AI is a viable alternative to high throughput screening: a 318-target study

: High throughput screening (HTS) is routinely used to identify bioactive small molecules. This requires physical compounds, which limits coverage of accessible chemical space. Computational approaches combined with vast on-demand chemical libraries can access far greater chemical space, provided that the predictive accuracy is sufficient to identify useful molecules. Through the largest and most diverse virtual HTS campaign reported to date, comprising 318 individual projects, we demonstrate that our AtomNet® convolutional neural network successfully finds novel hits across every major therapeutic area and protein class. We address historical limitations of computational screening by demonstrating success for target proteins without known binders, high-quality X-ray crystal structures, or manual cherry-picking of compounds. We show that the molecules selected by the AtomNet® model are novel drug-like scaffolds rather than minor modifications to known bioactive compounds. Our empirical results suggest that computational methods can substantially replace HTS as the first step of small-molecule drug discovery

Association of kidney disease measures with risk of renal function worsening in patients with type 1 diabetes

Background: Albuminuria has been classically considered a marker of kidney damage progression in diabetic patients and it is routinely assessed to monitor kidney function. However, the role of a mild GFR reduction on the development of stage 653 CKD has been less explored in type 1 diabetes mellitus (T1DM) patients. Aim of the present study was to evaluate the prognostic role of kidney disease measures, namely albuminuria and reduced GFR, on the development of stage 653 CKD in a large cohort of patients affected by T1DM. Methods: A total of 4284 patients affected by T1DM followed-up at 76 diabetes centers participating to the Italian Association of Clinical Diabetologists (Associazione Medici Diabetologi, AMD) initiative constitutes the study population. Urinary albumin excretion (ACR) and estimated GFR (eGFR) were retrieved and analyzed. The incidence of stage 653 CKD (eGFR < 60 mL/min/1.73 m2) or eGFR reduction > 30% from baseline was evaluated. Results: The mean estimated GFR was 98 \ub1 17 mL/min/1.73m2 and the proportion of patients with albuminuria was 15.3% (n = 654) at baseline. About 8% (n = 337) of patients developed one of the two renal endpoints during the 4-year follow-up period. Age, albuminuria (micro or macro) and baseline eGFR < 90 ml/min/m2 were independent risk factors for stage 653 CKD and renal function worsening. When compared to patients with eGFR > 90 ml/min/1.73m2 and normoalbuminuria, those with albuminuria at baseline had a 1.69 greater risk of reaching stage 3 CKD, while patients with mild eGFR reduction (i.e. eGFR between 90 and 60 mL/min/1.73 m2) show a 3.81 greater risk that rose to 8.24 for those patients with albuminuria and mild eGFR reduction at baseline. Conclusions: Albuminuria and eGFR reduction represent independent risk factors for incident stage 653 CKD in T1DM patients. The simultaneous occurrence of reduced eGFR and albuminuria have a synergistic effect on renal function worsening

Substrate targeting and inhibition of editing deaminases

Identification of small molecules against APOBEC3B The APOBECs are deaminases that act on DNA and RNA to restrict exogenous nucleic acids. Yet, the signature of their mutagenic activity –especially that of APOBEC3A and APOBEC3B- has been observed in the cancer genomes and their ability to increase the genetic heterogeneity of tumours has been linked to the onset of drug resistance in cancer. As such inhibition of their enzymatic activity represents a potential target for anticancer therapies. During my PhD I worked at the identification of APOBEC3B small-molecule inhibitors. To this aim, I used a computational approach to perform a virtual screening on large library of molecules to block APOBEC3B enzymatic activity. I then tested selected molecules from the virtual screening using biochemical assays to quantify their effect on APOBEC3B activity and their capacity to interfere with APOBEC3B binding to DNA. Through this, I was able to identify two small molecules that reduce the activity of this protein, which could provide basis for the development of the first drug for anti-APOBEC activity. Engineering ADAR2 to act on DNA Genome editing technologies have revolutionized our ability to target and modify the genomes of living cells and organisms. The fusion of AID/APOBECs to genome editing tools such as Cas9 allowed the development the first base editor, molecules that can be targeted to mutate specific cytosines. The pool of available Base Editors is in constant expansion as new molecules are developed to target DNA with more specificity and efficiency. As the only adenine-targeting Base Editor is based on TadA- an RNA deaminase-, I focused on the development of a A•T base editor based on the catalytic domain of ADAR2. Adenosine Deaminases Acting on RNA (ADARs), are editing enzymes that catalyse the C6 deamination of adenosine (A) to produce inosine (I) in double-stranded RNA. As human ADAR2 is able to target DNA/RNA hybrids, I first tried -without success- to use chimeras of n/dCas9 and the deaminase domain of ADAR2 to induce mutations in a fluorescent reporter. I then used a bacterial screen to select for mutants of ADAR2 that act on DNA. I selected a mutant that induces a mutator phenotype in bacteria and DNA damage in mammalian cells. I am currently working to engineer this mutant into a Base Editor suitable for biotechnological applications such as gene therapy, antiviral treatment and cancer therapy

Vascular Smooth Muscle Emilin-1 Is a Regulator of Arteriolar Myogenic Response and Blood Pressure

OBJECTIVE: Emilin-1 is a protein of elastic extracellular matrix involved in blood pressure (BP) control by negatively affecting transforming growth factor (TGF)-\u3b2 processing. Emilin1 null mice are hypertensive. This study investigates how Emilin-1 deals with vascular mechanisms regulating BP. METHODS AND RESULTS: This study uses a phenotype rescue approach in which Emilin-1 is expressed in either endothelial cells or vascular smooth muscle cells of transgenic animals with the Emilin1(-/-) background. We found that normalization of BP required Emilin-1 expression in smooth muscle cells, whereas expression of the protein in endothelial cells did not modify the hypertensive phenotype of Emilin1(-/-) mice. We also explored the effect of treatment with anti-TGF-\u3b2 antibodies on the hypertensive phenotype of Emilin1(-/-) mice, finding that neutralization of TGF-\u3b2 in Emilin1 null mice normalized BP quite rapidly (2 weeks). Finally, we evaluated the vasoconstriction response of resistance arteries to perfusion pressure and neurohumoral agents in different transgenic mouse lines. Interestingly, we found that the hypertensive phenotype was coupled with an increased arteriolar myogenic response to perfusion pressure, while the vasoconstriction induced by neurohumoral agents remained unaffected. We further elucidate that, as for the hypertensive phenotype, the increased myogenic response was attributable to increased TGF-\u3b2 activity. CONCLUSIONS: Our findings clarify that Emilin-1 produced by vascular smooth muscle cells acts as a main regulator of resting BP levels by controlling the myogenic response in resistance arteries through TGF-\u3b2