This is not going to be a sermon in which Doc Gumshoe once again attempts to convert the vaccine skeptics. Instead, we’ll be considering the evolution of vaccines, dating from the time when they weren’t called vaccines, and what the evolutionary process of vaccines points to, which, in the opinion of many eminent scientists, is very promising indeed.

It appears that we humans have known about immunity for quite a long time – a thousand years, at least. Our ancestors learned about immunity due to the horrible toll that smallpox exacted from populations in the Eastern hemisphere. In Africa, Asia, and Europe, smallpox epidemics regularly killed about a third of all infected people, and since a major fraction of the populace became infected during these epidemics, there were many, many deaths.

But one thing they learned about smallpox is that people who have had the disease and survive do not get infected again. This is true regardless of which of the two strains of smallpox cause the infection.

Smallpox is caused by one of the two variants of the variola virus – variola major and variola minor. Variola major causes the more severe disease, but variola minor, which causes a relatively mild version of smallpox, does confer immunity from the more severe disease.

In an effort to protect themselves against the consequences of a severe smallpox infection, many people were willing to infect themselves with variola, hoping that they would contract a mild form of the disease, thereby protecting themselves from severe infection and possibly death. What they did was to take really small amounts of the matter from the blisters of infected individuals and infect themselves. This was called “variolation,” and it was practiced in China as early as the 10th century, and also in parts of Africa. That was long before the emergence of vaccination, which didn’t happen until the end of the 18th century.

Perhaps we need to define some words here. The original meaning of “vaccine,” according to my Oxford English Dictionary, is “cowpox” – variola vaccinae – from vacca, the Latin for “cow.” The earliest use of the word cited in the OED is from 1799: “The certainty that having suffered the vaccine disease will prove a preservative from the infection of the small-pox.” (Med Journal I, 251)

In other words, if you’ve had cowpox, you won’t get smallpox. This had been observed for many years – milk-maids, who milked infected cows and developed the lesions and blisters of the cowpox, didn’t get infected with smallpox itself. That observation is what led to the establishment of vaccination. We’ll get to that later. But first, let’s take a look at some happenings before vaccination was discovered.

There were controversies about the practice of variolation. One of the prominent figures in these was the eminent New England clergyman, Cotton Mather. He was far from a milquetoast. He was central in the campaign to identify and bring to trial those women whom he (and others) called “witches.” In 1692, more than 200 so-called witches were accused and 19 were executed.

But he favored variolation. In Massachusetts at that time, African slaves that bore the scars of variolation were deemed more valuable, because they were immune from smallpox. In 1706, Mather purchased an African slave named Onesimus, who had experienced variolation as a child. Mather became interested in the practice, and, when a medical paper about variolation, written in 1714 by the Italian physician Emmanuel Timoni, reached the shores of Massachusetts, Mather read the paper and became committed to variolation. He pushed for the introduction of variolation in Boston in advance of the next smallpox epidemic, which he thought was sure to occur. There had been a smallpox outbreak in 1702, but Boston had been lucky in the years since then.

Boston got walloped by a bad smallpox epidemic in 1721. Mather publicly proposed the practice of variolation, for which he was roundly chastised. Some individuals characterized variolation as being “Negro-ish thinking” and stood by the principle that it was immoral to interfere with God’s judgment. More than half of Boston’s population of 11,000 were infected with smallpox, and of these about 14% died. But a Boston physician, Zamiel Boylston agreed with Mather and inoculated his own son and 248 other people in Boston who agreed to be inoculated. Of that cohort, only 2.5% died.

One would suppose that the 1721 Boston experience would be cited as evidence that variolation was a good idea, but the opposition did not give in. Benjamin Franklin, whom we now regard as an early advocate of scientific thinking, wrote articles in The New England Courant, published by his brother James, denouncing variolation. However, over the next fifteen years or so, Benjamin changed his views, although too late to variolate his four-year-old son, who died of smallpox. In his autobiography, Franklin articulated his regret that he had not attempted to prevent the disease in his son. He wrote, “In 1736 I lost one of my sons, a fine boy of four years old, by the small-pox, taken in the common way. I long regretted bitterly, and still regret, that I had not given it to him by inoculation. This I mention for the sake of parents who omit that operation.”

Another famous name comes up in the history of smallpox. North America (not yet the United States) experienced a severe smallpox epidemic starting in 1775. George Washington, himself a smallpox survivor, realized that smallpox running rampant through his troops would render them almost defenseless against the British, most of whom had been inoculated against the disease. Smallpox was a particularly menacing threat to troops in the field, crowded together in tents and in trenches, with minimal sanitation if any. After much debate and soul-searching, Washington went ahead with the decision to have the Continental Army inoculated. By that time, many prominent persons in the colonies had been variolated, including George’s wife, Martha. Washington wrote, “Necessity not only authorizes but seems to require the measure, for should the disorder infect the Army, we should have more to dread from it, than from the Sword of the Enemy.”

The switch from variolation with live smallpox virus to vaccination with the cowpox virus occurred just at the end of the 18th century. Edward Jenner in 1798 proposed that the cowpox – variola vaccinae – be employed. This was based on the previously- mentioned observation that milkmaids, who handled the udders of infected cows and themselves became infected with the cowpox, almost never contracted smallpox. It was concluded that cowpox infections made a person immune to being infected with smallpox. As a result of this discovery, smallpox itself has been eradicated. The last case of naturally occurring smallpox in the US was in 1948, and globally there have been zero cases of smallpox since 1980.

With your permission, Doc Gumshoe would like to look into the origins of the word “immunity,” which we use here to mean something like “highly effective protection against an infection.” But what it meant originally was “exemption from obligations to the municipality,” which could mean not being obligated to pay taxes or perform services of any kind. The source is the Latin immunis, which has nothing to do with disease. But there is a relationship there. In a sense, infectious diseases are part of the municipality. Smallpox, or COVID-19, or the flu, or the common cold, do not exist in a vacuum, circulating about in the external world waiting to pounce on us. Immunity means protection from a condition that co-exists with the human race. In a sense, the disease is present in the community. Picking that word was singularly apt.

The evolution of vaccines

Jenner demonstrated the efficacy of using the cowpox vaccine by inoculating a 13-year-old boy with matter from a cowpox pustule, and then deliberately exposing the boy to smallpox. The boy was not infected. Jenner also cited evidence from a group of other children who had been infected with cowpox and then exposed to smallpox; none of these children contracted smallpox.

For many years after the introduction of vaccination, material for the procedure came from the pustules of vaccinated persons. A tiny scratch would be performed on the upper arm of the patient, and a very small amount of the pus from another individual’s vaccination site would be rubbed into the scratch. Later, a method of producing larger quantities of vaccine was developed: the entire flank of a cow would be inoculated with the pox, thus providing an ongoing source for abundant quantities of cowpox pus for use as vaccines. The vaccine was sometimes kept in a solution with glycerol, which acted as a preservative. That method of vaccination against smallpox continued more or less unchanged for about 150 years.

The modern smallpox vaccine was conceived in the 1950s when a technique was developed to produce a heat-stable, freeze-dried vaccine. This process used centrifugation to create a suspension of the virus, which was then freeze-dried in ampules. This had the advantage of allowing long-term storage without refrigeration.

The smallpox vaccine was associated with many more adverse reactions than the vaccines used at present. The initial reaction to the vaccine, as I well remember, was a rather nasty lesion filled with pus at the inoculation site. You did not want to touch that lesion and then rub your eyes or any other part of your body, because that would prompt other nasty lesions to appear. The inoculation site would be covered by a bandage with a little celluloid bubble in it so that the pustule would be contained.

There were also some serious adverse reactions to the smallpox vaccine, most of which involved spread of the cowpox beyond the original inoculation site. More than one-third of vaccine recipients missed days of school or work because of vaccine-related side effects. Death following smallpox vaccination was exceedingly rare, but not unheard of. Estimates of the risk of death are generally smaller than one in a million vaccinations.

The success of the smallpox vaccine spurred investigation into the possibility that other infections could be prevented through the process of inoculating a person with a very small amount of the infective agent in the hope that the person’s body would in some way recognize the agent as being harmful and mount a defense. At that time, nothing was known about the specific mechanisms of immunity. All that scientists had to go on was the generally recognized experience that in the case of many common illnesses, once a person had been sick with that illness, that person would be spared future infections with that same illness. That experience led many parents to deliberately expose their children to a number of common illnesses such as the measles and chicken pox.

However, it was not until 1879 that the next great vaccine discovery occurred. That year, Louis Pasteur observed that cultures of a disease called chicken cholera had become weakened, due to having been left uncared for in a laboratory for some weeks. When chickens were inoculated with this attenuated culture, they developed very mild symptoms of chicken cholera, and were thereafter immune when challenged with the fully virulent strain.

Based on this chance observation, Pasteur devoted himself to the discovery of agents that would protect individuals from infection – what we now call vaccines. The first vaccine after Jenner’s smallpox vaccine was Pasteur’s vaccine for cholera in 1880. Pasteur had originated the idea that infection came from outside the body and was carried by tiny entities which he called “germs” – essentially the seeds of the infection. Prior to Pasteur, infections had been thought to originate spontaneously in the body.

Fairly soon after that, in 1885, Pasteur and Emile Roux introduced a vaccine for rabies. That was followed fairly shortly by vaccines for tetanus, typhoid fever, and the bubonic plague, developed by other scientists.

The 20th century saw a great deal of progress in the development of vaccines, starting with the first vaccine against tuberculosis in 1921. That was the bacillus Calmette-Guérin, (known as BCG) which induced immunity to the tuberculosis pathogen by challenging the patient with another microbe that was closely similar, but harmless. As it happens, BCG also minimizes severe disease in persons infected with the coronavirus that causes COVID.

About 25 new vaccines were introduced in the 20th century, including vaccines for diphtheria, scarlet fever, tetanus, whooping cough, yellow fever, cholera, influenza, polio, anthrax, measles, mumps, rubella, pneumonia, meningitis, hepatitis B, chicken pox, and Lyme disease. So far in the 21st century, vaccines have been developed for herpes zoster, malaria, dengue fever, Ebola, and most recently COVID-19. Also, improved pneumonia and influenza vaccines have emerged.

The effectiveness of vaccines varies widely. While the effectiveness of the smallpox and polio vaccines is nearly 100%, other vaccines have much spottier records. As you certainly know, the influenza vaccine needs to be reformulated every year, as the virus itself constantly mutates, and even then its effectiveness varies from year to year. The typhoid vaccine, which is recommended for travelers to countries where the disease is prevalent, is at most 80% effective, and has to be repeated every few years. On the whole, vaccines have proved to be highly effective in controlling disease. As we consider the diseases listed above, it is remarkable how many have been rendered extremely rare in the developed parts of the world because of the spread of vaccines.

Types of vaccines

First generation vaccines
As we saw, the first agents that conferred immunity were in themselves infectious, whether very small attenuated quantities of the actual pathogen, as in the material used in variolation, or a related pathogen that had been shown to confer immunity to the targeted pathogen, as in the original variola vaccinae, the cowpox. There are many vaccines that employ that same mode of action: the pathogen itself is diluted or attenuated to the point that it is no longer likely to cause serious infection, or the pathogen has been inactivated in some way. The combination measles-mumps-rubella vaccine, the chickenpox, and the hepatitis A vaccine are live attenuated vaccines, while the flu vaccine and several others consist of inactivated pathogens. Inactivated (killed) pathogens have an advantage in that they are less likely to cause infection than attenuated vaccines. But, on the other hand, they are expelled from the body more quickly and the degree of immunity they confer tends to be of shorter duration.

Second generation vaccines
A second generation of vaccines attempted to avoid the shortcomings of the earlier live or inactivated vaccines. To avoid the possibility that the vaccine itself might bring on the infection, researchers constructed vaccines from elements of the toxic substance, sometimes using synthetic proteins and copying the part of the pathogen which the host’s antibody recognizes and to which it binds. Development of these vaccines required specific understanding of the molecular structure of the pathogens. Vaccines of this second generation are classified as subunit, conjugated, and recombinant. Among the subunit vaccines are diphtheria, tetanus, and whooping cough; the streptococcus pneumoniae vaccine is classified as a conjugated vaccine; and the most successful example of a recombinant vaccine is the hepatitis B vaccine.

Third generation vaccines
Vaccines carrying the genetic information that encodes the antigen (that is the word that is used in vaccine discussions to denote the disease-causing entity) may be grouped under the term “genetic vaccines.” The encoding material stimulates the patient’s immune system to produce the recombinant protein which, in turn, stimulates the body to generate both the antibodies that directly attack the antigen, and also stimulates the generation of cellular immunity, carried by B cells and T cells. These vaccines may be said to constitute the third generation of vaccines.

Third generation vaccines, relying on the genetic material of the pathogen rather than killed or diluted pathogens or segmented pathogens, have a number of advantages over the previous vaccines. They are generally able to immunize against several different strains of an antigen; also, there is no possibility that these vaccines can revert to the original pathogen and result in infection and disease. Because these vaccines are entirely laboratory produced, the manufacture of genetic vaccines is relatively rapid and straightforward, unlike, for example, the flu vaccine, which requires time-consuming growth of the flu virus itself and then inactivation of the virus.

Even prior to the development of the COVID-19 vaccine, extensive research was underway on the possible use of genetic vaccines for hepatitis, HIV, flu, and the Ebola virus. I should point out that some writers on this subject refer to genetic vaccines as “DNA vaccines.” DNA is the entity that carries genetic information, but the COVID-19 vaccine, as everyone certainly knows, is an mRNA vaccine. Actually using DNA to make a vaccine would be exceedingly difficult. Human DNA consists of about 3 billion base pairs, while mRNA is only 29,900 bases.

The evolution of mRNA vaccines

The story begins back in 1869, when the Swiss scientist Friedrich Miescher spotted a little particle inside the nucleus of human white blood cells. He called it “nuclein,” and the particle was eventually named “deoxyribonucleic acid,” or DNA. It wasn’t until 1944 that a group of scientists at the Rockefeller Institute demonstrated that it was this substance that passed along the genetic material necessary for reproduction, and a few years later, in 1950, another scientist showed that the four bases in DNA – adenine, cytosine, guanine, and thymine – paired in specific ways. And in 1953, the discovery of the structure of DNA was published by James Watson and Francis Crick – the famous double helix in which strands of DNA were twisted together. That discovery, by the way, was in considerable part due to the work of Rosalind Franklin, who had captured X-ray diffraction images of DNA. When Watson and Crick got their Nobel Prize, Franklin was not mentioned.

Messenger RNA (mRNA) was discovered shortly after the structure of DNA was made known. These molecules had been identified as early as the 1960s as the entities that carried copies of segments of the DNA code to structures within cells (called ribosomes) that actually manufactured the DNA. Scientists began to explore the possibility that mRNA could command human cells to make parts of the virus that could be identified by the immune system. This could lead to an immune response to the virus itself. In the 1990s, the US government embarked on a mission to find a vaccine that would prevent AIDS, which was dangerously on the rise at that time. The idea was to identify and target the spikes on the HIV virus that enabled them to invade cells. So far, that approach has not worked for HIV.

Researchers continued to investigate the possibilities that injecting the mRNA of a pathogen into a human host could induce the host to manufacture the pathogen itself and thus provoke an immediate immune response. Many, many experiments failed. A problem seemed to be that the immune system recognized the mRNA itself as a pathogen and mobilized the forces to eliminate it.

Eventually, that specific problem was identified and addressed. The researchers discovered that immune cells protect their own mRNA with a specific chemical modification. So the researchers tried to duplicate that same chemical modification into the mRNA that they hoped to use as a vaccine. The modification was successful; the modified mRNA was taken up by the host cells.

A paper describing that discovery was presented to the journal Nature and Science in 2005 and initially rejected; it was eventually published in the less prestigious journal Immunity. However, the information in this paper was ignored for several years, except for a small group of scientists who continued to see highly important possibilities in mRNA as a platform for vaccines.

Actually creating a vaccine using mRNA required addressing several challenges, in addition to figuring out the precise mRNA sequence that would direct host cells to make the most effective version of the specific characteristics of the invading pathogen, so as to initiate an immune response at such time as the pathogen invaded the host. Another challenge was that cells in the host are typically encased in a lipid envelope which would need to be penetrated in order for the vaccine to do its work.

Among the many researchers whose work eventually led to the vaccine against SARS-CoV-2 were Dr Barney Graham and Dr Jason McLellan. They began working together in 2013, when Dr Graham, the senior member of the pair, suggested to Dr McLellan that they should work on the coronavirus. This may have seemed an odd choice at the time, since the coronavirus was mostly known as the cause of a form of the common cold. However, about that time another disease, also caused by a coronavirus, began spreading in the Middle East. The disease came to be named MERS – Middle East respiratory syndrome. And MERS was related to yet another disease caused by a coronavirus: SARS – Severe Acute Respiratory Syndrome.

Dr Graham and Dr McLellan had been doing some research on the MERS coronavirus for several years, although with no particular sense of urgency, since MERS had not become a major threat. That changed dramatically when, on December 31st, 2019, Dr Graham saw a news release reporting that a new pneumonia strain was spreading rapidly in Wuhan, China. He alerted Dr McLellan that they needed to watch the developments of this Wuhan disease closely. About a week later, the news emerged that the cause of the disease was a novel strain of the coronavirus.

In mid-January of 2020, Chinese researchers posted the genetic sequence of the Wuhan coronavirus online. Based on that information, the Graham-McLellan team focused on the coronavirus spikes and learned that these were encoded in mRNA molecules. On February 15, 2020, Drs Graham and McLellan published an online paper detailing the spike structure of the coronavirus which later came to be named SARS-CoV-2.

Within weeks, both Moderna and Pfizer-BioNTech had synthesized the mRNA molecules and applied the same modification that immune cells use to protect their own mRNA. These vaccine candidates were ready for clinical trials, as were non-mRNA vaccines from Johnson & Johnson, AstraZeneca, and Novavax. At that point, the government’s network of testing sites for HIV vaccines, which included about 100 sites, came into play. The team was led by Dr Larry Corey, and about 30,000 volunteers were recruited for the clinical trials.

As you know, results of the trial appeared in early November, 2020 – a very rapid achievement, compared with the multi-year span that other vaccines had taken from inception of research to the time when vaccines were ready for use. The news of the successful trials was made public on November 8th, 2020.

Doc Gumshoe got his first dose of the COVID-19 vaccine on January 28, 2021.

The role of gene editing in the development of COVID-19 vaccines

The term “CRISPR” has not appeared in this piece, but CRISPR is fundamental to gene editing, and the development of the mRNA vaccines that have done so much to control COVID-19 are based on gene editing. Here’s a bit about gene editing and CRISPR that was in a Doc Gumshoe on April 5th, 2021.

“The finding that opened the path to what we now call gene editing was the discovery that in those long strands of DNA, the gene sequences were separated by clusters of sequences that did not appear, at first glance, to convey any genetic information at all. We might call them nonsense sequences. They followed a certain pattern – they read the same way from either end: i.e., they were palindromic. They occurred at repeated intervals in the DNA strand. Thus, they came to be called Clustered Regularly Interspaced Palindromic Repeats – “CRISPR”. These sequences were discovered in 1980 and were originally dismissed as being of no interest – “junk DNA.”

It turned out that the CRISPR sequences, along with CRISPR-associated (CAS) enzymes are of vital importance in conferring immunity to repeated infection by a virus. This was discovered in research on bacteria, which also (of course!) have CRISPR in their genome. When a virus invades a bacterium, the CAS enzyme snips off pieces of the viral genome, and stores it in the CRISPR part of the bacterial genome so that the bacterium essentially remembers the virus that attacked it and thus mount a defense. A number of these enzymes have been discovered, but the one – CRISPR-Cas9 – that has up to this point been found to be most useful in gene editing was discovered in 2012 by Jennifer Doudna of the Broad Institute (Harvard / MIT) and Emmanuelle Charpentier of the Max Planck Institute for Infection Biology in Berlin.

CRISPR-Cas9 has been central in gene editing in the past seven years. The Cas9 enzyme makes it possible to “snip” entire gene sequences out of the genome without altering the gene itself. The Cas9 enzyme is guided by RNA, which has the capacity to locate specific DNA sequences. The enzyme binds to and breaks the CRISPR cluster on each end of the genetic material in a way that permits the genome to link together without a permanent break, resulting in the deletion, or “knock out”, of that gene. The word “snip” is of course used figuratively; there is no mechanical instrument that cuts the CRISPR cluster. The binding action of the enzyme separates the individual particles in the cluster. And the binding action is possible because of the configuration of the enzyme and the electric potential of its components.”

Scientists at Stanford have reported that they have developed a system using CRISPR that has the capacity to stop infection with 90% of all coronavirus strains, not only SARS-CoV-2. They have developed a technique called prophylactic antiviral CRISPR in human cells, dubbed PAC-MAN, which disables viruses by scrambling their genetic code.

PAC-MAN combines mRNA with the virus-killing enzyme Cas13. The mRNA directs Cas13 to destroy certain nucleotide sequences in the SARS-CoV-2 genome, effectively neutralizing it. PAC-MAN is enveloped in lipitoids, which are synthetic peptides that can deliver the CRISPR-Cas13-enabled package directly into cells.

Stanford researchers have also reported that PAC-MAN will very likely be useful against influenza. Their technology was not only effective against coronavirus strains – it also significantly lowered the viral load in human lung cells infected with the H1N1 strain of the flu.

There doesn’t seem to be any limit to the infectious diseases that can be controlled by means of mRNA vaccines. For a start, researchers are addressing the numerous diseases for which the vaccines that do exist are of limited efficacy. There are many such diseases, and the global toll these diseases exact in terms of sickness and death is enormous.

Developing vaccines against diseases that are not caused by infectious pathogens is another matter altogether. Heart disease, cancer, diabetes – these are not caused by germs. However, there are some forms of cancer that are directly associated with infectious diseases. Cancer of the cervix, as well as cancers of the head and neck are frequently cause by the human papilloma virus (HPV). Liver cancer can be the result of infection with the hepatitis B virus (HBV). Vaccines exist against both of these viruses and are effective in preventing those cancers. The bacillus Calmette-Guérin (BCG), the tuberculosis vaccine, is effective in the treatment of early stage bladder cancer. The sipuleucel-T vaccine was developed and received FDA approval in 2010 for the treatment of patients with advanced prostate cancer, because it has the capacity of targeting cancer cells based on the abnormally high quantities of prostatic acid phosphatase (PAP) that they emit.

The possibility of creating vaccines that target specific cancer cells based on specific genetic characteristics that can be addressed by mRNA appears to be wide-ranging and highly promising. We can conclude that vaccine research, based on the success of the COVID-19 vaccine, will increase in volume as well as scope, and with huge potential benefits.

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Yes, there’s a good deal of highly technical stuff in that Doc Gumshoe piece, and I thank you very much for persevering to the end. I am going to skip the chance to give you a short quiz; instead, I invite you to send along your comments, of any flavor – all are welcome! Best to all, Michael Jorrin (aka Doc Gumshoe)

[ed note: Michael Jorrin, who I dubbed “Doc Gumshoe” many years ago, is a longtime medical writer (not a doctor) and shares his commentary with Gumshoe readers a couple times a month. He does not generally write about the investment prospects of topics he covers, but has agreed to our trading restrictions.  Past Doc Gumshoe columns are available here.]