Characterisation of a common hotspot variant in acute intermittent porphyria sheds light on the mechanism of hydroxymethylbilane synthase function

Hydroxymethylbilane synthase (HMBS) is the third enzyme involved in haem biosynthesis, in which it catalyses the formation of tetrapyrrole 1-hydroxymethylbilane (HMB). In this process, HMBS binds four consecutive substrate molecules, creating the enzyme-intermediate complexes ES, ES2, ES3 and ES4. Pathogenic variants in the HMBS gene are associated with the dominantly inherited disorder acute intermittent porphyria. In this study, we have characterised the p.R26H variant to shed light on the role of Arg26 in the elongation mechanism of HMBS and to provide insights into its effect on the enzyme. With selected biophysical methods, we have been able to show that p.R26H forms a single enzyme-intermediate complex in the ES2-state. We were also able to demonstrate that the p.R26H variant results in an inactive enzyme, which is unable to produce the HMB product.publishedVersio

Acute Intermittent Porphyria: An Overview of Therapy Developments and Future Perspectives Focusing on Stabilisation of HMBS and Proteostasis Regulators

Acute intermittent porphyria (AIP) is an autosomal dominant inherited disease with low clinical penetrance, caused by mutations in the hydroxymethylbilane synthase (HMBS) gene, which encodes the third enzyme in the haem biosynthesis pathway. In susceptible HMBS mutation carriers, triggering factors such as hormonal changes and commonly used drugs induce an overproduction and accumulation of toxic haem precursors in the liver. Clinically, this presents as acute attacks characterised by severe abdominal pain and a wide array of neurological and psychiatric symptoms, and, in the long-term setting, the development of primary liver cancer, hypertension and kidney failure. Treatment options are few, and therapies preventing the development of symptomatic disease and long-term complications are non-existent. Here, we provide an overview of the disorder and treatments already in use in clinical practice, in addition to other therapies under development or in the pipeline. We also introduce the pathomechanistic effects of HMBS mutations, and present and discuss emerging therapeutic options based on HMBS stabilisation and the regulation of proteostasis. These are novel mechanistic therapeutic approaches with the potential of prophylactic correction of the disease by totally or partially recovering the enzyme functionality. The present scenario appears promising for upcoming patient-tailored interventions in AIP.publishedVersio

Suberin of Potato (Solanum tuberosum Var. Nikola): Comparison of the Effect of Cutinase CcCut1 with Chemical Depolymerization

Chemical and enzymatic depolymerizations of suberin isolated from potato peel (Solanum tuberosum var. Nikola) were performed under various conditions. Enzymatic hydrolysis with cutinase CcCut1 and chemical methanolysis with NaOMe of suberin yielded monomeric fragments, which were identified as TMS derivatives with GC-MS and GC-FID. The solid, hydrolysis-resistant residues were analyzed with solid state (13)C CPMAS NMR, FT-IR, and microscopic methods. Methanolysis released more CHCl(13)-soluble, material than the cutinase treatment when determined gravimetrically. Interestingly, cutinase-catalyzed hydrolysis produced higher proportions of aliphatic monomers than hydrolysis with the NaOMe procedure when analyzed by GC in the form of TMS derivatives. Monomers released by the two methods were mainly alpha,omega-dioic acids and omega-hydroxy acids, but the ratios of the detected monomers were different, at 40.0 and 32.7% for methanolysis and 64.6 and 8.2% for cutinase, respectively. Thus, cutinase CcCut1 showed higher activity toward ester bonds of alpha,omega-dioic acids than toward the bonds of omega-hydroxy acids. The most abundant monomeric compounds were octadec-9-ene-1,18-dioic acid and 18-hydroxyoctadec-9-enoic acid, which accounted for ca. 37 and 28% of all monomers, respectively. The results of the analyses of the chemical and enzymatic hydrolysis products were supported by the spectroscopic analyses with FT-IR and CPMAS (13)C NMR together with the analysis of the microstructures of the hydrolysis residues by light and confocal microscopy

No Association Between Ljungan Virus Seropositivity and the Beta-cell Damaging Process in the Finnish Type 1 Diabetes Prediction and Prevention Study Cohort

Background: Ljungan virus (LV) has not confirmed to associate with any human disease, but a possible connection with type 1 diabetes has been suggested. LV is a rodent-borne picornavirus that induces a diabetes-like condition in rodents. Approximately 30% of adults and 60% of children are seropositive in Finland. The Finnish Type 1 Diabetes Prediction and Prevention study enabled the use of very well characterized sample panels from children seroconverted to positivity for multiple islet autoantibodies during their prospective observation from birth; in addition, samples from age, sex, human leukocyte antigen (HLA), and residence area matched control children. Methods: We analyzed LV IgG seroprevalence in 102 case children (65 had also developed type 1 diabetes), in addition to nondiabetic control children. LV and human parechovirus (HPeV) immunofluorescence assays were used to analyze LV and HPeV-specific IgG from 102 plasma samples taken at the time of islet autoantibody appearance and from 204 samples from the matched control children. Results: Altogether 46.1% of the case and 50.7% of the control children were positive for LV IgG (odds ratio 0.8; 95% confidence interval, 0.47-1.36; P = 0.416) and 67.6% versus 79.8% were positive for HPeV IgG, respectively (odds ratio 0.49, 0.27-0.9, P = 0.023). Conclusions: Thus, no risk associations between LV or HPeV-specific IgG and islet autoimmunity were observed. However, a trend for significantly higher prevalence of HPeV antibodies in control children (P = 0.023) suggests a possible protective association of this virus with islet autoimmunity.Peer reviewe

VTT-006, an anti-mitotic compound, binds to the Ndc80 complex and suppresses cancer cell growth <i>in vitro</i>.

Hec1 (Highly expressed in cancer 1) resides in the outer kinetochore where it works to facilitate proper kinetochore-microtubule interactions during mitosis. Hec1 is overexpressed in various cancers and its expression shows correlation with high tumour grade and poor patient prognosis. Chemical perturbation of Hec1 is anticipated to impair kinetochore-microtubule binding, activate the spindle assembly checkpoint (spindle checkpoint) and thereby suppress cell proliferation. In this study, we performed high-throughput screen to identify novel small molecules that target the Hec1 calponin homology domain (CHD), which is needed for normal microtubule attachments. 4 million compounds were first virtually fitted against the CHD, and the best hit molecules were evaluated in vitro. These approaches led to the identification of VTT-006, a 1,2-disubstituted-tetrahydro-beta-carboline derivative, which showed binding to recombinant Ndc80 complex and modulated Hec1 association with microtubules in vitro. VTT-006 treatment resulted in chromosome congression defects, reduced chromosome oscillations and induced loss of inter-kinetochore tension. Cells remained arrested in mitosis with an active spindle checkpoint for several hours before undergoing cell death. VTT-006 suppressed the growth of several cancer cell lines and enhanced the sensitivity of HeLa cells to Taxol. Our findings propose that VTT-006 is a potential anti-mitotic compound that disrupts M phase, impairs kinetochore-microtubule interactions, and activates the spindle checkpoint

High intratumoral dihydrotestosterone is associated with antiandrogen resistance in VCaP prostate cancer xenografts in castrated mice

Antiandrogen treatment resistance is a major clinical concern in castration-resistant prostate cancer (CRPC) treatment. Using xenografts of VCaP cells we showed that growth of antiandrogen resistant CRPC tumors were characterized by a higher intratumor dihydrotestosterone (DHT) concentration than that of treatment responsive tumors. Furthermore, the slow tumor growth after adrenalectomy was associated with a low intratumor DHT concentration. Reactivation of androgen signaling in enzalutamide-resistant tumors was further shown by the expression of several androgen-dependent genes. The data indicate that intratumor DHT concentration and expression of several androgen-dependent genes in CRPC lesions is an indication of enzalutamide treatment resistance and an indication of the need for further androgen blockade. The presence of an androgen synthesis, independent of CYP17A1 activity, has been shown to exist in prostate cancer cells, and thus, novel androgen synthesis inhibitors are needed for the treatment of enzalutamide-resistant CRPC tumors that do not respond to abiraterone.Peer reviewe

Discovery and development of ODM-204: A Novel nonsteroidal compound for the treatment of castration-resistant prostate cancer by blocking the androgen receptor and inhibiting CYP17A1

We report the discovery of a novel nonsteroidal dual-action compound, ODM-204, that holds promise for treating patients with castration-resistant prostate cancer (CRPC), an advanced form of prostate cancer characterised by high androgen receptor (AR) expression and persistent activation of the AR signaling axis by residual tissue androgens. For ODM-204, has a dual mechanism of action. The compound is anticipated to efficiently dampen androgenic stimuli in the body by inhibiting CYP17A1, the prerequisite enzyme for the formation of dihydrotestosterone (DHT) and testosterone (T), and by blocking AR with high affinity and specificity. In our study, ODM-204 inhibited the proliferation of androgen-dependent VCaP and LNCaP cells in vitro and reduced significantly tumour growth in a murine VCaP xenograft model in vivo. Intriguingly, after a single oral dose of 10-30 mg/kg, ODM-204 dose-dependently inhibited adrenal and testicular steroid production in sexually mature male cynomolgus monkeys. Similar results were obtained in human chorionic gonadotropin-treated male rats. In rats, leuprolide acetate-mediated (LHRH agonist) suppression of the circulating testosterone levels and decrease in weights of androgen-sensitive organs was significantly and dose-dependently potentiated by the co-administration of ODM-204. ODM-204 was well tolerated in both rodents and primates. Based on our data, ODM-204 could provide an effective therapeutic option for men with CRPC.</p

The phylodynamics of SARS-CoV-2 during 2020 in Finland

Finland has had a low incidence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) infections as compared to most European countries. Here we report the origins and turnover of SARS-CoV-2 lineages circulating in Finland in 2020. SARS-CoV-2 introduced to Finland in January 2020 and spread rapidly across southern Finland during spring. We observed rapid turnover among Finnish lineages during this period. Clade 20C became the most prevalent among sequenced cases and was replaced by other strains in fall 2020. Bayesian phylogeographic reconstructions suggested 42 independent introductions into Finland during spring 2020, mainly from Italy, Austria, and Spain, which might have been the source for a third of cases. The investigations of the original introductions of SARS-CoV-2 to Finland during the early stages of the pandemic and of the subsequent lineage dynamics could be utilized to assess the role of transboundary movements and effects of early intervention and public health measures.Peer reviewe

Adrenals Contribute to Growth of Castration-Resistant VCaP Prostate Cancer Xenografts

The role of adrenal androgens as drivers for castration-resistant prostate cancer (CRPC) growth in humans is generally accepted; however, the value of preclinical mouse models of CRPC is debatable, because mouse adrenals do not produce steroids activating the androgen receptor. In this study, we confirmed the expression of enzymes essential for de novo synthesis of androgens in mouse adrenals, with high intratissue concentration of progesterone (P-4) and moderate levels of androgens, such as androstenedione, testosterone, and dihydrotestosterone, in the adrenal glands of both intact and orchectomized (ORX) mice. ORX alone had no effect on serum P-4 concentration, whereas orchectomized and adrenalectomized (ORX + ADX) resulted in a significant decrease in serum P-4 and in a further reduction in the Low levels of serum androgens (androstenedione, testosterone, and dihydrotestosterone), measured by mass spectrometry. In line with this, the serum prostate-specific antigen and growth of VCaP xenografts in mice after ORX + ADX were markedly reduced compared with ORX alone, and the growth difference was not abolished by a glucocorticoid treatment. Moreover, ORX + ADX altered the androgen-dependent gene expression in the tumors, similar to that recently shown for the enzalutamide treatment. These data indicate that in contrast to the current view, and similar to humans, mouse adrenals synthesize significant amounts of steroids that contribute to the androgen receptor dependent growth of CRPC.Peer reviewe