Some strains of tuberculosis bacteria have evolved to outwit every first line drug we have – requiring lengthy treatment with at least four medicines. With the antibiotic pipeline running dry, Crick researchers are trying to regain the upper hand.
One dot = one antibiotic pill. 54% of people with multidrug-resistant TB globally were being treated for 18-20 months in 2024, with many estimated to take around 4,500 pills, as represented here. (Source: WHO)
On the shelves behind cell biologist Max Gutierrez’s desk sit five conical flasks filled with thousands of multicoloured Smarties. They represent the sheer quantity of pills someone with the disease tuberculosis (TB), which primarily affects the lungs, will have to swallow over six months of treatment.
“This exhausting regime remains the best that modern medicine has to offer,” Max says. “Despite the need for new and better treatments, we’re facing bacteria that fight back and an economic climate where new drugs aren’t typically profitable.”
The Smarties symbolise antibiotics, one of science’s big success stories since the discovery of penicillin. Penicillin was discovered after the accidental contamination of a plate of Staphylococcus with a mould, under the watch of microbiologist and Nobel laureate Sir Alexander Fleming. The industrial production of penicillin by fellow prizewinners Howard Florey and Ernst Chain, and a subsequent suite of first-generation antibiotics, significantly reduced the likelihood of succumbing to bacterial infections or complications after routine surgery. It also built the foundations of the powerhouse pharmaceutical industry we know today. For a time, such discoveries seemed to guarantee a steady pipeline of new cures.
Eighty years later, Fleming might have expected penicillin to have been superseded by a smorgasbord of antibiotic drugs. The big killers of the past, like the bacteria behind urinary tract infections, syphilis or strep throat, are no longer a huge threat due to the development of antibiotics, but we must sober up to the fact that very few antibiotics with truly new mechanisms have emerged over the last few decades.
As of 2021, TB was responsible for 1.3 million deaths a year. But a comparable number of people, 1.14 million, died not from an infectious disease like TB itself, but from the adjacent problem of ‘antimicrobial resistance’: the ability of disease-causing microbes to evolve to resist drugs designed to kill them. The fear of resistance is not new; Fleming warned in his Nobel Prize acceptance speech that misusing penicillin would give bacteria the upper hand. How do we gain back control?
Our relationship with Mycobacterium tuberculosis (Mtb), the bacteria behind TB, has fascinated cell biologist Max over his twenty-year career. Worryingly, some strains of Mtb have acquired resistance to every drug used against them. With a nod to the flasks of Smarties behind him, Max describes that how we treat TB creates a perfect storm for resistance. “People have to take a concoction of drugs with side effects and there are financial and social barriers, even down to the pills being too big to swallow,” he says. “Imperfect adherence allows the emergence of bacteria with resistance genes which can then replicate unrestrained.”
The World Health Organisation’s ‘End TB Strategy’ declared in 2015 that it would pass with flying colours if it reduced the number of TB deaths by 95% and the incidence of TB by 90% by 2035. Max isn’t optimistic that we’ll get there.
He’s just returned from South Africa, meeting local communities through an initiative called Eh!Woza in Khayelitsha, a township in the suburbs of Cape Town with a high unemployment rate and overcrowding. As South Africa has a high number of TB cases for its population size, the initiative seeks to understand the barriers in accessing and maintaining treatment. “I was struck by how so many people who’d had TB really wanted assurance that they wouldn’t get it again,” says Max. “In an endemic setting, people are going to get re-infected, so they’ve got an increased chance of getting multidrug-resistant TB. We also have to face facts that if the disease doesn’t kill people, the toxicity of multiple treatment stints just might.”
It’s well understood that if you reduce poverty, the overall incidence of TB decreases, given that a multi-drug course over many months requires good healthcare and living standards. TB and poverty are so connected that there’s even a phrase, a ‘tuberculogenic environment’, for the deadly mix of overcrowding, poverty and weak health systems that leave people vulnerable to the spread of TB.
It’s also a political problem. Max explains that decades of instability in former Soviet Union countries have left healthcare systems vulnerable to new and reemerging diseases, like multidrug-resistant TB. “I met a TB researcher in South Africa from Ukraine,” Max reflects. “Understandably, antimicrobial resistance isn’t a huge priority given the war and other insurmountable pressures, so sadly cases of multidrug-resistant TB are now on the rise. I’m not sure this is really being talked about.”
Beyond thinking about the sociopolitical problems, daunting biological challenges have kept Max busy since his early days as a biochemistry student. Fresh from an internship at a winery in Argentina, a pivotal moment defined his future career. “We were just becoming aware of the microbiome and a more harmonious relationship with bacteria inside the body,” he says. “I learnt that bacteria can even get inside cells, which really fascinated me: surely it meant that’s how they might be able to evade treatment.”
Mtb takes hold in the cells lining the air sacs in the lungs. The immune system responds as it should, dispatching cells built to swallow and destroy pathogens called macrophages. Mtb has evolved to fight against this too. A hallmark of TB infection, clumps of macrophages, dead bacteria and debris from infected cells form as ‘granulomas’ at the site of the infection. “Mtb has to be very adaptable to the biochemically diverse environments inside lung cells and macrophages,” Max says. “This ability to manipulate diverse conditions is likely why TB is so hard to treat.”
To find out where Mtb hides inside cells, his team uses a new generation of electron microscopy, where electrons fired at a sample can form an image. Volume electron microscopy involves doing this methodically in layers through a tissue and then building back the individual slices to give the 3D shape. It’s the only way to see the individual compartments, like the mitochondria or the nucleus, inside a cell at the nanometre scale. And, for Max, it’s the only way to spy on the TB-host relationship up close.
A few years ago, he developed a new imaging method which combined the resolving power of volume electron microscopy with traditional fluorescent microscopy and drug imaging. This not only allows a zoomed-in snapshot inside a cell, but also allows Max and his team to mark features of interest.
“We analysed lung tissue from mice with TB who had been treated with the antibiotic bedaquiline, and saw that the drug didn’t reach all infected cells. Even when it did reach them, it wasn’t always where it needed to be,” says Max. “So even if the drug was able to cross the cell membrane, it could be nowhere near the bacteria it was meant to kill.” This finding points to a crucial bottleneck: the need to ensure that any new drugs developed actually reach their target.
More work with Max’s imaging method showed that bedaquiline accumulates in lipid droplets. His conclusion was that antibiotics in these fatty reservoirs could be transferred to bacteria, if they consumed the lipid droplets. It could also inspire future drug design to make sure that antibiotics can exist inside a fatty environment. High-resolution imaging, Max argues, was not just useful; it was the only way to see what was happening at all.
Mtb has evolved to keep its friends close and its enemies closer. A landmark study from Max’s lab involved live-imaging of a host macrophage and the Mtb inside it, finding that the macrophage creates an acidic environment in an attempt to restrict the bacteria, but Mtb can maintain its own pH independently to buffer the acidity. Understanding the need to interfere with the bacteria’s pH could help inspire further drug design.
By becoming adapted to a single host, a bacterial interloper can learn the host’s ways and how to manipulate them. In response, a macrophage tries to dispose of the pathogen through an innate immune response. It packages the bacterium in a membrane bubble called a phagosome and drags it toward a lysosome, the cell’s internal incinerator, which breaks down and destroys the contents.
Not only can Mtb disrupt the recycling machinery, it can also push macrophages into a type of uncontrolled cell death called necrosis, which leads to membrane rupture and tissue damage. This allows it to be released from the exploding cell: not at all helpful for controlling infection.
For this reason, Max’s team are advocates for improving the body’s ability to clear waste, finding that two key genes for waste removal are critical for controlling an infection. “Encouraging host cells to more efficiently remove damaged parts, like Mtb-containing phagosomes, could help antibiotics work better, or one day be a treatment in its own right,” Max concludes.
In a moment of reflection in the Crick’s café, chemist and geneticist Eachan Johnson agrees that antimicrobial resistance is a sociopolitical problem. But he believes that, even if we took away the inequities, some infections would remain resistant and untreatable.
Like Max, Eachan also works on mycobacteria, the family that includes Mtb, but also Mycobacterium abscessus, a hard-to-treat pathogen which is an increasingly prominent threat to people with cystic fibrosis. Unlike Max, Eachan focuses on what’s happening within the bacteria themselves.
“The variety in bacterial responses to drugs is so fascinating,” says Eachan. “Mycobacterium abscessus is inherently drug resistant, whereas Mtb is susceptible on paper but develops resistance one by one to most drugs we have. And how each species deals with drugs is so different – some degrade it once it’s inside the cell whereas others batten down the hatches and stop it getting in in the first place.”
Bacteria often become resistant by passing genes which confer a resistant property between themselves, through what’s known as mobile genetic elements. This includes the gene for beta-lactamase, an enzyme that degrades beta-lactam antibiotics like penicillin. This skillsharing between pathogens allows a species to become equipped with a gene it wouldn’t otherwise have.
Mtb is different. “It doesn’t do gene-swapping,” says Eachan. “It picks up resistance by acquiring random but beneficial mutations in its genome. Just 4,000 genes, compared with 20,000 in humans, contain the recipe for resistance to drugs of the past, present and future.”
Mtb’s streamlined genome isn’t gratuitous: by abandoning gene swapping, it’s shed unnecessary genes, honing its genome for one host only. Given how long a TB infection lasts, Mtb colonies that acquire mutations with no clear fitness advantage are simply selected against. Colonies with useful mutations – allowing them to alter a drug target or build ‘efflux pumps’ to discharge antibiotics from the cell – can expand.
Antibiotic discovery has cycled in waves of hopefulness and then disappointment. The first wave relied on chemistry, but the isolation of penicillin sparked a second wave, a golden era, which focused on finding natural products. The problem is that, if it exists in nature, chances are bacteria know about it too. Case in point, genes encoding resistance to beta-lactam antibiotics have been found in 30,000 year-old samples despite beta-lactams only coming online in the early twentieth century.
After the boom in genetic sequencing in the past two decades, a third wave of antibiotic discovery favoured high-throughput screening to find molecules that would neatly block bacterial proteins. A largely failed endeavour, the third wave ended in a discovery bottleneck with few new drugs in the last 10 years working by a new mechanism. For Eachan, this shows that cleverer chemistry is needed to bring an idea for a drug into reality.
“We need to find compounds that are a bit sneakier with unpredictable mechanisms,” he says. “This could involve blocking an enzyme somewhere other than its active site or producing toxic chemicals, called reactive oxygen species, when the drug comes into contact with a bacterial protein.”
Eachan’s lab’s approach sits at the interface of chemistry and genetics, building on that third era of antibiotic research hoping that more modern techniques can deliver results, focusing on TB but also on other hard-to-treat bacterial diseases. In a new project, he’s building chemical libraries of millions of potential molecules that can target fundamental bacterial proteins in Klebsiella pneumoniae, which causes a range of serious infections, including potentially fatal sepsis in newborns. Computational methods will get those 100,000 down to the most promising few hundred.
“We’ll use genetic editing to dial down the expression of every gene in bacteria one by one, making hundreds of mutant strains, and treating them with the potential drug candidates to see which strains are most affected,” says Eachan. “It’s based on the idea that quietening some genes will cause minimal effects to the bacteria, allowing them to continue to spread. We want to avoid developing drugs that target these genes, instead focusing on essential genes that the bacteria can’t live without.”
All current antibiotics in clinical use inhibit biological processes, such as penicillin stopping bacteria building their cell walls. This year, Max’s lab turned that logic on its head, reporting for the first time that boosting synthesis of a key nutrient drove Mtb bacteria into a kind of metabolic overdrive, consuming resources faster than they could be replenished. The bacteria essentially burned out.
It’s precisely this kind of lateral thinking that drew chemical biologist Ester Morreale to the field. Ester, who now runs her own lab at the Crick, can trace her research agenda to a lecture in Dundee she attended as a postdoctoral researcher – a talk that, for her, reframed what fighting back against bacteria even meant.
Still clear in Ester’s memory is structural biologist Tim Clausen describing how he’d found a mechanism in Bacillus subtilis where proteins are marked for degradation by phosphorylation. Until this moment, a drug discovery technology called ‘targeted protein degradation’ had only been outlined in human cells, as a way for cells to tag disease-causing proteins so they can be degraded by complex protein-hoovering machines. It was being tested for cancer treatment, at the time a proof-of-concept enterprise to see whether marking key cancer proteins for removal could halt uncontrolled tumour growth.
The fruits of this labour are now in clinical trials for cancer treatment, but working out that bacteria use a small tag to degrade their own proteins, that could be manipulated too, involved a return to the drawing board. “Bacteria tag and degrade proteins using totally different machinery to human cells, so we couldn’t just translate what we’d learnt in cancer,” says Ester. “This accounts for the lag of several decades in understanding protein degradation tags in microbes.”
As part of her research before joining the Crick, Ester worked with Tim to weaponise this mechanism. They engineered small molecules, dubbed BacPROTACs, that trigger degradation: each one latches onto a critical bacterial protein and drags it to the cell’s disposal machinery, which shreds it. The bacterium, deprived of a protein it cannot do without, stops replicating.
“If we can artificially bring a critical bacterial protein to the cell’s recycling centre then we’ve got a new way to damage a pathogen and stop it replicating,” Ester says. “We tested the degraders in a species of Mycobacterium, finding that they did degrade a protein of interest very well in a test tube, and relatively well inside bacteria.”
Ester is now working on designing new BacPROTACs, improving their abilities inside living bacteria, mainly focusing on Mtb. Excitingly, her team at the Crick has managed to develop degraders that are more effective at killing Mtb than the corresponding conventional inhibitors of the proteins in question. Her team’s search for similar mechanisms in other bacteria like E. coli or Staphylococcus aureus has begun, but it’s an uphill battle, as protein degradation is specific for each type of bacteria.
There is also the problem of engaging the pharmaceutical industry. Ester remains optimistic, but this clearly weighs on her mind. “The degraders we have in the lab right now aren’t drugs; they would need teams working on their optimisation for clinical use,” she says. “Industry investment would help to progress these molecules into potential drug candidates as this isn’t something that academia can do alone.”
Even the most promising ideas, however, must confront a harsh economic reality. Eachan describes the industry’s predicament as ‘use it and lose it’: a new antibiotic, once approved, is held in reserve rather than prescribed, unable to be used because of the inevitable resistance that will develop to it. Companies have spent a billion dollars developing what amounts to an expensive insurance policy. Many small biotech companies that get an antibiotic approved go out of business in a year.
All three researchers are acutely aware of the need for more investment, to quieten some of the nervousness around using these precious medicines. That means more money for basic research and incentives for ‘big pharma’, such as tax considerations, longer patents, improved clinical trial design, or speedier approval, as we’ve seen in the success stories of antiretrovirals or the COVID-19 vaccination.
Incidentally, what about vaccines? “They’re gold standard,” says Eachan. “But I think the ones that can be made easily have been; we’ve got the low-hanging fruit. And with TB it’s even harder, because previous infection doesn’t protect you against a future infection. Instead, I put hope in combinations, mixing drugs in a smart way that reduces treatment times and improves compliance.”
Eachan reflects that the atmosphere at infectious disease conferences can verge on doom and gloom. But there’s reason to be hopeful. Both Ester and Eachan have joined big global consortiums, proof that funding bodies and philanthropists have renewed interest in antimicrobial research. Ester also urges that some healthy competition can go a long way. “I’ve started to see more people pick up protein degradation at conferences, and yes, they’ll be our competitors, but we have to think of the big picture and the benefit to society if we’re all focusing on the same problem,” she says. “We can also encourage people to go into the field not just because it’s a pressing problem – it’s interesting, challenging chemistry that draws in clever people.”
Back at the Crick, Max is still plagued by the question of why it takes six months to clear TB. His findings that the antibiotics we’re relying on might not even reach their targets is worrying, but also gives hope. “We may not need to completely reinvent the wheel: maybe we haven’t yet unlocked the full power of the drugs already in our arsenal,” he says.
The breadth of antimicrobial research suggests, however, that no one approach is likely to be enough. As techniques improve, such as those used by Max, Eachan and Ester, and their colleagues, the solution might lie in a combination of finding new drugs, developing better protocols for existing drugs and stopping bacteria becoming resistant in the first place. One day, the number of drugs needed to treat TB could plummet, doing away with those five flasks in Max’s office.
Eachan allows himself one slightly bleak thought about the short-term nature of success. “Say we’ve ‘cracked’ TB,” he considers. “Our standards will then improve which means something else will take its place.”
The bacteria, in other words, will keep evolving, and so must we. Fleming’s warning of the impermanence of antibiotic effectiveness rings true as the three scientists continue the search for both the underlying principles of resistance and the as-yet-undiscovered chemical remedies to combat it. The challenge is not to win the contest between medicine and bacteria just once, but to change the rules faster than the bacteria can evolve.
