Publications by authors named "Soldano Ferrone"

317 Publications

A Pan-Histone Deacetylase Inhibitor Enhances the Antitumor Activity of B7-H3-specific CAR-T Cells in Solid Tumors.

Clin Cancer Res 2021 Apr 2. Epub 2021 Apr 2.

Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Sun Yat-sen Memorial Hospital

Purpose: The limited efficacy of chimeric antigen receptor (CAR) T cell therapies with solid malignancies prompted us to test whether epigenetic therapy could enhance the antitumor activity of B7-H3 CAR T cells with several solid cancer types.

Experimental Design: We evaluated B7-H3 expression in many human solid cancer and normal tissue samples. The efficacy of the combinatorial therapy with B7-H3.CAR-T cells and the deacetylase inhibitor SAHA with several solid cancer types and the potential underlying mechanisms were characterized with and experiments.

Results: B7-H3 is expressed in most of the human solid tumor samples tested, but exhibits a restricted expression in normal tissues. B7-H3.CAR-T cells selectively killed B7-H3 expressing human cancer cell lines A low dose of SAHA upregulated B7-H3 expression in several types of solid cancer cells at the transcriptional level and B7-H3.CAR expression on human transgenic T cell membrane. In contrast, the expression of immunosuppressive molecules, such as CTLA-4 and TET2, by T cells was downregulated upon SAHA treatment. A low dose of SAHA significantly enhanced the antitumor activity of B7-H3.CAR-T cells with solid cancers and , including orthotopic PDX and metastatic models treated with autologous CAR-T cell infusions.

Conclusions: Our results show that our novel strategy which combines SAHA and B7-H3.CAR-T cells enhances their therapeutic efficacy with solid cancers and justify its translation to a clinical setting.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-2487DOI Listing
April 2021

Disulfiram Acts as a Potent Radio-Chemo Sensitizer in Head and Neck Squamous Cell Carcinoma Cell Lines and Transplanted Xenografts.

Cells 2021 Feb 28;10(3). Epub 2021 Feb 28.

Department of Otolaryngology, Head and Neck Surgery, Charité-Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Berlin Institute of Health, Charité Campus Benjamin Franklin, Hindenburgdamm 30, 12203 Berlin, Germany.

The poor prognosis of locally advanced and metastatic head and neck squamous cell carcinoma (HNSCC) is primarily mediated by the functional properties of cancer stem cells (CSCs) and resistance to chemoradiotherapy. We investigated whether the aldehyde dehydrogenase (ALDH) inhibitor disulfiram (DSF) can enhance the sensitivity of therapy. Cell viability was assessed by the 1-(4,5-dimethylthiazol-2-yl)-3,5-diphenylformazan (MTT) and apoptosis assays, and the cell cycle and reactive oxygen species (ROS) levels were evaluated by fluorescence-activated cell sorting (FACS). The radio-sensitizing effect was measured by a colony formation assay. The synergistic effects were calculated by combination index (CI) analyses. The DSF and DSF/Cu inhibited the cell proliferation (inhibitory concentration 50 (IC) of DSF and DSF/Cu were 13.96 μM and 0.24 μM). DSF and cisplatin displayed a synergistic effect (CI values were < 1). DSF or DSF/Cu abolished the cisplatin-induced G2/M arrest (from 52.9% to 40.7% and 41.1%), and combining irradiation (IR) with DSF or DSF/Cu reduced the colony formation and attenuated the G2/M arrest (from 53.6% to 40.2% and 41.9%). The combination of cisplatin, DSF or DSF/Cu, and IR enhanced the radio-chemo sensitivity by inducing apoptosis (42.04% and 32.21%) and ROS activity (46.3% and 37.4%). DSF and DSF/Cu enhanced the sensitivity of HNSCC to cisplatin and IR. Confirming the initial data from patient-derived tumor xenograft (PDX) supported a strong rationale to repurpose DSF as a radio-chemosensitizer and to assess its therapeutic potential in a clinical setting.
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http://dx.doi.org/10.3390/cells10030517DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7999545PMC
February 2021

Targeting Radiation-Resistant Prostate Cancer Stem Cells by B7-H3 CAR T Cells.

Mol Cancer Ther 2021 03;20(3):577-588

Division of Surgical Oncology, Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts.

Radiotherapy (RT) is a key treatment for prostate cancer. However, RT resistance can contribute to treatment failure. Prostate cancer stem cells (PCSCs) are radioresistant. We recently found that fractionated irradiation (FIR) upregulates expression of the immune checkpoint B7-H3 (CD276) on PCSCs and bulk cells in each prostate cancer cell line tested. These findings prompted us to investigate whether B7-H3 targeting chimeric antigen receptor (CAR) T cells, which may abrogate function of an immune checkpoint and mediate lysis of targeted cells, can target RT-resistant PCSCs and . B7-H3 expression is naturally higher on PCSCs than bulk prostate cancer cells and cytotoxicity of B7-H3 CAR T cells to PCSCs is more potent than to bulk prostate cancer cells. Furthermore, FIR significantly upregulates B7-H3 expression on PCSCs and bulk prostate cancer cells. The duration of FIR or single-dose irradiation-induced further upregulation of B7-H3 on bulk prostate cancer cells and PCSCs lasts for up to 3 days. B7-H3 CAR T-cell cytotoxicity against FIR-resistant PCSCs at a low effector to target ratio of 1:1 was assessed by flow cytometry and sphere formation assays. Further upregulation of B7-H3 expression by FIR made PCSCs even more sensitive to B7-H3 CAR T-cell-mediated killing. Consequently, the FIR and B7-H3 CAR T-cell therapy combination is much more effective than FIR or CAR T cells alone in growth inhibition of hormone-insensitive prostate cancer xenografts in immunodeficient mice. Our work provides a sound basis for further development of this unique combinatorial model of RT and B7-H3 CAR T-cell therapy for prostate cancer. SIGNIFICANCE: We demonstrate that FIR significantly upregulates B7-H3 expression by RT-resistant PCSCs and bulk cells; cytotoxicity of B7-H3 CAR T cells to FIR-treated PCSCs is potent and results in significantly improved antitumor efficacy in mice.
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http://dx.doi.org/10.1158/1535-7163.MCT-20-0446DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7952034PMC
March 2021

Proteomic profile of melanoma cell-derived small extracellular vesicles in patients' plasma: a potential correlate of melanoma progression.

J Extracell Vesicles 2021 Feb 11;10(4):e12063. Epub 2021 Feb 11.

UPMC Hillman Cancer Center University of Pittsburgh Cancer Institute Pittsburgh Pennsylvania USA.

Molecular profiling of small extracellular vesicles (sEV) isolated from plasma of cancer patients emerges as promising strategy for biomarkers discovery. We investigated the proteomic profiles of sEV immunoselected using anti-CSPG4 antibodies from 15 melanoma patients' plasma. The proteomes of sEV separated into melanoma cell-derived (MTEX) and non-malignant cell-derived (NMTEX) were compared using high-resolution mass spectrometry. Paired analysis identified the MTEX-associated profile of 16 proteins that discriminated MTEX from NMETEX. We also identified the MTEX profile that discriminated between seven patients with no evidence of melanoma (NED) after therapy and eight with progressive disease (PD). Among 75 MTEX proteins overexpressed in PD patients, PDCD6IP (ALIX) had the highest discriminating value, while CNTN1 (contactin-1) was upregulated only in MTEX of NED patients. This is the first report documenting that proteomes of tumour-derived sEV in patients' plasma discriminate cancer from non-cancer and identify proteins with potential to serve as prognostic biomarkers in melanoma.
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http://dx.doi.org/10.1002/jev2.12063DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7876545PMC
February 2021

Spatial Analysis and Clinical Significance of HLA Class-I and Class-II Subunit Expression in Non-Small Cell Lung Cancer.

Clin Cancer Res 2021 Feb 18. Epub 2021 Feb 18.

Department of Pathology, Yale University School of Medicine, New Haven, Connecticut.

Purpose: To analyze the distribution, associated immune contexture, and clinical significance of human leukocyte antigen (HLA) class-I and HLA class-II subunits in non-small cell lung cancer (NSCLC).

Experimental Design: Using spatially resolved and quantitative multiplexed immunofluorescence we studied the tumor/stromal tissue distribution, cancer cell-specific defects, and clinicopathologic/survival associations of β2 microglobulin (β2M), HLA-A, and HLA-B,-C heavy chains, as well as HLA class-II β chain in >700 immunotherapy-naïve NSCLCs from four independent cohorts. Genomic analysis of HLA genes in NSCLC was performed using two publicly available cohorts.

Results: Cancer cell-specific downregulation of HLA markers was identified in 30.4% of cases. β2M was downregulated in 9.8% (70/714), HLA-A in 9% (65/722), HLA-B,-C in 12.1% (87/719), and HLA class-II in 17.7% (127/717) of evaluable samples. Concurrent downregulation of β2M, HLA-B,-C, and HLA class-II was commonly identified. Deleterious mutations in HLA genes were detected in <5% of lung malignancies. Tumors with cancer cell-specific β2M downregulation displayed reduced T cells and increased natural killer (NK)-cell infiltration. Samples with cancer cell HLA-A downregulation displayed modest increase in CD8 T cells and NK-cell infiltration. Samples with cancer cell-selective HLA-B,-C or HLA class-II downregulation displayed reduced T cells and NK-cell infiltration. There was limited association of the markers with clinicopathologic variables and KRAS/EGFR mutations. Cancer cell-selective downregulation of the HLA subunits was associated with shorter overall survival.

Conclusions: Our results reveal frequent and differential defects in HLA class-I and HLA class-II protein subunit expression in immunotherapy-naïve NSCLCs associated with distinct tumor microenvironment composition and patient survival.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-3655DOI Listing
February 2021

Proceedings From the First International Workshop at Sidra Medicine: "Engineered Immune Cells in Cancer Immunotherapy (EICCI): From Discovery to Off-the-Shelf Development", 15-16 February 2019, Doha, Qatar.

Front Immunol 2020 14;11:589381. Epub 2021 Jan 14.

Research Department, Sidra Medicine, Doha, Qatar.

The progress in the isolation and characterization of tumor antigen (TA)-specific T lymphocytes and in the genetic modification of immune cells allowed the clinical development of adoptive cell therapy (ACT). Several clinical studies highlighted the striking clinical activity of T cells engineered to express either Chimeric Antigen (CAR) or T Cell (TCR) Receptors to target molecularly defined antigens expressed on tumor cells. The breakthrough of immunotherapy is represented by the approval of CAR-T cells specific for advanced or refractory CD19 B cell malignancies by both the Food and Drug Administration (FDA) and the European Medicinal Agency (EMA). Moreover, advances in the manufacturing and gene editing of engineered immune cells contributed to the selection of drug products with desired phenotype, refined specificity and decreased toxicity. An important step toward the optimization of CAR-T cell therapy is the development of "off-the shelf" T cell products that allow to reduce the complexity and the costs of the manufacturing and to render these drugs available for a broad number of cancer patients. The Engineered Immune Cells in Cancer Immunotherapy (EICCI) workshop hosted in Doha, Qatar, renowned experts, from both academia and industry, to present and discuss the progress on both pre-clinical and clinical development of genetically modified immune cells, including advances in the "off-the-shelf" manufacturing. These experts have addressed also organizational needs and hurdles for the clinical grade production and application of these biological drugs.
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http://dx.doi.org/10.3389/fimmu.2020.589381DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7874217PMC
January 2021

Modifications to the Framework Regions Eliminate Chimeric Antigen Receptor Tonic Signaling.

Cancer Immunol Res 2021 Feb 5. Epub 2021 Feb 5.

Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina.

Chimeric antigen receptor (CAR) tonic signaling, defined as spontaneous activation and release of proinflammatory cytokines by CAR-T cells, is considered a negative attribute because it leads to impaired antitumor effects. Here, we report that CAR tonic signaling is caused by the intrinsic instability of the mAb single-chain variable fragment (scFv) to promote self-aggregation and signaling via the CD3ζ chain incorporated into the CAR construct. This phenomenon was detected in a CAR encoding either CD28 or 4-1BB costimulatory endodomains. Instability of the scFv was caused by specific amino acids within the framework regions (FWR) that can be identified by computational modeling. Substitutions of the amino acids causing instability, or humanization of the FWRs, corrected tonic signaling of the CAR, without modifying antigen specificity, and enhanced the antitumor effects of CAR-T cells. Overall, we demonstrated that tonic signaling of CAR-T cells is determined by the molecular instability of the scFv and that computational analyses of the scFv can be implemented to correct the scFv instability in CAR-T cells with either CD28 or 4-1BB costimulation.
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http://dx.doi.org/10.1158/2326-6066.CIR-20-0451DOI Listing
February 2021

Preclinical Evaluation of B7-H3-specific Chimeric Antigen Receptor T Cells for the Treatment of Acute Myeloid Leukemia.

Clin Cancer Res 2021 Feb 2. Epub 2021 Feb 2.

Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, North Carolina.

Purpose: The development of safe and effective chimeric antigen receptor (CAR) T-cell therapy for acute myeloid leukemia (AML) has largely been limited by the concomitant expression of most AML-associated surface antigens on normal myeloid progenitors and by the potential prolonged disruption of normal hematopoiesis by the immunotargeting of these antigens. The purpose of this study was to evaluate B7-homolog 3 (B7-H3) as a potential target for AML-directed CAR T-cell therapy. B7-H3, a coreceptor belonging to the B7 family of immune checkpoint molecules, is overexpressed on the leukemic blasts of a significant subset of patients with AML and may overcome these limitations as a potential target antigen for AML-directed CAR-T therapy.

Experimental Design: B7-H3 expression was evaluated on AML cell lines, primary AML blasts, and normal bone marrow progenitor populations. The antileukemia efficacy of B7-H3-specific CAR-T cells (B7-H3.CAR-T) was evaluated using coculture models and xenograft models of disseminated AML, including patient-derived xenograft models. The potential hematopoietic toxicity of B7-H3.CAR-Ts was evaluated using colony formation assays and in a humanized mouse model.

Results: B7-H3 is expressed on monocytic AML cell lines and on primary AML blasts from patients with monocytic AML, but is not significantly expressed on normal bone marrow progenitor populations. B7-H3.CAR-Ts exhibit efficient antigen-dependent cytotoxicity and in xenograft models of AML, and are unlikely to cause unacceptable hematopoietic toxicity.

Conclusions: B7-H3 is a promising target for AML-directed CAR-T therapy. B7-H3.CAR-Ts control AML and have a favorable safety profile in preclinical models.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-2540DOI Listing
February 2021

Human hepatitis B virus negatively impacts the protective immune cross-talk between natural killer and dendritic cells.

Hepatology 2021 Jan 22. Epub 2021 Jan 22.

Laboratory of Immunology and Biotherapy, University of Messina, Messina, Italy.

NK cells play a crucial role in the clearance of human viruses but their activity is significantly impaired in chronic hepatitis B infected patients (CHB). Cooperation with dendritic cells (DCs) is pivotal for obtaining optimal NK cell antiviral function, thus we investigated whether HBV might impact the ability of DCs to sustain NK cell functions. Human DCs were poor stimulators of IFN-γ production by NK cells when exposed to HBV, while maintained capability to trigger NK cell cytotoxicity. HBV prevented DC maturation but did not affect their expression of HLA class I, thus allowing DCs to evade NK cell lysis. Tolerogenic features of DCs exposed to HBV were further supported by their increased expression of IL-10 and immunosuppressive enzyme indoleamine-2, 3-dioxygenase that contributed to the impairment of DC-mediated NK cell IFN-γ production and proliferation respectively. HBV could also inhibit the expression of inducible immunoproteasome (iP) subunits on DCs. In fact, NK cells could induce iP subunit expression on DCs but they failed in the presence of HBV. Remarkably, circulating BDCA1 DCs isolated from CHB patients were functionally compromised, hence altering, in turn, NK cell responses. The abnormal NK/DC interplay caused by HBV may significantly impair the efficacy of antiviral immune response in CHB patients.
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http://dx.doi.org/10.1002/hep.31725DOI Listing
January 2021

A monocentric phase I study of vemurafenib plus cobimetinib plus PEG-interferon (VEMUPLINT) in advanced melanoma patients harboring the V600BRAF mutation.

J Transl Med 2021 01 6;19(1):17. Epub 2021 Jan 6.

Istituto Nazionale Tumori-IRCCS-Fondazione G Pascale, Naples, Italy.

Background: Studies carried out in vitro and in a mouse model have shown that BRAF inhibitors enhance the effects of IFN-α on BRAFV600E melanoma cells through the inhibition of ERK. Therefore, the combination of vemurafenib and IFN-α in patients with BRAFV600E melanoma may provide therapeutic benefits; MEK inhibition may prevent the reactivation of the MAPK pathway induced by BRAF inhibitor resistance.

Patients And Methods: In a phase I study, adult patients with advanced BRAFV600-mutated melanoma were treated with vemurafenib + PEG-IFN-α-2b or vemurafenib + cobimetinib + PEG-IFN-α-2b, to assess the safety of the combination and the upregulation of IFN-α/β receptor-1 (IFNAR1).

Results: Eight patients were treated; 59 adverse events with four serious ones (three related to study treatments) were reported. Patients with a pre-treatment IFNAR1 expression on ≤ 35% melanoma cells had a median progression-free survival of 12.0 months (range: 5.6-18.4 months) and a median overall survival of 31.0 months (range: 19.8-42.2 months), while patients with a pre-treatment IFNAR1 expression on > 35% of melanoma cells had a median progression-free survival of 4.0 months (range: 0-8.8; p = 0.03), and a median overall survival of 5 months (p = 0.02). Following treatment, responders had higher levels of growth-suppressor genes, including GAS1 and DUSP1, and genes involved in a metabolically robust immune response, including FAP.

Conclusion: Our study supports the overall safety of the vemurafenib + PEG-IFN-α-2b + cobimetinib combination. IFNAR1 expression levels correlated with response to treatment, including survival. Vemurafenib + PEG-IFN-α-2b + cobimetinib would have difficulty finding a niche in the current treatment scenario for advanced melanoma, but we speculate that our findings may contribute to identify subjects particularly responsive to treatment.

Trial Registration: The study was registered at clinicaltrials.gov (NCT01959633). Registered 10 October 2013, https://clinicaltrials.gov/ct2/show/NCT01959633.
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http://dx.doi.org/10.1186/s12967-020-02680-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7789377PMC
January 2021

B7-H3 targeted antibody-based immunotherapy of malignant diseases.

Expert Opin Biol Ther 2020 Dec 21:1-16. Epub 2020 Dec 21.

Department of Surgery, Massachusetts General Hospital/Harvard Medical School , Boston, MA, USA.

: Recent advances in immuno-oncology and bioengineering have rekindled the interest in monoclonal antibody (mAb)-based immunotherapies for malignancies. Crucial for their success is the identification of tumor antigens (TAs) that can serve as targets. B7-H3, a member of the B7 ligand family, represents such a TA. Although its exact functions and receptor(s) remain unclear, B7-H3 has predominantly a pro-tumorigenic effect mainly by suppressing the anti-tumor functions of T-cells. : Initially we present a historical perspective on TA-specific antibodies for diagnosis and treatment of malignancies. Following a description of the TA requirements to be an attractive antibody-based immunotherapy target, we show that B7-H3 fulfills these criteria. We discuss its structure and functions. In a review and pooled analysis, we describe the limited B7-H3 expression in normal tissues and estimate B7-H3 expression frequency in tumors, tumor-associated vasculature and cancer initiating cells (CICs). Lastly, we discuss the association of B7-H3 expression in tumors with poor prognosis. : B7-H3 is an attractive target for mAb-based cancer immunotherapy. B7-H3-targeting strategies are expected to be highly effective and - importantly - safe. To fully exploit the diagnostic and therapeutic potential of B7-H3, its expression in pre-malignant lesions, serum, metastases, and CICs requires further investigation.
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http://dx.doi.org/10.1080/14712598.2021.1862791DOI Listing
December 2020

Defective HLA Class I Expression and Patterns of Lymphocyte Infiltration in Chordoma Tumors.

Clin Orthop Relat Res 2020 Dec 11. Epub 2020 Dec 11.

S. S. Patel, Orthopaedic Spine Service, Department of Orthopaedic Surgery, Washington University School of Medicine, St. Louis, MO, USA S. S. Patel, S. P. Nota, S. Ferrone, J. H. Schwab, Orthopaedic Oncology Service, Department of Orthopaedic Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA F. Sabbatino, X. Wang, S. Ferrone, Surgical Oncology Service, Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA G. P. Nielsen, V. Deshpande, Department of Pathology, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA.

Background: There are no effective systemic therapies for chordoma. The recent successes of immunotherapeutic strategies in other cancers have resulted in a resurgence of interest in using immunotherapy in chordoma. These approaches rely on a functional interaction between the host's immune system and the expression of tumor peptides via the human leukocyte antigen (HLA) Class I antigen. It is not known whether chordoma cells express the HLA Class I antigen.

Questions/purposes: (1) Do chordoma tumors exhibit defects in HLA Class I antigen expression? (2) What is the pattern of lymphocyte infiltration in chordoma tumors?

Methods: Patients with chordoma treated at Massachusetts General Hospital between 1989 and 2009 were identified with permission from the institutional review board. Of the 75 patients who were identified, 24 human chordoma tumors were selected from 24 distinct patients based on tissue availability. Histology slides from these 24 formalin-fixed paraffin-embedded chordoma tissue samples were deparaffinized using xylene and ethanol and underwent heat-induced antigen retrieval in a citrate buffer. Samples were incubated with monoclonal antibodies directed against HLA Class I antigen processing machinery components. Antibody binding was detected via immunohistochemical staining. Staining intensity (negative, weakly positive, strongly positive) was assessed semiquantitatively and the percentage of chordoma cells stained for HLA Class I antigen subunits was assessed quantitatively. Hematoxylin and eosin-stained histology slides from the same 24 chordoma samples were assessed qualitatively for the presence of tumor-infiltrating lymphocytes and histologic location of these lymphocytes. Immunohistochemical staining with monoclonal antibodies directed against CD4 and CD8 was performed in a quantitative manner to identify the lymphocyte subtype present in chordoma tumors. All results were scored independently by two investigators and were confirmed by a senior bone and soft tissue pathologist.

Results: Seven of 24 chordoma samples exhibited no staining by the anti-HLA-A heavy chain monoclonal antibody HC-A2, two had weak staining intensity, and eight had a heterogeneous staining pattern, with fewer than 60% of chordoma cells exhibiting positive staining results. Four of 24 samples tested were not stained by the anti-HLA-B/C heavy chain monoclonal antibody HC-10, five had weak staining intensity, and 11 displayed a heterogeneous staining pattern. For the anti-β-2-microglobulin monoclonal antibody NAMB-1, staining was detected in all samples, but 11 had weak staining intensity and four displayed a heterogeneous staining pattern. Twenty-one of 24 samples tested had decreased expression in at least one subunit of HLA Class I antigens. No tumors were negative for all three subunits. Lymphocytic infiltration was found in 21 of 24 samples. Lymphocytes were primarily found in the fibrous septae between chordoma lobules but also within the tumor lobules and within the fibrous septae and tumor lobules. Twenty-one of 24 tumors had CD4+ T cells and 11 had CD8+ T cells.

Conclusion: In chordoma tissue samples, HLA Class I antigen defects commonly were present, suggesting a mechanism for escape from host immunosurveillance. Additionally, nearly half of the tested samples had cytotoxic CD8+ T cells present in chordoma tumors, suggesting that the host may be capable of mounting an immune response against chordoma tumors. The resulting selective pressure imposed on chordoma tumors may lead to the outgrowth of chordoma cell subpopulations that can evade the host's immune system.

Clinical Relevance: These findings have implications in the design of immunotherapeutic strategies for chordoma treatment. T cell recognition of tumor cells requires HLA Class I antigen expression on the targeted tumor cells. Defects in HLA Class I expression may play a role in the clinical course of chordoma and may account for the limited or lack of efficacy of T cell-based immunity triggered by vaccines and/or checkpoint inhibitors.
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http://dx.doi.org/10.1097/CORR.0000000000001587DOI Listing
December 2020

The SPPL3-Defined Glycosphingolipid Repertoire Orchestrates HLA Class I-Mediated Immune Responses.

Immunity 2021 Jan 2;54(1):132-150.e9. Epub 2020 Dec 2.

Department of Immunopathology, Sanquin Research, Amsterdam, the Netherlands; Landsteiner Laboratory, Amsterdam UMC, University of Amsterdam, Amsterdam, the Netherlands; Cancer Center Amsterdam, Amsterdam, the Netherlands. Electronic address:

HLA class I (HLA-I) glycoproteins drive immune responses by presenting antigens to cognate CD8 T cells. This process is often hijacked by tumors and pathogens for immune evasion. Because options for restoring HLA-I antigen presentation are limited, we aimed to identify druggable HLA-I pathway targets. Using iterative genome-wide screens, we uncovered that the cell surface glycosphingolipid (GSL) repertoire determines effective HLA-I antigen presentation. We show that absence of the protease SPPL3 augmented B3GNT5 enzyme activity, resulting in upregulation of surface neolacto-series GSLs. These GSLs sterically impeded antibody and receptor interactions with HLA-I and diminished CD8 T cell activation. Furthermore, a disturbed SPPL3-B3GNT5 pathway in glioma correlated with decreased patient survival. We show that the immunomodulatory effect could be reversed through GSL synthesis inhibition using clinically approved drugs. Overall, our study identifies a GSL signature that inhibits immune recognition and represents a potential therapeutic target in cancer, infection, and autoimmunity.
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http://dx.doi.org/10.1016/j.immuni.2020.11.003DOI Listing
January 2021

Peritumoral Immune Infiltrate as a Prognostic Biomarker in Thin Melanoma.

Front Immunol 2020 29;11:561390. Epub 2020 Sep 29.

Department of Medicine, Surgery and Dentistry "Scuola Medica Salernitana", University of Salerno, Baronissi, Italy.

Thin melanomas are tumors less than 1 mm thick according to Breslow classification. Their prognosis is in most cases excellent. However, a small subset of these tumors relapses. These clinical findings emphasize the need of novel prognostic biomarkers to identify this subset of tumors. Characterization of tumor immune microenvironment (TIME) is currently investigated as a prognostic and predictive biomarker for cancer immunotherapy in several solid tumors including melanoma. Here, taking into account the limited availability of tumor tissues, by characterizing some of the characteristics of TIME such as number of infiltrating lymphocytes, HLA class I antigen and PD-L1 expression, we show that number of infiltrating CD8+ and FOXP3+ T cells as well as CD8+/FOXP3+ T cell ratio can represent a useful prognostic biomarker in thin melanoma. Although further investigations in a larger patient cohort are needed, these findings have potential clinical significance since they can be used to define subgroups of thin melanoma patients who have a worse prognosis and might need different treatment modalities.
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http://dx.doi.org/10.3389/fimmu.2020.561390DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7550791PMC
September 2020

B7-H3: An Attractive Target for Antibody-based Immunotherapy.

Clin Cancer Res 2021 Mar 13;27(5):1227-1235. Epub 2020 Oct 13.

Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts.

The recent impressive clinical responses to antibody-based immunotherapy have prompted the identification of clinically relevant tumor antigens that can serve as targets in solid tumors. Among them, B7-H3, a member of the B7 ligand family, represents an attractive target for antibody-based immunotherapy, it is overexpressed on differentiated malignant cells and cancer-initiating cells, with limited heterogeneity, and high frequency (60% of 25,000 tumor samples) in many different cancer types, but has a limited expression at low level in normal tissues. In nonmalignant tissues, B7-H3 has a predominantly inhibitory role in adaptive immunity, suppressing T-cell activation and proliferation. In malignant tissues, B7-H3 inhibits tumor antigen-specific immune responses, leading to a protumorigenic effect. B7-H3 also has nonimmunologic protumorigenic functions, such as promoting migration and invasion, angiogenesis, chemoresistance, and endothelial-to-mesenchymal transition, as well as affecting tumor cell metabolism. As a result, B7-H3 expression in tumors is associated with poor prognosis. Although experimental B7-H3 silencing reduces cancer cell malignant potential, there has been limited emphasis on the development of B7-H3-blocking antibodies, most likely because the B7-H3 receptor remains unknown. Instead, many antibody-based strategies utilizing distinct effector mechanisms to target B7-H3-expressing cancer cells have been developed. These strategies have demonstrated potent antitumor activity and acceptable safety profiles in preclinical models. Ongoing clinical trials are assessing their safety and efficacy in patients. Identification of the B7-H3 receptor will improve our understanding of its role in tumor immunity, and will suggest rational strategies to develop blocking antibodies, which may enhance the therapeutic efficacy of tumor immunity.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-2584DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7925343PMC
March 2021

The HDAC Inhibitor Domatinostat Promotes Cell-Cycle Arrest, Induces Apoptosis, and Increases Immunogenicity of Merkel Cell Carcinoma Cells.

J Invest Dermatol 2021 Apr 28;141(4):903-912.e4. Epub 2020 Sep 28.

Department of Translational Skin Cancer Research (TSCR), German Cancer Consortium (DKTK), Partner Site Essen, German Cancer Research Center, Heidelberg, Germany; Department of Dermatology, University Hospital Essen, Essen, Germany. Electronic address:

Merkel cell carcinoma (MCC) is a rare, highly aggressive skin cancer for which immune modulation by immune checkpoint inhibitors shows remarkable response rates. However, primary or secondary resistance to immunotherapy prevents benefits in a significant proportion of patients. For MCC, one immune escape mechanism is insufficient for recognition by T cells owing to the downregulation of major histocompatibility complex I surface expression. Histone deacetylase inhibitors have been demonstrated to epigenetically reverse the low major histocompatibility complex I expression caused by the downregulation of the antigen-processing machinery. Domatinostat, an orally available small-molecule inhibitor targeting histone deacetylase class I, is currently in clinical evaluation to overcome resistance to immunotherapy. In this study, we present preclinical data on domatinostat's efficacy and mode of action in MCC. Single-cell RNA sequencing revealed a distinct gene expression signature of antigen processing and presentation, cell-cycle arrest, and execution phase of apoptosis on treatment. Accordingly, functional assays showed that domatinostat induced G2M arrest and apoptosis. In the surviving cells, antigen-processing machinery component gene transcription and translation were upregulated, consequently resulting in increased major histocompatibility complex I surface expression. Altogether, domatinostat not only exerts direct antitumoral effects but also restores HLA class I surface expression on MCC cells, therefore, restoring surviving MCC cells' susceptibility to recognition and elimination by cognate cytotoxic T cells.
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http://dx.doi.org/10.1016/j.jid.2020.08.023DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7987731PMC
April 2021

NK-Cell-Mediated Targeting of Various Solid Tumors Using a B7-H3 Tri-Specific Killer Engager In Vitro and In Vivo.

Cancers (Basel) 2020 Sep 18;12(9). Epub 2020 Sep 18.

Department of Medicine, Division of Hematology, Oncology, and Transplantation, University of Minnesota, Minneapolis, MN 55455, USA.

We improved the bispecific antibody platform that primarily engages natural killer (NK) cells to kill cancer cells through antibody-dependent cellular cytotoxicity (ADCC) by adding IL-15 as a crosslinker that expands and self-sustains the effector NK cell population. The overall goal was to target B7-H3, an established marker predominantly expressed on cancer cells and minimally expressed on normal cells, and prove that it could target cancer cells in vitro and inhibit tumor growth in vivo. The tri-specific killer engager (TriKE) was assembled by DNA shuffling and ligation using DNA encoding a camelid anti-CD16 antibody fragment, a wild-type IL-15 moiety, and an anti-B7-H3 scFv (clone 376.96). The expressed and purified cam1615B7H3 protein was tested for in vitro NK cell activity against a variety of tumors and in vivo against a tagged human MA-148 ovarian cancer cell line grafted in NSG mice. cam1615B7H3 showed specific NK cell expansion, high killing activity across a range of B7-H3+ carcinomas, and the ability to mediate growth inhibition of aggressive ovarian cancer in vivo. cam1615B7H3 TriKE improves NK cell function, expansion, targeted cytotoxicity against various types of B7-H3-positive human cancer cell lines, and delivers an anti-cancer effect in vivo in a solid tumor setting.
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http://dx.doi.org/10.3390/cancers12092659DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7564091PMC
September 2020

CSPG4-Specific CAR.CIK Lymphocytes as a Novel Therapy for the Treatment of Multiple Soft-Tissue Sarcoma Histotypes.

Clin Cancer Res 2020 Dec 8;26(23):6321-6334. Epub 2020 Sep 8.

Candiolo Cancer Institute, FPO-IRCCS, Candiolo, Turin, Italy.

Purpose: No effective therapy is available for unresectable soft-tissue sarcomas (STS). This unmet clinical need prompted us to test whether chondroitin sulfate proteoglycan 4 (CSPG4)-specific chimeric antigen receptor (CAR)-redirected cytokine-induced killer lymphocytes (CAR.CIK) are effective in eliminating tumor cells derived from multiple STS histotypes and in immunodeficient mice.

Experimental Design: The experimental platform included patient-derived CAR.CIK and cell lines established from multiple STS histotypes. CAR.CIK were transduced with a retroviral vector encoding second-generation CSPG4-specific CAR (CSPG4-CAR) with 4-1BB costimulation. The functional activity of CSPG4-CAR.CIK was explored , in two- and three-dimensional STS cultures, and in three STS xenograft models.

Results: CSPG4-CAR.CIK were efficiently generated from patients with STS. CSPG4 was highly expressed in multiple STS histotypes by analysis and on all 16 STS cell lines tested by flow cytometry. CSPG4-CAR.CIK displayed superior cytolytic activity against multiple STS histotypes as compared with paired unmodified control CIK. CSPG4-CAR.CIK also showed strong antitumor activity against STS spheroids; this effect was associated with tumor recruitment, infiltration, and matrix penetration. CSPG4-CAR.CIK significantly delayed or reversed tumor growth in three STS xenograft models (leiomyosarcoma, undifferentiated pleomorphic sarcoma, and fibrosarcoma). Tumor growth inhibition persisted for up to 2 weeks following the last administration of CSPG4-CAR.CIK.

Conclusions: This study has shown that CSPG4-CAR.CIK effectively targets multiple STS histotypes and in immunodeficient mice. These results provide a strong rationale to translate the novel strategy we have developed into a clinical setting.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-0357DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7710537PMC
December 2020

Perspectives in melanoma: meeting report from the "Melanoma Bridge" (December 5th-7th, 2019, Naples, Italy).

J Transl Med 2020 09 7;18(1):346. Epub 2020 Sep 7.

Cancer Diagnosis Program, Division of Cancer Treatment and Diagnosis, NCI, Bethesda, MD, USA.

The melanoma treatment landscape changed in 2011 with the approval of the first anti-cytotoxic T-lymphocyte-associated protein (CTLA)-4 checkpoint inhibitor and of the first BRAF-targeted monoclonal antibody, both of which significantly improved overall survival (OS). Since then, improved understanding of the tumor microenvironment (TME) and tumor immune-evasion mechanisms has resulted in new approaches to targeting and harnessing the host immune response. The approval of new immune and targeted therapies has further improved outcomes for patients with advanced melanoma and other combination modalities are also being explored such as chemotherapy, radiotherapy, electrochemotherapy and surgery. In addition, different strategies of drugs administration including sequential or combination treatment are being tested. Approaches to overcome resistance and to potentiate the immune response are being developed. Increasing evidence emerges that tissue and blood-based biomarkers can predict the response to a therapy. The latest findings in melanoma research, including insights into the tumor microenvironment and new biomarkers, improved understanding of tumor immune response and resistance, novel approaches for combination strategies and the role of neoadjuvant and adjuvant therapy, were the focus of discussions at the Melanoma Bridge meeting (5-7 December, 2019, Naples, Italy), which are summarized in this report.
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http://dx.doi.org/10.1186/s12967-020-02482-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7487701PMC
September 2020

IL15 Stimulation with TIGIT Blockade Reverses CD155-mediated NK-Cell Dysfunction in Melanoma.

Clin Cancer Res 2020 Oct 26;26(20):5520-5533. Epub 2020 Jun 26.

Division of Hematology/Oncology, Department of Medicine, University of Pittsburgh, School of Medicine, Pittsburgh, Pennsylvania.

Purpose: Natural killer (NK) cells play a critical role in tumor immunosurveillance. Multiple activating and inhibitory receptors (IR) regulate NK-cell-mediated tumor control. The IR T-cell immunoglobulin and ITIM domain (TIGIT) and its counter-receptor CD226 exert opposite effects on NK-cell-mediated tumor reactivity.

Experimental Design: We evaluated the frequency, phenotype, and functions of NK cells freshly isolated from healthy donors and patients with melanoma with multiparameter flow cytometry. We assessed TIGIT and CD226 cell surface expression and internalization upon binding to CD155. We evaluated the role of IL15 and TIGIT blockade in increasing NK-cell-mediated cytotoxicity and in two mouse models.

Results: NK cells are present at low frequencies in metastatic melanoma, are dysfunctional, and downregulate both TIGIT and CD226 expression. As compared with TIGIT NK cells, TIGIT NK cells exhibit higher cytotoxic capacity and maturation, but paradoxically lower cytotoxicity against CD155 MHC class I-deficient melanoma cells. Membrane bound CD155 triggers CD226 internalization and degradation, resulting in decreased NK-cell-mediated tumor reactivity. IL15 increases TIGIT and CD226 gene expression by tumor-infiltrating NK cells (TiNKs) and, together with TIGIT blockade, increases NK-cell-mediated melanoma cytotoxicity and decreases tumor metastasis in two mouse melanoma models. Specific deletion of TIGIT on transferred NK cells enhances the antimetastatic activity of IL15, while CD226 blockade decreases the effects of IL15 and TIGIT blockade.

Conclusions: Our findings support the development of novel combinatorial immunotherapy with IL15 and TIGIT blockade to promote NK-cell-mediated destruction of MHC class I-deficient melanoma, which are refractory to CD8 T-cell-mediated immunity..
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http://dx.doi.org/10.1158/1078-0432.CCR-20-0575DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8045409PMC
October 2020

A fast, simple, and cost-effective method of expanding patient-derived xenograft mouse models of pancreatic ductal adenocarcinoma.

J Transl Med 2020 06 24;18(1):255. Epub 2020 Jun 24.

Division of Surgical Oncology, Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA.

Background: Patient-derived xenograft (PDX) mouse models of cancer have been recognized as better mouse models that recapitulate the characteristics of original malignancies including preserved tumor heterogeneity, lineage hierarchy, and tumor microenvironment. However, common challenges of PDX models are the significant time required for tumor expansion, reduced tumor take rates, and higher costs. Here, we describe a fast, simple, and cost-effective method of expanding PDX of pancreatic ductal adenocarcinoma (PDAC) in mice.

Methods: We used two established frozen PDAC PDX tissues (derived from two different patients) and implanted them subcutaneously into SCID mice. After tissues reached 10-20 mm in diameter, we performed survival surgery on each mouse to harvest 90-95% of subcutaneous PDX (incomplete resection), allowing the remaining 5-10% of PDX to continue growing in the same mouse.

Results: We expanded three consecutive passages (P1, P2, and P3) of PDX in the same mouse. Comparing the times required for in vivo expansion, P2 and P3 (expanded through incomplete resection) grew 26-60% faster than P1. Moreover, such expanded PDX tissues were successfully implanted orthotopically into mouse pancreases. Within 20 weeks using only 14 mice, we generated sufficient PDX tissue for future implantation of 200 mice. Our histology study confirmed that the morphologies of cancer cells and stromal structures were similar across all three passages of subcutaneous PDX and the orthotopic PDX and were reflective of the original patient tumors.

Conclusions: Taking advantage of incomplete resection of tumors associated with high local recurrence, we established a fast method of PDAC PDX expansion in mice.
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http://dx.doi.org/10.1186/s12967-020-02414-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7315507PMC
June 2020

Improving the Clinical Significance of Preclinical Immunotherapy Studies through Incorporating Tumor Microenvironment-like Conditions.

Clin Cancer Res 2020 Sep 22;26(17):4448-4453. Epub 2020 Jun 22.

Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts.

Frequently, the results generated when testing novel antitumor immunotherapies do not correlate with data collected in models and/or in clinical settings. It is our hypothesis that this discrepancy is caused by the use of conditions, such as normoxia, a two-dimensional surface, optimal growth media, and lack of cell complexity and heterogeneity. These conditions do not accurately reflect the tumor microenvironment (TME) that the tested immunotherapeutic strategies experience While there are many variables which can have an impact upon the antitumor efficacy of an immunotherapy, the immunosuppressive TME is one in which several of the conditions commonly found can be mimicked These conditions, which include hypoxia, low pH, low glucose, presence of adenosine, cell complexity and heterogeneity, as well as the three-dimensional structure of TME, can all affect immune cell-tumor cell interactions. Here, we discuss the impact that these conditions, either individually or in combination, can have on these interactions. Furthermore, we propose that performing assays under TME-like conditions improves the clinical relevance of the yielded results. This, in turn, contributes to accelerate the speed, reduce the cost, and increase efficiency of screening novel immunotherapies and eventually the development of prospective clinical trials.
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http://dx.doi.org/10.1158/1078-0432.CCR-20-0358DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7483623PMC
September 2020

Role of the anatomic site in the association of HLA class I antigen expression level in metastases with clinical response to ipilimumab therapy in patients with melanoma.

J Immunother Cancer 2020 06;8(1)

Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, USA.

Background: The clinical response to immune checkpoint inhibitors (ICIs) in only part of the treated patients, in conjunction with the potentially serious side effects associated with this type of therapy, has emphasized the need to identify biomarkers to select patients who may benefit from ICI treatment. The aim of our study was to test human leukocyte antigen (HLA) class I and II expression in melanoma metastases as potential biomarkers of response to ipilimumab and survival in patients with metastatic melanoma, since these molecules play a crucial role in the interactions of malignant cells with host's immune system.

Materials And Methods: HLA class I and II antigen expression level in pretreatment surgical tissue samples (50 lymph node and 35 cutaneous or subcutaneous metastases) from 30 patients was analyzed by immunohistochemical staining with monoclonal antibodies. Expression levels were correlated to intratumoral density of lymphocytes expressing cluster of differentiation (CD)8, CD45RO, CD4, forkhead box P3 (FOXP3) and/or programmed cell death protein 1 (PD-1), to clinical response to treatment, and to patients' survival.

Results: HLA class I antigen expression level in lymph node metastases, but not in cutaneous or subcutaneous metastases was significantly correlated to density of CD8 and CD45RO T cells and of lymphocytes expressing PD-1, as well as to clinical response and to patients' survival.

Conclusions: Our results corroborate the role of HLA class I expression level (alone or in combination with T-cell density values) as a predictive biomarker of response to ipilimumab in patients with melanoma. In addition, our results show that this association is influenced by the anatomic site of the metastasis used to measure the HLA class I antigen expression level.
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http://dx.doi.org/10.1136/jitc-2019-000209DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7304850PMC
June 2020

Tumor Microenvironment Immune Response in Pancreatic Ductal Adenocarcinoma Patients Treated With Neoadjuvant Therapy.

J Natl Cancer Inst 2021 Feb;113(2):182-191

Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA.

Background: Neoadjuvant folinic acid, fluorouracil, irinotecan, and oxaliplatin (FOLFIRINOX) and chemoradiation have been used to downstage borderline and locally advanced pancreatic ductal adenocarcinoma (PDAC). Whether neoadjuvant therapy-induced tumor immune response contributes to the improved survival is unknown. Therefore, we evaluated whether neoadjuvant therapy induces an immune response towards PDAC.

Methods: Clinicopathological variables were collected for surgically resected PDACs at the Massachusetts General Hospital (1998-2016). Neoadjuvant regimens included FOLFIRINOX with or without chemoradiation, proton chemoradiation (25 Gy), photon chemoradiation (50.4 Gy), or no neoadjuvant therapy. Human leukocyte antigen (HLA) class I and II expression and immune cell infiltration (CD4+, FoxP3+, CD8+, granzyme B+ cells, and M2 macrophages) were analyzed immunohistochemically and correlated with clinicopathologic variables. The antitumor immune response was compared among neoadjuvant therapy regimens. All statistical tests were 2-sided.

Results: Two hundred forty-eight PDAC patients were included. The median age was 64 years and 50.0% were female. HLA-A defects were less frequent in the FOLFIRINOX cohort (P = .006). HLA class II expression was lowest in photon and highest in proton patients (P = .02). The FOLFIRINOX cohort exhibited the densest CD8+ cell infiltration (P < .001). FOLFIRINOX and proton patients had the highest CD4+ and lowest T regulatory (FoxP3+) cell density, respectively. M2 macrophage density was statistically significantly higher in the treatment-naïve group (P < .001) in which dense M2 macrophage infiltration was an independent predictor of poor overall survival.

Conclusions: Neoadjuvant FOLFIRINOX with or without chemoradiation may induce immunologically relevant changes in the tumor microenvironment. It may reduce HLA-A defects, increase CD8+ cell density, and decrease T regulatory cell and M2 macrophage density. Therefore, neoadjuvant FOLFIRINOX therapy may benefit from combinations with checkpoint inhibitors, which can enhance patients' antitumor immune response.
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http://dx.doi.org/10.1093/jnci/djaa073DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7850539PMC
February 2021

Targeting the innate immunoreceptor RIG-I overcomes melanoma-intrinsic resistance to T cell immunotherapy.

J Clin Invest 2020 08;130(8):4266-4281

Department of Dermatology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.

Understanding tumor resistance to T cell immunotherapies is critical to improve patient outcomes. Our study revealed a role for transcriptional suppression of the tumor-intrinsic HLA class I (HLA-I) antigen processing and presentation machinery (APM) in therapy resistance. Low HLA-I APM mRNA levels in melanoma metastases before immune checkpoint blockade (ICB) correlated with nonresponsiveness to therapy and poor clinical outcome. Patient-derived melanoma cells with silenced HLA-I APM escaped recognition by autologous CD8+ T cells. However, targeted activation of the innate immunoreceptor RIG-I initiated de novo HLA-I APM transcription, thereby overcoming T cell resistance. Antigen presentation was restored in interferon-sensitive (IFN-sensitive) but also immunoedited IFN-resistant melanoma models through RIG-I-dependent stimulation of an IFN-independent salvage pathway involving IRF1 and IRF3. Likewise, enhanced HLA-I APM expression was detected in RIG-Ihi (DDX58hi) melanoma biopsies, correlating with improved patient survival. Induction of HLA-I APM by RIG-I synergized with antibodies blocking PD-1 and TIGIT inhibitory checkpoints in boosting the antitumor T cell activity of ICB nonresponders. Overall, the herein-identified IFN-independent effect of RIG-I on tumor antigen presentation and T cell recognition proposes innate immunoreceptor targeting as a strategy to overcome intrinsic T cell resistance of IFN-sensitive and IFN-resistant melanomas and improve clinical outcomes in immunotherapy.
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http://dx.doi.org/10.1172/JCI131572DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7410049PMC
August 2020

Correction to: Induction of immunogenic cell death in radiation-resistant breast cancer stem cells by repurposing anti-alcoholism drug disulfiram.

Cell Commun Signal 2020 Apr 2;18(1):53. Epub 2020 Apr 2.

Division of Surgical Oncology, Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, USA.

An amendment to this paper has been published and can be accessed via the original article.
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http://dx.doi.org/10.1186/s12964-020-00567-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7115073PMC
April 2020

Induction of immunogenic cell death in radiation-resistant breast cancer stem cells by repurposing anti-alcoholism drug disulfiram.

Cell Commun Signal 2020 03 5;18(1):36. Epub 2020 Mar 5.

Division of Surgical Oncology, Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, USA.

Background: The current successful clinical use of agents promoting robust anti-tumor immunity in cancer patients warrants noting that radiation therapy (RT) induces immunogenic cell death (ICD) of tumor cells, which can generate anti-tumor immune responses. However, breast cancer stem cells (BCSCs) are resistant to RT and RT alone usually failed to mount an anti-tumor immune response.

Methods: High aldehyde dehydrogenase activity (ALDH) and CD44/CD24/ESA cancer cells, previously shown to have BCSC properties, were isolated from human MDA-MB-231 and UACC-812 breast cancer cell lines by flow cytometer. Flow sorted BCSCs and non-BCSCs were further tested for their characteristic of stemness by mammosphere formation assay. Induction of ICD in BCSCs vs. non-BCSCs in response to different in vitro treatments was determined by assessing cell apoptosis and a panel of damage-associated molecular pattern molecules (DAMPs) by flow and enzyme-linked immunosorbent assay (ELISA).

Results: We found that ionizing radiation (IR) triggered a lower level of ICD in BCSCs than non-BCSCs. We then investigated the ability of disulfiram/cooper (DSF/Cu) which is known to preferentially induce cancer stem cells (CSCs) apoptosis to enhance IR-induced ICD of BCSCs. The results indicate that DSF/Cu induced a similar extent of IDC in both BCSCs and non-BCSCs and rendered IR-resistant BCSCs as sensitive as non-BCSCs to IR-induced ICD. IR and DSF/Cu induced ICD of BCSCs could be partly reversed by pre-treatment of BCSCs with a reactive oxygen species (ROS) scavenger and XBP1s inhibitors.

Conclusion: DSF/Cu rendered IR-resistant BCSCs as sensitive as non-BCSCs to IR-induced ICD. Our data demonstrate the potential of IR and DSF/Cu to induce ICD in BCSCs and non-BCSCs leading to robust immune responses against not only differentiated/differentiating breast cancer cells but also BCSCs, the root cause of cancer formation, progression and metastasis.
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http://dx.doi.org/10.1186/s12964-019-0507-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7057578PMC
March 2020

lncRNA CISAL Inhibits BRCA1 Transcription by Forming a Tertiary Structure at Its Promoter.

iScience 2020 Feb 11;23(2):100835. Epub 2020 Jan 11.

Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation of Sun Yat-Sen Memorial Hospital, Guangzhou 510120, China; Department of Oral and Maxillofacial Surgery, Sun Yat-Sen Memorial Hospital of Sun Yat-Sen University, Guangzhou 510120, China. Electronic address:

Cisplatin-based neoadjuvant chemotherapy has been shown to improve survival in patients with squamous cell carcinoma (SCC), but clinical biomarkers to predict chemosensitivity remain elusive. Here, we show the long noncoding RNA (lncRNA) LINC01011, which we termed cisplatin-sensitivity-associated lncRNA (CISAL), controls mitochondrial fission and cisplatin sensitivity by inhibiting BRCA1 transcription in tongue SCC (TSCC) models. Mechanistically, we found CISAL directly binds the BRCA1 promoter and forms an RNA-DNA triplex structure, sequestering BRCA1 transcription factor-GABPA away from the downstream regulatory binding region. Importantly, the clinical relevance of these findings is suggested by the significant association of CISAL and BRCA1 expression levels in TSCC tumors with neoadjuvant chemosensitivity and overall survival. We propose a new model where lncRNAs are tethered at gene promoter by RNA-DNA triplex formation, spatially sequestering transcription factors away from DNA-binding sites. Our study uncovers the potential of CISAL-BRCA1 signaling as a potential target to predict or improve chemosensitivity.
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http://dx.doi.org/10.1016/j.isci.2020.100835DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7033639PMC
February 2020

Melanoma cell-derived exosomes in plasma of melanoma patients suppress functions of immune effector cells.

Sci Rep 2020 01 9;10(1):92. Epub 2020 Jan 9.

Department of Pathology, University of Pittsburgh School of Medicine and UPMC Hillman Cancer Center, Pittsburgh, PA, 15213, USA.

Melanoma patients' plasma contains exosomes produced by malignant and normal cells. Plasma exosomes were isolated and separated by immunocapture into two fractions: melanoma cell-derived exosomes (MTEX) and normal cell-derived exosomes (non-MTEX). Immunosuppressive effects of MTEX on primary human immune cells were evaluated. Exosomes were isolated from plasma of 12 melanoma patients and six healthy donors (HDs). Expression levels of 19 immunoregulatory proteins in MTEX, non-MTEX and HDs exosomes were evaluated by on-bead flow cytometry. Functional/phenotypic changes induced in CD8 T or natural killer (NK) cells by MTEX or non-MTEX were compared. Plasma protein levels were higher in patients than HDs (P < 0.0009). In patients, MTEX accounted for 23-66% of total exosomes. MTEX were enriched in immunosuppressive proteins (P = 0.03). MTEX, but not HDs exosomes, inhibited CD69 expression (P ≤ 0.0008), induced apoptosis (P ≤ 0.0009) and suppressed proliferation (P ≤ 0.002) in CD8 T cells and downregulated NKG2D expression in NK cells (P = 0.001). Non-MTEX were enriched in immunostimulatory proteins (P = 0.002) and were only weakly immunosuppressive. Elevated MTEX/total exosome ratios and, surprisingly, non-MTEX ability to induce apoptosis of CD8 T cells emerged as positive correlates of disease stage. MTEX emerge as the major mechanism of tumor-induced immune suppression and as an underestimated barrier to successful melanoma immunotherapy.
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http://dx.doi.org/10.1038/s41598-019-56542-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6952363PMC
January 2020