Publications by authors named "Alessandra Maria Storaci"

8 Publications

  • Page 1 of 1

V-ATPase controls tumor growth and autophagy in a Drosophila model of gliomagenesis.

Autophagy 2021 May 12:1-11. Epub 2021 May 12.

Department of Biosciences, Università Degli Studi Di Milano, Milan, Italy.

Glioblastoma (GBM), a very aggressive and incurable tumor, often results from constitutive activation of EGFR (epidermal growth factor receptor) and of phosphoinositide 3-kinase (PI3K). To understand the role of autophagy in the pathogenesis of glial tumors , we used an established model of glioma based on overexpression in larval glial cells of an active human and of the PI3K homolog . Interestingly, the resulting hyperplastic glia express high levels of key components of the lysosomal-autophagic compartment, including vacuolar-type H-ATPase (V-ATPase) subunits and ref(2)P (refractory to Sigma P), the homolog of SQSTM1/p62. However, cellular clearance of autophagic cargoes appears inhibited upstream of autophagosome formation. Remarkably, downregulation of subunits of V-ATPase, of , or of the Tor (Target of rapamycin) complex 1 (TORC1) component prevents overgrowth and normalize ref(2)P levels. In addition, downregulation of the V-ATPase subunit reduces Akt and Tor-dependent signaling and restores clearance. Consistent with evidence in flies, neurospheres from patients with high V-ATPase subunit expression show inhibition of autophagy. Altogether, our data suggest that autophagy is repressed during glial tumorigenesis and that V-ATPase and MTORC1 components acting at lysosomes could represent therapeutic targets against GBM.
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http://dx.doi.org/10.1080/15548627.2021.1918915DOI Listing
May 2021

Interplay Between V-ATPase G1 and Small EV-miRNAs Modulates ERK1/2 Activation in GBM Stem Cells and Nonneoplastic Milieu.

Mol Cancer Res 2020 11 4;18(11):1744-1754. Epub 2020 Aug 4.

Division of Pathology, Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico, Milan, Italy.

The ATP6V1G1 subunit (V1G1) of the vacuolar proton ATPase (V-ATPase) pump is crucial for glioma stem cells (GSC) maintenance and tumorigenicity. Moreover, V-ATPase reprograms the tumor microenvironment through acidification and release of extracellular vesicles (EV). Therefore, we investigated the role of V1G1 in GSC small EVs and their effects on primary brain cultures. To this end, small EVs were isolated from patients-derived GSCs grown as neurospheres (NS) with high (V1G1-NS) or low (V1G1-NS) V1G1 expression and analyzed for V-ATPase subunits presence, miRNA contents, and cellular responses in recipient cultures. Our results show that NS-derived small EVs stimulate proliferation and motility of recipient cells, with small EV derived from V1G1-NS showing the most pronounced activity. This involved activation of ERK1/2 signaling, in a response reversed by V-ATPase inhibition in NS-producing small EV. The miRNA profile of V1G1-NS-derived small EVs differed significantly from that of V1G1-NS, which included miRNAs predicted to target MAPK/ERK signaling. Mechanistically, forced expression of a MAPK-targeting pool of miRNAs in recipient cells suppressed MAPK/ERK pathway activation and blunted the prooncogenic effects of V1G1 small EV. These findings propose that the GSC influences the brain milieu through a V1G1-coordinated EVs release of MAPK/ERK-targeting miRNAs. Interfering with V-ATPase activity could prevent ERK-dependent oncogenic reprogramming of the microenvironment, potentially hampering local GBM infiltration. IMPLICATIONS: Our data identify a novel molecular mechanism of gliomagenesis specific of the GBM stem cell niche, which coordinates a V-ATPase-dependent reprogramming of the brain microenvironment through the release of specialized EVs.
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http://dx.doi.org/10.1158/1541-7786.MCR-20-0078DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7642188PMC
November 2020

MFF Regulation of Mitochondrial Cell Death Is a Therapeutic Target in Cancer.

Cancer Res 2019 Dec 3;79(24):6215-6226. Epub 2019 Oct 3.

Prostate Cancer Discovery and Development Program.

The regulators of mitochondrial cell death in cancer have remained elusive, hampering the development of new therapies. Here, we showed that protein isoforms of mitochondrial fission factor (MFF1 and MFF2), a molecule that controls mitochondrial size and shape, that is, mitochondrial dynamics, were overexpressed in patients with non-small cell lung cancer and formed homo- and heterodimeric complexes with the voltage-dependent anion channel-1 (VDAC1), a key regulator of mitochondrial outer membrane permeability. MFF inserted into the interior hole of the VDAC1 ring using Arg225, Arg236, and Gln241 as key contact sites. A cell-permeable MFF Ser223-Leu243 d-enantiomeric peptidomimetic disrupted the MFF-VDAC1 complex, acutely depolarized mitochondria, and triggered cell death in heterogeneous tumor types, including drug-resistant melanoma, but had no effect on normal cells. In preclinical models, treatment with the MFF peptidomimetic was well-tolerated and demonstrated anticancer activity in patient-derived xenografts, primary breast and lung adenocarcinoma 3D organoids, and glioblastoma neurospheres. These data identify the MFF-VDAC1 complex as a novel regulator of mitochondrial cell death and an actionable therapeutic target in cancer. SIGNIFICANCE: These findings describe mitochondrial fission regulation using a peptidomimetic agent that disturbs the MFF-VDAC complex and displays anticancer activity in multiple tumor models..
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http://dx.doi.org/10.1158/0008-5472.CAN-19-1982DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6911621PMC
December 2019

Mitochondrial fission factor is a novel Myc-dependent regulator of mitochondrial permeability in cancer.

EBioMedicine 2019 Oct 18;48:353-363. Epub 2019 Sep 18.

Prostate Cancer Discovery and Development Program, USA; Immunology, Microenvironment and Metastasis Program, The Wistar Institute, Philadelphia, PA 19104, USA. Electronic address:

Background: Mitochondrial functions are exploited in cancer and provide a validated therapeutic target. However, how this process is regulated has remained mostly elusive and the identification of new pathways that control mitochondrial integrity in cancer is an urgent priority.

Methods: We studied clinically-annotated patient series of primary and metastatic prostate cancer, representative cases of multiple myeloma (MM) and publicly available genetic databases. Gene regulation studies involved chromatin immunoprecipitation, PCR amplification and Western blotting of conditional Myc-expressing cell lines. Transient or stable gene silencing was used to quantify mitochondrial functions in bioenergetics, outer membrane permeability, Ca homeostasis, redox balance and cell death. Tumorigenicity was assessed by cell proliferation, colony formation and xenograft tumour growth.

Findings: We identified Mitochondrial Fission Factor (MFF) as a novel transcriptional target of oncogenic Myc overexpressed in primary and metastatic cancer, compared to normal tissues. Biochemically, MFF isoforms, MFF1 and MFF2 associate with the Voltage-Dependent Anion Channel-1 (VDAC1) at the mitochondrial outer membrane, in vivo. Disruption of this complex by MFF silencing induces general collapse of mitochondrial functions with increased outer membrane permeability, loss of inner membrane potential, Ca unbalance, bioenergetics defects and activation of cell death pathways. In turn, this inhibits tumour cell proliferation, suppresses colony formation and reduces xenograft tumour growth in mice.

Interpretation: An MFF-VDAC1 complex is a novel regulator of mitochondrial integrity and actionable therapeutic target in cancer.
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http://dx.doi.org/10.1016/j.ebiom.2019.09.017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6838406PMC
October 2019

Specific V-ATPase expression sub-classifies IDHwt lower-grade gliomas and impacts glioma growth in vivo.

EBioMedicine 2019 Mar 5;41:214-224. Epub 2019 Feb 5.

Division of Pathology, Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico, Milan, Italy; Department of Pathophysiology and Transplantation, Università degli Studi di Milano, Milan, Italy; Fondazione Istituto Nazionale Genetica Molecolare 'Romeo ed Enrica Invernizzi', Milan, Italy. Electronic address:

Background: Cancer cells use specific V-ATPase subunits to activate oncogenic pathways. Therefore, we investigated V-ATPase deregulation in aggressive gliomas and associated signaling.

Methods: V-ATPase genes expression and associated pathways were analyzed in different series of glioma available from public databases, as well as in patients' cohort. Activation of pathways was analyzed at gene and protein expression levels. A genetic model of glioma in Drosophila melanogaster and mice with GBM patients-derived orthotopic xenografts were used as in vivo models of disease.

Findings: GBM and recurrent gliomas display a specific V-ATPase signature. Such signature resolves the heterogeneous class of IDH-wild type lower-grade gliomas, identifying the patients with worse prognosis independently from clinical and molecular features (p = 0·03, by Cox proportional-hazards model). In vivo, V-ATPase subunits deregulation significantly impacts tumor growth and proliferation. At the molecular level, GBM-like V-ATPase expression correlates with upregulation of Homeobox genes.

Interpretation: Our data identify a V-ATPase signature that accompanies glioma aggressiveness and suggest new entry points for glioma stratification and follow-up. FUND: This work was supported by Fondazione Cariplo (2014-1148 to VV), Fondazione IRCCS Ca' Granda, and Fondazione INGM Grant in Molecular Medicine 2014 (to VV).
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http://dx.doi.org/10.1016/j.ebiom.2019.01.052DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6441867PMC
March 2019

A GBM-like V-ATPase signature directs cell-cell tumor signaling and reprogramming via large oncosomes.

EBioMedicine 2019 Mar 6;41:225-235. Epub 2019 Feb 6.

Division of Pathology, Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico, Milan, Italy; Department of Pathophysiology and Transplantation, Università degli Studi di Milano, Milan, Italy; Fondazione Istituto Nazionale Genetica Molecolare 'Romeo ed Enrica Invernizzi', Milan, Italy. Electronic address:

Background: The V-ATPase proton pump controls acidification of intra and extra-cellular milieu in both physiological and pathological conditions. We previously showed that some V-ATPase subunits are enriched in glioma stem cells and in patients with poor survival. In this study, we investigated how expression of a GBM-like V-ATPase pump influences the non-neoplastic brain microenvironment.

Methods: Large oncosome (LO) vesicles were isolated from primary glioblastoma (GBM) neurospheres, or from patient sera, and co-cultured with primary neoplastic or non-neoplastic brain cells. LO transcript and protein contents were analyzed by qPCR, immunoblotting and immunogold staining. Activation of pathways in recipient cells was determined at gene and protein expression levels. V-ATPase activity was impaired by Bafilomycin A1 or gene silencing.

Findings: GBM neurospheres influence their non-neoplastic microenvironment by delivering the V-ATPase subunit V1G1 and the homeobox genes HOXA7, HOXA10, and POU3F2 to recipient cells via LO. LOs reprogram recipient cells to proliferate, grow as spheres and to migrate. Moreover, LOs are particularly abundant in the circulation of GBM patients with short survival time. Finally, impairment of V-ATPase reduces LOs activity.

Interpretation: We identified a novel mechanism adopted by glioma stem cells to promote disease progression via LO-mediated reprogramming of their microenvironment. Our data provide preliminary evidence for future development of LO-based liquid biopsies and suggest a novel potential strategy to contrast glioma progression. FUND: This work was supported by Fondazione Cariplo (2014-1148 to VV) and by the Italian Minister of Health-Ricerca Corrente program 2017 (to SF).
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http://dx.doi.org/10.1016/j.ebiom.2019.01.051DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6441844PMC
March 2019

Correction: SNP rs12435998, a predictor of glioblastoma survival and response to radio-chemotherapy.

Oncotarget 2018 08 24;9(66):32731. Epub 2018 Aug 24.

Institute for Genetic and Biomedical Research, National Research Council, Milan 20138, Italy.

[This corrects the article DOI: 10.18632/oncotarget.3611.].
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http://dx.doi.org/10.18632/oncotarget.26066DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6135690PMC
August 2018

SEL1L SNP rs12435998, a predictor of glioblastoma survival and response to radio-chemotherapy.

Oncotarget 2015 May;6(14):12452-67

Institute for Genetic and Biomedical Research, National Research Council, Milan 20138, Italy.

The suppressor of Lin-12-like (C. elegans) (SEL1L) is involved in the endoplasmic reticulum (ER)-associated degradation pathway, malignant transformation and stem cells. In 412 formalin-fixed and paraffin-embedded brain tumors and 39 Glioblastoma multiforme (GBM) cell lines, we determined the frequency of five SEL1L single nucleotide genetic variants with regulatory and coding functions by a SNaPShot™ assay. We tested their possible association with brain tumor risk, prognosis and therapy. We studied the in vitro cytotoxicity of valproic acid (VPA), temozolomide (TMZ), doxorubicin (DOX) and paclitaxel (PTX), alone or in combination, on 11 GBM cell lines, with respect to the SNP rs12435998 genotype. The SNP rs12435998 was prevalent in anaplastic and malignant gliomas, and in meningiomas of all histologic grades, but unrelated to brain tumor risks. In GBM patients, the SNP rs12435998 was associated with prolonged overall survival (OS) and better response to TMZ-based radio-chemotherapy. GBM stem cells with this SNP showed lower levels of SEL1L expression and enhanced sensitivity to VPA.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4494950PMC
http://dx.doi.org/10.18632/oncotarget.3611DOI Listing
May 2015
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