Overexpression Of CD44 Research Articles

CD109 facilitates progression and 5-fluorouracil resistance of nasopharyngeal carcinoma

CD109 involves in human cancers, but its role and mechanism in the chemosensitivity of nasopharyngeal carcinoma (NPC) cells is yet to be investigated. Our investigation discovered that the CD109 had abnormally higher expression in the NPC tissues and cells. By depleting or supplementing CD109 in the NPC cells, the aggressive capabilities of NPC cells were suppressed or augmented, respectively. Moreover, silencing of CD109 promoted NPC cells chemosensitivity to 5-FU, and also inhibited 5-FU-mediated cell biological behaviors, facilitating cell apoptosis. Furthermore, CD109 was found to regulate the activities of AKT/mTOR signaling in the NPC cells. The tumor-promotive effect of CD109 overexpression was partially annulled by AKT/mTOR signaling inhibition. Finally, we also observed that tumor growth was retarded in the xenograft mice model and activity of AKT/mTOR signaling was reduced in the CD109 deficient group. Our data indicated that CD109, could be a therapeutic biomarker candidate in NPC treatment.

CD147 mediates epidermal malignant transformation through the RSK2/AP-1 pathway

BackgroundMalignant transformation of the epidermis is an essential process in the pathogenesis of cutaneous squamous-cell carcinoma (cSCC). Although evidence has demonstrated that CD147 plays key roles in various tumors, the role of CD147 in epidermal malignant transformation in vivo remains unclear.MethodsEpidermal CD147-overexpression or knockout (EpiCD147-OE or EpiCD147-KO) transgenic mouse models were generated for in vivo study. RNA-sequencing and q-PCR were performed to identify the differentially expressed genes. Immunohistochemistry and flow cytometry were performed to investigate the role of CD147 in regulating myeloid-derived suppressor cells (MDSCs). Immunoprecipitation, EMSA and ChIP assays were performed to investigate the mechanism of CD147 in cell transformation.ResultsWe found that specific overexpression of CD147 in the epidermis (EpiCD147-OE) induces spontaneous tumor formation; moreover, a set of chemokines and cytokines including CXCL1, which play essential function in MDSC recruitment, were significantly upregulated in EpiCD147-OE transgenic mice. As expected, overexpression of CD147 in the epidermis remarkably facilitated tumorigenesis by increasing the rate of tumor initiation and the number and size of tumors in the DMBA/TPA mouse model. Interestingly, the expression of CXCL1 and the infiltration of MDSCs were dramatically increased in EpiCD147-OE transgenic mice. Our findings also showed that knockdown of CD147 attenuated EGF-induced malignant transformation as well as CXCL1 expression in HaCaT cells. Consistently, CD147 was found overexpressed in cutaneous squamous cell carcinoma (cSCC), and positively related with the expression of CD33, a myeloid-associated marker. We further identified RSK2, a serine/threonine kinase, as an interacting partner of CD147 at the binding site of CD147D207-230. The interaction of CD147 and RSK2 activated RSK2, thus enhancing AP-1 transcriptional activation. Furthermore, EMSAs and ChIP assays showed that AP-1 could associate with the CXCL1 promoter. Importantly, RSK2 inhibitor suppressed the tumor growth in DMBA/TPA mouse model by inhibiting the recruitment of MDSCs.ConclusionOur findings demonstrate that CD147 exerts a key function in epidermal malignant transformation in vivo by activating keratinocytes and recruiting MDSCs via the RSK2/AP-1 pathway.

Ezrin and CD44 participate in the internalization process of Coxiella burnetii into non-phagocytic cells.

Ezrin protein is involved in the interaction of actin cytoskeleton with membrane receptors such as CD44. It regulates plasma membrane dynamics and intracellular signaling. Coxiella burnetii, the etiologic agent of Q fever, is internalized into host cell through a poorly characterized molecular mechanism. Here we analyzed the role of ezrin and CD44 in the C. burnetii internalization by HeLa cells. The knockdown of ezrin and CD44 inhibited the bacterial uptake. Interestingly, at early stages of C. burnetii internalization, ezrin was recruited to the cell membrane fraction and phosphorylated. Moreover, the overexpression of non-phosphorylatable and phosphomimetic ezrin mutants decreased and increased the bacterial entry, respectively. A decrease in the internalization of C. burnetii was observed by the overexpression of CD44 truncated forms containing the intracellular or the extracellular domains. Interestingly, the CD44 mutant was unable to interact with ERM proteins decreased the bacterial internalization. These findings demonstrate the participation of ezrin in the internalization process of C. burnetii in non-phagocytic cells. Additionally, we present evidence that CD44 receptor would be involved in that process.

Hyaluronic acid-coated ultrasmall BiOI nanoparticles as a potentially targeted contrast agent for X-ray computed tomography

Commercial X-ray computed tomography (CT) contrast agents (CAs) are not appropriate for multicolor CT imaging and are limited due to a lack of tumor targeting. In this contribution, to favor the combination of high Z elements like bismuth and iodine for a broad range of X-ray photon energy, BiOI nanoparticles (NPs) are synthesized with hyaluronic acid (HA) coating to target the tumor cells with CD44 overexpression. The crystal structure of BiOI NPs is determined by X-ray powder diffraction, and the size of NPs is determined by a transmission electron microscope at about 14 ± 3 nm. The dynamic diameter of the NPs (DLS) in PBS buffer was measured at about 100 nm. Fourier transform infrared spectroscopy and thermal gravimetric analysis confirm the coating of BiOI NPs with HA. The stability of the NPs was also investigated via UV–vis spectroscopy. The immunocytochemistry process is performed to investigate the targeting ability of synthesized NPs on the human colon cancer cells with CD44 antigen. Finally, the X-ray attenuation of prepared HA-coated BiOI NPs is higher than Iohexol as the commercially available iodinated contrasting agent at the same concentrations, which can be administrated at much lower doses to achieve similar contrast to Iohexol.

CD147 Targeting by AC-73 Induces Autophagy and Reduces Intestinal Fibrosis Associated with TNBS Chronic Colitis.

Intestinal fibrosis is a common complication of inflammatory bowel diseases. Medical treatment of intestinal fibrosis is an unmet therapeutic need. CD147 overexpression can induce myofibroblast differentiation associated with extracellular matrix deposition, favouring the development of fibrosis. To understand whether CD147 may promote intestinal fibrosis, we analysed its expression and blocked its function by using its specific inhibitor AC-73 [3-{2-[([1,1'-biphenyl]-4-ylmethyl) amino]-1-hydroxyethyl} phenol] in the murine TNBS [trinitrobenzenesulfonic acid]-chronic colitis model associated with intestinal fibrosis. TNBS chronic colitis was induced by weekly intrarectal administration of escalating doses of TNBS. Ethanol-treated and untreated mice were used as controls. Separated groups of TNBS, ethanol-treated or untreated mice received AC-73 or vehicle administered intraperitoneally from day 21 to day 49. At day 49, mice were killed, and colons collected for histological analysis, protein and RNA extraction. CD147, α-SMA and activated TGF-β1 protein levels, CD147/ERK/STAT3 signalling pathway and autophagy were assessed by Western blot, collagen and inflammatory/fibrogenic cytokines mRNA tissue content by quantitative PCR. In mice with chronic TNBS colitis, CD147 protein level increased during fibrosis development in colonic tissue, as compared to control mice. CD147 inhibition by AC-73 treatment reduced intestinal fibrosis, collagen and cytokine mRNA tissue content, without significant modulation of activated TGF-β1 protein tissue content. AC-73 inhibited CD147/ERK1/2 and STAT3 signalling pathway activation and induced autophagy. CD147 is a potential new target for controlling intestinal fibrosis and its inhibitor, AC-73, might represent a potential new anti-fibrotic therapeutic option in IBD.

Tristetraprolin regulates phagocytosis through interaction with CD47 in head and neck cancer.

CD47 is expressed in all human cancer cells, including head and neck cancer, and initiates a signaling cascade to inhibit macrophage phagocytosis. However, the mechanism underlying CD47 overexpression has not been elucidated in radioresistant head and neck cancer. The present study demonstrated that decreased Tristetraprolin (TTP) expression induced a sustained overexpression of CD47 using reverse transcription-quantitative PCR and western blotting, and that CD47 overexpression prevented phagocytosis using a phagocytosis assay in a radioresistant HN31R cell line. Subsequently, using TTP transfection, RNA interference, duel-luciferase assay and EMSA, it was revealed that TTP transfection enhanced phagocytosis through degradation of CD47 mRNA by directly binding to CD47 AREs within the CD47 3'UTR. Based on our previous study, methylation-specific PCR and western blotting revealed that DNMT1 was overexpressed in radioresistant HN31R cell line and TTP expression was decreased epigenetically by DMNT1 associated DNA methylation. Overall, these findings provided novel insight into the role of TTP as a biomarker of CD47-positive head and neck cancer patients.

Abstract 6280: Preclinical development of a bispecific antibody-trap selectively targeting CD47 and HER2 for the treatment of breast as well as gastric cancer

Abstract Trastuzumab executes anti-tumor efficacy mainly via antibody-dependent cell cytotoxicity (ADCC). However due to the fact that majority of the populations including patients with cancers carry a CD16A-158F allele which has low binding affinity to the Fc fragment of trastuzumab, response to trastuzumab treatment is hindered. Furthermore, overexpression of CD47 on cancer cells prevent macrophages from activation via Fc-FcγRIIa interaction due to the so called “Don’t eat me” signal. We therefore designed a bispecific mAb-trap designated as IMM2902. Extensive in vitro as well as in vivo characterization demonstrated that IMM2902 binds to both CD47 and HER2 with high affinity (KD = 5.8x10-10M for HER2; KD = 8.9x10-9M for CD47) and has stronger ADCC activity against NCI-N87 (EC50 = 0.054nM for IMM2902, v.s. 0.868nM for trastuzumab). In vivo efficacy studies in several xenograft tumor models revealed a robust anti-tumor activity as reflected by the complete elimination of established tumors in BT-474 breast cancer and NCI-N87 gastric xenograft models even at the dose as low as 3.5mg/kg. Interestingly, even in Herceptin-resistant breast tumor model, when administered at a dose of 10mg/kg, the established HCC-1954 tumors were also eliminated. Immunohistochemistry staining of the tumor tissues demonstrated an accelerated HER2 degradation which is highly correlated with tumor volume. IMM2902 has no impact on hemagglutination, nor does it have significant hemotoxicity following single as well as multiple administrations in non-human primate animals at different dosage. Our study suggests that IMM2902 has the potential to be an alternative promising treatment option for patients with her2-positive cancers refractory to trastuzumab treatment. Citation Format: Wenzhi Tian, Song Li, Dianze Chen, Dandan Liu, Huiqin Guo, Chunmei Yang, Li Zhang, Wei Zhang, Xiaoping Tu, Liang Peng, Gui Zhao, Ruliang Zhang, Fan Zhang. Preclinical development of a bispecific antibody-trap selectively targeting CD47 and HER2 for the treatment of breast as well as gastric cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6280.

Abstract 5196: CD47 Blockade potentiates immunotherapy of durvalumab against cutaneous T cell lymphoma

Abstract Immune checkpoint (IC) blockade for programmed cell death 1 (PD1) and PD-Ligand 1 (PD-L1) has shown promising and durable therapeutic outcomes in cutaneous T-cell lymphoma (CTCL), but resistance and/or relapses are common. While research focused on improving adaptive immune functions in CTCL, we investigated the role of 1) the “do not eat me signal” (CD47), and PD-L1, as a dual (innate and adaptive IC)-targeting strategy in CTCL. RNA sequencing and flow cytometric analysis showed that CTCL tumor cells and CTCL cell lines (MyLa, Hut78, HH, and H9 cells) overexpressed CD47 and PD-L1 compared with healthy control, and CD47 positively correlated with PD-L1 in CTCL patients. Overexpression of CD47 and PD-L1 was induced by high MYC expression in CTCL tumor cells lines. In addition, the signal-regulatory protein (SIRP)α receptor for CD47 and PD-L1 were significantly more abundant on macrophages, dendritic cells and natural killer cells in CTCL patients than healthy control by flow cytometric analysis. Furthermore, the RNA sequencing data indicated that the M2 macrophages, immature dendritic cells, and inhibitory receptors expressed natural killer cells in CTCL patients were higher than in healthy control. Notably, TTI-621 (SIRPαFc) treatment decreased M2 macrophage, immature dendritic cells, and inhibitory receptors expressed natural killer cells in CTCL patients at the end of the treatment, compared to baseline. In vitro, TTI-621 treatment increased the macrophage phagocytic activity compared to untreated control. Moreover, TTI-621 synergized with an anti-PD-L1 antibody (durvalumab) to reprogram M2-like TAMs, induced by MyLa supernatant, to M1-like phenotypes. The simultaneous blockade of both CD47 and PD-L1 inhibited growth of CTCL cell lines more potently than each single antibody alone or blank control in vitro. Enhanced CD8+ T cell mediated killing was detected using a chromium-release assay following dual blockade of in CTCL cell lines, suggesting anti-CD47 and anti-PD-L1 may synergistically facilitate elimination of CTCL tumor cells through specific pathways. RNA-sequencing analysis indicated that these effects were mediated by cell death related pathways such as apoptosis, autophagy, and necroptosis. Collectively, our findings demonstrated that CD47 and PD-L1 are critical regulators of innate and adaptive immune surveillance in CTCL and that dual targeting of CD47 and PD-L1 will provide insight into tumor immunotherapy to improve tumor control in CTCL. Citation Format: Zhen Han, Mingye Feng, Xiwei Wu, Chingyu Su, Yate-Ching Yuan, Hanjun Qin, James F. Sanchez, Jasmine Zain, Steven T. Rosen, Christiane Querfeld. CD47 Blockade potentiates immunotherapy of durvalumab against cutaneous T cell lymphoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5196.

Abstract CT503: A phase I/Ib study of the safety and preliminary efficacy of NZV930 alone and in combination with spartalizumab and/or taminadenant in patients (pts) with advanced malignancies

Abstract Background: The over-expression of CD73 has been linked to many cancers. NZV930, an investigational, CD73-targeting monoclonal antibody (mAb), forms bidentate structures with the CD73 dimer, shutting down the catalytic activity of cell surface and soluble CD73. Taminadenant is an investigational A2AR antagonist and spartalizumab is an anti-PD-1 mAb. Preclinical studies provide compelling antitumor activity when combining CD73, A2AR, and PD-1 blockade. Methods: This first-in-human study (NCT03549000) comprised dose-escalation and dose-expansion parts. Here, we present the dose-escalation results of the four treatment (tx) arms: NZV930 single agent (SA; 60-1000 mg Q2W), NZV930 (200-600 mg Q2W) + spartalizumab (400 mg Q4W) or taminadenant (80-240 mg Q2W), or in combination with spartalizumab (400 mg Q4W) + taminadenant (80-240 mg BID). Eligible pts had selected malignancies, and disease progression following standard therapy. Primary objectives included assessing safety and tolerability and determining the recommended dose for expansion (RDE) of NZV930 SA and its combinations. Secondary objectives were to determine efficacy, characterize the pharmacokinetics (PKs) of NZV930 SA and its combinations, and assess the immunogenicity of spartalizumab and NZV930. Biomarker tumor samples were obtained at screening, C2D1, and C4D1. Results: As of June 10, 2021, 105 pts were enrolled for dose escalation, including pts with CRC (n=63), ovarian cancer (n=15), pancreatic cancer (n=11), NSCLC (n=8), TNBC (n=5), RCC (n=2), and prostate cancer (n=1). Most pts (81%) had been treated with ≥3 lines of prior therapy. In total, 98 pts discontinued, mainly due to disease progression (72%); six were ongoing. At least one tx-related adverse event (TRAE) was reported by 89 pts (85%); these were mostly grade 1-2. A triad of headache, fever, and nausea/emesis was observed after the first infusion of NZV930. Most common TRAEs (≥30%) were headache (67%), nausea and vomiting (32% each), and pyrexia (30%). At least one grade ≥3 TRAE was reported in 15 pts (14%); headache (4%) was most common, accounting for 4/7 DLTs (all grade ≥3). There were no tx-related deaths. Overall, no responses were reported; 12 pts (11%) had stable disease and 69 pts (66%) had progressive disease. NZV930 has good PK properties, with target-mediated drug disposition at lower doses, which normalize to typical IgG-like PKs at higher doses. Biomarker data confirm CD73 target engagement in plasma. Adenosine pathway modulation in tumors and plasma was seen post-NZV930 tx. Conclusions: NZV930 reduced adenosine levels in plasma and tumors. The RDE of NZV930 SA or in combination with spartalizumab and/or taminadenant is 600 mg Q2W with a step-up dosing regimen. NZV930 SA and its combinations exhibited a tolerable safety profile in multiple advanced solid tumors and showed good PK properties. Citation Format: Siqing Fu, Udai Banerji, Philippe L. Bedard, Aitana Calvo Ferrándiz, Alberto Chiappori, Jayesh Desai, Rahima Jamal, Desamparados Roda Perez, Noboru Yamamoto, Erica Vieira, Gabrielle Wong, Kulandayan K. Subramanian, Shunguang Wang, Britta Mueller, Javier A. Otero, Si-Lin Koo. A phase I/Ib study of the safety and preliminary efficacy of NZV930 alone and in combination with spartalizumab and/or taminadenant in patients (pts) with advanced malignancies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr CT503.

Abstract 907: CD133 stimulates cell proliferationviaupregulation of amphiregulin in melanoma

Abstract Melanoma, a malignant transformation of melanocytes, accounts for 90% of skin cancer deaths due to its ability to efficiently metastasize to other organs. CD133, also known as Prominin1, is a pentaspan glycosylated transmembrane protein that is overexpressed in cancer stem cells from various tumors, including melanoma, and has been associated with tumor recurrence, chemoresistance, and metastasis. To characterize and target CD133-positive cancer stem cells, patient-derived melanoma cell lines BAKP and POT, harboring BRAFWT/NRASQ61K and BRAFWT/NRASQ61R driver mutations, respectively, were co-transduced with a Tet activator (rtTA3) and a Tet-on vector expressing CD133 (TRE3G-CD133) to produce stable doxycycline (Dox)-inducible cell lines. These cell lines were exposed to Dox (1µg/mL) to induce CD133 expression for 24 h; mRNA was extracted and subjected to RNA-seq analysis, followed by bioinformatic analyses, which revealed genes that are significantly up- or down-regulated by Dox-induced CD133 expression. Among the most significantly altered genes was amphiregulin (AREG), which increased 7-fold with Dox induction. These results were validated in both BAKP and POT cell lines by qPCR and immunoblot analysis. AREG binds EGFR and activates RAS/RAF/MEK/ERK and PI3K/AKT signaling pathways, leading to cell proliferation, survival, and migration. Consistently, cell cycle and cell growth analyses revealed that Dox-induced CD133 expression increases the percent of cells in S-phase of the cell cycle, leading to increased cell growth in BAKP cells. The role of CD133 in cell proliferation via AREG upregulation was further studied with immunofluorescent and flow cytometry analysis of proliferation markers PCNA, Ki67, as well as BrdU incorporation into newly synthesized DNA in Dox-induced BAKP cells, with or without siRNA knockdown of AREG. AREG may play an integral role in melanoma progression due to its vital function in cell proliferation, serving as an attractive target for novel therapeutics in melanoma and cancers exhibiting overexpression of both CD133 and AREG. Further studies using specific AREG inhibitors or RNAi knockdown in melanoma cell lines is currently ongoing, and can further elucidate molecular mechanisms highlighting the role(s) of CD133 and AREG in melanoma cancer stem cells. Citation Format: Cynthia M. Simbulan-Rosenthal, Nusrat Islam, Yoga Haribabu, Ryyan Alobaidi, Li-Wei Kuo, Azadeh Shalamzari, Adam Leibowitz, Joanna Moulton, Dean S. Rosenthal. CD133 stimulates cell proliferationviaupregulation of amphiregulin in melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 907.

Amelioration of experimental tendinopathy by lentiviral CD44 gene therapy targeting senescence-associated secretory phenotypes

CD44 exerts anti-senescence effects in many disease models. We examined senescence in tendinopathy and the effect of CD44 on senescence-associated secretory phenotypes (SASPs). Senescent markers were determined in human tendinopathic long head of bicep (LHB) and normal hamstring tendons. CD44 gene transfer in rat tendinopathic tenocytes stimulated with interleukin (IL)-1β and a rat Achilles tendinopathy model were performed using lentiviral vectors. Expression levels of p53, p21, and p16 and senescence-associated β-galactosidase (SA-β-gal) activity were positively correlated with the severity of human tendinopathy and were higher in rat and human tendinopathic tenocytes than in normal controls. CD44 overexpressed tenocyte transfectants exhibited reduced levels of IL-6, matrix metalloproteinases (MMPs), cyclooxygenase (COX)-2, p53, p21, p16, SA-β-gal, and phospho-nuclear factor (NF)-κB, whereas their collagen type I alpha 1 (COL1A1) and tenomodulin (tnmd) levels were increased when compared with control transfectants under IL-1β-stimulated conditions. In the animal model, CD44 overexpression lowered the ultrasound and histology scores and expression levels of the senescent and SASP markers COX-2 and phospho-NF-κB. Bromodeoxyuridine (BrdU)- and tnmd-positive cell numbers were increased in the LVCD44-transduced tendinopathic tendons. Senescence is positively correlated with tendinopathic severity, and CD44 overexpression may protect the tendinopathic tendons from SASPs via anti-inflammation and maintenance of extracellular matrix homeostasis.

IMMU-21. Targeting the adenosinergic immune suppression pathway in high grade glioma synergizes with innate immune checkpoint blockade

Abstract Immune response in the tumor microenvironment is modulated by the conversion of eATP to AMP to adenosine via the ecto-enzymes CD39 and CD73. Overexpression of CD73 in tumors leads to an increase in extracellular adenosine concentration and decrease in eATP, resulting in immune suppression. In this study, CD73 knockouts were created in adult and pediatric glioma cell lines (DIPG17, T3691, BT245) and used to quantify the effects of CD73 knockout on macrophage phagocytosis in the presence of extracellular AMP. In cell line DIPG17, CD73 knockout significantly increased phagocytosis with and without external AMP compared to the WT. In the WT condition, addition of AMP significantly reduced phagocytosis, while this decrease was not significant in the CD73-KO. The addition of anti-CD47 antibody significantly increased phagocytosis with external AMP in the WT condition. Similar results were obtained with cell line T3691. However, CD73 knockout combined with anti-CD47 treatment and external AMP showed significantly greater phagocytosis than the WT condition. This suggests that a combination of anti-CD73 and anti-CD47 treatment may be more effective in the tumor microenvironment. Lastly, in cell line BT245, CD73 knockout significantly increased phagocytosis as seen in the other cell lines. However, no significant difference was observed between WT and CD73-KO with extracellular AMP or anti-CD47. In vivo studies were conducted in orthotopic xenograft mouse models with DIPG17 CD73 knockout cells. Combination treatment with anti-CD47 antibody significantly decreased tumor burden and prolonged survival in the CD73-KO tumors compared to anti-CD47 treated DIPG13-WT tumors. We conclude that the CD73 adenosinergic pathway and the CD47-SIRPα pathway may present a target for immunotherapy in pediatric and adult gliomas.

Lemzoparlimab (lemzo) with venetoclax (ven) and/or azacitidine (aza) in patients (pts) with acute myeloid leukemia (AML) or myelodysplastic syndromes (MDS): A phase 1b dose escalation study.

TPS7067 Background: Despite treatment advances, pts with AML or higher-risk MDS who are ineligible for standard intensive treatments still have poor survival, highlighting the need for novel therapies. Overexpression of CD47 is common in leukemic stem cells and AML blasts and correlates with poor clinical outcomes. Lemzo is an anti-CD47 antibody with red blood cell–sparing properties. Treatment with ven plus aza has shown favorable safety and efficacy in older/unfit pts with AML and higher-risk MDS. Blocking CD47 is hypothesized to hypersensitize AML cells to the antitumor activity of ven and aza. This study will evaluate the safety and dose-limiting toxicities (DLTs) of lemzo with ven + aza for pts with treatment-naïve AML, as well as lemzo with aza ± ven for treatment-naïve higher-risk MDS. Methods: This phase 1b, open-label, dose-escalation study (NCT04912063) is enrolling adults with: (1) treatment-naïve AML with adverse cytogenetic/molecular risk not suitable for induction therapy, with an Eastern Cooperative Oncology Group Performance Status (ECOG-PS) 0–2 (aged ≥75 years) or 0–3 (aged ≥18–74 years) excluding acute promyelocytic leukemia; or (2) treatment-naïve higher-risk MDS (Revised International Prognostic Scoring Score > 3) with < 20% bone marrow blasts, ECOG-PS 0–2, and no immediately planned stem cell transplant. For each 28-day cycle, aza is administered subcutaneously or intravenously (IV) daily for 7 days within the first 9 days (7-0-0 or 5-2-2 schedule); ven is administered orally daily on days 1–28 (AML, following dose ramp-up) or days 1–14 (ven-containing MDS cohorts). Lemzo is administered IV at a schedule that is to be determined in this study. Dose escalation has Bayesian optimal interval design and may be expanded to investigate alternate dosing for lemzo. Dose expansion will initiate at recommended phase 2 dose. Treatment discontinuation criteria are unacceptable toxicity, progressive disease, lack of partial/complete remission or clinical benefit within 6 cycles, or at physician’s discretion. Pts who discontinue study treatment without progression will continue with posttreatment follow-up; pts who progress will enter survival follow-up. The primary endpoints are DLTs of lemzo. Secondary endpoints for both cohorts include best overall responses of complete remission, duration of response, event-free survival, and overall survival. Exploratory biomarker endpoints are included. Safety assessments include adverse event (AE, graded per National Cancer Institute Common Terminology Criteria for AEs v5.0) monitoring, physical examinations, vital signs, electrocardiograms, and laboratory tests. Response rates will be analyzed with estimates and 95% confidence intervals based on exact binomial distribution. Time-to-event endpoints will be analyzed using Kaplan–Meier methodology. Clinical trial information: NCT04912063.

TTI-622-01: A phase 1a/1b dose-escalation and expansion trial of TTI-622 in patients with advanced hematologic malignancies, including diffuse large B-cell lymphoma (DLBCL).

TPS7595 Background: CD47 is an innate immune checkpoint that binds signal regulatory protein alpha (SIRPα) and delivers a "don’t eat me" signal to suppress macrophage phagocytosis. Overexpression of CD47 on cancer cells serves as a mechanism of immune surveillance evasion, and is associated with poor prognosis in both hematologic and solid malignancies. TTI-622 is a fusion protein consisting of the CD47-binding domain of human SIRPα linked to the Fc region of human IgG4. It is designed to enhance phagocytosis and antitumor activity by preventing CD47 from delivering its inhibitory signal as well as generating a moderate pro-phagocytic signal via IgG4 Fc. Importantly, unlike many CD47-blocking agents, TTI-622 does not bind to human red blood cells. Preclinical studies demonstrate that TTI-622 induces macrophage-mediated phagocytosis of different malignant cell lines, including DLBCL cells, decreases tumor growth and improves survival in a DLBCL xenograft tumor model. Anti-CD47 antibody enhances rituximab stimulated macrophage-mediated phagocytosis of non-GCB DLBCL cell lines (Bouwstra et al, Cancer Immunol Res. 2019). The ongoing phase 1a part of this study has been previously described. Here we describe 2 cohorts within the phase 1b part of the study that are intended to determine the safety and preliminary efficacy of TTI-622 when given in combination with anti-CD20 targeting agent in patients with CD20+ relapsed/refractory (RR) DLBCL. Methods: TTI-622-01 is a multi-center Phase 1a/1b study. Phase 1a was designed to determine the MTD, pharmacokinetics (PK), pharmacodynamics, and preliminary antitumor activity of QW, Q2W, and Q3W single-agent TTI-622 in R/R lymphoma using a 3+3 dose escalation schema. Phase 1b, ongoing, will determine the safety, recommended dose and preliminary efficacy of TTI-622 in combination with select approved anticancer treatments for patients with hematological malignancies including, but not limited to anti-CD20 therapy in patients with CD20+ RR DLBCL. Secondary objectives are to further characterize safety, PK and immunogenicity of TTI-622 when combined with approved therapies. Patients will be enrolled in 2 cohorts exploring different doses of TTI-622 in combination with anti-CD20 therapy. Cohorts will open in a staggered manner. In each cohort 3 patients will be dosed and followed for 28 days before expanding enrolment to additional 27 patients per cohort. Key eligibility criteria include: age ≥18 years; relapsed and/or refractory disease after ≥1 prior line of therapy; not eligible for or have progressed after high dose chemotherapy (HDT)/auto-SCT; ≥1 site of measurable disease (Lugano 2014 classification); ECOG PS ≤2; adequate organ functions, no known CNS involvement; no prior anti-CD47 or anti-SIRPα therapy. Patient recruitment is planned or ongoing at 40 sites worldwide. Clinical trial information: NCT03530683.

CD44 potentiates hepatocellular carcinoma migration and extrahepatic metastases via the AKT/ERK signaling CXCR4 axis.

BackgroundCell adhesion molecule cluster of differentiation 44 (CD44) plays a significant role in cancer cell local invasion, intravasation, migration, and the establishment of metastatic lesions. However, little is known about the underlying mechanism of how CD44 regulates hepatocellular carcinoma (HCC) extrahepatic metastasis (EHM).MethodsThe expression of CD44 in HCC tissues and cell lines was detected through western blot and immunohistochemistry (IHC). Through gain- and loss-of-function assays, we examined the oncogenic roles of CD44 in regulating HCC cell growth and metastasis in vitro and in vivo. To identify the potential mechanism, we employed quantitative real-time polymerase chain reaction, and western blot.ResultsIn this study, CD44 was highly expressed in HCC cells and HCC-patient specimens that exhibited high malignancy potential. The overall survival (OS) was worse and the cumulative recurrence rate was higher in HCC patients with CD44 overexpression than those with low levels of CD44 expression. Our in-vitro and in-vivo experiments showed that CD44 downregulation reduced HCC cell colony formation, migration, and invasion, and HCC tumor growth and metastasis, and that the pro-metastasis effect of CD44 was mediated by the protein kinase B (AKT)/extracellular signal-regulated kinase (ERK) signaling-chemokine receptor C-X-C chemokine receptor type 4 (CXCR4) axis. The reported capacity of CD44 to induce CXCR4 expression and increase the propensity of tumors to invade and metastasize to distant organs is consistent with the aggressive clinical characteristics of HCCs.ConclusionsCD44 could represent a future therapeutic target for EHM.

EHD1 promotes the cancer stem cell (CSC)-like traits of glioma cells via interacting with CD44 and suppressing CD44 degradation.

Plenty of evidence has shown that endocytosis plays a key role in cancer progression; however, its effects in the progression of cancer stem cells (CSCs) are still fragmentary. In the present study, we firstly identified that mammalian Eps15 homology domain protein 1 (EHD1), an endocytic and metastasis-associated gene, was upregulated in the 3D non-adherent spheres derived from glioma cells compared to that in the corresponding parental cells. Further functional experiments revealed that EHD1 knockdown reduced the CSC-like traits of glioma cells, which were evident by the decrease of sphere-formation ability, ALDH1 activity, and CSC markers' expression. Additionally, EHD1 knockdown attenuated the tumor-initiating ability of glioma cells in vivo. Furthermore, it was shown that EHD1 bound to CD44, enhanced CD44 stability, and prevented its total ubiquitination. Indeed, overexpression of CD44 rescued the inhibitory effects of EHD1 knockdown on the CSC-like traits of glioma cells. Finally, through the online dataset analysis, we found that EHD1 indeed exhibited a higher level in glioma tissues relative to that in normal tissues, and a positive correlation with CSC markers' expression in glioma tissues. Notably, EHD1 expression was negatively correlated with the overall survival and relapse-free survival of glioma patients. Thus, this work indicates that EHD1 might be a potent target for glioma progression, especially through breaking the EHD1-CD44 interaction.

Functional expression of CD73 on human natural killer cells.

The production of adenosine by CD73 on cancer cells in the tumor microenvironment is a recognized immunosuppressive mechanism contributing to immune evasion in many solid tumors. While NK cells have been purported to overexpress CD73 under certain conditions, this phenomenon has remained elusive and unclear. We have found that while NK cells are able to upregulate expression of CD73 on their surface when exposed to CD73+ cancer cells, this upregulation is not universal, nor is it often substantial. Rather, our data point to the extent of CD73 expression on NK cells to be both cancer-specific and environmentally-driven, and largely limited in intensity. We found that NK cell overexpression of CD73 responds to the level of CD73 on cancer cells and is enhanced in hypoxia. Interestingly, human CD73+ NK cells appear hyperfunctional in vitro compared to CD73- NK cells, suggesting that CD73 expression could be a bystander of NK cell activation. In addition, glioblastoma patient data show that tumor-infiltrating NK cells express CD73 variably, depending on donor, and present lower expression of CD16, alongside patient-specific changes in CEACAM1, CXCR3 and TIM-3, suggesting some functional changes in NK cell responses associated with expression of CD73 on NK cells in vivo. Taken together, our study is the first to show that while NK cells are largely resistant to the upregulation of CD73, CD73 expression is inducible on NK cells in response to CD73 on cancer cells, and these cells are associated with distinct functional signatures.

Sialylation of CD44 by ST8sia6 is required for recognition by the inhibitory receptor Siglec-7

Abstract Glycans decorate almost all proteins and lipids on the cell surface, and they play a critical role in cell-cell interactions. Terminal sialic acid addition to glycans helps the immune system distinguish between “self” and “non-self.” The enzyme ST8sia6 adds alpha-2,8 disialic acids to cell surface receptors, and this sialic acid addition promotes binding to the inhibitory receptor Siglec-E. We have previously shown ST8sia6 overexpression in tumors accelerates tumor growth in a Siglec-E dependent manner. Proteomic analysis in murine tumor lines identified CD44 as a target for ST8sia6. CD44 expression on tumors is associated with metastasis and chemoresistance. Since humans and mice have different sialic acid structures, we examined whether ST8sia6 makes CD44 a ligand to the ortholog of Siglec-E in human cells, Siglec-7. When HEK293T cells overexpress ST8sia6 and CD44 together, Siglec-7 binding was significantly increased compared to ST8sia6 or CD44 overexpression alone. Thus, ST8sia6 has a conserved role from mouse to human in decorating CD44 with sialic acid. Our study proposes that ST8sia6 sialylation of CD44 could suppress immune responses and enables cancer progression in human. Supported by grant: 2T32AI007425-23 to DJF and R01 CA243545-01S1 to V.S.S.

Biomarkers expression among paired serous ovarian cancer primary lesions and their peritoneal cavity metastases in treatment-naïve patients: A single-center study.

BackgroundHigh‐grade serous ovarian carcinoma (HGSOC), the most common histologic subtype of ovarian epithelial cancer, is associated with treatment resistance, enhanced recurrence rates, and poor prognosis. HGSOCs often metastasize to the peritoneal cavity, while fluid cytology examination could identify such metastases. This retrospective study aimed to identify potential biomarker discrepancies between paired HGSOC primary tissues and metastatic peritoneal fluid cytology samples, processed as cell blocks (CBs).MethodsTwenty‐four pairs of formalin‐fixed, paraffin‐embedded primary tissues and metastatic CBs from an equal number of treatment‐naïve patients were used, and immunohistochemistry (IHC) for epidermal growth factor receptor (EGFR), human epidermal growth factor receptor, programmed cell death‐1 ligand 1 (PD‐L1), and CD147 was applied.Results13/24 pairs showed discordant EGFR IHC results; in all these 13 patients, EGFR was positive (≥1+ membranous staining intensity found in at least 10% of the cancer cells) in the peritoneal, yet negative in the primary tissue samples. Notably, EGFR IHC was positive in 15/24 of the metastatic, whereas in just 2/24 of the primary HGSOC samples (p < 0.001). Although most PD‐L1 results were concordant, 5/24 and 6/24 pairs exhibited discordant results when stained with the E1L3N and 22C3 clones, respectively. Lastly, CD147 overexpression was found more often in the metastatic rather than the matched primary HGSOCs stained with CD147, though the difference was not significant.ConclusionsCytology from effusions could be considered for biomarker testing when present, even when tissue from the primary cancer is also available and adequately cellular, as it could provide additional information of potential clinical significance.

HSP90 promotes radioresistance of cervical cancer cells via reducing FBXO6-mediated CD147 polyubiquitination.

HSP90 inhibition might be a promising strategy to overcome the radioresistance of some cancers. In the current study, we further explored the mechanisms of HSP90 in regulating the radiosensitivity of cervical cancer cells. Bioinformatic analysis was performed based on data from TCGA‐CESC. Cellular and molecular studies were conducted using CaSki and SiHa and the derived radioresistant (RR) subclones. Through a proteomics screen, we identified HSP90 chaperones (both HSP90α and HSP90β) as CD147‐binding partners supporting its stabilization. Targeting HSP90 sensitized CaSki‐RR and SiHa‐RR cancer cells to irradiation partially through CD147 destabilization. Mechanistically, HSP90 interacts with FBXO6 and reduces FBXO6‐mediated proteasomal degradation of CD147. Enforced FBXO6 overexpression also sensitized CaSki‐RR and SiHa‐RR cancer cells to irradiation. These effects were enhanced using 17‐AAG treatment but were weakened by CD147 overexpression. Survival analysis further confirmed the association between high FBXO6 expression and favorable progression‐free survival among patients with cervical cancer. In conclusion, this study showed that HSP90 promotes radioresistance of cervical cancer cells partially via reducing FBXO6 mediated CD147 polyubiquitination. These findings help to explain why HSP90 inhibitor exerts radio‐sensitizing effects in cervical cancer.