries, including the paperless clinical trial.

 

For more information about Target Health contact Warren Pearlson (212-681-2100 ext. 104). For additional information about software tools for paperless clinical trials, please also feel free to contact Dr. Jules T. Mitchel or Ms. Joyce Hays. The Target Health software tools are designed to partner with both CROs and Sponsors. Please visit the Target Health Website, and if you like the weekly newsletter, ON TARGET, you’ll love the Blog.

 

Joyce Hays, Founder and Editor in Chief of On Target

Jules Mitchel, Editor

 

Test Your Knowledge of the Blood-Brain-Barrier

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The blood-brain barrier (BBB) is a highly selective permeability barrier that separates the circulating blood from the brain extracellular fluid (BECF) in the central nervous system (CNS). The BBB is formed by capillary endothelial 1) ___, which are connected by tight junctions with an extremely high electrical resistivity. The BBB allows the passage of water, some gases, and lipid soluble molecules by passive diffusion, as well as the selective transport of molecules such as glucose and amino acids that are crucial to neural function. On the other hand, the BBB may prevent the entry of lipophilic, potential neurotoxins by way of an active transport mechanism mediated by P-glycoprotein. Astrocytes are necessary to create the BBB. A small number of regions in the brain, including the circumventricular organs (CVOs), do not have a BBB.

 

The BBB occurs along all capillaries and consists of tight junctions around the capillaries that do not exist in normal circulation. Endothelial cells restrict the diffusion of microscopic objects (e.g., bacteria) and large or hydrophilic molecules into the cerebrospinal 2) ___ (CSF), while allowing the diffusion of small hydrophobic molecules (O2, CO2, hormones). Cells of the barrier actively transport metabolic products such as glucose across the barrier with specific proteins. This barrier also includes a thick basement membrane and astrocytic endfeet. This “barrier“ results from the selectivity of the tight junctions between endothelial cells in CNS vessels that restricts the passage of solutes. At the interface between blood and the brain, endothelial cells are stitched together by these tight junctions, which are composed of smaller subunits, frequently biochemical dimers, that are transmembrane proteins such as occludin, claudins, junctional adhesion molecule (JAM), or ESAM, for example. Each of these transmembrane proteins is anchored into the endothelial cells by another protein complex that includes zo-1 and associated proteins.

 

The BBB is composed of high-density cells restricting passage of substances from the 3) ___ much more than do the endothelial cells in capillaries elsewhere in the body. Astrocyte cell projections called astrocytic feet (also known as “glia limitans“) surround the endothelial cells of the BBB, providing biochemical support to those cells. The BBB is distinct from the quite similar blood-cerebrospinal-fluid barrier, which is a function of the choroidal cells of the choroid plexus, and from the blood-retinal barrier, which can be considered a part of the whole realm of such barriers. Several areas of the human brain are not on the brain side of the BBB. Some examples of this include the circumventricular organs, the roof of the third and fourth ventricles, capillaries in the pineal gland on the roof of the diencephalon and the pineal gland. The pineal gland secretes the hormone melatonin “directly into the systemic circulation“, thus 4) ___ is not affected by the BBB.

 

Originally, experiments in the 1920s showed that the BBB was still immature in newborns. The reason for this mistake was an error in methodology (the osmotic pressure was too high and the delicate embryonal capillary vessels were partially damaged). It was later shown in experiments with a reduced volume of the injected liquids that the markers under investigation could not pass the BBB. It was reported that those natural substances such as albumin, alpha-1-fetoprotein or transferrin with elevated plasma concentration in the 5) ___ could not be detected outside of cells in the brain. The transporter P-glycoprotein exists already in the embryonal endothelium. The measurement of brain uptake of acetamide, antipyrine, benzyl alcohol, butanol, caffeine, cytosine, diphenyl hydantoin, ethanol, ethylene glycol, heroin, mannitol, methanol, phenobarbital, propylene glycol, thiourea, and urea in ether-anesthetized newborns vs. adult rabbits shows that newborn rabbit and adult rabbit brain endothelia are functionally similar with respect to lipid-mediated permeability. These data confirmed no differences in permeability could be detected between newborn and adult BBB capillaries. No difference in brain uptake of glucose, amino acids, organic acids, purines, nucleosides, or choline was observed between adult and newborn rabbits. These experiments indicate that the newborn BBB has restrictive properties similar to that of the 6) ___. In contrast to suggestions of an immature barrier in young animals, these studies indicate that a sophisticated, selective BBB is operative at birth.

 

The BBB acts very effectively to protect the brain from many common bacterial infections. Thus, infections of the brain are very rare. Infections of the brain that do occur are often very serious and difficult to treat. Antibodies are too large to cross the blood-brain 7) ___, and only certain antibiotics are able to pass. In some cases, a pharmacologic agent has to be administered directly into the cerebrospinal fluid. However, drugs delivered directly to the CSF do not effectively penetrate into the brain tissue itself, possibly due to the tortuous nature of the interstitial space in the brain.

 

The BBB becomes more permeable during inflammation. This allows some antibiotics and phagocytes to move across the BBB. However, this also allows bacteria and viruses to infiltrate the BBB. An exception to the bacterial exclusion is the diseases caused by spirochetes, such as Borrelia, which causes Lyme disease, and Treponema pallidum, which causes syphilis. These harmful bacteria seem to breach the BBB by physically tunneling through the blood vessel walls. There are also some biochemical poisons that are made up of large molecules that are too big to pass through the BBB. This was especially important in more primitive times when people often ate contaminated food. Neurotoxins such as Botulinum in the food might affect peripheral nerves, but the BBB can often prevent such 8) ___ from reaching the central nervous system, where they could cause serious or fatal damage.

 

The BBB is formed by the brain capillary endothelium and excludes from the brain ~100% of large-molecule neurotherapeutics and more than 98% of all small-molecule drugs. Overcoming the difficulty of delivering therapeutic agents to specific regions of the brain presents a major challenge to treatment of most brain disorders. In its neuroprotective role, the BBB functions to hinder the delivery of many potentially important diagnostic and therapeutic agents to the brain. Therapeutic molecules and antibodies that might otherwise be effective in diagnosis and therapy do not cross the BBB in adequate amounts. Mechanisms for drug targeting in the brain involve going either “through“ or “behind“ the BBB. Modalities for drug delivery/Dosage form through the BBB entail its disruption by osmotic means; biochemically by the use of vasoactive substances such as bradykinin; or even by localized exposure to high-intensity focused ultrasound (HIFU). Other methods used to get through the BBB may entail the use of endogenous transport systems, including carrier-mediated transporters such as glucose and amino acid carriers; receptor-mediated transcytosis for insulin or transferrin; and the blocking of active efflux transporters such as p-glycoprotein. However, vectors targeting BBB transporters, such as the transferrin receptor, have been found to remain entrapped in brain endothelial cells of capillaries, instead of being ferried across the BBB into the cerebral parenchyma. Methods for drug 9) ___ behind the BBB include intracerebral implantation (such as with needles) and convection-enhanced distribution. Mannitol can be used in bypassing the BBB.

 

Nanotechnology may also help in the transfer of drugs across the BBB. Recently, researchers have been trying to build liposomes loaded with nanoparticles to gain access through the BBB. More research is needed to determine which strategies will be most effective and how they can be improved for patients with brain tumors. The potential for using BBB opening to target specific agents to brain tumors has just begun to be explored. Delivering drugs across the BBB is one of the most promising applications of nanotechnology in clinical neuroscience. Nanoparticles could potentially carry out multiple tasks in a predefined sequence, which is very important in the delivery of drugs across the BBB. A significant amount of research in this area has been spent exploring methods of nanoparticle-mediated delivery of antineoplastic drugs to tumors in the central nervous system. For example, radio labeled polyethylene glycol coated hexadecylcyanoacrylate nanospheres targeted and accumulated in a rat gliosarcoma. However, this method is not yet ready for clinical trials, due to the accumulation of the nanospheres in surrounding healthy tissue. It should be noted that vascular endothelial cells and associated pericytes are often abnormal in tumors and that the BBB may not always be intact in brain tumors. Also, the basement membrane is sometimes incomplete. Other factors, such as astrocytes, may contribute to the resistance of brain tumors to therapy.

 

Peptides are able to cross the BBB through various mechanisms, opening new diagnostic and therapeutic avenues. However, their BBB transport data are scattered in the literature over different disciplines, using different methodologies reporting different influx or efflux aspects. Therefore, a comprehensive BBB peptide database (Brainpeps) was constructed to collect the BBB data available in the literature. Brainpeps currently contains BBB transport information with positive as well as negative results. The database is a useful tool to prioritize peptide choices for evaluating different BBB responses or studying quantitative structure-property (BBB behavior) relationships of peptides. Because a multitude of methods have been used to assess the BBB behavior of compounds, we classified these methods and their responses. Moreover, the relationships between the different BBB transport methods have been clarified and visualized. Casomorphin is a heptapeptide and could be able to pass the BBB.

 

Paul Ehrlich, a Nobel Prize winning bacteriologist, was studying staining, a procedure that is used in many microscopic studies to make fine biological structures visible using chemical dyes. As Ehrlich injected some of these dyes (notably the aniline dyes that were then widely used), the dye stained all of the organs of some kinds of animals except for their brains. At that time, Ehrlich attributed this lack of staining to the brain simply not picking up as much of the dye. However, in a later experiment in 1913, the dye was injected into the cerebro-spinal fluids of animals’ brains directly. It was found that in this case the brains did become dyed, but the rest of the body did not. This clearly demonstrated the existence of some sort of compartmentalization between the two. At that time, it was thought that the blood vessels themselves were responsible for the barrier, since no obvious membrane could be found. The concept of the 10) ___-brain barrier (then termed hematoencephalic barrier) was proposed by a Berlin physician, Lewandowsky, in 1900. It was not until the introduction of the scanning electron microscope to the medical research fields in the 1960s that the actual membrane could be observed and proved to exist.

 

ANSWERS: 1) cells; 2) fluid; 3) bloodstream; 4) melatonin; 5) newborn; 6) adult; 7) barrier; 8) toxins; 9) delivery; 10) blood

 

Paul Ehrlich

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Paul Ehrlich in 1908

 

Paul Ehrlich (14 March 1854- 20 August 1915) was a German Jewish physician and scientist who worked in the fields of hematology, immunology, and chemotherapy. He invented the precursor technique to Gram staining bacteria. The methods he developed for staining tissue made it possible to distinguish between different type of blood cells, which led to the capability to diagnose numerous blood diseases. His laboratory discovered arsphenamine (Salvarsan), the first effective medicinal treatment for syphilis, thereby initiating and also naming the concept of chemotherapy. Ehrlich popularized the concept of a “magic bullet“. He also made a decisive contribution to the development of an antiserum to combat diphtheria and conceived a method for standardizing therapeutic serums. For providing a theoretical basis for immunology as well as for his work on serum valency, Ehrlich was awarded the Nobel Prize for Physiology or Medicine in 1908 together with Elie Metchnikoff. He was the founder and first director of what is now known as the Paul Ehrlich Institute.

 

Born 14 March 1854 in Strehlen near Breslau, Paul Ehrlich was the second child of Ismar and Rosa Ehrlich. His father was a distiller of liqueurs and the royal lottery collector in Strehelen, a town of some 5,000 inhabitants in the province of Lower Silesia, now in Poland. His grandfather Heymann Ehrlich had been a fairly well off distiller and tavern manager. Ismar Ehrlich was the leader of the local Jewish community. After elementary school, Paul attended the time-honored secondary school Maria-Magdalenen-Gymnasium in Breslau, where he met Albert Neisser, who later became a professional colleague. As a schoolboy (inspired by his cousin Karl Weigert who owned one of the first microtomes), he became fascinated by the process of staining microscopic tissue substances. He retained that interest during his subsequent medical studies at the universities of Breslau, Strasbourg, Freiburg im Breisgau and Leipzig. After obtaining his doctorate in 1882, he worked at the Charite in Berlin as an assistant medical director under Theodor Frerichs, the founder of experimental clinical medicine, focusing on histology, hematology and color chemistry (dyes). He married Hedwig Pinkus in 1883. The couple had two daughters, Stephanie and Marianne.

 

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Commemorative plaque at Bergstra?e 96 in Berlin-Steglitz, where Ehrlich lived and worked from 1890 to 1899

 

After completing his clinical education and habilitation at the prominent Charite medical school and teaching hospital in Berlin in 1886, Ehrlich traveled to Egypt and other countries in 1888 and 1889, one reason being to cure a case of tuberculosis, with which he had become infected in the laboratory. On his return he established a private medical practice and small laboratory in Berlin-Steglitz. In 1891, Ehrlich received a call from Robert Koch to join the staff at his Berlin Institute of Infectious Diseases, where in 1896 a new institute was established for Ehrlich’s specialization, the Institute for Serum Research and Testing (Institut fur Serumforschung und Serumprufung), whose director he became.

 

 

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Ehrlich’s grave in the Jewish cemetery on Rat-Beil-Strasse in Frankfurt am Main

 

In 1899 his institute moved to Frankfurt am Main and was renamed the Institute of Experimental Therapy (Institut fur experimentelle Therapie). One of his important collaborators there was Max Neisser. In 1906 Ehrlich became the director of the Georg Speyer House in Frankfurt, a private research foundation affiliated with his institute. Here he discovered in 1909 the first drug to be targeted against a specific pathogen: Salvarsan, a treatment for syphilis, which was at that time one of the most lethal and infectious diseases in Europe. Among the foreign guest scientists working with Ehrlich were two Nobel Prize winners, Henry Hallett Dale and Paul Karrer. The institute was renamed Paul Ehrlich Institute in Ehrlich’s honor in 1947. In 1914 Ehrlich signed the controversial Manifesto of the Ninety-Three which was a defense of Germany’s World War I politics and militarism. On 17 August 1915 Ehrlich suffered a heart attack and died on 20 August in Bad Homburg vor der Hohe. Wilhelm II the German emperor, wrote in a telegram of condolence, “I, along with the entire civilized world, mourn the death of this meritorious researcher for his great service to medical science and suffering humanity; his life’s work ensures undying fame and the gratitude of both his contemporaries and posterity“.Paul Ehrlich was buried at the Jewish cemetery on Rat-Beil-Strasse in Frankfurt am Main (Block 114 N).

 

In the early 1870s, Ehrlich’s cousin Karl Weigert was the first person to stain bacteria with dyes and to introduce aniline pigments for histological studies and bacterial diagnostics. During his studies in Strassburg under the anatomist Heinrich Wilhelm Waldeyer, Ehrlich continued the interest started by his cousin in pigments and staining tissues for microsopic study. He spent his eighth university semester in Freiburg im Breisgau investigating primarily the red dye dahlia (monophenylrosanilin), giving rise to his first publication. In 1878 he followed his dissertation supervisor Julius Friedrich Cohnheim to Leipzig and that year obtained a doctorate with a dissertation entitled “Contributions to the Theory and Practice of Histological Staining“ (Beitraege zur Theorie und Praxis der histologischen Faerbung).

 

 

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Photo of cultured mast cells at 100X stained with Tol Blue

 

One of the most outstanding results of his dissertation investigations was the discovery of a new cell type. Ehrlich discovered in the protoplasm of supposed plasma cells a granulate which could be made visible with the help of an alkaline dye. He thought this granulate was a sign of good nourishment and accordingly named these cells mast cells, (from the German word for an animal-fattening feed, Mast). This focus on chemistry was unusual for a medical dissertation. In it Ehrlich presented the entire spectrum of known staining techniques and the chemistry of the pigments employed. While he was at the Charite Ehrlich elaborated the differentiation of white blood cells according to their different granules. A precondition was a dry specimen technique, which he also developed. A drop of blood placed between two glass slides and heated over a Bunsen burner fixed the blood cells but enabled them still to be stained. Ehrlich used both alkaline and acid dyes, and also created new, “neutral“ dyes. For the first time this made it possible to differentiate the lymphocytes among the leucocytes (white blood cells). By studying their granulation he could distinguish between nongranular lymphocytes, mono- and poly-nuclear leucocytes, eosinophil granulocytes, and mast cells.

 

Starting in 1880 Ehrlich also studied red blood cells. He demonstrated the existence of nucleated red blood cells, which he subdivided into normoblasts, megaloblasts, microblasts and poikiloblasts; he had discovered the precursors of erythrocytes. Ehrlich thus also laid the basis for the analysis of anemias, after he had created the basis for systematizing leukemias with his investigation of white blood cells. His duties at the Charite included analyzing patients’ blood and urine specimens. In 1881 he published a new urine test which could be used to distinguish various types of typhoid from simple cases of diarrhea. The intensity of staining made possible a disease prognosis. The pigment solution he used is known today as Ehrlich’s reagent. Ehrlich’s great achievement, but also a source of problems during his further career, was that he had initiated a new field of study interrelating chemistry, biology and medicine. Much of his work was rejected by the medical profession, which lacked the requisite chemical knowledge. It also meant that there was no suitable professorship in sight for Ehrlich.

 

When a student in Breslau, the pathologist Julius Friedrich Cohnheim gave Ehrlich an opportunity to conduct extensive experiments, and also introduced him to Robert Koch, who was at the time a district physician in Wollstein, Posen Province. In his spare time, Koch had clarified the life cycle of the anthrax pathogen and contacted Ferdinand Cohn who was quickly convinced by Koch’s work and introduced him to his Breslau colleagues. From 30 April to 2 May 1876 Koch presented his investigations in Breslau, which the student Paul Ehrlich was able to experience. On 24 March 1882 Ehrlich was present when Robert Koch, working since 1880 at the Imperial Public Health Office (Kaiserliches Gesundheitsamt) in Berlin, presented the lecture in which he reported how he was able to identify the tuberculosis pathogen. Ehrlich later described this lecture as his “greatest experience in science“. Already the day after Koch’s lecture Ehrlich had made an improvement to Koch’s staining method, which Koch unreservedly welcomed. From this date on, the two men were bound in friendship. In 1887 Ehrlich became an unsalaried lecturer in internal medicine (Privatdozent fur Innere Medizin) at Berlin University, and in 1890 took over the tuberculosis station at a public hospital in Berlin-Moabit at Koch’s request. This was where Koch’s alleged tuberculosis therapeutic agent tuberculin was under study, and Ehrlich also had himself injected with it. (Eventually, Paul Ehrlich overcame the tuberculosis disease, with which he was infected in Egypt.) In the ensuing tuberculin scandal, Ehrlich tried to support Koch and stressed the value of tuberculin for diagnostic purposes. In 1891 Koch invited Ehrlich to work at the newly founded Institute of Infectious Diseases (Institut fur Infektionskrankheiten – now the Robert Koch Institute) at Friedrich-Wilhelms-Universit?t (now Humboldt University) in Berlin. Koch was unable to give him any remuneration, but did offer him full access to laboratory staff, patients, chemicals and laboratory animals, which Ehrlich always remembered with gratitude.

 

Ehrlich had started his first experiments on immunization already in his private laboratory. He accustomed mice to the poisons ricin and abrin. After feeding them with small but increasing dosages of ricin he ascertained that they had become “ricin-proof“. Ehrlich interpreted this as immunization and observed that it was abruptly initiated after a few days and was still in existence after several months, but mice immunized against ricin were just as sensitive to abrin as untreated animals. This was followed by investigations on the “inheritance“ of acquired immunity. It was already known that in some cases after a smallpox or syphilis infection, specific immunity was transmitted from the parents to their offspring. Ehrlich rejected inheritance in the genetic sense because the offspring of a male mouse immunized against abrin and an untreated female mouse were not immune to abrin. He concluded that the fetus was supplied with antibodies via the pulmonary circulation of the mother. This idea was supported by the fact that this “inherited immunity“ decreased after a few months. In another experiment he exchanged the offspring of treated and untreated female mice. The mice which were nursed by the treated females were protected from the poison, providing the proof that antibodies can also be conveyed in breast milk. Ehrlich also researched autoimmunity, calling it “horror autotoxicus“.

 

Emil Behring had worked at the Berlin Institute of Infectious Diseases until 1893 on developing an antiserum for treating diphtheria and tetanus but with inconsistent results. Koch suggested that Behring and Ehrlich cooperate on the project. This joint work was successful to the extent that Ehrlich was quickly able to increase the level of immunity of the laboratory animals based on his experience with mice. Clinical tests with diphtheria serum early in 1894 were successful and in August the chemical company Hoechst started to market Behring’s “Diphtheria Remedy synthesized by Behring-Ehrlich“. The two discoverers had originally agreed to share any profits after the Hoechst share had been subtracted. Their contract was changed several times and finally Ehrlich was eventually pressured into accepting a profit share of only 8%. Ehrlich resented what he considered as unfair treatment and his relationship with Behring was thereafter problematic, a situation which later escalated over the issue of the valency of tetanus serum. Ehrlich recognized that the principle of serum therapy had been developed by Behring and Kitasato. But he was of the opinion that he had been the first to develop a serum which could also be used on humans and that his role in developing the diphtheria serum had been insufficiently acknowledged. Behring on his part schemed against Ehrlich at the Prussian Ministry of Culture, and from 1900 Ehrlich refused to collaborate with him. Only von Behring received the first Nobel Prize in Medicine, in 1901, for contributions to research on diphtheria.

 

Since antiserums were an entirely new type of medicine whose quality was highly variable, a government system was established to guarantee their safety and effectiveness. From 1 April 1895 only government-approved serum could be sold in the German Reich. The testing station for diphtheria serum was provisionally housed at the Institute of Infectious Diseases. On the initiative of Friedrich Althoff, an Institute of Serum Research and Testing (Institut fur Serumforschung und Serumprufung) was established in 1896 in Berlin-Steglitz, with Paul Ehrlich as director (which required him to cancel all his contracts with Hoechst). In this function and as honorary professor at Berliner University he had annual earnings of 6000 marks, approximately the salary of a university professor. In addition to a testing department the institute also had a research department. In order to determine the effectiveness of diphtheria antiserum, an invariable concentration of diphtheria toxin was required. Ehrlich discovered that the toxin being used was perishable, in contrast to what had been assumed, which for him led to two consequences: He did not use the toxin as a standard, but instead a serum powder developed by Behring, which had to be dissolved in liquid shortly before use. The strength of a test toxin was first determined in comparison with this standard. The test toxin could then be used as a reference for testing other serums. For the test itself, toxin and serum were mixed in a ratio so that their effects just cancelled each other when injected into a guinea pig. But as there was a large margin in determining whether symptoms of illness were present, Ehrlich established an unambiguous target: the death of the animal. The mixture was to be such that the test animal would die after four days. If it died earlier, the serum was too weak and was rejected. Ehrlich claimed to have made the determination of the valency of serum as accurate as it would be with chemical titration. This again demonstrates his tendency to quantify the life sciences.

 

Influenced by the mayor of Frankfurt am Main, Franz Adickes, who endeavored to establish science institutions in Frankfurt in preparation of the founding of a university, Ehrlich’s institute moved to Frankfurt In 1899 and was renamed the Royal Prussian Institute of Experimental Therapy (Koniglich Preussisches Institut fur Experimentelle Therapie). The German quality-control methodology was copied by government serum institutes all over the world, and they also obtained the standard serum from Frankfurt.

 

After diphtheria antiserum, tetanus serum and various bactericide serums for use in veterinary medicine were developed in rapid sequence. These were also evaluated at the institute, as was tuberculin and later on various vaccines. Ehrlich’s most important colleague at the institute was the Jewish doctor and biologist Julius Morgenroth.

 

 

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Paul Ehrlich around 1900 in his Frankfurt office

 

This research inspired Ehrlich in 1897 to develop his famous side-chain theory. As he saw it, the reaction between a toxin and the operative components of a serum is a chemical reaction. He explained the toxic effect using the example of tetanus toxin. He postulated that cell protoplasm contains special structures which have chemical side chains (today’s term is macromolecules) to which the toxin binds, affecting function. If the organism survives the effects of the toxin, the blocked side-chains are replaced by new ones. This regeneration can be trained, the name for this phenomenon being immunization. If the cell produces a surplus of side chains, these might also be released into the blood as antibodies. In the following years Ehrlich expanded his side chain theory using concepts (“amboceptors,“ “receptors of the first, second and third order“, etc.) which are no longer customary. Between the antigen and the antibody he assumed there was an additional immune molecule, which he called an “additive“ or a “complement“. For him, the side chain contained at least two functional groups.

 

In 1901 the Prussian Ministry of Finance criticized Ehrlich for exceeding his budget and as a consequence reduced his income. In this situation Althoff arranged a contact with Georg Speyer, a Jewish philanthropist and joint owner of the bank house Lazard Speyer-Ellissen. The cancerous disease of Princess Victoria, the widow of the German Emperor Friedrich II, had received much public attention and prompted a collection among wealthy Frankfurt citizens, including Speyer, in support of cancer research. Ehrlich had also received from the German Emperor Wilhelm II a personal request to devote all his energy to cancer research. Such efforts led to the founding of a department for cancer research affiliated with the Institute of Experimental Therapy; the chemist Gustav Embden, among others worked there. Ehrlich informed his sponsors that cancer research meant basic research, and that a cure could not be expected soon. Among the results achieved by Ehrlich and his research colleagues was the insight that when tumors are cultivated by transplanting tumor cells, their malignancy increases from generation to generation. If the primary tumor is removed, then metastasis precipitously increases. Ehrlich applied bacteriological methods to cancer research. In analogy to vaccination he attempted to generate immunity to cancer by injecting weakened cancer cells. Both in cancer research and chemotherapy research (see below) he introduced the methodologies of Big Science. In 1885 Ehrlich?s monograph “The Need of the Organism for Oxygen“, appeared, which he also submitted as a habilitation thesis. In it he introduced the new technology of in vivo staining. One of his findings was that pigments can only be easily assimilated by living organisms if they are in granular form. He injected the dyes alizarin blue and indophenol blue into laboratory animals and established that after their death various organs had been colored to different degrees. In organs with high oxygen saturation indophenol was retained; in organs with medium saturation indophenol was reduced, but not Alizarin blue. And in areas with low oxygen saturation both pigments were reduced. With this work Ehrlich also formulated the conviction which guided his research: that all life processes can be traced to processes of physical chemistry occurring in the cell.

 

Methylene blue

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Staining in vivo with methylene blue of a cell from the mucous membrane of a human mouth

 

In the course of his investigations Ehrlich came across methylene blue, which he regarded as particularly suitable for staining bacteria (later, Robert Koch also used methylene blue as a dye in his research on the tuberculosis pathogen). In Ehrlich’s view, an added benefit was that methylene blue also stained the long appendages of nerve cells, the axons. He initiated a doctoral dissertation on the subject, but did not follow up the topic himself. It was the opinion of the neurologist Ludwig Edinger that Ehrlich had thereby opened up a major new topic in the field of neurology. After mid-1889, when Ehrlich was unemployed, he privately continued his research on methylene blue. His work on in vivo staining gave him the idea of using it therapeutically. Since the parasite family of Plasmodiidae – which includes the malaria pathogen – can be stained with methylene blue, he thought it could possibly be used in the treatment of malaria. In the case of two patients so treated at the city hospital in Berlin-Moabit, their fever indeed subsided and the malaria plasmodia disappeared from their blood. Ehrlich obtained methylene blue from the company Meister Lucius & Bruning AG (later renamed Hoechst AG), which started a long collaboration with this company. Already before the Institute of Experimental Therapy had moved to Frankfurt, Ehrlich had resumed work on methylene blue. After the death of Georg Speyer, Speyer’s widow Franziska Speyer, desired to do something in his memory and endowed the Georg-Speyer House, which was erected next door to Ehrlich’s institute. As director of the Georg Speyer House Ehrlich transferred his chemotherapeutic research there. He was looking for an agent which was as good as methylene blue but without its side effects. His model was on the one hand the impact of quinine on malaria, and on the other hand, in analogy to serum therapy, he thought there must also be chemical pharmaceuticals which would have just as specific an effect on individual diseases. His goal was to find a “Therapia sterilisans magna“, in other words a treatment that could kill all disease pathogens.

 

 

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Dr. Paul Ehrlich and Dr. Sahachiro Hata

 

As a model for experimental therapy Ehrlich used a guinea pig trypanosoma disease and tested out various chemical substances on laboratory animals. The trypanosomes could indeed be successfully killed with the dye trypan red. From 1906, he intensively investigated atoxyl and had it tested by Robert Koch along with other arsenic compounds during Koch’s sleeping sickness expedition of 1906/07. Although the name literally means “nonpoisonous“, atoxyl damages especially the optic nerve. Ehrlich elaborated the systematic testing of chemical compounds in the sense of “screening“, as now practiced in the pharmaceutical industry. He discovered that Compound 418, Arsenophenylglycine, had an impressive therapeutic effect and had it likewise tested in Africa. With the support of his assistant Sahachiro Hata, Ehrlich discovered in 1909 that Compound 606, Arsphenamine effectively combatted “spirillum“ spirochaetes bacteria, one of whose subspecies causes syphilis. The compound proved to have few side effects in human trials, and the spirochetes disappeared in seven syphilis patients after this treatment. After extensive clinical testing (all the participants had the negative example of tuberculin in mind) the Hoechst company began to market the compound toward the end of 1910 under the name “Salvarsan“. This was the first agent with a specific therapeutic effect to be created on the basis of theoretical considerations. Salvarsan proved to be amazingly effective, particularly when compared with the conventional therapy of mercury salts. Manufactured by Hoechst AG, Salvarsan became the most widely prescribed drug in the world. It was the most effective drug for treating syphilis until penicillin became available in the 1940s. Salvarsan required improvement as to side effects and solubility and was replaced in 1911 with Neosalvarsan. Ehrlich’s work illuminated the existence of the BBB.

 

The medication triggered off the so-called “Salvarsan war“. On the one hand there was hostility on the part of those who feared a resulting moral break down of sexual inhibitions. Ehrlich was also accused, with clearly anti-Semitic undertones, of excessively enriching himself. In addition, Ehrlich’s associate, Paul Uhlenhuth claimed priority in discovering the drug. Because some people died during the clinical testing, he was even accused of “stopping at nothing“. In 1914, one of the most prominent of the accusers was convicted of criminal libel at a trial for which Ehrlich was called to testify. Though Ehrlich was exonerated thereby, the ordeal threw him into a depression from which he never recovered. Ehrlich reasoned that if a compound could be made that selectively targeted a disease-causing organism, then a toxin for that organism could be delivered along with the agent of selectivity. Hence, a “magic bullet“ (magische Kugel, his term for an ideal therapeutic agent) would be created that killed only the organism targeted. The concept of a “magic bullet“ was to some extent realized by the invention of monoclonal antibodies as they provide a very specific binding affinity.

 

 

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West German postage stamp (1954) commemorating Paul Ehrlich and Emil von Behring

 

Already in 1910, a street was named after Ehrlich in Frankfurt-Sachsenhausen. He was honored by the Prussian government in 1911, when he was elected to the highest rank of Excellency in the Privy Medical Council.

 

During the Third Reich, Ehrlich’s achievements were ignored while Emil Adolf von Behring was stylized as the ideal Aryan scientist, and the street named after Ehrlich was given another name. Shortly after the end of the war the name Paul-Ehrlich-Strasse was reinstated and today numerous German cities have streets named after Paul Ehrlich. West Germany issued a postage stamp in 1954 on the 100th anniversary of the births of Paul Ehrlich (14 March 1854) and Emil von Behring (15 March 1854). A 200 Deutsche Mark bank note featured Paul Ehrlich.

 

 

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1996 series 200 Deutsche Mark banknote

 

His name is also borne by many schools and pharmacies, by the Paul-Ehrlich-Gesellschaft fur Chemotherapie e. V. (PEG) in Frankfurt am Main, and the Paul-Ehrlich-Klinik in Bad Homburg vor der Hoehe. The Paul Ehrlich and Ludwig Darmstaedter Prize is the most distinguished German award for biomedical research. A European network of PhD studies in Medicinal Chemistry has been named after him (Paul Ehrlich MedChem Euro PhD Network). The Anti-Defamation League awards a Paul Ehrlich-Gunther K. Schwerin Human Rights Prize and a crater of the moon was named after Paul Ehrlich in 1970. Ehrlich’s life and work was featured in the 1940 U.S. film Dr. Ehrlich’s Magic Bullet with Edward G. Robinson in the title role. It focused on Salvarsan (arsphenamine, “compound 606“), his cure for syphilis. Since the Nazi government was opposed to this tribute to a Jewish scientist, attempts were made to keep the film a secret in Germany.

 

Eating Habits, Body Fat Related to Differences in Brain Chemistry

 

According to the Centers for Disease Control and Prevention, with more than one-third of U.S. adults being obese, obesity-related conditions now include heart disease, type 2 diabetes and certain types of cancer, some of the leading causes of preventable death. Putting this together with the fact that dopamine, a chemical messenger in the brain influences reward, motivation and habit formation, this is starting to reveal an intriguing story.

 

The obesity epidemic is believed to be driven by a food environment that promotes consumption of inexpensive, convenient, high-calorie, palatable foods. Individual differences in obesity susceptibility or resistance to weight loss may arise because of alterations in the neurocircuitry supporting food reward and eating habits. In particular, dopamine signaling in the ventromedial striatum is thought to encode food reward and motivation, whereas dopamine in the dorsal and lateral striatum orchestrates the development of eating habits.

 

As a result, a study published online in Molecular Psychiatry (9 September 2014), measured striatal dopamine D2-like receptor binding potential (D2BP) using positron emission tomography (PET) with [18F]fallypride in 43 human subjects with body mass indices (BMI) ranging from 18 to 45?kg?m-2. The Pet scans evaluated the sites in the brain where dopamine was able to act. Study participants followed the same eating, sleeping and activity schedule. Tendency to overeat in response to triggers in the environment was determined from a detailed questionnaire.

 

Results showed that opportunistic eating behavior and BMI were both positively associated with D2BP in the dorsal and lateral striatum, whereas BMI was negatively associated with D2BP in the ventromedial striatum. These results suggest that obese people have alterations in dopamine neurocircuitry that may increase their susceptibility to opportunistic overeating while at the same time making food intake less rewarding, less goal directed and more habitual. Whether or not the observed neurocircuitry alterations pre-existed or occurred as a result of obesity development, they may perpetuate obesity given the omnipresence of palatable foods and their associated cues.

 

The study did not demonstrate cause and effect among habit formation, reward, dopamine activity, eating behavior and obesity. Future research will examine dopamine activity and eating behavior in people over time as they change their diets, physical activity, and their weight.

 

Plugging Into a Learning Brain

 

The human brain contains nearly 86 billion neurons, which communicate through intricate networks of connections. Understanding how they work together during learning can be challenging. Brain-computer interfaces seek to turn thoughts into action. With small surgically implanted electrodes, researchers can simultaneously monitor the electrical activity of hundreds of neurons. A computer converts the signals into commands to move an external device, such as a robotic arm or a computer cursor. Brain-computer interfaces are being developed to help paralyzed patients as well as to study the function of healthy brains.

 

According to a study performed in monkeys and published online in Nature (28 August 2014), learning is easier when it only requires nerve cells to rearrange existing patterns of activity than when the nerve cells have to generate new patterns. The study combined two innovative technologies, brain-computer interfaces and machine learning, to study patterns of activity among neurons in monkey brains as the animals learned to use their thoughts to move a computer cursor.

 

For the study, the research team used brain-computer interfaces in two animals to examine learning in the motor cortex, a part of the brain that controls movement. The firing patterns of the neurons they recorded were used to control a computer cursor. As the animals learned to move the cursor to a designated spot on the monitor, the computer used machine learning to map brain cell activity to cursor movement. Machine learning is a method of programming a computer to learn and constantly adjust its commands based on previous data or experience. In this case, it created a feedback loop between the animal and the computer, which improved the animal’s ability to use its thoughts to move the cursor.

 

At first, the authors noticed that the ensemble of neurons recorded in each animal had a small set of natural, or favored, firing patterns that were used to move the cursor, which they called the “intrinsic manifold.“ After determining the intrinsic manifold, the team reprogrammed the map between neural activity and cursor movement. For instance, if a firing pattern originally caused the cursor to move to the top of the screen, then the interface would move the cursor to the bottom. The team then observed whether the animals could learn to generate the appropriate neural activity patterns to compensate for the changes. According to the authors, it’s as if a computer mouse was turned upside down in a person’s hand and asked him to click on an icon, except the mouse is entirely within the subject’s brain.

 

The authors discovered that the monkeys easily relearned how to move the cursor if they could use patterns within the intrinsic manifold in new ways. In contrast, learning was more difficult when the interface required patterns of neural activity that were outside of the intrinsic manifold. According to the authors, it appears that the brain sets constraints on the speed with which we learn new things and that by characterizing those constraints might enable us to predict which skills will be quicker to learn, and which might take longer. The authors speculated that, for humans, thinking outside the box requires more difficult changes in neural activity.

 

FDA Approves Contrave for Weight-Management

 

BMI, which measures body fat based on an individual’s weight and height, is used to define the obesity and overweight categories. According to the Centers for Disease Control and Prevention, more than one-third of adults in the United States are obese.

 

The FDA has approved Contrave (naltrexone hydrochloride and bupropion hydrochloride extended-release tablets) as treatment option for chronic weight management in addition to a reduced-calorie diet and physical activity. The drug is approved for use in adults with a body mass index (BMI) of 30 or greater (obesity) or adults with a BMI of 27 or greater (overweight) who have at least one weight-related condition such as high blood pressure (hypertension), type 2 diabetes, or high cholesterol (dyslipidemia).

 

Contrave is a combination of two FDA-approved drugs, naltrexone and bupropion, in an extended-release formulation. Naltrexone is approved to treat alcohol and opioid dependence. Bupropion is approved to treat depression and seasonal affective disorder and as an aid to smoking cessation treatment. The effectiveness of Contrave was evaluated in multiple clinical trials that included approximately 4,500 obese and overweight patients with and without significant weight-related conditions treated for one year. All patients received lifestyle modification that consisted of a reduced- calorie diet and regular physical activity.

 

Results from a clinical trial that enrolled patients without diabetes showed that patients had an average weight loss of 4.1% over treatment with placebo (inactive pill) at one year. In this trial, 42% of patients treated with Contrave lost at least 5% of their body weight compared with 17% of patients treated with placebo. Results from another clinical trial that enrolled patients with type 2 diabetes showed that patients had an average weight loss of 2% over treatment with placebo at one year. In this trial, 36% of patients treated with Contrave lost at least 5% of their body weight compared with 18% of patients treated with placebo.

 

Patients using Contrave at the maintenance dose should be evaluated after 12 weeks to determine if the treatment is working. If a patient has not lost at least 5% of baseline body weight, Contrave should be discontinued, as it is unlikely that the patient will achieve and sustain clinically meaningful weight loss with continued treatment. Because it contains bupropion, Contrave has a boxed warning to alert health care professionals and patients to the increased risk of suicidal thoughts and behaviors associated with antidepressant drugs. The warning also notes that serious neuropsychiatric events have been reported in patients taking bupropion for smoking cessation.

 

Contrave can cause seizures and must not be used in patients who have seizure disorders. The risk of seizure is dose-related. Contrave should be discontinued and not restarted in patients who experience a seizure while being treated with Contrave. Contrave can also raise blood pressure and heart rate and must not be used in patients with uncontrolled high blood pressure. The clinical significance of the increases in blood pressure and heart rate observed with Contrave treatment is unclear, especially for patients with heart-related and cerebrovascular (blood vessel dysfunction impacting the brain) disease, since patients with a history of heart attack or stroke in the previous six months, life-threatening arrhythmias, or congestive heart failure were excluded from the clinical trials. Blood pressure and pulse should be measured prior to starting the drug and should be monitored at regular intervals, particularly among patients with controlled high blood pressure prior to treatment.

 

Other products containing bupropion should not be taken along with Contrave. The drug should not be used in patients who have eating disorders (bulimia or anorexia nervosa). Contrave should also not be taken by patients who are using opioids or treatments for opioid dependence, or who are experiencing acute opiate withdrawal. Patients undergoing an abrupt discontinuation of alcohol, benzodiazepines, barbiturates and antiepileptic drugs should not take Contrave. Women who are pregnant or trying to become pregnant should not take Contrave.

 

The most common adverse reactions reported with Contrave include nausea, constipation, headache, vomiting, dizziness, insomnia, dry mouth, and diarrhea.

 

The FDA is requiring the following post-marketing requirements:

  • a cardiovascular outcomes trial to assess the cardiovascular risk associated with Contrave use;
  • two efficacy, safety, and clinical pharmacology studies in pediatric patients (one in patients 12 to 17 years of age, and one in patients 7 to 11 years of age);
  • a nonclinical (animal) juvenile toxicity study with a particular focus on growth and development as well as behavior, learning, and memory;
  • a study to evaluate the effect of Contrave on cardiac conduction;
  • clinical trials to evaluate dosing in patients with hepatic or renal impairment;
  • a clinical trial to evaluate the potential for interactions between Contrave and other drugs.

 

Contrave is distributed by Takeda Pharmaceuticals America Inc. of Deerfield, Illinois for Orexigen Therapeutics, Inc. of La Jolla, California. Orexigen licensed North American Contrave rights to Takeda Pharmaceuticals.

 

Creamy Blueberry Angel

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If you like your healthy blueberries – melt-in-your-mouth-gooey – then this is the dessert for you! Plus, this is a low calorie, dessert, especially if you make your own angel food cake and don’t use a store bought mix. ©Joyce Hays, Target Health Inc.

 

Ingredients

Blueberry Filling:

12 ounces frozen blueberries

2 Tablespoons granulated Splenda

2 Tablespoons cornstarch

1/4 cup cold water

Squeeze of fresh lemon juice (about 1/2 tablespoon)

 

Cake-Cut-Into-Cubes and Cream: 

Baked, cooled and cubed angel food cake

16 ounces Tofutti, softened to room temperature

2/3 cup almond milk or evaporated milk

2/3 cup granulated Splenda

 

Whipped Cream Topping:

1 1/2 cups fat-free Cool Whip

2 teaspoons powdered Splenda, or 2 packets

 

Directions

First make the vanilla angel food cake. See the directions below. While baking and then, while cake is cooling off, make the other parts of this dessert. If you use a store bought mix or buy a ready-made angel food cake, the calories will be much higher.

 

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Stirring the blueberry mixture. This whole recipe is very easy. ©Joyce Hays, Target Health Inc.

 

For the blueberry filling: in a medium saucepan, combine the blueberries, sugar, cornstarch, water and lemon juice. Bring the mixture to a simmer and cook until thickened, 5-7 minutes, stirring often. Remove from the heat and let cool to room temperature.

 

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Here, I’m about to add the cubes to the creamy mixture. ©Joyce Hays, Target Health Inc.

 

For the cake and cream layer: In a blender or with an electric mixer (handheld or stand mixer), whip together the cream cheese, half-and-half or evaporated milk and sugar until smooth and creamy. Transfer the mixture to a bowl if you used a blender.

 

Now, fold in the angel food cake cubes. Keep in mind (from the note above) that you may not use all the cake cubes, especially if using an angel food cake mix. Add cake cubes until they are all thickly coated with a layer of cream. If making this in advance, it can dry out if there is too much angel food cake added.

 

For the Cool Whip Topping: Whisk together the Cool Whip and powdered sugar until fully incorporated. Set aside in fridge until ready to use.

 

 

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This is the very first layer of the cake cubes, slathered on all sides, with the creamy mixture. ©Joyce Hays, Target Health Inc.

 

To Assemble: in a trifle dish or in a large glass bowl, spread half of the angel food cake mixture as bottom layer. Top with half of the blueberries, spreading evenly across, and then spread half of the Cool Whip mixture. Repeat the layers a second time.

 

Cover and refrigerate at least 2 hours or up to 24 hours. Serve chilled in pretty dessert dishes or on a dessert plate with a few fresh blueberries as garnish.

 

 

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A total of 4 layers have already been done here. This is going to be the 2nd layer of the blueberry mixture about to be smoothed over the 2nd layer of creamy cake cubes. On top of these blueberries will be the last layer of Cool Whip. Then this bowl goes into the fridge for at least 2 hours or overnight.

 

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©Joyce Hays, Target Health Inc.

 

Here’s the bowl after 2 hours in fridge, with two people dining on dessert.   LOL BTW, this cake gets even better with time. Also, the next time I make this recipe, I’m gonna try putting the final layers into a square glass container, so that after left in the fridge for several days, I can cut it like a layer cake.

 

Vanilla Angel Food Cake Recipe

 

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I used a square cake pan. Here the cake has been cooled and cut into cubes. ©Joyce Hays, Target Health Inc.

 

Vanilla Version

Ingredients

Dry ingredients:

1 cup almond flour

3/4 cup granulated Splenda

1/2 teaspoon salt

For the egg white mixture:

3/4 cup granulated Splenda

12 large egg whites (make sure not to get any of the shell or egg yolk in with the whites or they won’t beat-to-peak properly)

1 teaspoon vanilla extract

1 1/2 teaspoon cream of tartar

 

DIRECTIONS

Preheat the oven to 325 degrees.

 

In a medium bowl, whisk together the flour, cocoa powder (if using), sugar and salt and set aside.

 

In another bowl place the egg whites and add the vanilla. With a hand mixer (or with a stand mixer), beat the egg whites and vanilla on medium-high until the mixture is just frothy, about one minute.

 

Sprinkle the cream of tartar on the top of the foamy egg whites and continue beating on medium-high until soft peaks form, another 2-3 minutes.

 

To the egg whites, add the sugar 1/4 cup at a time until fully incorporated. Continue beating until the whites are stiff and glossy. This may take several minutes, depending on the type of mixer you are using.

 

Now, with a whisk, gently fold the dry ingredients into the beaten egg whites. Pour the batter evenly into an ungreased angel food cake pan and smooth the top with a rubber spatula. Place the cake on a rack in the center of the oven and bake for 40-45 minutes, until the top of the cake is golden brown and the cake springs back when lightly touched and the cracks are dry to the touch.

 

Place the cake (still in the cake pan) upside down on cooking rack, to cool.

 

When cake is cool, slide a knife around the edges of the pan and gently remove the cake.

 

After removing cake from pan, allow it to cool even more. When cake is cool to your touch, cut the whole cake into cubes, about 1 inch by 1 inch. You will be using these cake cubes in the Tofutti part of the recipe.

 

Be careful with the cream/cake mixture – only add cake cubes while they are all evenly coated with a thick layer of cream. Too many cake cubes and it might dry out (especially if it is made in advance). Speaking of making it ahead, this can be assembled and refrigerated up to 24 hours in advance.

 

Note: To make a chocolate version, substitute 1/4 cup cocoa powder for 1/4 cup of the flour.

 

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©Joyce Hays, Target Health Inc.

 

He had just come back from a meeting in DC and had a veggie burger on the train. He was tired and not hungry. We poured some wine. He wanted red, I wanted white. I put dinner away and pulled the Blueberry Angel Dessert out of the fridge. We dined on dessert.

 

Any greater proof of the (so-called) pudding?

 

We polished off the two dessert dishes in photo above (top of this section) and then dug into the bowl.

 

A cautionary note: because this whole evening was play-as-you-go, no attention was paid to choosing the wine. As a result, both wines did not “go“ with the dessert. If there had been some semblance of a plan, we would have had a liqueur like Maraschino or Amaretto. But we had each other, and that was what really mattered.

 

 

From Our Table to Yours!

 

Bon Appetit !