Strain diversity in Mycobacterium tuberculosis (Mtb) underlies distinct clinical presentations and outcomes, but the range of drug susceptibility phenotypes among clinical isolates is poorly understood. We aimed to identify drug response patterns in phylogenetically diverse clinical isolates to combination treatment. We selected 13 strains out of 641 drug-sensitive clinical isolates that capture local and global phylogenetic diversity and included Erdman ATCC-35801 as a reference. We treated each strain with 10 single drugs, 45 drug pairs, and 20 three-way combinations in standard and cholesterol-rich media. Mtb clinical strains displayed a broad range of drug response phenotypes across the 65 drug combinations and two metabolic conditions tested, with the most effective drug pairs (based on potency and synergy) varying by strain and metabolic condition. Within our 14-strain panel, strains that were less sensitive to single drugs were also less sensitive to combination treatment, with very few exceptions. For all drug combinations tested, the variation in combination potency was driven primarily by variation among genetically related strains, rather than between strains belonging to disparate lineages. Preclinical regimen design should reflect the diversity of Mtb clinical strains; our data suggest that selecting strains based on the range of drug response phenotypes displayed, rather than by genetic diversity alone, may better account for pathogen diversity. Our findings also show that constituent drug pairs of high-order combinations can be differentially effective against Mtb adapted to different carbon sources. Selection of these pairs should likely involve multiple factors including the infecting strain, metabolic niche, and drug response metrics.

Twelve pairs of baseline and post-exposure Pseudomonas aeruginosa isolates from patients treated with ceftazidime-avibactam were evaluated to define mechanisms of treatment-emergent resistance. Resistance was associated with amino acid substitutions in ampC and OXA β-lactamases, or mutations in regulatory genes conferring hyper-production of AmpC and MexAB-OprM efflux pumps. Cross-resistance was common between ceftazidime-avibactam and ceftolozane-tazobactam, less common for imipenem-relebactam and cefepime-zidebactam, and lowest for cefiderocol. These findings have important implications for sequential treatment of P. aeruginosa infections.

Nacubactam is a novel bicyclic non-β-lactam β-lactamase inhibitor that acts as a penicillin-binding protein (PBP-2) active antibacterial, almost exclusively cleared by renal excretion. This study assessed the effect of renal function and hemodialysis on nacubactam pharmacokinetics following a single intravenous (IV) 1 g infusion. Part 1 included three subjects with normal renal function, and six each with mild, moderate, or severe renal impairment; Part 2 included eight subjects with end-stage renal disease undergoing hemodialysis. We observed a strong linear relationship between renal function and both nacubactam total clearance and renal clearance. Compared with subjects with normal renal function, mean nacubactam total clearance was 49%, 73%, and 85% lower among subjects with mild, moderate, and severe renal impairment, respectively, and renal clearance was 42%, 70%, and 90% lower. Nacubactam systemic exposure was correspondingly greater in subjects with impaired renal function. Compared with the normal renal function group, AUC0-inf was 2-fold, 3.7-fold, and 6.6-fold higher in the mild, moderate, and severe impairment groups, respectively. Similar results were observed across four different methods of estimating renal clearance. Nacubactam was cleared by hemodialysis, with more than 60% of a single dose eliminated during a 4-h dialysis session. Renal impairment had no apparent effect on the safety profile of nacubactam, and a single 1 g dose was well tolerated in all groups. These data can be used to underwrite nacubactam dosing recommendations for patients with impaired renal function, including those receiving renal replacement therapy.CLINICAL TRIALSThis study is registered with ClinicalTrials.gov as NCT02975388.

Management of pulmonary mucormycosis includes intravenous liposomal amphotericin-B (L-AmB), followed by posaconazole or isavuconazole. We aimed to determine the optimal L-AmB duration before switching to triazole therapy. Using data from a retrospective study in France, we performed an emulated trial evaluating stopping L-AmB after 14 days (short treatment) versus continuing L-AmB (long treatment). The benefit of combination L-AmB therapy was also investigated. A cloning, censoring, and weighting approach was used to account for immortal time and indication biases. Ninety-four cases of pulmonary mucormycosis were included, of which 35% had dissemination. Eighty-nine patients received L-AmB first-line therapy for a median of 25 (IQR: 12-54) days, including 18 patients as combination therapy. L-AmB was discontinued for adverse events in 16/89 patients (18%). Treatment was switched to posaconazole (n = 38) or isavuconazole (n = 9) in 47 cases and was switched back to L-AmB due to disease progression in 11/47 (23%) cases. Overall, the 180-day mortality was 54%. Diagnosis in the ICU, dyspnea at diagnosis, disseminated disease, and ground-glass opacities on chest CT scan were associated with increased mortality. No difference in 180-day survival was found between patients receiving short or long L-AmB treatment (HR=0.80, 95% CI [0.29-1.99]) and between patients receiving L-AmB alone or in combination (HR=1.14, 95% CI [0.56-2.32]) in emulated trials, although confidence intervals were wide. Switching to triazole therapy after 14 days of L-AmB treatment was as effective as longer durations while reducing L-AmB side effects. Combination therapy did not improve prognosis. These results could inform randomized clinical trials assessing different first-line therapeutic strategies for pulmonary mucormycosis.

Human babesiosis is an emerging tick-borne disease caused by Babesia parasites, most notably Babesia microti and Babesia duncani in North America and Babesia divergens in Europe. Infections can be severe or persistent or relapse despite treatment, and current therapeutic options remain limited, underscoring the urgent need for new and effective treatment options. Acridone derivatives originally developed as potent antimalarial agents against multiple life cycle stages of the malaria parasites were evaluated for their antibabesial activity using continuous in vitro culture systems of B. duncani and B. divergens. Lead candidates were assessed for selectivity against human cell lines to establish preliminary safety profiles, and select compounds were further advanced into preliminary in vivo efficacy studies using murine models of B. duncani and B. microti babesiosis to assess their therapeutic potential. A set of prioritized acridone derivatives demonstrated potent in vitro activity against both B. duncani and B. divergens and displayed favorable selectivity indices relative to human cell lines. However, in vivo evaluation of representative compounds did not achieve parasite clearance in murine models. Structure-activity relationship (SAR) analyses highlight key structural features that are critical for maintaining antibabesial potency and offer guidance for further lead optimization. Acridone derivatives show strong in vitro antibabesial activity and represent promising lead chemotypes for therapeutic development. To advance these candidates, future studies focused on optimizing their pharmacokinetic properties and evaluating synergistic combination regimens will be essential for progressing toward effective treatments for human babesiosis.

Malaria, particularly by Plasmodium vivax, remains a significant public health challenge in endemic regions. P. vivax has the dormant liver-stage hypnozoites that can reactivate, causing relapses, even after the initial blood-stage infection has been cleared. The prevention of these relapses hinges on the use of 8-aminoquinolines such as primaquine (PQ) and tafenoquine (TQ). However, the efficacy of these treatments is associated with Cytochrome P450 2D6 (CYP2D6) activity, which influences drug metabolism. This case-control study aimed to evaluate the relationship between CYP2D6 activity and the efficacy of TQ and PQ in preventing P. vivax relapses. Patients with P. vivax malaria were included in a nested case-control cohort study conducted at the Fundação de Medicina Tropical Dr. Heitor Vieira Dourado (Manaus, AM, Brazil). Participants were classified as cases (recurrent malaria within 180 days) or controls (patients without recurrence). They were treated with PQ or TQ. Epidemiological data were collected from participants, and blood samples were obtained for CYP2D6 genotyping by real-time PCR. The results showed that CYP2D6 polymorphisms did not affect TQ efficacy (P = 0.172). However, participants treated with PQ had a higher risk of recurrence until six months (P = 0.016). Those with a predicted intermediate or poor metabolizer phenotype (gIM + gPM) were more likely to experience recurrence compared to normal metabolizers (gNM) (P = 0.036). Our findings indicate that CYP2D6 does not impact the efficacy of TQ, whereas P. vivax recurrence is associated with predicted metabolic phenotype, with increased risk among patients treated with PQ, suggesting that CYP2D6 variability influences treatment response, particularly in intermediate/poor metabolizers.

ABSTRACT Antimicrobial central venous catheters (CVCs) help prevent CVC-related bloodstream infection (CRBSI), and their effectiveness depends on sustaining broad antimicrobial activity in serum. A new CVC containing minocycline, rifampin, and chlorhexidine (MRC) was recently approved, complementing existing minocycline-rifampin (MR) and chlorhexidine-silver-sulfadiazine (CSiSz) CVCs. This study compared the elution profiles of minocycline, rifampin, and chlorhexidine from these three CVC types in serum and assessed resulting antimicrobial durability against CRBSI pathogens. MRCs were prepared by impregnation with minocycline and rifampin, followed by sequential chlorhexidine coating; commercial MR and CSiSz CVCs served as comparators. CVCs were immersed in plasma for 24 h, and then in serum for 3 weeks. At baseline and weekly intervals, CVC segments were analyzed for antimicrobial content and challenged with clinical CRBSI isolates. MRC retained higher minocycline and rifampin levels than MR and higher chlorhexidine levels than CSiSz after 3 weeks in serum. Correspondingly, MRC demonstrated superior inhibition of gram-negative colonization ( Klebsiella pneumoniae , Escherichia coli , Acinetobacter baumannii ) and Enterococcus faecalis at week 3 ( P ≤ 0.028). They also showed greater activity against Candida species (spp.) ( C. albicans , C. glabrata ) at weeks 2 and 3 ( P ≤ 0.005). MR and MRC CVCs completely prevented methicillin-resistant Staphylococcus aureus and S. epidermidis colonization for 3 weeks, outperforming CSiSz ( P = 0.003). In conclusion, slow elution from MRC CVCs resulted in significantly enhanced antimicrobial durability against gram-negative bacteria, E. faecalis , and Candida spp. MR and MRC CVCs provided equivalent, sustained protection against Staphylococcal colonization, both surpassing CSiSz CVCs.

ABSTRACT Olorofim is a novel oral antifungal agent which selectively inhibits the fungal enzyme dihydroorotate dehydrogenase found in molds including Aspergillus , Scedosporium , Lomentospora , and Scopulariopsis and in dimorphic fungi such as Coccidioides . Based on in vitro studies, clinical pharmacology, and pharmacokinetic (PK) data from the Phase 2b FORMULA study (F901318/0032) as well as physiologically based PK (PBPK) modeling of drug interactions, the drug’s CYP-mediated interactions are now well understood. Olorofim is mainly metabolized by cytochrome P450 (CYP) CYP3A4, with minor contributions from CYP2C8 and CYP2C9. In vitro studies found that it inhibits CYP3A4, weakly inhibits CYP2D6 and CYP2C8, and weakly induces CYP1A2 and CYP2B6. Phase I studies confirm it is predominantly cleared by CYP3A4 and weakly inhibits CYP3A4. In the Phase 2b FORMULA study, co-administered sensitive CYP3A4 substrates required limited dose reductions when using olorofim. When used with strong CYP3A4 inhibitors, dual moderate inhibitors like fluconazole, or CYP3A4 inducers, dose adjustments were needed for olorofim. Physiologically based PK (PBPK) modeling predicted moderate effects with CYP3A4 inhibitors or inducers and negligible effects with weak CYP3A4 inhibitors or strong inhibitors of CYP2D6, CYP2C8, and CYP2C9. Overall, the drug’s interaction profile is predictable and manageable, making it a suitable candidate drug to use in treating patients with invasive fungal disease (IFD), who often require polypharmacy. CLINICAL TRIALS This study is registered with ClinicalTrials.gov as NCT02680808 , NCT02730442 , NCT02737371 , NCT04171739 , NCT03340597 , and NCT03583164 .

Severe acute respiratory syndrome coronaviruses use the ACE2 receptor for viral entry while downregulating its activity, potentially leading to hypertension and major organ injuries. Dual-action technologies based on soluble ACE2 aimed to neutralize the virus while restoring ACE2's normal enzymatic activity. Here, we describe a novel general molecular design, VHHSpike-ACE2ECD-VHHAlbumin, in the toolbox of ACE2-centric therapeutic modalities. The optimized nanobody module VHHSpike afforded strong pan-specific binding against the entire sarbecoviral clade. This correlated with potent in vitro neutralization of pseudotyped virus variants of concern, with IC50 values in the picomolar range. Exogenous enzymatic activity was provided by the ACE2ECD module, which also contributed binding avidity via intrinsic homodimerization. Persistence of enzymatic activity in circulation was increased in vivo via the nanobody module VHHAlbumin optimized for serum albumin binding. Single-dose therapeutic administration of lead compound 72opt-ACE2-R28 demonstrated virus neutralization in lungs of hamsters at day 5 post-infection with SARS-CoV-2. In hypertensive mice maintained under continuous injection of angiotensin II, a single dose of 72opt-ACE2-R28 normalized systolic blood pressure, maintaining a 30 mmHg reduction after 24 h. Overall, encouraging coronavirus neutralization and hypertension reduction showed magnitudes and timeframes appropriate for treatment of typical acute infections. We discuss molecular bases of interactions with coronavirus spike protein molecules and future scale-up manufacturability toward clinical development of this modular design scaffold with high potential against emerging SARS-CoV-2 variants.