Break the bug, spare the child. The partial attenuation of Small Hydrophobic (SH) gene deleted RSV is associated with elevated IL-1β responses.

Maurice Hilleman
Hero of the 20th Century

Many of our best (and oldest) vaccines are based upon a principle called live attenuation. This is where we have somehow modified the infectious agent so that it is less infectious, allowing the body to kill it and generate an immune memory in order to be fighting fit on exposure to the real thing. Historically, this process was achieved empirically (by trial and error), either by using an equivalent bug from a related animal for example – Cow Pox/ Small Pox. Alternatively the bugs can be serially passaged – grown in conditions that cause them to lose the genes that makes them infectious– e.g BCG/ TB . Or it can be done in a more targeted way, for example the live flu vaccine was developed by selection for growth at 32°C – closer to the temperature of the nose than the lung and therefore limiting infection to the nose causes colds at worst rather than lung infections. The master attenuator was a microbiologist working at Merck called Maurice Hilleman, who is probably the most influential person you have never heard of – saving more lives in the 20^th century than any other person. I just want to emphasize his importance, because his record is often under stated, Hilleman invented 8 of the 14 vaccines we use today and yet, I had to google him to remind myself of his name.

However, we are now using more targeted approaches to attenuate viruses and bacteria. This is based on our improved understanding about how pathogens are able to infect people. For example what do they look for on the surface of cells to invade them, how do they coerce the machinery of the cell to make copies of themselves rather than more cells and critically how do they hide from the immune response. All human viruses have evolved ways of escaping the immune response (immune evasion), viruses that are not able to escape our immune response are not able to infect us – that’s why for example we don’t get myxamatosis from rabbits. In our recently published study in the Journal of Virology, we discovered a new immune evasion function for a gene in respiratory syncytial virus (RSV). RSV is a really important disease in children, causing 160,000 deaths worldwide and hospitalising 1% of all children under 1 in the UK (including my son).

The gene we were interested in is called SH (or small hydrophobic gene – sadly whilst drosophila geneticists get to call genes things like sonic the hedgehog and LUSH, we get names based on the structure or function, or sometimes just the order they were found). It is believed to make a small hydrophobic protein (can you see what we did there!) which folds up to make a pore or tube like structure. Based on other studies using similar proteins from other viruses, we hypothesized that this protein would actually alert the immune system, so we were surprised to find that infection with RSV lacking SH (RSV ΔSH) led to MORE of specific type of signal rather than less. We then saw that if this signal was blocked, the virus – which previously grew less well in lungs, grew to the same level as unchanged virus. We think this might be important both in our understanding about viral biology and possibly in developing strategies to make targeted vaccines. We were supported in this work by two grant programs from the EU, Aditec and Biovacsafe, which has enabled a lot of the work in the lab to be performed.

What's the difference between a Jelly fish and a vaccine?

A Comparison of Red Fluorescent Proteins to Model DNA Vaccine Expression by Whole Animal In Vivo Imaging

PLOS One, June 2015

Vaccines are an incredibly potent tool in our arsenal to prevent infections. Simplistically they work by exposing the immune system to a safe part of the infectious bacteria or virus and evoking a memory response. When the vaccinated individual encounters the actual bug in real life, this memory allows the immune system to recognise and respond to the bug faster, thus preventing the infection before it takes hold. A critical step in vaccine development is the identification, isolation and mass production of the parts of the virus and bacteria that are able to evoke an immune memory. These components are often made up of proteins, which can be problematic and expensive to make to a standard that is acceptable for use as a medicine. Recently an alternative approach has been developed called DNA vaccination, which we describe in this review. DNA, the material of genes, contains the coding message for the production of proteins by cells. In the 1990’s a number of groups made the breakthrough observation, that if you inject DNA encoding proteins from a pathogen into a mouse, the mouse will make the protein in its cells and generate an immune response to this protein. This technique has the potential to significantly change vaccines because it is cheaper, easier and faster to make, for example, if a new viral epidemic occurs e.g. Ebola or MERS-CoV the time to make and test a DNA vaccine could be significantly shorter and therefore the spread of the disease would be reduced.

The rainbow explosion of fluorescent proteins, From Roger Tsien's Nobel Prize Talk.

However, DNA vaccines have performed poorly in early phase clinical trials (i.e. they work really well in mice, but poorly in people). There have been a number of reasons proposed for this failure, but one possibility is that, after immunisation, the genes encoded by the DNA vaccine are not turned into protein by the patient (technically referred to as poor expression). In our recent paper, we set out to address this problem in our study, using a novel technology called in vivo imaging, which uses the proteins that allow jellyfish to glow in the dark and fireflies to flash at night. The original fluorescent protein was green (named green fluoresecent protein), but since then a whole colour chart of proteins has been developed with names like tdTomato and mPlum. If the proteins are injected into mice, they can, using a special camera, be seen. The colour of the protein has a significant impact on our ability to see it in mice, blue and green proteins work poorly because the skin has evolved to reflect light in these wavelengths as a defence against damage caused by UV light. Red proteins tend to be better and in our recently published paper we tested a range of red proteins delivered as DNA, called tdTomato, mCherry, mKatushka and tdKatushka. We observed that only tdTomato gave us a strong enough signal to see in the mice and that there are number of confounding problems with the approach – pens we use to identify individual mice fluoresce in the same colour as mouse poo.

An Unusual Location for Scientific Training!

Delivering the DNA by different routes also had an impact on the brightness with DNA injected into muscles producing a brighter signal than DNA injected into the skin. We also looked into alternate methods of delivering vaccines trying to more directly access cells involved in the immune response. One approach we tried was tattooing, which is exactly as it sounds, we tattooed DNA directly onto the skin of mice using the same system as you might get an anchor or ex-girlfriend’s name tattooed on your arm. This came as a kit and to get training, Katya (the lead author) had to get help from the local tattoo parlour on Praed St! Delivering the DNA that expressed the red proteins by tattoo failed to induce any signal, but when we used it to deliver a vaccine, the mice were protected against influenza infection. However, this result needs some caution as the equivalent body surface area in a person would be the entire upper thigh!

Visualising Luciferase transfection in vivo

This work was supported by the Medical Research Council (MRC) as part of a CASE studentship (collaborative awards in science and engineering). These awards are a collaboration between academia and industry, with the industrial partner providing a novel product or technology and expertise in that area and the academic providing the capacity to perform in depth, slightly more esoteric research. The scheme is mutually beneficial as the academic partner gets a student to pursue novel areas and the industrial partner gets to explore angles that would not necessarily fit within the usual timeline constraints of getting products to market. We have been collaborating with a biotech startup called Touchlight Genetics who have a novel process for generating DNA without bacteria. As part of this collaboration, we have previously shown that the DNA is as effective as conventional bacteriaderived DNA and we are working to develop new vaccines with the technology. Overall, we conclude that imaging protein expression from DNA may be useful for some applications, but is poorly predictive of vaccine efficacy.

#DistractinglySexy. Gender Bias in academia.

Distractingly Sexy

An incorrect assertion

“'Let me tell you about my trouble with girls.Three things happen when they are in the lab: you fall in love with them, they fall in love with you, and when you criticise them, they cry." Prof Tim Hunt (attrib).

The first I knew about this story was an email from my Dad saying to be careful of what you say and to whom and the trouble you can get yourself.

Sensible Advice.

That I will now ignore and blindly enter the minefield of writing about women in science as a man. I cannot fully appreciate all of the challenges associated facing women in the workplace and there are obviously better informed people than me who have done more reading and research on both the problems and the solutions. So why write this? Because the system is broken and we need to fix it, because I am inclined to value my own opinion and want to share it (one of the defining features of being an academic) and because I want to add my voice to people saying things need to change.

The fault in our stats

 

Figure 1. Staff at UK HE providers by occupation, age and sex.

Taken from Staff in higher education 2013/14 (https://www.hesa.ac.uk/pr212)

The comments attributed to Professor Hunt reflect a wrong, but not unique view in science and therefore underlie a massive problem in academia. This bias is then further exacerbated by the uneven numbers of women and men in senior academic posts. This data from HESA (Higher education statistics agency) demonstrates the current situation. 45% of academic staff were female, but at senior levels women made up 22% of professors, and 33% of other senior academic staff. The majority (63%) of non-academic staff in 2013/14 were women, including 54% of managerial, professional and technical staff and 82% of clerical staff.

Why do things have to be like this?

A question is why is there such an imbalance? In part it may reflect a historical bias, with the environment in the 1980’s and 1990’s being even less supportive than it is now. I had the privilege of knowing Prof Ita Askonas, who passed away a couple of years ago, a truly remarkable, supportive academic mentor, who rose to the top of her field in a generation when this was far from the norm, but she was one of a very small handful of female academics in her generation. There is some evidence of an upward trend – in 2004 only 21% of clinical academics were women, slowly rising to 26% in 2011 and 28% in 2013, with professorships at 11% in 2004 and at 17% in 2013 (medschools.ac.uk), but it is slow progress.

But historical bias is not the whole story and there are other factors. I would put children and childcare near the top of the list. There is still a perceived bias in expectation towards mums taking time off rather than dads. This is a whole complicated issue and which someone should address separately in a well rounded, but witty blog post . The MRC’s recent change to make fellowship eligibility independent of time post-PhD is a big positive step in this regards. There are some well described unconscious biases in recruitment, job advert writing, interviewing. There are self-reported biases in science by academics of both sexes (good nature op/ed article here). But the prevalence of gender stereotyping may unfortunately be propagating this bias. For example, Larry Summers (then President of Harvard) infamously commented on aptitude and gender (for fuller details see here) and a very brief dip into the interweb brings up, for example, an article about the coconut genomes from The Journal of Proteomics. An unlikely source of sexism, until you see the graphical abstract depicting a woman holding up two coconuts against her chest for no obvious reason (from http://www.stemwomen.net/recognising-sexism/).

I will re-iterate, opinions about the relative aptitudes of people based on what they are, are wrong. People make good (and terrible) scientists, regardless of race, sex, sexual orientation or background. It is also wrong to stereotype about lab personalities, for any given “but type X people are more likely to behave in Y way”, I can give you multiple examples of the opposite. These are wrong views and as trainers of the next generation (and parents of the one after that), we need to dispel them. At the end of the day, science – and the world that depends on scientific progress, needs the best qualified/most able people to deliver it.

It’s my laboratoire

The fact is that for multiple reasons it is harder to be a woman in science. Therefore things need to be done to level the playing field or we will not get the best people. However, it is easy to see why some people may not (consciously or subconsciously) want the status quo to change. Full disclosure: I am a white, privately educated, Oxbridge graduate, man. If 2 men are appointed to every 1 woman, then my chances are much improved. Any levelling of the playing field, could make the path harder for me and those like me. Whilst I assume my genius, social skills and zingy personality got me where I am, I have had every possible advantage from a loving home to extra golf lessons. There are some schemes and support networks, for example Athena Swan and awards targeted at women, or women coming back from childcare breaks but they are few and far between. Sadly, it is not uncommon to hear young male academics/ post docs/ PhD students complaining about how unfair these schemes that preferentially support women are. Interestingly these type of comments tend to tail off with age as these same young men see the problems their friends and loved ones go through juggling children and work. Optimistically, I think these complaints are a direct consequence of the terrible funding situation rather than the next generation of bigots being made. There is so little money available, that any factor that puts you at a perceived disadvantage seems outrageous. I do my best to point out the error of their ways, but it is a view point that needs to be changed. It probably doesn’t help that one of the schemes is sponsored by Loreal (brilliant Mitchell and Webb sketch on this).

We found love in a hopeless place

Was there any truth in the comment: "you fall in love with them, they fall in love with you". Yes, some people do fall in love in lab settings – I met my wife doing my PhD and many of my friends met their partners in science. But then again, many of my other friends met their partners in the army, being doctors, accountants, lawyers etc and then others met in bars, on Tinder, Grinder and Gap years in Africa. If you put people together, some of them will end up sleeping with each other.

Boys don’t Cry

Science is stressful, repetitive and mostly doesn’t work. PhD’s are more so, because you are learning and feel under a time pressure. Hindsight and experience are potent – of course I wouldn’t have done it that way, I would have included that control, I would have put the right chemicals in in the right order. Except when I am actually doing the labwork and adding my own unique cock ups to our long list of ways to fail at ELISA (we are currently at reason 20 – wrong species: long story/ different blog). Bosses are unsympathetic, under stress themselves and often just had a grant rejected. Even worse, Bosses also have a rose tinted view of how easy lab work was in their day, which with incomplete hindsight it possibly was, what was cutting edge to me is now provided in a kit to my students. All of this can add up to toxic interpersonal interactions. Some people cry, some people smash keyboards, things happen, to everyone. So again, an incomplete observation.

Amazing pithy conclusion

Sadly I don’t have one. There is a sex bias in academia (and many professions). It may (or may not) be improving. Changes need to be made, to make it fair so that selection is entirely on aptitude and ability, because science as a career is hard enough, without disadvantaging 50% of scientists. On the minus side there is still bias out there: on the plus side (some) people on twitter are provocatively funny (#Distractinglysexy), reminding us that the more we discuss workplace biases and educate the next generation the less likely it is to be perpetuated.

Post:Script Backlash against 140 characters

I have slightly amended this piece since I first posted it, due to the toxic backlash against Professor Hunt and the complexities of understanding what someone said when you only see a small snippet of it. I put this down to my naivety of social media and not truly understanding the impact it can have until I read Jon Ronson's brilliant - So you've been publicly shamed. The changes I made have left the sentiment intact (academia should be more fair), but in a less accusatory tone.