Video-immunotherapy: a New Approach in Breast Cancer Treatment

Primary outcomes

1. Change from baseline in Depression Anxiety Stress Scale (DASS-42) total score

   Time frame: Experimental and control group patients will answer questionnaires to monitor their psychological status and physiological symptoms 3 times in total: baseline (month 0), during the treatment (month 3) and at the end of treatment (month 5-6).

   The Depression Anxiety Stress Scale (DASS-42) is a self-reported questionnaire designed to measure negative emotional states across three subscales: depression, anxiety, and stress. The total score is calculated by summing all 42 items.

   Minimum value: 0 Maximum value: 126 (42 items × 0-3 Likert scale)

   Score interpretation: Higher scores indicate worse psychological distress.

2. Change from baseline in Edmonton Symptom Assessment System-revised (ESAS-r) total symptom score

   Time frame: Baseline (Month 0), during treatment (Month 3), and end of treatment (Month 5-6)

   The Edmonton Symptom Assessment System-revised (ESAS-r) is a patient-reported outcome measure assessing the severity of common cancer-related symptoms, including pain, fatigue, nausea, depression, anxiety, drowsiness, appetite, well-being, shortness of breath, and an optional additional symptom. Each symptom is rated on a numeric scale.

   Minimum value: 0 Maximum value: 100 (10 symptoms × 0-10 numeric rating scale)

   Score interpretation: Higher scores indicate worse symptom burden.

3. Change from baseline in frequency of immune cell subsets in peripheral blood and tumor tissue

   Time frame: Peripheral blood: Baseline (Month 0) and end of treatment (Month 5-6) Tumor tissue: End of treatment (Month 5-6)

   The frequency of immune cell subsets involved in breast cancer immunity will be assessed by flow cytometry. The analyzed cell populations include CD4⁺ T cells, CD8⁺ T cells, regulatory T cells (Treg), NK cells, monocytes in peripheral blood and tumor tissue, and myeloid-derived suppressor cells, as well as M1 and M2 macrophages in tumor tissue only. Cell frequencies will be reported as a percentage of CD45⁺ live cells.

   Unit of Measure: Percentage (%)

4. Change from baseline in activation marker expression on immune cell subsets

   Time frame: Peripheral blood: Baseline (Month 0) and end of treatment (Month 5-6) Tumor tissue: End of treatment (Month 5-6)

   Activation status of immune cells will be evaluated by flow cytometry based on the expression of activation markers. NK cells and T cell subsets will be assessed using CD25, CD69, and HLADR expression, while monocytes and macrophages will be assessed using CD86 and HLADR expression. Activation will be reported as median fluorescence intensity (MFI).

   Unit of Measure: Median fluorescence intensity (MFI)

5. Change from baseline in tumor size measured by ultrasound and mammography for the assessment of the effects on disease progression

   Time frame: Baseline (Month 0), mid-treatment (Month 3), and end of treatment (Month 5-6)

   Tumor size will be assessed using standard clinical imaging methods, including ultrasound and mammography, to evaluate disease progression in patients receiving the UMAY video-immunotherapy or placebo control intervention.

   Tumor size will be recorded as the largest tumor diameter (in millimeters), according to routine clinical assessment.

   Unit of Measure: Millimeters (mm)