Biomarkers in Patients With Metastatic Breast Cancer

Procedure:

• Verification of the inclusion / non-inclusion criteria
• Signing the consent form
• Surgical, Medical and family history, ongoing treatments
• Clinical Examination1
• Radiological examinations 2
• TNM Classification (Appendix 3)
• Standard biochemical assessment 3
• Inflammatory assessment (CRP, LDH, IL6)
• Hematological assessment (NFS, platelets)
• Tumor markers (ACE, CA15-3)
• Histological and molecular examination 4:

• Expression PD-L1 by IHC (Immuno-Histochemistry)
• TILs by IHC
• - MSI status: MMR * proteins (Mismatch Repair Protein) (Appendix 4) by NGS via TruSight™ Oncology 500 Kit (Illumina) , if PD-L1>1%
• Score HRD : by NGS via TruSight™ Oncology 500 Kit (Illumina) , if PD-L1>1%
• TMB mutational burden by next generation sequencing "Next Generation Sequencing" (NGS) via TruSight™ Oncology 500 Kit (Illumina) if PD-L1>1%
• tDNA by digital PCR
• Radiological evaluation5
• Indication of surgery? (if yes type of surgery to be specified)
• Evaluation of adverse events / effects: Tolerance of the previous treatment
• Completion the source documents and the eCRF

Clinical Exam1:

• General State according to the WHO classification or according to the Karnowsky Index
• Breast examination: inspection (symmetry, appearance of the skin), palpation (size of the tumor, presence of mammary discharge).
• Examination of the axillary and supraclavicular lymph node areas
• Abdominal examination
• Pulmonary auscultation
• Skeleton examination Radiological examination2:
• Mammography and echo-mammary
• Thoraco-abdomino-pelvic CT scan
• Bone scintigraphy
• Cardiac ultrasound

Standard biochemical balance3:

Blood group, creatininemia, blood ionogram, calcemia, glycemia, bilirubin, transaminases, gamma-GT, alkaline phosphatases, total cholesterol, triglycerides, uric acid, albuminemia.

Histological examination4:

• Biopsy with part number assigned by the laboratory, name of the laboratory
• Histological type
• Grade of Scarff-Bloom and Richardson (SBR)
• Estrogen receptors, progesterone receptors
• Her2Neu
• Ki67
• PD-L1
• TILs
• MMR

Molecular Examination4:

• MSI status by pentaplex PCR and capillary electrophoresis by NGS platform via the TruSight™ kit Oncology 500 (Illumina)
• HRD Score by NGS platform via the TruSight™ kit Oncology 500 (Illumina)
• Mutational burden TMB by NGS platform via the TruSight™ kit Oncology 500 (Illumina)
• Circulating tumor DNA by digital PCR Radiological assessment5:
• If the stage is locally advanced (appendix 3): Breast ultrasound +/- Mammography +/- Breast MRI
• If metastatic stage: Thoraco-abdomino-pelvic CT scan 12 | 27 Visit details

• Initial visit D0 (V1)
• Signature of informed consent form
• Verification of inclusion / non-inclusion criteria
• Medical, surgical and family history
• Current treatments
• Clinical examination1
• Radiological examinations2
• TNM classification
• Standard biochemical assessment3
• Inflammatory assessment (CRP, LDH, IL6)
• Hematological assessment (NFS, platelets)
• Tumor markers (ACE, CA15-3)
• Histological examination with immunohistochemistry and molecular4:
• Biopsy with part number assigned by the lab, name of the lab
• Histological type
• SBR grade
• Estrogen receptors, progesterone receptors
• Her2Neu
• Ki67
• PD-L1
• TILs
• MMR
• MSI
• HRD
• TMB mutational charge
• ctDNA o Completion of source documents and eCRF

• Visit after 3 chemotherapy treatment cycles (V2)

• Current treatments
• Clinical examination1
• Hematological assessment (NFS, platelets)
• Tumor markers (ACE, CA15-3)
• Radiological assessment5
• Evaluation of adverse events / effects: Tolerance of the previous treatment
• Completion of source documents and eCRF
• Assessment visit (V3) o Current treatments

o Clinical examination1

• TNM classification
• Inflammatory assessment (CRP, LDH, IL6)
• Hematological assessment (NFS, platelets)
• Tumor markers (ACE, CA15-3)
• Histological examination with immunohistochemistry and molecular4:
• Biopsy with part number assigned by the lab, name of the lab
• Histological type
• SBR grade 13 | 27
• Estrogen receptors, progesterone receptors
• Her2Neu
• Ki67
• PD-L1
• TILs
• ctDNA

o Radiological assessment5

o Assessment of adverse events / effects

o Fill in the source files and the eCRF

• Follow-up visit at 3 months (V4)

o Current treatments

o Clinical examination1

o Tumor markers (ACE, CA15-3)

o Radiological examinations2

o Evaluation of adverse events / effects

• Completion of source documents and eCRF
• Follow-up visit at 6 months (V5) o Current treatments o Clinical examination1 o Tumor markers (ACE, CA15-3)

• Radiological examinations2
• Evaluation of adverse events / effects
• Completion of source documents and eCRF

Inclusion criteria

• Age ≥ 18 years old.
• Signed informed consent
• Histological evidence of breast carcinoma.
• Locally advanced (stage III inoperable) or metastatic (stage IV) carcinoma, the metastases of which have been diagnosed straight away or after adjuvant or neoadjuvant treatment.
• No treatment for metastatic disease.
• Metastatic disease: histological evidence of metastasis
• Patients who will receive systemic treatment (chemotherapy, targeted therapy, hormone therapy)
• Life expectancy greater than 3 months

Exclusion criteria

• Non-epithelial breast cancer
• Non-biopsiable metastases
• Patients who have received before systemic treatment for metastatic disease (chemotherapy, targeted therapy, hormone therapy)
• Patients with a contraindication to chemotherapy, and / or hormone therapy, and

/ or targeted therapy.

• Foreign patients