Do antibiotic combinations proposed from in vitro studies lead to changes in treatments?
– by Angela Huttner

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Yes, they do. That is the simple answer to the question I will be focusing on in this viewpoint. The next question is more difficult: Should they? Should an antibiotic combination be given to a patient because it was seen to be effective in in vitro experiments? The answer there is neither simple nor satisfying, but it is clear.

In vitro studies remove the complexity and nuance of a messy world to provide simple but important mechanistic information. In the reductive world of bacterium versus antibacterial, we gain valuable insights into which antibiotics work to kill or inhibit the growth of bacteria, which do not, and which work even better when applied in combination.

The urge to add a second antibiotic

In the clinic, when your previously healthy patient enters septic shock, those insights make it hugely tempting to add a second antibiotic. There is evidence that combination therapy is more effective at bacterial destruction than your single antibiotic alone. Meanwhile, the additional agent is likely to be market-approved with a known safety profile, making risks to your patient seem minimal. In those challenging moments when you watch your patient deteriorate and fall out of your control, it may feel almost unethical to deny her this one last chance. So you add the antibiotic. A few hours later, your patient’s blood pressure begins to stabilize. A few hours more, and she no longer requires supplemental oxygen. Still later, she has left the intensive-care unit and will make a full recovery.

That is a story whose simplicity matches that of the stories told by in vitro experiments. All of them tell a truth, one truth, but none of them tell the whole truth. In in vitro experiments, important variables are left out of the equation. In clinical anecdotes such as this one, important biases are left in.

The truest answer to this question is that we need more clinical data. We should not be fooled by our anecdotal experiences, and we should not let them stand in for clinical evidence.

The whole truth is that combination therapy certainly puts patients under higher selection pressure for resistant bacterial strains and increased risk for side effects, and that we do not yet know how or whether most antibiotic combinations improve clinical outcomes. The part of the Hippocratic oath, which instructs to ‘first do no harm’, should push us clinicians to produce that missing clinical evidence. This applies especially to those of us in teaching hospitals, where clinical research can be integrated into daily clinical activity.

What in vitro studies can miss

By definition, in vitro observations leave out important elements of the story, a chief one being host factors. Immune responses are not assessed. The dynamic conditions of the anatomic site of infection are also typically omitted: environmental pH, which influences antibiotic activity, the abundance or dearth of iron, necessary for bacterial survival, and so on.^1,2 While host factors are not accounted for, many bacterial factors cannot be fully captured either. The development of biofilms and their subsequent effects may take more time than that allotted for the in vitro experiment. Bacterial escape mechanisms, such as persistence and tolerance, may not be picked up in in vitro experiments.³ Finally, antibiotic properties are also not fully accounted for.⁴ How well does the antibiotic penetrate the tissue where the infection is occurring? How protein-bound will the antibiotic be in vivo? Indeed, well-powered randomized clinical trials to date have not returned positive results for combination therapy. The AIDA trial could not show a benefit in adding meropenem to colistin for severe infections due to carbapenem-resistant Gram-negative organisms.⁵ The CAMERA2 trial could not show a benefit in adding an anti-staphylococcal beta-lactam antibiotic to vancomycin or daptomycin for staphylococcal bloodstream infections; indeed, the trial had to be stopped early because of increased kidney injury in the combination group.⁶ The SAFO⁷ and ARREST⁸ trials could not show a benefit in adding fosfomycin to cloxacillin or rifampicin to standard antibiotic therapy, respectively, for these infections either.

Should an antibiotic combination be given to a patient because it was seen to be effective in in vitro experiments? The answer there is neither simple nor satisfying, but it is clear.

Clinical efficacy is not the only uncertainty. The safety profile of two antibiotics in combination is not simply the sum of their individual safety profiles, as demonstrated by the CAMERA2 trial. As another example, combining intravenous fosfomycin with, say, flucloxacillin would subject patients to higher levels of fluid overload that could much more easily tip them into pulmonary edema than either antibiotic alone.⁹

What we really need

The truest answer to this question is that we need more clinical data. We should not be fooled by our anecdotal experiences, and we should not let them stand in for clinical evidence. Our biases—and our wishful thinking—may lead us to think that our one-off clinical interventions are safe and effective. In the particular scenario described above, the patient was not responding to antibiotic monotherapy because she had not yet entered steady state. She began to improve once she was in steady state and the single antibiotic had reached therapeutic levels in tissue. The second antibiotic was not going to, and did not, change her trajectory.

We need more clinical evidence from randomized trials with sufficient statistical power to confirm (or refute) any clinical synergy with combination therapy. In Geneva, we will soon launch a randomized trial comparing ceftazidime alone to ceftazidime with fosfomycin in patients with severe bacterial infections.¹⁰ We hope for superior clinical outcomes which is why we will include a placebo and blind both ourselves and participants. We also know, however, that we may not see them— and may well see increased side effects instead.

Yet no matter the outcome, at the end of the trial we will know the answer to “Should they?” That is, we will know it for that particular antibiotic combination. The answer is not simple or satisfying—or ‘one size fits all’. The answer is that we need good-quality clinical data for each combination proposed by those in vitro studies before we can know what should, or should not, be done for our patients.

References: 

1. Kincses A, Racz B, Baaity Z, et al. The Relationship between Antibiotic Susceptibility and pH in the Case of Uropathogenic Bacteria. Antibiotics. 10(12) 1431.
2. Vollenweider V, Rehm K, Chepkirui C, et al. Antimicrobial activity of iron-depriving pyoverdines against human opportunistic pathogens. Elife. 13:RP92493.
3. Eisenreich W, Rudel T, Heesemann J, Goebel W. Link Between Antibiotic Persistence and Antibiotic Resistance in Bacterial Pathogens. Front Cell Infect Microbiol. 2022;12:900848.
4. Tangden T. Combination antibiotic therapy for multidrug-resistant Gram-negative bacteria.
   Ups J Med Sci. 119(2):149-53.
5. Paul M, Daikos GL, Durante-Mangoni E, et al. Colistin alone versus colistin plus meropenem for treatment of severe infections caused by carbapenem-resistant Gram-negative bacteria: an open-label, randomised controlled trial. 18(4):391-400.
6. Tong SYC, Lye DC, Yahav D, et al. Effect of Vancomycin or Daptomycin With vs Without an Antistaphylococcal beta-Lactam on Mortality, Bacteremia, Relapse, or Treatment Failure in Patients With MRSA Bacteremia: A Randomized Clinical Trial. JAMA. 323(6):527-537.
7. Grillo S, Pujol M, Miro JM, et al. Cloxacillin plus fosfomycin versus cloxacillin alone for methicillin-susceptible Staphylococcus aureus bacteremia: a randomized trial. Nat Med. 29(10):2518-2525.
8. Thwaites GE, Scarborough M, Szubert A, et al. Adjunctive rifampicin to reduce early mortality from Staphylococcus aureus bacteraemia: the ARREST RCT. Health Technol Assess. 22(59):1-148.
9. Zirpe KG, Mehta Y, Pandit R, et al. A Real-world Study on Prescription Pattern of Fosfomycin in Critical Care Patients. Indian J Crit Care Med. Sep 25(9):1055-1058.
10. JPI-AMR (2023) CAVIFOS, a JPI-AMR project. [Accessed 2024-07-08].