Classical Hodgkin's Lymphoma (cHL) is a rare but highly treatable malignancy of the immune system, primarily affecting young adults. Despite significant therapeutic advancements, frontline treatment failure occurs in up to 30% of cases, with relapse or refractory disease affecting over 50% of these patients. The main therapeutic challenge in cHL remains achieving an optimal balance between disease control and reducing long-term adverse effects. Current prognostic tools only partially capture patient heterogeneity, and cHL continues to evolve spatially and temporally throughout the course of the disease. Personalized treatment strategies require novel integrated tools that better monitor tumor complexity and anticipate disease progression.

Fluorodeoxyglucose positron emission tomography (FDG-PET) has improved risk stratification in cHL, as metabolic response during or after chemotherapy strongly correlates with disease progression and survival. However, FDG-PET has limitations, including the absence of standardized criteria and the necessity to initiate treatment before response assessment. To overcome these limitations, molecular profiling and radiomic analysis of baseline FDG-PET data may provide deeper insights into tumor biology, improving prognostic accuracy.

This observational study aims to dissect the genetic and phenotypic heterogeneity of cHL at diagnosis and during disease evolution, with the goal of identifying novel prognostic biomarkers. These findings could lead to better treatment personalization, increasing cure rates while minimizing treatment-related toxicity. The study is based on the hypothesis that correlating DNA profiling at diagnosis, gene expression, and radiomic features may enable the identification of high-risk signatures, refining prognostic models in cHL. Additionally, liquid biopsy represents a non-invasive method for assessing tumor mutational complexity. The analysis of circulating DNA (cDNA) throughout disease progression could provide insights into genetic evolution and help predict overt progression before clinical manifestations occur.

The primary objective is to define the genetic mutational profile of cHL at disease progression. As secondary objectives, it will evaluate whether liquid biopsy can accurately recapitulate the genetic heterogeneity observed in tumor tissue, determine the predictive accuracy of liquid biopsy in anticipating disease progression, and correlate genomic and radiomic features with patient outcomes to refine risk stratification and therapeutic decision-making.

By integrating molecular and imaging-based biomarkers, this study aims to enhance personalized treatment strategies, improve risk-adapted therapeutic approaches, and ultimately optimize curability and quality of life for patients with cHL.

This study has both retrospective and prospective components. The prospective part includes patients with Classical Hodgkin's Lymphoma (cHL) who have completed active treatment and entered follow-up (Cohort A), as well as patients with a histological confirmation of relapse or progression (Cohort B). The retrospective cohort (Cohort C) consists of 235 consecutive cHL patients collected from the archives of the Hematology Unit of AUSL-IRCCS between 2004 and 2019, along with up to 250 cHL patients diagnosed between 2016 and 2021 from other Italian hematology centers.

Inclusion Criteria:

• Age >18 anni
• Written informed consent signed

Cohort A

• Age >18 years
• Histologically confirmed diagnosis of relapsed/refractory classical Hodgkin lymphoma identified during induction or follow-up
• Available formalin-fixed, paraffin-embedded (FFPE) biopsy at diagnosis and at the time of progression/relapse
• Available plasma sample at progression (before the beginning of salvage therapy)
• Available FDG-PET evaluation at study enrollment
• Available clinical, laboratory, and radiologic data at diagnosis and relapse

Cohort B

• Diagnosis of classical Hodgkin lymphoma
• Completion of first-line standard systemic treatment (chemotherapy-based or chemoradiotherapy combined modality)
• Available plasma sample at the end of treatment (at least 30 days from the last chemotherapy)
• Available FFPE biopsy at diagnosis
• No further treatment planned
• Available clinical, laboratory, and radiologic data at diagnosis and response evaluation
• Patient's willingness to undergo 6 months follow-up plasma sample collection and to attend regular follow-up

Cohort C

• Histologically confirmed diagnosis of classical Hodgkin lymphoma
• Standard treatment as per available guidelines (e.g., ESMO guidelines)
• Available treatment data, response, and follow-up data
• Available FFPE biopsy at diagnosis
• Available FDG-PET evaluation at study enrollment

Exclusion Criteria:

• Patients with nodular lymphocyte predominant Hodgkin lymphoma are not eligible; all other subtypes including nodular sclerosis, lymphocyte-depleted, lymphocyte-rich, and mixed cellularity Hodgkin lymphoma may be enrolled.
• Active HIV, HBV, HCV viral infection
• Concomitant neoplasm not treated with a curative aim

The study will involve three patient cohorts, with participants recruited from multiple centers across Italy during routine follow-up visits.

Cohort A will include 70 patients with classical Hodgkin lymphoma (cHL) who have a histological confirmation of relapse or progression. These patients will be enrolled at the time of disease relapse/progression, and plasma samples for ctDNA analysis will be collected concurrently.

Cohort B will consist of 200 cHL patients who have completed active treatment and entered the follow-up phase. Plasma samples will be collected from these patients at the time of enrollment (P0) and again at a six-month follow-up (P6FU). This cohort will be followed for a minimum of two years from the time of enrollment.

Cohort C is a retrospective cohort that will be divided into two subgroups:

Training cohort: 235 consecutive cHL patients collected from the archives of the Hematology Unit of AUSL-IRCCS between 2004 and 2019. This cohort is representative of the ge