Tuesday, February 4, 2020

The Biology of the Coronavirus

The global spread of viral pneumonia associated with the so-called “Wuhan coronavirus” appears to be reaching pandemic proportions. Given this distressing reality, it is important to more fully understand the biology of this virus.

Viruses represent a class of infectious agents that pose interesting challenges as witnessed by the HIV/AIDS virus that is the causative agent of the devastating acquired immunodeficiency syndrome (AIDS) that targets a particularly important cell type in the human adaptive immune system – the so-called, “T-helper cells (CD4).” Viruses possess the unusual property of being inert when outside a living cell. However, once they gain access to a living cell, the infective agent – either ribonucleic acid (RNA) or deoxyribonucleic acid (DNA) - commandeers the host cellular machinery to make copies of itself. This process can lead either to the ultimate death of the target cell or can result in a transformation of the host DNA that in some instances can lead to cancer – T-cell leukemia (HTLV-1 virus) and Cervical cancer (Human Papilloma virus – HPV) being some important examples.

Coronaviruses are a family of enveloped, single-stranded, positive-strand RNA viruses that are classified within the Nidovirales order. The name coronavirus was coined on account of the corona-like appearance of this virus as viewed under the electron microscope (See accompanying image). The existence of this type of virus was first reported in 1949 and the molecular mechanisms of both replication and disease formation was well-studied in the 1970s. The coronavirus family comprises pathogens that infect many animal species. Coronaviruses have been shown to be the causative agents for acute and chronic respiratory, enteric and central nervous system (CNS) diseases. This family of viruses have been associated with infectious disease in mouse (murine), a pig (porcine) transmissible gastroenteritis (TGEV), cow (bovine) and a bird (avian) bronchitis (BCoV). The first example of a potentially life-threatening human emerging coronavirus was the acute respiratory syndrome coronavirus (SARS-CoV).

Until 2003 the human coronavirus was only known to produce cold-like symptoms. This changed with the onset of severe acute respiratory syndrome (SARS) and now the Wuhan coronavirus that is apparently spread readily from human to human. These kinds of changes in infectivity are not unusual in the evolution of a virus; since, the infective material is prone to mutation. It is for this reason that it has proven exceedingly difficult to come up with an effective vaccine against HIV/AIDS.

Structurally, a virus particle consists of a protein outer coat that interacts and fuses with the cell membrane of the host cell. This is generally followed by the transfer of the infective agent – in this case the RNA of the coronavirus. Once this RNA enters the host cell, it directs the replication of its viral RNA and interferes with host cell processes. These tasks are accomplished through the transcription of viral RNA into proteins that exploit the cell’s protein synthesis “machinery.” It seems that coronavirus contains 7 genes – each gene having the blueprints for the production of a unique protein. One of the products of these genes is the so-called “spike” protein that plays a role in attaching to the host cell and has been shown to play a major part in the virus’ pathogenicity. The end result of this process is the formation of multiple copies of the virus followed by the death of the host cell and subsequent release of the new viruses into the extra-cellular environment.

The global nature of this threat can be circumvented by a number of different approaches - the first being isolating infected individuals and thereby thwarting the spread of the disease to others. It is likely that the virus is spread through aerosols as a result of coughing from infected individuals. This standard epidemiological approach is made particularly difficult given the reality of the constant movement of people to all areas of the globe.

However, it is also imperative that research efforts be directed towards developing a vaccine in order to assist the human immune system in its attempt to destroy the coronavirus once it has gained entry into the host. In regard to SARS, several studies were directed towards the development of active immunization strategies. These included Inactivated virions, recombinant antigen, DNA vaccines, and adenoviral vectors as well as other avenues of research. Undoubtedly, these kinds of studies will continue with added urgency.

Monday, January 20, 2020

Resistance to the anti-malarial drug Artemisinin in Malaria Parasites

Image showing human red blood cells infected with Plasmodium falciparum

Resistance of the anti-malarial drug Artemisinin in Malaria ParasitesMalaria continues to be a scourge in many parts of the world. The problem is particularly acute in Africa. Malaria is a pervasive illness characterized by high fevers, shaking chills, flu-like symptoms, and anemia. It is caused by a parasite referred to as Plasmodium falciparum. Plasmodium is carried by the Anopheles mosquito prevalent in the tropics.

The drug, Artemisinin (ART) has proven to be an effective drug against the malarial parasite – plasmodium falciparum. However, the parasite has apparently developed an immunity to this efficacious drug. It is, therefore, imperative that the mechanism of this resistance be more fully understood if an effective remedy is to be found. The collaborative efforts from research investigators at the Bernhard Nocht Institute of Tropical Medicine in Hamburg, Germany and the Department of Molecular Biology at Radbound University in the Netherlands have helped elucidate this mechanism.

The life-threatening aspect of infection by the Plasmodium falciparum parasite is the capacity of this parasite to continuously multiply within human red blood cells. Residing within human red blood cells, these parasites actively breakdown hemoglobin thereby obviating its capacity to deliver oxygen to the body’s tissues. Artemisinin has been long regarded as a first-line drug. However, ART resistance has manifested itself as a decreased susceptibility of young ring-stage parasites to a short pulse of this drug.

ART resistance has been shown to be associated with point mutations in the parasite’s so-called, Kelch propeller protein (Kelch13). However, the precise mechanism of this resistance to ART was essentially unknown. Although cellular stress, reduced protein translation and altered DNA replication had been implicated, the role of Kelch13 within the parasitic cell remained enigmatic. Here, the authors report an entire pathway in ART resistance and the Kelch13-dependent mechanism that effectively describes the reduced susceptibility to ART in resistant parasites.

The investigators in this extensive study, “show that Kelch13 defines an endocytosis pathway required for the uptake of host cell hemoglobin and its subsequent breakdown and that this pathway is critical for ART resistance. Their data indicate that Kelch13 and its compartment proteins mediate resistance upstream of both, drug activation and action. They have proposed a model where Kelch13 and its compartment proteins control endocytosis levels, thereby influencing the amount of hemoglobin available for degradation and hence the concentration of active drug.”

As a result of these findings that help elucidate the mechanism of ART resistance in malarial parasites, the authors conclude that, “We envisage that the mechanism of ART resistance indicated by this work will aid in finding ways to antagonize it. It may also inform the choice of ART partner drugs, particularly as hemoglobin digestive processes are the target of existing drugs.”

Tuesday, October 29, 2019

Microbiota and Immunity in Healthy Human Organisms

As a result of the millions of years of the evolution of the human species, the human body has established a commensal association with microbiota (microorganisms). This mutually beneficial relationship is essential for the maintenance of the health of the human host. However, it appears that when the natural barriers are disrupted, disease can result. The normal balance (homeostasis) between microbiota and host is apparently maintained by a robust immune system. The nature of this natural process, however, is poorly understood.

The current evidence is that specialized immunocompetent cells are implicated in this process. These are referred to as mucosal-associated invariant T (MAIT) cells and that these MAIT cells recognize and react to metabolites that are that are a byproduct of microbial metabolism. These metabolites are believed to play a role in microbial defense.

Drs Constantinides Legoux and their colleagues have reported that commensal bacteria exert control of the development of MAIT cells in the thymus (the organ involved in the development and proliferation of T cells) and their subsequent expansion within mucosal tissue. Additionally, the development of MAIT cells depends upon exposure in early life to well-defined microbial communities. Furthermore, it appears that a distinct subset of MAIT cells is actively involved in wound healing.

What makes MAIT cells distinct is the fact that in their activity they do not recognize the major histocompatibility complex (MHC) molecules like classic T cells. Instead, they are stimulated by nonpeptide (non-protein) antigens such as vitamin B2 precursor derivatives that are produced by many bacteria bound to an MHC-like protein referred to MR1.C-like H

Several recent studies point to an even broader range of activity for MAIT cells including: the control of bacterial, fungal and viral infections, a role in autoimmune disease and possible involvement in the immune processes involved in attacking the proliferation of tumor cells.

The results of extensive study of MAIT cells coming from the laboratories of Constantinides and Legoux have established that MAIT cells require a community of microbiota to enable their development in the thymus and insure their expansion into specific tissues. It seems that, in fact, newborns are rapidly colonized by a diverse complement of species and strains demonstrating the key role that these microorganisms play in establishing the healthy status of the rapidly developing new human. Furthermore, in humans, it appears that the frequency and localization of MAIT cells throughout the organism changes over a lifetime and is apparently diminished in the elderly population.

In summary, the diagram below illustrates the intimate relationship that exists between the development and function of MAIT cells and the resident commensal microbiota on barrier tissues such as the skin. These finding also emphasize the key role that these organisms play in the health of individuals beginning at birth and proceeding throughout an individual lifetime.

Sunday, August 11, 2019

Cancers Caused by Contagious Agents

We do not generally regard cancer as being contagious. Although this is generally t rue, there are a growing number of cancers that have been shown to be caused by contagious agents, especially by specific viruses. The types of cancers caused by such agents are shown along with the microorganism implicated.

Cancer Causative Agent

Stomach (Gastric) Cancer H. Pylori - Bacterium
Cervical Cancer Human Papilloma Virus (HPV)
T-Cell Leukemia HTLV-1 – related to the AIDS virus
Burkitt’s Lymphoma Epstein Barr Virus (EBV)
Kaposi’s Sarcoma HTLV-3 – AIDS Virus
Primary Liver Cancer Hepatitis B Virus (HBV)

In regards to the data listed in the table above, there is additional information associated with them:

• H.Pylori is a highly specialized and fairly ubiquitous bacterium that can survive in the harsh acidic environment of the stomach; it is found in the mucus layer on the inside of the stomach within those individuals who are infected. For this reason, it is particularly difficult to treat with antibiotics. It is found in about 2/3 of the world’s population. It has also been implicated in the onset of chronic gastritis and peptic ulcers.
• HPV has recently been implicated as the causative agent for cervical cancer. Cervical cancer is prevalent in sexually active women. A vaccine against this virus has recently been developed. It has been shown to be highly effective in preventing the onset of this cancer in young women.
• EBV, the virus that causes Burkitt’s lymphoma found predominantly in Africa, is the same virus that causes mononucleosis in individuals in the West. The reason for this distinct difference in disease outcomes is poorly understood, but a genetic basis for this difference is likely.
• The AIDS virus is not directly responsible for the onset of Kaposi’s sarcoma. The disease is manifested in AIDS patients on account of their highly suppressed immune system due to infection with the AIDS virus that make AIDS patients especially susceptible to this cancer.
• Once an individual is infected with HBV as a result of close human contact with an infected individual, the virus can infect the liver without any noticeable symptoms. This can be the case for many years before the onset of liver cancer. HBV is believed to account for 80% of the reported cases of primary liver cancer (cancer that originates in the liver). It is a deadly cancer. Fortunately, there is a vaccine against HBV infection, but there is currently no vaccine to prevent infection by the Hepatitis C Virus (HCV) that is also known to cause primary liver cancer.

Control of Inflammation following Injury or Infection

The natural immune responses that are elicited after an insult to the body as a result of injury or infection from a pathogen, is both rapid and powerful. Initially inflammatory cells are mobilized to the site of the trauma. The predominant cell types that are recruited are macrophages and neutrophils that release free radical reactive oxygen and nitrogen species (RONS) that are lethal to invading organisms. However, these chemical moieties are so powerful that they can also kill and mutate the surrounding normal tissue. Although there are naturally induced anti-inflammatory responses that are also initiated, the optimum balance is difficult to achieve. In some diseases, such as ulcerative colitis and rheumatoid arthritis, such anti-inflammatory responses are unavailable. Therefore, for this reason medical intervention in the form of medication becomes appropriate. 

There are, of course, many different anti-inflammatory medications currently available. Recently Dr. Torkild Visnes and his colleagues from the Karolinska Institutet in Sweden in collaboration with the University of Texas Medical Branch, Uppsala University and Stockholm University have discovered a new methodology for enhancing the anti-inflammatory response. In order to understand the investigator’s approach, we need to examine the rationale for this research in greater detail. 

Normally the enzyme 8-oxyguanine DNA glycosylase 1 (OGG1) functions as a DNA repair enzyme that both recognizes and repairs the nucleotide base excision repair of 7,8 dihydro-8-oxoguanine (8-oxoG) that represents one of the major types of DNA damage produced by RONS. Paradoxically, the binding of OGG1 to 8-oxoG, facilitates the action of the NF-kB transcription factor that promotes the activation of quiescent chemokine and cytokine genes that subsequently leads to the inflammatory response and the subsequent release of RONS at the site of trauma. 

Visnes and his group have identified a small molecule (TH5487) that binds to the active site of OGG1 and effectively blocks its repair capabilities on account of the fact that inhibited OGG1 cannot bind to that G-rich region of the DNA leads to the activation of the NF-kB transcription factor that promotes proinflammatory genes. In fact, TH5487 has been shown to inhibit this process in mouse and human lung epithelial cells in vitro and the TNF-induced neutrophil inflammation in the in vivo mouse model. 

This research is of value since it elucidates a molecular mechanism that demonstrates a connection between the normal DNA repair function of OGG1 and the inflammatory response and has discovered a small molecule inhibitor of this process.

Saturday, August 10, 2019

A Study in the Evolution of a Transmissible Cancer

There is a canine-related cancer that is sexually transmitted from one animal to another. It is the canine transmissible venereal tumor (CTVT) that presents as genital tumors. It has been shown that this cancer spreads through the transfer of living cancer cells (see image below) through coitus. These CTVT cells function as a unicellular, asexually reproducing (but sexually transmitted) pathogen. In addition, CTVT cells are genetically aberrant in terms of chromosome number – studies have established that instead of the normal 78 chromosomes present in the canine species these cells have a chromosome number in the range of 57-64. This type of cancer represents an exceedingly rare etiology in that the cancer is spread through the transfer of the cancer cell itself.

CTVT Cells
This disease exists worldwide and apparently has the longest known and most prolific evolutionary lineage. This reality provides an opportunity to explore the mechanism of the evolution of cancer over the long term. For this reason, Adrian Baez Ortega, a bioinformatician, and her colleagues from the Transmissible Cancer Group at Cambridge University, UK, have done a detailed analysis of genetic sequence data from 546 individual CTVT tumors taken from diseased animals. Their focus in this analysis was to identify somatic single-nucleotide variants (SNVs) from the exomes – areas of the genetic material that are involved in the active production of proteins. The aim of this intense research study was to help to uncover the mutational events and genetic signatures of the evolutionary selection process over the thousands of years.

One of the predominant findings of this analysis was that this lineage arose from its originator canine individual between 4000 and 8500 years ago most likely from Asia with the most recent ancestor of the current global distribution estimated to have lived about 1900 years ago. There does not appear to be any positive selection for this lineage. In addition, the author of this study concludes that, “a highly context-specific mutational pattern named signature A was identified, which was active in the past but ceased to operate about 1000 years ago. BP, years before present.”

These results provide an insight into the progression of an unusual type of cancer that is transmissible through the transfer of the cancer cell itself rather than through a vector such as a virus – human T Cell Leukemia being one particular example. It also is a demonstration of the power of the widely used tools of research regarding the discovery of the underlying genetic mechanisms involved in disease processes.

Friday, June 7, 2019

Senescent Cells and Human Longevity

Although the average human lifespan has increased substantially over time due to the improvement in living conditions made possible by advances in public health, sanitation, medicine, etc., there is no selective advantage afforded by a longer life once the reproductive period has passed. Consequently, there is a normal and gradual deterioration of the tissues that is age-related.

Furthermore, there is a natural process referred to as cellular senescence – cells undergoing this change are no longer able to divide. Cellular senescence confers a reproductive advantage for the individual in that it helps block cancer cell proliferation; however, overtime it results in an increasing abundance of senescent cells (SNC) within the tissues. It seems that in animal studies, using the mouse model, in which the SNCs are selectively eliminated (senolysis),the median lifespan of individual test mice is extended, and the frequency of age-related diseases has been shown to be diminished. This result has encouraged the search for and development of drugs that selectively target SNCs.

In terms of this research, it is vitally important to discover the actual mechanisms that underly cellular senescence. In studies using cells grown in culture, it has been shown that SNCs are in a state of permanent cell cycle arrest. This state is apparently initiated and maintained by the p53-p21 retinoblastoma (RB) and p16-RB tumor suppressor pathways. The factors that can trigger this process are –
  • Oxidative stress
  • Shortening to telomeres – repetitive DNA sequences at the ends of chromosomes that afford protection
  • Prolonged mitotic activity
  • DNA errors during replication
  • Mitochondrial impairment.
SNCs produce a so-called, “Senescence associated secretory phenotype” (SASP). SASP has been shown to negatively impact normal tissue architecture through a variety of processes including the onset of fibrosis and the inhibition of stem cell functionality. Although SASP has a protective function in regard to the development of cellular neoplasm, in later life it does not seem to provide a protective function against the onset of cancer. This raised the possibility that the selective elimination of SNCs from older patients might exert an anti-cancer effect.

Given this data, it would seem that therapies that can effectively eliminate SNCs might produce a two-fold health advantage by increasing longevity and by decreasing the onset of cancer in later life. Encouraging results from animal model studies have shown that drugs that target those pathways that block apoptosis – programmed cell death – promote senolysis and afford an anti-cancer potential. In regard to future research, this may provide a very fruitful line of enquiry.