Explore the vast range of titles available.

The spread of insecticide resistance in Anopheles mosquitoes and drug resistance in Plasmodium parasites is contributing to a global resurgence of malaria, making the generation of control tools that can overcome these roadblocks an urgent public health priority. We recently showed that the transmission of Plasmodium falciparum parasites can be efficiently blocked when exposing Anopheles gambiae females to antimalarials deposited on a treated surface, with no negative consequences on major components of mosquito fitness. Here, we demonstrate this approach can overcome the hurdles of insecticide resistance in mosquitoes and drug resistant in parasites. We show that the transmission-blocking efficacy of mosquito-targeted antimalarials is maintained when field-derived, insecticide resistant Anopheles are exposed to the potent cytochrome b inhibitor atovaquone, demonstrating that this drug escapes insecticide resistance mechanisms that could potentially interfere with its function. Moreover, this approach prevents transmission of field-derived, artemisinin resistant P . falciparum parasites ( Kelch13 C580Y mutant), proving that this strategy could be used to prevent the spread of parasite mutations that induce resistance to front-line antimalarials. Atovaquone is also highly effective at limiting parasite development when ingested by mosquitoes in sugar solutions, including in ongoing infections. These data support the use of mosquito-targeted antimalarials as a promising tool to complement and extend the efficacy of current malaria control interventions.

Autophagy-based targeted degradation offers a powerful complement to proteasomal degradation leveraging the capacity and versatility of lysosomes to degrade complex cargo. However, it remains unclear which components of the autophagy-lysosomal pathway are most effective for targeted degradation. Here, we describe two orthogonal induced-proximity strategies to identify autophagy effectors capable of degrading organelles and soluble targets. Recruitment of autophagy cargo receptors, ATG8-like proteins, or the kinases ULK1 and TBK1 is sufficient to trigger mitophagy, while only autophagy cargo receptors capable of self-oligomerization degrade soluble cytosolic proteins. We further report a single-domain antibody against p62 and its use as a heterobifunctional degrader to clear mitochondria. Fusing the p62 single-domain antibody to PINK1 enables selective targeting of damaged mitochondria. Our study highlights the importance of avidity for targeted autophagy and suggests that autophagy cargo receptors are attractive entry points for the development of heterobifunctional degraders for organelles or protein aggregates. Using proximity-based screening, protein engineering, and structural analysis, this study describes the development of a p62-based biodegrader for the clearance of organelles and aggregated proteins by autophagy-targeted degradation.

Targeted protein degradation (TPD) is a novel strategy for developing therapeutics against pathogens. Prior to causing malaria, Plasmodium parasites replicate within hepatocytes as liver stages, surrounded by a parasitophorous vacuole membrane (PVM). We hypothesized that TPD can be employed to trigger host-driven degradation of essential liver stage PVM proteins and lead to parasite death. To explore this, we took advantage of the proteolysis-targeting-chimera HaloPROTAC3, a molecule that recruits the host von Hippel-Lindau (VHL) E3 ligase to the HaloTag (HT). Parasites expressing HT fused to the host cytosol-exposed domain of the PVM protein UIS4 (UIS4-HT) were generated in Plasmodium berghei and Plasmodium cynomolgi , but only P. berghei UIS4-HT enabled productive liver stage infection experiments in vitro. Although HaloPROTAC3 triggered the degradation of HT proteins in host cells, it had no impact on the survival of P. berghei UIS4-HT liver stages. Furthermore, HaloPROTAC3 bound to P. berghei UIS4-HT but did not recruit VHL or trigger ubiquitination of the PVM. Overall, although this study did not establish whether host-driven TPD can degrade Plasmodium PVM proteins, it highlights the challenges of developing TPD approaches against novel targets and offers insights for advancing this therapeutic strategy against pathogens.

The protozoan parasite can induce amebic colitis and amebic liver abscess. First-line drugs for the treatment of amebiasis are nitroimidazoles, particularly metronidazole. Metronidazole has side effects and potential drug resistance is a concern. Schistosomiasis, a chronic and painful infection, is caused by various species of the flatworm. There is only one partially effective drug, praziquantel, a worrisome situation should drug resistance emerge. As many essential metabolic pathways and enzymes are shared between eukaryotic organisms, it is possible to conceive of small molecule interventions that target more than one organism or target, particularly when chemical matter is already available. Farnesyltransferase (FT), the last common enzyme for products derived from the mevalonate pathway, is vital for diverse functions, including cell differentiation and growth. Both and genomes encode FT genes. In this study, we phenotypically screened and with the established FT inhibitors, lonafarnib and tipifarnib, and with 125 tipifarnib analogs previously screened against both the whole organism and/or the FT of and . For , we also explored whether synergy arises by combining lonafarnib and metronidazole or lonafarnib with statins that modulate protein prenylation. We demonstrate the anti-amebic and anti-schistosomal activities of lonafarnib and tipifarnib, and identify 17 tipifarnib analogs with more than 75% growth inhibition at 50 μM against . Apart from five analogs of tipifarnib exhibiting activity against both and , 10 additional analogs demonstrated anti-schistosomal activity (severe degenerative changes at 10 μM after 24 h). Analysis of the structure-activity relationship available for the FT suggests that FT may not be the relevant target in and . For , combination of metronidazole and lonafarnib resulted in synergism for growth inhibition. Also, of a number of statins tested, simvastatin exhibited moderate anti-amebic activity which, when combined with lonafarnib, resulted in slight synergism. Even in the absence of a definitive molecular target, identification of potent anti-parasitic tipifarnib analogs encourages further exploration while the synergistic combination of metronidazole and lonafarnib offers a promising treatment strategy for amebiasis.

Malaria is a devastating disease that affects hundreds of millions of people worldwide, leading to nearly 600,000 deaths in 2023. Plasmodium falciparum parasites are the causative agent of over 90% of malaria cases and are transmitted to people by the bite of an infectious Anopheles female mosquito. Vector control strategies that target these mosquitoes have been instrumental to control efforts, and the extensive rollout of insecticide treated bed nets has contributed to substantial reductions in malaria prevalence and mortality since the turn of the century. However, high levels of insecticide resistance have emerged in anophelines, and to-date, insecticide-resistant mosquitoes have been identified in over 80% of malaria endemic countries. This widespread resistance jeopardizes the efficacy of current vector control tools and has contributed to the recent plateau in malaria cases worldwide. New interventions are thus urgently needed.In this dissertation, I describe our efforts to develop a novel transmission blocking strategy that circumvents insecticide resistance by targeting parasites directly with antimalarial compounds during their development in the mosquito. In Chapter 1, I review the current state of malaria control and promising new tools, with an emphasis on vector control. I introduce the concept of mosquito-targeted antiplasmodials and outline key features for their successful implementation. In Chapter 2, I identify suitable targets for this strategy by performing an in vivo screen of antiplasmodial compounds in mosquitoes, optimize key inhibitors for improved uptake and activity, and assess their resistance propensity and transmissibility. In Chapter 3, I test the feasibility of incorporating hit compounds in bed net-like materials, determine their antiplasmodial activity in insecticide-resistant mosquitoes, and assess their activity across sporogonic development. Finally, in Chapter 4 I discuss implications of this work and directions for further development of this strategy. Altogether, this dissertation significantly expands our understanding of mosquito-targeted antiplasmodials and demonstrates their potential for malaria control in the context of widespread insecticide resistance.

In the search for new anti-schistosomal agents, a series of fifteen ortho-nitrobenzyl derivatives was assayed in vitro against both the schistosomulum (somule) and adult forms of Schistosoma mansoni. Compounds 8 and 12 showed significant activity against somules at low micromolar concentrations, but none was active against adults. The SAR demonstrated that the compounds most active against the parasite were mutagenic to the human cell line RKO-AS45-1 only at concentrations 10- to 40-fold higher than the worm-killing dose. Given their electrophilicity, compounds were also screened as inhibitors of the S. mansoni cysteine protease (cathepsin B1) in vitro. Amides 5 and 15 exhibited a modest inhibition activity with values of 55.7 and 50.6 % at 100 µM, respectively. The nitrobenzyl compounds evaluated in this work can be regarded as hits in the search for more active and safe anti-schistosomal agents.

The human malaria parasite, , relies exclusively on mosquitoes for transmission. Once ingested during blood feeding, most parasites die in the mosquito midgut lumen or during epithelium traversal . How surviving ookinetes interact with midgut cells and form oocysts is poorly known, yet these steps are essential to initiate a remarkable growth process culminating in the production of thousands of infectious sporozoites . Here, using single-cell RNA sequencing of both parasites and mosquito cells across different developmental stages and metabolic conditions, we unveil key transitions and mosquito-parasite interactions occurring in the midgut. Functional analyses uncover processes regulating oocyst growth and identify the transcription factor SIP2 as essential for sporozoite infection of human hepatocytes. Combining shared mosquito-parasite barcode analysis with confocal microscopy, we reveal that parasites preferentially interact with midgut progenitor cells during epithelial crossing, potentially using their basal location as an exit landmark. Additionally, we show tight connections between extracellular late oocysts and surrounding muscle cells that may ensure parasites adherence to the midgut. We confirm our major findings in several mosquito-parasite combinations, including field-derived parasites. Our study provides fundamental insight into the molecular events characterizing previously inaccessible biological transitions and mosquito-parasite interactions, and identifies candidates for transmission-blocking strategies.

Abstract The spread of insecticide resistance in Anopheles mosquitoes and drug resistance in malaria parasites is contributing to a global resurgence of malaria, and the generation of control tools that can overcome these issues is an urgent public health priority. We recently demonstrated that the transmission of Plasmodium falciparum parasites by Anopheles gambiae can be efficiently blocked when female mosquitoes contact the antimalarial atovaquone deposited on a treated surface, with no negative consequences on mosquito fitness. Here, we demonstrate that the transmission-blocking efficacy of mosquito-targeted atovaquone is maintained when highly insecticide resistant, field-derived Anopheles mosquitoes are exposed to this antimalarial, indicating that insecticide resistance mechanisms do not interfere with drug function. Moreover, this approach prevents transmission of field P. falciparum isolates, including an artemisinin resistant Kelch13 C580Y mutant, demonstrating that this strategy could prevent the spread of parasite mutations that induce resistance to front-line antimalarials. Finally, we show that ingestion of atovaquone in sugar solution is highly effective at limiting parasite development, including in ongoing infections. These data support the use of mosquito-targeted antimalarial interventions to potentiate and extend the efficacy of our best malaria control tools. Competing Interest Statement The authors have declared no competing interest.

Introduction Intraligamentary correctional operations like a high tibial osteotomy were performed in genua valga to prevent later medial gonarthrosis especially in younger patients. An unwanted effect of this method seems to be the inferiorization of the patella. This is feared because of the complications in case of subsequent alloarthroplasty. Besides the classical Coventry method as a subtractive osteotomy the hemicallotasis has been established as a sustainable additive procedure. This means a gradual open wedge correction using an external fixateur. Objective The aim of this study was to determine the position of the patella pre- and postoperatively and in follow-ups with subtractive versus additive intraligamentary high tibial osteotomies on the basis of five radiological parameters. It was expected that an additive osteotomy leads to an inferiorized patella position whereas a subtractive osteotomy leads postoperative to a higher position of the tibia. Method Between 1990 and 2001, 54 patients (61 legs) had undergone an operation due to a genu varum either by the subtractive osteotomy ( n  = 30) according to Coventry’s method or the additive gradually hemicallotasis ( n  = 31) with an external fixator. Results In coherence with the Coventry’s osteotomy a significant inferiorization of the postoperative patella position with all five radiological parameters was observed, the hemicallotasis showed no operation-related significant alteration of the patella height. Instancing the Insall–Salvati Index there were four (12.9%) preoperative and three (9.7%) postoperative patella baja positions detected. Along with the subtractive osteotomy there were 5 preoperative patellae baja (16.7%) and 11 postoperative patellae baja (36.7%) positions. Furthermore a significant interrelation was noticed between the extent of the correctional angle and the postoperative alteration of the patella. Conclusion The results are surprising, contrary was expected. First this can be explained by its gradual, additive correctional property in contrast to the spontaneous correction by the conventional method according to Coventry, second by the postoperative treatment, which allows an early mobilization and active remedial gymnastics, provided an impact resistant osteosynthesis by a fixateur externe is given. In the case of the additive hemicallotasis an intraligamentary osteotomy is recommended. Technically expensive step cuts in order to osteotomize below the tuberositas tibiae are not necessary. Due to the low quota of complications and the small operative expense the continuous distraction is preferential to ad hoc correction. A postoperative patella baja position has not to be afraid in hemicallotasis.

Harnessing autophagy for targeted degradation is a promising extension to proteasome-based 18 targeted protein degradation because of the capacity and versatility of lysosomes to degrade large and 19 complex cargo, thus broadening the scope of therapeutic targets. While small-molecule degraders 20 recruiting the autophagy machinery to targets are starting to emerge, it remains unclear which component 21 of the autophagy lysosomal pathway is most efficacious to induce selective target degradation. Here, we 22 describe two orthogonal induced-proximity strategies to identify and prioritize autophagy effectors that are 23 sufficient to degrade organelles as well as soluble targets. We show that induced proximity of different 24 effectors such as autophagy cargo receptors, ATG8-like proteins or the kinases ULK1 and TBK1 are 25 sufficient to trigger mitophagy. In contrast, self-oligomerizing autophagy cargo receptors outperform ATG8-26 like effectors and autophagy-related kinases in clearing a soluble cytosolic protein. We further explore the 27 importance of avidity for targeted degradation via autophagy and reveal that the PB1 domain of p62 fused 28 to a LIR peptide is a minimal degron to induce the degradation of mitochondria as well as cytosolic proteins. 29 By developing a novel and highly selective intrabody against the autophagy cargo receptor p62 into a 30 heterobifunctional degrader, we demonstrate that recruitment of endogenous p62 is sufficient to clear 31 mitochondria. This biodegrader, however, is unable to induce degradation of soluble cytosolic proteins due 32 to its inhibitory effect on p62 self-oligomerization. Our study highlights the importance of avidity and 33 suggests that autophagy cargo receptors are attractive entry points for the development of 34 heterobifunctional degraders for complex targets such as organelles or protein aggregates.Competing Interest StatementZ.T, D.M., C.M., N.S., G.S., L.T., A.P., C.S.M., N.G., M.V., S.S.m A.L., J.-Y.C., L.F., P.H., C.B., F.V., S.G., M.M., F.F., G.M., and B.N. are employees of Novartis Pharma. B.G., L.E., and R.M. are former employees of Novartis.