The Copenhagen Actinic Keratosis Study (COAKS). A decentralised clinical trial to evaluate tolerability, safety and efficacy of daily field-directed topical treatment with cytosolic phospholipase A2 inhibitor, AVX001, in participants with actinic keratosis: protocol for a randomised controlled phase I/IIa trial.

INTRODUCTION: Actinic keratosis (AK) is the most common precancerous skin condition caused by long-term UV exposure. Given the high recurrence rate of 15%-53%, identifying safe and effective treatment options is warranted. AVX001, a cytosolic phospholipase A2α (cPLA2α) enzyme inhibitor, is a novel anti-inflammatory drug for field-directed, self-administered, topical therapy of AK. METHODS AND ANALYSIS: This study is a single-centre, randomised, vehicle-controlled, double-blind, parallel-group hybrid clinical trial in adults with multiple AK lesions Olsen grade 1 or 2. The hybrid design combines decentralised participant tasks and assessments with conventional in-clinic visits. Recruitment using targeted advertising on social media and eligibility prescreening are conducted via the Studies&Me online recruitment platform. Participants (n=60) are randomly assigned to 1 of 3 treatment arms: AVX001 gel 1%, AVX001 gel 3% or vehicle gel. The trial consists of a 4-week treatment period with daily field-directed topical application of the gel and an 8-week follow-up period. Participants attend in-clinic visits at baseline, week 4 and week 12. The remote participant trial tasks include questionnaires and upload of smartphone-obtained photos of the treated skin area using a study-specific web-based app. Both remote and in-clinic assessments of safety and efficacy will be performed. The primary objective is to evaluate the local tolerability of daily application of AVX001 gel (1% or 3%) compared with vehicle gel. Secondary objectives include safety, efficacy, dose-response efficacy relationship, treatment satisfaction and cosmetic outcome. Exploratory objectives include evaluations of tolerability and efficacy assessed by dermatologists using smartphone photos uploaded by participants, comparisons of in-clinic and remote assessments and assessment of AK-related skin changes by non-invasive optical imaging. ETHICS AND DISSEMINATION: Approved by the Ethics Committee of the Capital Region of Denmark (H-21018064) and the Danish Medicines Agency (2021032485). Results will be submitted for publication in peer-reviewed scientific journals. TRIAL REGISTRATION NUMBERS: 2021-000934-32; NCT05164393.

Inhibition of Cytosolic Phospholipase A2α Induces Apoptosis in Multiple Myeloma Cells.

Cytosolic phospholipase A2α (cPLA2α) is the rate-limiting enzyme in releasing arachidonic acid and biosynthesis of its derivative eicosanoids. Thus, the catalytic activity of cPLA2α plays an important role in cellular metabolism in healthy as well as cancer cells. There is mounting evidence suggesting that cPLA2α is an interesting target for cancer treatment; however, it is unclear which cancers are most relevant for further investigation. Here we report the relative expression of cPLA2α in a variety of cancers and cancer cell lines using publicly available datasets. The profiling of a panel of cancer cell lines representing different tissue origins suggests that hematological malignancies are particularly sensitive to the growth inhibitory effect of cPLA2α inhibition. Several hematological cancers and cancer cell lines overexpressed cPLA2α, including multiple myeloma. Multiple myeloma is an incurable hematological cancer of plasma cells in the bone marrow with an emerging requirement of therapeutic approaches. We show here that two cPLA2α inhibitors AVX420 and AVX002, significantly and dose-dependently reduced the viability of multiple myeloma cells and induced apoptosis in vitro. Our findings implicate cPLA2α activity in the survival of multiple myeloma cells and support further studies into cPLA2α as a potential target for treating hematological cancers, including multiple myeloma.

cPLA2α Enzyme Inhibition Attenuates Inflammation and Keratinocyte Proliferation.

As a regulator of cellular inflammation and proliferation, cytosolic phospholipase A2 α (cPLA2α) is a promising therapeutic target for psoriasis; indeed, the cPLA2α inhibitor AVX001 has shown efficacy against plaque psoriasis in a phase I/IIa clinical trial. To improve our understanding of the anti-psoriatic properties of AVX001, we sought to determine how the compound modulates inflammation and keratinocyte hyperproliferation, key characteristics of the psoriatic epidermis. We measured eicosanoid release from human peripheral blood mononuclear cells (PBMC) and immortalized keratinocytes (HaCaT) and studied proliferation in HaCaT grown as monolayers and stratified cultures. We demonstrated that inhibition of cPLA2α using AVX001 produced a balanced reduction of prostaglandins and leukotrienes; significantly limited prostaglandin E2 (PGE2) release from both PBMC and HaCaT in response to pro-inflammatory stimuli; attenuated growth factor-induced arachidonic acid and PGE2 release from HaCaT; and inhibited keratinocyte proliferation in the absence and presence of exogenous growth factors, as well as in stratified cultures. These data suggest that the anti-psoriatic properties of AVX001 could result from a combination of anti-inflammatory and anti-proliferative effects, probably due to reduced local eicosanoid availability.

Defining estrogenic mechanisms of bisphenol A analogs through high throughput microscopy-based contextual assays.

Environmental exposures to chemically heterogeneous endocrine-disrupting chemicals (EDCs) mimic or interfere with hormone actions and negatively affect human health. Despite public interest and the prevalence of EDCs in the environment, methods to mechanistically classify these diverse chemicals in a high throughput (HT) manner have not been actively explored. Here, we describe the use of multiparametric, HT microscopy-based platforms to examine how a prototypical EDC, bisphenol A (BPA), and 18 poorly studied BPA analogs (BPXs), affect estrogen receptor (ER). We show that short exposure to BPA and most BPXs induces ERα and/or ERß loading to DNA changing target gene transcription. Many BPXs exhibit higher affinity for ERß and act as ERß antagonists, while they act largely as agonists or mixed agonists and antagonists on ERα. Finally, despite binding to ERs, some BPXs exhibit lower levels of activity. Our comprehensive view of BPXs activities allows their classification and the evaluation of potential harmful effects. The strategy described here used on a large-scale basis likely offers a faster, more cost-effective way to identify safer BPA alternatives.

Control of expression of the lectin-like protein Reg-1 by gastrin: role of the Rho family GTPase RhoA and a C-rich promoter element.

The expression of members of the Reg family of secreted lectin-like proteins is increased in response to stress, inflammation and damage in many tissues. In the stomach, Reg is located in enterochromaffin-like cells, where its expression is stimulated by the gastric hormone gastrin. We have examined the mechanisms by which gastrin stimulates expression of Reg-1. Deletional mutations of 2.1 to 0.1 kb of the rat Reg-1 promoter in a luciferase reporter vector were transiently transfected into gastric cancer AGS-G(R) cells. All promoter fragments tested showed similar relative increases in luciferase expression in response to gastrin (1 nM). The response to gastrin of the smallest (104 bp) construct was 4.2+/-0.4-fold over basal. These responses were reduced by Ro-32-0432, a protein kinase C inhibitor, by C3-transferase, a Clostridium botulinum toxin and a selective inhibitor of the Rho family GTPase RhoA, and by co-transfection with a dominant negative form of RhoA. Co-transfection with a constitutively active form of RhoA stimulated expression 11.6+/-1.7-fold over basal. Mutations through the 104 bp construct identified a C-rich element (C-79CCCTCCC-72) required for responses to gastrin, PKC (protein kinase C) and L63RhoA (the constitutively active form of human RhoA protein containing a glutamine-to-leucine substitution at position 63). EMSAs (electrophoretic-mobility-shift assays) using nuclear extracts of control and gastrin-stimulated AGS-G(R) cells and a probe spanning -86 to -64 bp revealed multiple binding proteins. There was no effect of gastrin on the pattern of binding. Supershift assays indicated that transcription factors Sp1 and Sp3 bound the C-rich sequence. We conclude that gastrin stimulates Reg expression via activation of PKC and RhoA, that a C-rich region (-79 to -72) is critical for the response and that Sp-family transcription factors bind to this region of the promoter.