First name
Letizia
Last name
Amadori
Year of Study
Thesis Title
Identification of molecular mechanisms responsible for degradation of the tumor suppressor protein NUMB in cancer
Thesis Abstract
NUMB was initially described as a cell fate determinant involved in neurogenesis. More recently, NUMB has been implicated in different types of human cancers, in which it has a tumor suppressor role. My research has been focused on the identification of the molecular players responsible for NUMB degradation, which is due to exaggerated ubiquitination and could be restored by proteasome inhibition by MG-132, in breast and lung tumors.
We have assessed an siRNA high-throughput ELISA-based screening for detection of NUMB protein increase upon gene silencing in a NUMB-deficient breast cancer cell line, MDA-MB-361. We have started screening a library containing all human E3-ligase families first and, after two rounds of screening, we validated RBX1 and FBXW8 proteins as putative NUMB downregulators both in breast and lung tumors from human patients, in which NUMB basal expression is low respect to their normal counterpart and could be restored to normal levels after both RBX1 and FBXW8 silencing. These results have opened the possibility that the mechanism responsible for NUMB degradation in NUMB-deficient tumors is a cullin-RING ligase complex-dependent one.
We have also demonstrated that the in vivo treatment of immunocompromised mice with the proteasome inhibitor MG-132 inhibits the growth of NUMB-deficient breast tumors and not that of NUMB- proficient ones through the restoration of NUMB protein levels. This evidence pointed out that targeting loss of NUMB could be used as an anti-cancer therapy for NUMB-deficient breast tumors. We are also performing experiments to demonstrate that the silencing of RBX1 and FBXW8 could inhibit NUMB-deficient tumors' growth potential as well.
We have assessed an siRNA high-throughput ELISA-based screening for detection of NUMB protein increase upon gene silencing in a NUMB-deficient breast cancer cell line, MDA-MB-361. We have started screening a library containing all human E3-ligase families first and, after two rounds of screening, we validated RBX1 and FBXW8 proteins as putative NUMB downregulators both in breast and lung tumors from human patients, in which NUMB basal expression is low respect to their normal counterpart and could be restored to normal levels after both RBX1 and FBXW8 silencing. These results have opened the possibility that the mechanism responsible for NUMB degradation in NUMB-deficient tumors is a cullin-RING ligase complex-dependent one.
We have also demonstrated that the in vivo treatment of immunocompromised mice with the proteasome inhibitor MG-132 inhibits the growth of NUMB-deficient breast tumors and not that of NUMB- proficient ones through the restoration of NUMB protein levels. This evidence pointed out that targeting loss of NUMB could be used as an anti-cancer therapy for NUMB-deficient breast tumors. We are also performing experiments to demonstrate that the silencing of RBX1 and FBXW8 could inhibit NUMB-deficient tumors' growth potential as well.
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