Diabetic individuals with COVID-19 exhibit reduced efficacy of gliptins in inhibiting dipeptidyl peptidase 4 (DPP4). A suggested explanation for increased COVID-19 susceptibility in patients with type 2 diabetes mellitus (T2DM).

AIMS: Dipeptidyl peptidase 4 (DPP4) has been proposed as a coreceptor for SARS-CoV-2 cellular entry. Considering that type 2 diabetes mellitus (T2DM) has been identified as the most important risk factor for SARS-CoV-2, and that gliptins (DPP4 inhibitors) are a prescribed diabetic treatment, this study aims to unravel the impact of DPP4 in the intersection of T2DM/COVID-19. MATERIALS AND METHODS: We analyzed 189 serum human samples, divided into six clinical groups (controls, T2DM, T2DM + gliptins, COVID-19, COVID-19 + T2DM, and COVID-19 + T2DM + gliptins), measuring DPP4 protein concentration and activity by Western blot, ELISA, and commercial activity kits. The obtained results were verified in Huh-7 cellular models. KEY FINDINGS: Both DPP4 concentration and activity were decreased in COVID-19 patients, and as in T2DM patients, compared to controls. Despite these lower levels, the ratio of DPP4 activity/concentration in COVID-19 sera was the highest (0.782 ± 0.289 µU/ng vs. 0.547 ± 0.050 µU/ng in controls, p < 0.0001), suggesting a compensating mechanism in these patients. Supernatants of Huh-7 cells incubated with COVID-19 serum showed a consistent and significantly lower DPP4 concentration and activity. Furthermore, COVID-19 + T2DM + gliptins patients showed a higher serum DPP4 concentration and activity than T2DM + gliptin subjects (p < 0.05), indicating that sera from COVID-19 convalescents interfere with gliptins. SIGNIFICANCE: Either SARS-CoV-2 or some metabolites present in the sera of COVID-19-convalescent patients interact with soluble DPP4 or even gliptins themselves since the inhibitory effect of gliptins on DPP4 activity is being prevented. The interactions between DPP4, gliptins, and SARS-CoV-2 should be further elucidated to reveal the mechanism of action for these interesting observations.

Resistance to 2-Hydroxy-Flutamide in Prostate Cancer Cells Is Associated with the Downregulation of Phosphatidylcholine Biosynthesis and Epigenetic Modifications.

In this study, we examined the metabolic adaptations of a chemoresistant prostate cancer cell line in comparison to a sensitive cell line. We utilized prostate cancer LNCaP cells and subjected them to a stepwise increase in the antiandrogen 2-hydroxy-flutamide (FLU) concentration to generate a FLU-resistant cell line (LN-FLU). These LN-FLU cells displayed characteristics of cancer stem cells, exhibited drug resistance, and showed a significantly reduced expression of Cyclin D1, along with the overexpression of p16, pointing to a proliferation arrest. In comparing the cancer stem-like LN-FLU cells to the LNCaP cells, we observed a decrease in the expression of CTP-choline cytidylyl transferase α (CCTα), as well as a decline in choline kinase, suggesting altogether a downregulation of the phosphatidylcholine biosynthetic pathway. In addition, we found decreased levels of the protein methyl transferase PRMT2 and the upregulation of the histone deacetylase Sirtuin1 (Sirt1). Analysis of the human prostate cancer samples revealed similar results in a population with high expressions of the stem cell markers Oct4 and ABCB1A1. Our findings suggest that the adaptation of prostate cancer cells to antiandrogens could induce reprogramming into stem cells that survive in a low phosphocholine metabolism and cell cycle arrest and display drug resistance.

Transcriptional inaccuracy threshold attenuates differences in RNA-dependent DNA synthesis fidelity between retroviral reverse transcriptases.

In M13mp2 lacZα forward mutation assays measuring intrinsic fidelity of DNA-dependent DNA synthesis, wild-type human immunodeficiency virus type 1 (HIV-1) RTs of group M/subtype B previously showed >10-fold higher error rates than murine leukaemia virus (MLV) and avian myeloblastosis virus (AMV) RTs. An adapted version of the assay was used to obtain error rates of RNA-dependent DNA synthesis for several RTs, including wild-type HIV-1BH10, HIV-1ESP49, AMV and MLV RTs, and the high-fidelity mutants of HIV-1ESP49 RT K65R and K65R/V75I. Our results showed that there were less than two-fold differences in fidelity between the studied RTs with error rates ranging within 2.5 × 10-5 and 3.5 × 10-5. These results were consistent with the existence of a transcriptional inaccuracy threshold, generated by the RNA polymerase while synthesizing the RNA template used in the assay. A modest but consistent reduction of the inaccuracy threshold was achieved by lowering the pH and Mg2+ concentration of the transcription reaction. Despite assay limitations, we conclude that HIV-1BH10 and HIV-1ESP49 RTs are less accurate when copying DNA templates than RNA templates. Analysis of the RNA-dependent mutational spectra revealed a higher tendency to introduce large deletions at the initiation of reverse transcription by all HIV-1 RTs except the double-mutant K65R/V75I.

Viral reverse transcriptases.

Reverse transcriptases (RTs) play a major role in the replication of Retroviridae, Metaviridae, Pseudoviridae, Hepadnaviridae and Caulimoviridae. RTs are enzymes that are able to synthesize DNA using RNA or DNA as templates (DNA polymerase activity), and degrade RNA when forming RNA/DNA hybrids (ribonuclease H activity). In retroviruses and LTR retrotransposons (Metaviridae and Pseudoviridae), the coordinated action of both enzymatic activities converts single-stranded RNA into a double-stranded DNA that is flanked by identical sequences known as long terminal repeats (LTRs). RTs of retroviruses and LTR retrotransposons are active as monomers (e.g. murine leukemia virus RT), homodimers (e.g. Ty3 RT) or heterodimers (e.g. human immunodeficiency virus type 1 (HIV-1) RT). RTs lack proofreading activity and display high intrinsic error rates. Besides, high recombination rates observed in retroviruses are promoted by poor processivity that causes template switching, a hallmark of reverse transcription. HIV-1 RT inhibitors acting on its polymerase activity constitute the backbone of current antiretroviral therapies, although novel drugs, including ribonuclease H inhibitors, are still necessary to fight HIV infections. In Hepadnaviridae and Caulimoviridae, reverse transcription leads to the formation of nicked circular DNAs that will be converted into episomal DNA in the host cell nucleus. Structural and biochemical information on their polymerases is limited, although several drugs inhibiting HIV-1 RT are known to be effective against the human hepatitis B virus polymerase. In this review, we summarize current knowledge on reverse transcription in the five virus families and discuss available biochemical and structural information on RTs, including their biosynthesis, enzymatic activities, and potential inhibition.