Autophagy as Therapeutic Target in Relapsed/refractory Human Lymphomas

Project location: ITALY, Rome, Milan
Project start date: October 2014 - Project end date: October 2017
Project number: 2014-033
Beneficiary: Arcobaleno ONLUS

 
The project assesses the effects of new therapeutic strategies in the light of the cellular mechanisms involved. It  also aims at providing novel biomarkers of relevance as bioindicators of progression and response to therapy.

This is a medical research project aimed at investigating the role autophagy in affecting the therapeutic response of patients with relapsed/refractory lymphoma to molecularly targeted agents in order to clarify the relevance of autophagy as well as to develop new therapeutic strategies exploiting the modulation of autophagy as a therapeutic challenge.

Autophagy is a cellular degradation pathway for the clearance of damaged or superfluous proteins and organelles. The recycling of these intracellular constituents also serves as an alternative energy source during periods of metabolic stress to maintain homeostasis and viability. In tumour cells with defects in apoptosis, autophagy allows prolonged survival. Paradoxically, autophagy defects are associated with increased tumorigenesis, but the mechanism behind this has not been determined. Recent evidence suggests that autophagy provides a protective function to limit tumour necrosis and inflammation, and to mitigate genome damage in tumour cells in response to metabolic stress.
Lymphoma is the third most common childhood cancer. Approximately 60% of pediatric lymphomas are non-Hodgkin's lymphomas, with the remainder being Hodgkin's lymphomas. In the adults NHL represents 89% of all lymphomas while HL represents 11% of all lymphoma diagnoses. NHL and HL are genetically heterogeneous diseases with a remarkable ability to develop adaptive strategies to cytotoxic or pro-apoptotic challenges. Thus, molecular abnormalities with a key role in the pathogenesis and progression of lymphoma represent attractive targets for exploiting new therapeutic strategies.
Despites therapeutic improvements achieved in the last two decades, significant proportions of lymphomas are not cured with currently available therapeutic strategies, suggesting that new treatments are required particularly for high-risk, relapsed or refractory patients.  The detection of autophagy determinants of possible use in the clinical practice as progression markers or drug response bioindicators will also be a key point of the project. Both non-Hodgkin’s lymphoma (NHL) and Hodgkin’s lymphoma (HL) will be considered.

In the recent years several studies have been carried out by both experimental and clinical points of view dealing with chemical or biological agents capable of activating tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). This member of the tumor necrosis factor (TNF) superfamily was suggested to act as an apoptosis inducer for cancer cells sparing non-tumor cell targets. Unfortunately, clinical benefits deriving from this cell death based approach appeared as limited. In fact, cancer cell susceptibility was variable so that phase I/II clinical trials, carried out by using different ligands such as recombinant soluble TRAIL, specific agonistic antibodies or TRAIL-armed cells (CD34+ cells), although encouraging, gave conflicting results: oncological patients with the same diagnosis were either susceptible or refractory.
It has been reported that TRAIL is able to induce autophagy in certain cancer cells, protecting them by blunting TRAIL’s cytotoxicity and possibly contributing to TRAIL resistance. However, how TRAIL induces autophagy has not been elucidated.
In a recent work we demonstrated that cell susceptibility to TRAIL ligation could be due to the presence of TRAIL death receptors (DR) 4 and 5 in the membrane lipid microdomains (Marconi et al. 2013). Results obtained from a panel of cancer cell lines of hematologic origin as well as from normal cells (lymphocytes, fibroblasts, endothelial cells) demonstrated that recombinant soluble TRAIL, agonistic antibodies or TRAIL-armed cells (CD34+ cells) were effective only if administered to target cells that embed DR4 into lipid microdomains. These results suggest that lipid microdomains could exert a catalytic role for DR4-mediated death triggering.
Based on these results obtained by our group and on our in vitro and ex vivo preliminary results, the project will refer to two main tasks.

•    Task 1: IN VITRO MODULATION OF TRAIL RECEPTORS LOCALIZATION INSIDE OR OUTSIDE MEMBRANE MICRODOMAIS IN HUMAN LYMPHOMA-DERIVED CELLS.
Drugs used in the proposed project will include the PI3K/Akt/MTOR inhibitor perifosine and minerval that activates sphingomyelin synthase (SMS), which results in an important increase in the levels of membrane sphingomyelin (SM) and diacylglycerol (DAG), which have been found to be significantly reduced in cancer cell membranes, used alone or in combination with each other. Initial studies of different human lymphoma-derived cell lines will be carried out in order:
(i) to characterize the modulation of cell death processes (apoptosis, autophagy, the shift autophagy/apoptosis) by the compounds under investigation;
(ii)    to verify whether autophagy plays a role in the survival of treated cells.
(iii)     to explore the relationship between the specific localization of TRAIL receptors and autophagy induction.
•    Task 2: BLOOD SAMPLING FROM RELAPSED/REFRACTORY LYMPHOMA PATIENTS.
Blood samples will be obtained for ex vivo studies from patients with relapsed/refractory lymphoma. According to the experimental design, lymphocytes isolated from peripheral blood samples will be treated with the same protocol used with cell lines.
To obtain new and accurate information on the issues listed above, combined cellular and molecular biology approaches will be carried out. The Istituto Superiore di Sanità, the Humanitas Institute and the Bambino Gesù Pediatric Hospital facilities will provide the environment and the know-how necessary to carry out the proposed research. The main final goal of the project will be to determine the possible prognostic and therapeutic tools deriving from the autophagic machinery targeted agents.

 

Tasks

 36 months

(semesters)

 

  I

 

  II

  III

 

 IV

 

  V

 

 VI

TASK 1. IN VITRO MODULATION OF TRAIL RECEPTORS LOCALIZATION INSIDE OR OUTSIDE MEMBRANE MICRODOMAIS IN HUMAN LYMPHOMA-DERIVED CELLS

 

 

 

 

 

 

Characterization and modulation of cell death processes

 

 

 

 

 

 

 

Evaluation of the role of autophagy in the survival of cells treated with TRAIL

 

 

 

 

 

 

 

Relationship between TRAILRs localization and autophagy induction

 

 

 

 

 

 

TASK 2. BLOOD SAMPLING FROM RELAPSED/REFRACTORY LYMPHOMA PATIENTS.

 

 

 

 

 

 

Patients selection

 

 

 

 

 

 

Characterization of TRAILRs localization and modulation of cell death processes in isolated PBMC

 

 

 

 

 

 

 

Evaluation of the role of autophagy in the survival of PBMC treated with TRAIL

 

 

 

 

 

 

 

 

 

Relationship between TRAILRs localization and autophagy induction in isolated PBMC

 

 

 

 

 

 

 


 The Nando Peretti Foundation has awarded a grant for this project.

think global, act local
you are here: Home  > Projects:  Medical research or Europe  (or Both)  > 2014-033  > Project Description