PhD in BIOCHEMISTRY AND MOLECULAR BIOLOGY

email: caccuri@uniroma2.it
phone: 3315365482
building: torre E sud
room: 204

CURRICULUM VITAE DI ANNA MARIA CACCURI
CURRENT POSITION: Full Professor of Biochemistry at the Department of Chemical Sciences and Technologies of the University of Rome "Tor Vergata"
RESEARCH ACTIVITY
Since 1983, the scientific activity of Anna Maria Caccuri has focused on the study of the structural and functional properties of enzymes belonging to the class of glutathione S-transferases (GSTs). In this period of time, she has carried out kinetic and spectroscopic studies on GSTs of class Alpha, Mu and Pi, and contributed to the understanding of the roles played by some amino acid residues in maintaining the catalytically competent structure of these GST isoenzymes. These studies have revealed the existence of a cooperative interaction between the GST subunits, involving the binding site of the substrate GSH. She has also investigated the kinetic mechanism of GSTs in the presence of the new substrate 7-chloro-4-nitrobenzo-2-oxo-1,3-diazole, contributing to the development of a more sensitive assay to measure GST activity.
For 20 years, Anna Maria Caccuri has carried out studies comparing the major human GSTs with those most phylogenetically distant. This includes defining the catalytic mechanism of GST of Lucilia cuprina, an insect that infests sheep farms in Australia. She studied the catalytic mechanism of the human Theta isoenzyme (GSTT2-2), an ancestral enzyme characterized by considerable structural rigidity and low catalytic efficiency. She clarified the catalytic mechanism of GSTB1-1 isoenzyme (class Beta) of bacterial origin and of the human GSTZ1-1 isoenzyme (class Zeta). All the work cited above, has highlighted the different roles played, in the course of evolution, by some conserved amino acid residues in the GST active site. The data above revealed an evolutionary process in the GST superfamily. Starting from the bacterial isoenzyme Beta, which is similar to the redox glutaredoxin enzymes, this process generated the human isoenzymes Alpha, Mu and Pi, characterized by increased structural flexibility and catalytic efficiency.
Over the years, the role played by GSTs in the onset of drug resistance in tumors has become increasingly evident. This has prompted the search for new efficient inhibitors of these enzymes to counteract the phenomenon.
Since 2002, Anna Maria Caccuri has been involved in the design, synthesis and characterization of new selective inhibitors of GSTs described in the United States patent (US 8,796,317 B2 of 05/08/2014). The mechanism of action of these compounds differs from those of other known inhibitors that target GSTs. In fact, these molecules are not GSH peptidomimetic analogues but derivatives of 7-nitro-2,1,3-benzoxadiazole. These aromatic compounds are recognized as substrates by GST and conjugated with GSH giving rise, in the active site, to a sigma-complex intermediate which inhibits the Pi and Mu GST isoforms very efficiently (Kd = 10-9-10-11 M). The inhibition is highly selective since the inhibitor is formed by the same target enzyme.
Subsequently, Anna Maria Caccuri has extended her research field by studying the mechanism of action of these new molecules, in particular of 6-(7-nitro-2,1,3-benzoxadiazol-4-ylthio)hexanol (NBDHEX), at cellular level. Her research found that the binding of NBDHEX to GSTP1-1, the GST isoform overexpressed in many tumors, induces the dissociation of the protein from the complex with c-Jun N-terminal kinase (JNK). This triggers the activation of JNK, followed by tumor cell death by apoptosis. NBDHEX can therefore be considered an activator of the apoptotic pathway mediated by the MAP kinase JNK.
Furthermore, she established that NBDHEX is not recognized as a substrate by the P-glycoprotein and MRP1 (multidrug-resistance associated protein) membrane pumps, which are among the major factors responsible for drug resistance in cancer cells. The compound is therefore not actively expelled from tumor cells by these pumps, thus proving highly pro-apoptotic even against tumors resistant to many chemotherapeutic drugs including vinblastine, doxorubicin, cisplatin, methotrexate, and etoposide.
Anna Maria Caccuri has investigated, directly or in collaboration with other research groups, the NBDHEX efficacy on a panel of tumor cell lines of different origin, both sensitive and resistant to the most common chemotherapeutic agents. In particular, her work evaluated the efficacy of NBDHEX on leukemia (K562, CCRF-CEM, CEM-VBL10, CEM-VBL100, HL60, HL60 / DNR and HL60 / ADR, both sensitive and resistant to vinblastine, daunorubicin and adriamycin), small cell lung cancer (GLC-4, H69, H69AR both sensitive and resistant to adriamycin), hepatic carcinoma (HepG2), osteosarcoma (U-2OS, Saos-2 both sensitive and resistant to cisplatin, methotrexate and adriamycin), Ewing sarcoma (TC -71, 6647, SK-ES-1), rhabdomyosarcoma (SJRH30, SJ-RH4, RMZ-RC2, RD/18), mesothelioma (MSTO-211H, MPP89, MM-B1 and Mero 48a) and melanoma (Me501, A375, B16) cell lines. The compound was found to be very active on most of these cell lines, revealing IC50 values in the nano- micromolar concentration range. Her findings indicate that NBDHEX may have a cytotoxic or cytostatic action which is effective against tumor cells and their chemo-resistant variants.
Drug-combination studies, whether her own or in collaboration, have shown synergistic activity with cisplatin, adriamycin, vincristine and temozolomide, suggesting the use of NBDHEX in combination with these drugs for the treatment of osteosarcoma, Ewing's sarcoma, mesothelioma and melanoma.
Such studies have also demonstrated the antitumor efficacy of NBDHEX in vivo in mouse models of human melanoma, Ewing's sarcoma and osteosarcoma. In these experiments, NBDHEX was not toxic but was well tolerated.
In another collaboration, the crystal structures of the molecular complexes between NBDHEX and the GSTs of class Pi (GSTP1-1) and Mu (GSTM2-2) were solved. These studies have provided a detailed characterization, at a molecular level, of the interaction between NBDHEX and the active site of these GST isoforms, resulting in the design and synthesis of new NBDHEX derivatives.
The new compounds are described in the Italian patent n ° 0001412189 of 11/20/2014. While these new NBDHEX derivatives maintain the nitrobenzoxadiazole scaffold, which guarantees an effective interaction with the active site of GSTs, they also contain a polar or charged group which improves their solubility in aqueous solution. Many of these new GST inhibitors are characterized by a greater selectivity towards the GSTP1-1 isoform, highly expressed in tumors.
In particular, the compound MC3181 is 50 times more soluble in water than NBDHEX and highly effective against human melanoma cells carrying the BRAFV600E mutation, both sensitive and resistant to vemurafenib. MC3181 also shows antitumor activity in vivo, against both types of tumor, after i.v. or oral administration. Furthermore, the in vivo treatment is completely tolerated and does not cause organotoxicity.
Over the last few years, the work of Anna Maria Caccuri has led to the synthesis and characterization of the new analogues MC3479 e MC3626. These compounds are phosphate monoesters of NBDHEX and MC3181 characterized by very good solubility in aqueous solutions. Furthermore, these compounds have proved safe and effective when administered intravenously or orally as an aqueous solution to mice xenografted with A375 human melanoma tumor cells.
Collectively, these findings support the potential use of 7-nitro-2,1,3-benzoxadiazole derivatives in novel therapies for the treatment of different types of tumors, in particular of melanoma.
In 2012, Anna Maria Caccuri obtained experimental evidence for the interaction between GSTP1-1 and TNF receptor-associated factor 2 (TRAF2) and for the involvement of this protein in the cellular response to the treatment with NBDHEX and its derivatives. In particular, she demonstrated that NBDHEX, MC3181 and other analogues are able to dissociate the complex between GSTP1-1 and TRAF2, thereby leading to prolonged JNK activation and cell death by apoptosis.
Anna Maria Caccuri's research activity has therefore focused on TRAF2 and, in particular, on the finding that the C-terminal truncated form of TRAF2 (residues 310-501) is able to bind unilamellar vesicles containing lipid rafts and induce the formation of intra-luminary vesicles. Based on this evidence, Anna Maria Caccuri has started a line of research aimed at studying the properties of the C-terminal domain of TRAF2 and its possible involvement in the formation of multivesicular bodies and / or in the autophagic process. To this end, she is conducting research on TRAF2 knockout cell lines or on cells transfected with the C-terminal portion of TRAF2. In the hope it will result in new strategies for drug-delivery, she is also studying how best to exploit the TRAF2 C-terminal domain’s ability to induce endovesiculation.
Since 2016, Anna Maria Caccuri has collaborated with the IRCCS Foundation "G.B. Bietti” in studying the molecular mechanisms involved in the pathogenesis of retinal neurodegenerative diseases and to clarify the role played in these processes by the proteins GSTP1-1 and TRAF2.