Flavonoids represent a potential source of new antitrypanosomatidic leads. Starting from a library of natural products, we combined target-based screening on pteridine reductase 1 with phenotypic screening on Trypanosoma brucei for hit identification. Flavonols were identified as hits, and a library of 16 derivatives was synthesized. Twelve compounds showed EC50 values against T. brucei below 10 μM. Four X-ray crystal structures and docking studies explained the observed structure-activity relationships. Compound 2 (3,6-dihydroxy-2-(3-hydroxyphenyl)-4H-chromen-4-one) was selected for pharmacokinetic studies. Encapsulation of compound 2 in PLGA nanoparticles or cyclodextrins resulted in lower in vitro toxicity when compared to the free compound. Combination studies with methotrexate revealed that compound 13 (3-hydroxy-6-methoxy-2-(4-methoxyphenyl)-4H-chromen-4-one) has the highest synergistic effect at concentration of 1.3 μM, 11.7-fold dose reduction index and no toxicity toward host cells. Our results provide the basis for further chemical modifications aimed at identifying novel antitrypanosomatidic agents showing higher potency toward PTR1 and increased metabolic stability.

Adenoid cystic carcinoma (ACC) is a rare cancer that preferentially occurs in the head and neck, breast, as well as in other sites. It is an aggressive cancer with high rates of recurrence and distant metastasis. Patients with advanced disease are generally incurable due to the lack of effective systemic therapies. Activation of the master transcriptional regulator MYB is the genomic hallmark of ACC. MYB activation occurs through chromosomal translocation, copy number gain or enhancer hijacking, and is the key driving event in the pathogenesis of ACC. However, the functional consequences of alternative mechanisms of MYB activation are still uncertain. Here, we show that overexpression of MYB or MYB-NFIB fusions leads to transformation of human glandular epithelial cells in vitro and results in analogous cellular and molecular consequences. MYB and MYB-NFIB expression led to increased cell proliferation and upregulation of genes involved in cell cycle control, DNA replication, and DNA repair. Notably, we identified the DNAdamage sensor kinase ATR, as a MYB downstream therapeutic target that is overexpressed in primary ACCs and ACC patient-derived xenografts (PDXs). Treatment with the clinical ATR kinase inhibitor VX-970 induced apoptosis in MYBpositive ACC cells and growth inhibition in ACC PDXs. To our knowledge, ATR is the first example of an actionable target downstream of MYB that could be further exploited for therapeutic opportunities in ACC patients. Our findings may also have implications for other types of neoplasms with activation of the MYB oncogene.

Graphical abstract

The mechanisms underlying the drug resistance of Leishmania spp. are manifold and not completely identified. Apart from the highly conserved multidrug resistance gene family known from higher eukaryotes, Leishmania spp. also possess genus-specific resistance marker genes. One of them, ARM58, was first identified in Leishmania braziliensis using a functional cloning approach, and its domain structure was characterized in L. infantum. Here we report that L. infantum ARM58 is part of a gene cluster at the telomeric end of chromosome 34 also comprising the neighboring genes ARM56 and HSP23. We show that overexpression of all three genes can confer antimony resistance to intracellular amastigotes. Upon overexpression in L. donovani, ARM58 and ARM56 are secreted via exosomes, suggesting a scavenger/secretion mechanism of action. Using a combination of functional cloning and next-generation sequencing, we found that the gene cluster was selected only under antimonyl tartrate challenge and weakly under Cu 2؉ challenge but not under sodium arsenite, Cd 2؉ , or miltefosine challenge. The selective advantage is less pronounced in intracellular amastigotes treated with the sodium stibogluconate, possibly due to the known macrophage-stimulatory activity of this drug, against which these resistance markers may not be active. Our data point to the specificity of these three genes for antimony resistance. P arasitic protozoa of the genus Leishmania, order Trypanosomatida, are responsible for the various clinical manifestations of leishmaniasis. The disease is transmitted by the bite of sandflies, with 2 million new infections occurring per year, and occurs in 98 countries on five continents (1). Leishmania spp. exist in two morphologically distinct life cycle stages. In the transmitting sandflies, the flagellated and elongated promastigotes proliferate attached to the epithelium of the digestive tract until they reach a high density. A switch in the surface molecule composition then causes detachment, and the so-called metacyclic promastigotes are free to invade a mammalian host when the sandfly takes a blood meal. Inside the mammalian skin, the parasites are quickly taken up by antigen-presenting cells, such as dendritic cells, neutrophilic granulocytes, and macrophages, and establish themselves in these cells as small, ovoid, aflagellated amastigotes. The resulting destruction of infected macrophages causes inflammatory immune responses and the concomitant immune pathologies.Depending on the infecting species and on the host's immune status, Leishmania-related pathologies range from localized, ulcerating skin lesions (cutaneous leishmaniasis [CL]) and diffuse cutaneous lesion formation (diffuse cutaneous leishmaniasis [DCL]) to mucocutaneous leishmaniasis (MCL) and, lastly, a generalized infection known as kala azar (visceral leishmaniasis [VL]). VL is invariably lethal in untreated cases, while MCL can also have lethal outcomes due to secondary infections of the nasopharyngeal area.As there is no vaccine against Leishmania a...