Non-interventional Translational Study in Mild, Moderate and Severe Acne Patients

Acne lesion counts

The number of total lesions, inflammatory lesions (papules and pustules), and non-inflammatory lesions (open and closed comedones) on the face will be counted. All lesions on the face should be counted, including those on the nose. Counts of nodulocystic lesion (nodules and cysts) will be reported separately and are not to be included in the inflammatory or non-inflammatory lesion counts.

Investigator's Global Assessment (IGA) Scale

The overall severity of facial acne will be assessed using the 5-point IGA scale (Table 6) (15). Each grade is defined by a distinct and clinically relevant morphologic description that minimizes interobserver variability. The IGA severity score assessments should be performed exclusively by site staff qualified as IGA rater. Investigators or designee should use the IGA scale provided in the study protocol throughout the study.

Patient Reported Outcome Measures

With the objective of evaluating the impact of different acne severities in the quality of life, participants will be asked to complete 2 quality of life (QoL) questionnaires: the Skindex-29 and the Acne-specific Quality of Life (Acne-QoL). The Skindex-29 questionnaire is a validated health-related quality of life self-administered questionnaire to assess the impact of skin diseases on patient's quality of life (17). Skindex-29 consists of 30 items, distributed into 3 domains (symptoms, emotion and function. Each item is scored on a 5-point Likert scale, from 1 to 5. Scores are standardized to 100, with lower score indicating higher levels of QoL. The Acne-QoL is a validated health-related quality of life self-administered questionnaire to assess quality of life among participants with facial acne (16). Acne-QoL consists of 19 items organized into 4 domains: self-perception, role-social, role-emotional and acne symptoms. Each item is scored

Facial Photographs

The investigator or designee will take digital photographs of the participants' face using Canfield Facial Image Photography Device. Photographs will be taken under standardized conditions. Images will be captured, viewed, and uploaded using Canfield Facial Image Photography Device software which automatically checksums, encrypts, packages, and transfers the data to secure, validated and compliant web servers hosted by Canfield Clinic. Deidentified images will be used to assess the skin condition of each participant and be the input data for the AI-based tool to derive acne severity.

Immunogenicity Assessments

1.Surface Binding (SB) assays using a fluorescence cytometer will be used to determine the binding efficiency of serum antibodies to the cell surface of live C. acnes bacteria. 2. Opsonophagocytic killing (OPK) assays will be used to determine the ability of antibodies to induce opsonization of C. acnes bacteria. Bound antibodies mobilize and activate immune cells against the bacterium, leading to a reduction in the bacterial cell numbers. 3. Sheep Red Blood Cells Co-Hemolysis Neutralization Assay will be used to measure serological titers of neutralizing recombinant C. acnes CAMP2 polypeptide (CAMP2)-specific antibodies. 4. Antigen-specific Antibody Titers to further explore the humoral immune response elicited against C. acnes (e.g. C. acnes types IA1 \[NCTC737\], IB \[KPA171202\]), IgG, IgA titres and IgG subclasses distribution will be measured.

Immunogenicity Assessments

5\. Antibody-dependent complement deposition titer documents the ability of C. acnes-specific Ig to activate the complement system via the classical pathway. For tested serum samples, the method consists of measuring the antibody-driven recruitment and deposition of the complement components C3b/iC3b on live C. acnes bacterial using flow cytometry. 6. Cell-Mediated Immunity against C. acnes antigens cytokine assessment in supernatant from whole blood culture, the cytokine responses in whole blood will be measured by either multiplex bead array assay or MSD ECL-based assay.7.Immunophenotyping of Peripheral Blood Mononuclear Cells (PBMCs) T cell profiling by flow cytometry subsequent to stimulation. PBMCs isolated from participants will be thawed and cultured with relevant reagents and then evaluated using cytometry, using BD analyzers, spectral flow and/or CyTOF as required as required to determine activated T cell signatures.

Immunogenicity Assessments

8\. Peripheral blood cytokine concentration will be measured in the serum of participants by either multiplex bead array assay or MSD ECL-based assay or appropriate proteomics profiling platforms such as NULISA or OLINK. 9\. Genomewide genetic analysis, DNA will be extracted from PBMCs isolated from the blood collected from each participant. This DNA will then be used for genetic analysis using genomewide genotyping arrays such as Infinium (Illumina).

METAGENOMIC AND METATRANSCRIPTOMIC ASSESSMENTS

To characterize the microbial strains, microbiome composition and transcriptional activity, skin swabs will be collected from the participants' forehead and cheeks. For sequencing assays, material from swabs will be dislodged and submerged in DNA/RNA shield and stored at -80°C prior to nucleic acid extraction. Microbial isolation will be done according to procedures described in 4.6. Analyses such as microbiome metagenomics profiling (taxonomic profiles, relative abundance, diversity, richness) and amplicon sequencing of isolates (C. acnes Single Locus Sequence Type) could be performed alongside similar analysis from follicular extracts. Alongside this metatranscriptomic analysis of collected microbial material will be undertaken. Collected samples will be processed and stored at sites according to the best practice for each material type prior to downstream analytical assays.