Reduction In Tumor Volume Research Articles

Sustainable Nanomedicine: Enhancement of Asplatin’s Cytotoxicity In Vitro and In Vivo Using Green-Synthesized Zinc Oxide Nanoparticles Formed via Microwave-Assisted and Gambogic Acid-Mediated Processes

Chemoresistance encountered using conventional chemotherapy demands novel treatment approaches. Asplatin (Asp), a novel platinum (IV) prodrug designed to release cisplatin and aspirin in a reductive environment, has demonstrated high cytotoxicity at reduced drug resistance. Herein, we investigated the ability of green-synthesized nanocarriers to enhance Asp’s efficacy. Zinc oxide nanoparticles (ZnO-NPs) were synthesized using a green microwave-assisted method with the reducing and capping agent gambogic acid (GA). These nanoparticles were then loaded with Asp, yielding Asp@ZnO-NPs. Transmission electron microscopy was utilized to study the morphological features of ZnO-NPs. Cell viability studies conducted on MDA-MB-231 breast cancer cells demonstrated the ability of the Asp@ZnO-NPs treatment to significantly decrease Asp’s half-maximal inhibitory concentration (IC50) (5 ± 1 µg/mL). This was further demonstrated using flow cytometric analysis that revealed the capacity of Asp@ZnO-NPs treatment to significantly increase late apoptotic fractions. Furthermore, in vivo studies carried out using solid Ehrlich carcinoma-bearing mice showed significant tumor volume reduction with the Asp@ZnO-NPs treatment (156.3 ± 7.6 mm3), compared to Asp alone (202.3 ± 8.4 mm3) and untreated controls (342.6 ± 10.3 mm3). The histopathological analysis further demonstrated the increased necrosis in Asp@ZnO-NPs-treated group. This study revealed that Asp@ZnO-NPs, synthesized using an eco-friendly approach, significantly enhanced Asp’s anticancer activity, offering a sustainable solution for potent anticancer formulations.

NIMG-84. BELZUTIFAN REDUCTION OF CEREBRAL BLOOD FLOW IN VHL-ASSOCIATED HEMANGIOBLASTOMA PREDICTS RESPONSE TO TREATMENT

Abstract Hemangioblastoma (HMB) is a vascular-origin tumor occurring primarily in the cerebellum. Treatment typically involves surgical resection of the solid part of the tumor or stereotactic radiosurgery. An emerging treatment for hemangioblastoma is belzutifan (BZF), a hypoxia-inducible factor 2a (HIF2a) inhibitor. A recent trial found BZF to control hemangioblastoma in patients with von Hippel Lindau (vHL) syndrome. Perfusion MRI techniques including arterial spin labelling (ASL) and can be used to determine relative cerebral blood flow (rCBF). No prior studies have investigated rCBF in hemangioblastoma with relation to belzutifan, and the mechanism of action of belzutifan on hemangioblastoma is not well understood. Here, we aim to measure the response in rCBF in patients with vHL-associated hemangioblastoma treated with belzutifan. This retrospective cohort study constituted a group of seven patients with vHL-associated hemangioblastoma treated with Belzutifan at a daily oral dose of 120mg. Perfusion MRI with ASL sequences were acquired as part of the tumor imaging protocol. We quantified rCBF of the tumors prior to treatment, following intiation of BZF, and at most recent follow-up. Statistics were performed using R studio. The long axis diameter, volume of the solid tumor, rCBF mean, rCBF max, N mean, N max of lesions all decreased after Belzutifan treatment (p<0.001). While the long axis diameter of the cyst in cystic lesions did not show statistically significant change, the volume of cysts decreased significantly on Belzutifan (p=0.016). Spearman’s rank correlation analysis demonstrated association between change in rCBF mean, rCBF max, N mean and N max to % reduction in tumor volume. In addition, change in rCBF mean, change in rCBF max and change in N max all showed statistically significant association on multiple linear regression. Finally, early change in CBF and N predicted further volumetric response to treatment with belzutifan. We demonstrate that vHL-associated hemangioblastoma demonstrates decrease in rCBF in conjunction with and predictive of decreased volume following treatment with belzutifan.

NIMG-46. UNDERSTANDING PERMEABILITY CHANGES IN VESTIBULAR SCHWANNOMAS AS PART OF THE DYNAMIC RESPONSE TO RADIOSURGERY USING GOLDEN-ANGLE RADIAL SPARSE PARALLEL IMAGING, A RETROSPECTIVE STUDY

Abstract Vestibular schwannomas demonstrate different responses after stereotactic radiosurgery (SRS), commonly including a transient loss of internal enhancement on post contrast T1-weighted MRI thought to be due to an early reduction in tumor vascularity. We used dynamic contrast-enhanced based golden-angle radial sparse parallel (GRASP) MRI to characterize the vascular permeability changes underlying this phenomenon, with correlations to long-term tumor regression. Consecutive patients with vestibular schwannoma who underwent SRS between 2017-2019, had a transient loss of enhancement after SRS, and had long-term longitudinal GRASP studies (6, 12, and 24-36 months) were included in this retrospective cohort analysis (n = 19). Using GRAVIS, an analysis pipeline for GRASP studies, we extracted the key parameters normalized to the venous sinus from a region of interest within the tumor. The peak, area under the curve (AUC), and wash-in phase slope were significantly reduced at 6, 12, and 24 months after SRS (corrected p < 0.05), even while the internal enhancement returned in the tumors. Larger pre-SRS tumors were more likely to have a greater reduction in peak (p = 0.013) and AUC (p = 0.029) at 6 months. In a subset of patients (N = 13) with long-term follow-up, the median percentage reduction in tumor volume was 58% at median 62 months. These patients showed a strong correlation between changes in peak, AUC, and wash-in phase slope at 6 months and tumor volume at last follow-up. After SRS and loss of internal contrast uptake within the vestibular schwannomas, a slow vascular permeability dynamic found on GRASP studies persisted, suggesting the internal development of scar tissue. Moreover, the degree of reduction in GRASP parameters at 6 months positively predicted the degree of tumor regression long-term.

EXTH-46. TESTING PRECISION GENOMICS-GUIDED THERAPY IN A MODEL OF ANAPLASTIC PLEOMORPHIC XANTHOASTROCYTOMA: DUAL INHIBITION OF CDK4/6 AND MEK

Abstract Typical survival for pediatric high-grade gliomas remains less than 18 months despite recent improved understanding of the molecular drivers of these tumors. Hyperactivating MAPK and CDK4/6 pathway mutations are common and targetable alterations implicated in tumorigenesis and malignant transformation in pediatric glioma. We have established and characterized a novel patient-derived xenograft (PDX) model, RHT128, from a pediatric patient diagnosed with the high-grade glioma anaplastic pleomorphic xanthoastrocytoma (APXA). The molecular landscape of PDX RHT128 exhibits molecular fidelity to the patient’s tumor. Clinical precision genomics analysis of the tumor revealed a novel BRAF chromosomal rearrangement and CDKN2A/B deletion. Based on this molecular signature, the patient was treated with MEK inhibitor trametinib as a monotherapy and, following progression of disease, with CDK4/6 inhibitor ribociclib. However, the tumor continued to progress. In this study our objective is to simultaneously target the CDK4/6 and MAPK pathways in RHT128 and determine to what extent this combination therapy minimizes emergence of therapeutic resistance. Single-agent efficacy assessments in a subcutaneous RHT128 model, showing significant dose-dependent reduction in tumor volume after treatment with abemaciclib (p<0.05), palbociclib (p<0.01), and the blood-brain barrier-permeable MEK inhibitor mirdametinib (p<0.0001). Analysis of the global kinome in CDK4/6 inhibitor-treated PDX tissues compared to vehicle treatment using multiplexed-inhibitor bead chromatography–mass spectrometry demonstrated effective inhibition of CDK4/6. Moreover, this analysis revealed dose-dependent alterations in the MAPK and PI3K pathways and modulation of critical neurotransmitter pathways in treated tissues. Future studies will explore efficacy of these MEK and CDK4/6 inhibitors in combination and in our BRAF wild-type and BRAF V600E variant APXA models to gain further mechanistic insights. The pervasiveness of alterations to the MAPK and CDK4/6 pathways in pediatric gliomas make this approach promising for further study in a broader range of these deadly tumors.

CNSC-59. EPIGENETIC POTENTIATION OF ANTI-TUMOR IMMUNE RESPONSES USING VIRAL MIMICRY IN GLIOMAS

Abstract Clinical trials using immune checkpoint inhibition(ICI) have traditionally failed in glioblastoma (GBM). Viral mimicry, which augments anti-tumor immune responses and sensitizes response to immunotherapy in other cancers, involves the epigenetic activation of endogenous retroelements (REs). REs are silenced via the HUSH complex and H3K9me3. This process is mediated by ZNF638. We aimed to elucidate the role of viral mimicry in enhancing ICI through epigenetic reprogramming of the HUSH complex. We demonstrated that RE expression among 48 superfamilies inversely correlated with ZNF638 expression in gliomas, based on data from 71 newly-diagnosed GBMs. Using transcriptional deconvolution, we showed ZNF638 negatively correlates with innate antiviral immune response signatures(TLR3)(RTLR3=-0.300,pTLR3=0.00006). We validated these in-silico results in pure glioma cell lines, patient-derived GBM neurospheres, and syngeneic murine models. ZNF638 knockdown induced intracellular RE-mediated dsRNA signaling cascades via RIG-I and TLR3. This knockdown significantly increased PD-L1 expression(p<0.001), reduced H3K9 trimethylation, and enhanced cytoplasmic dsRNA accumulation in glioma cell lines. In patient-derived GBM neurospheres, ZNF638 knockdown upregulated immune and antiviral programs. Additionally, knockdown resulted in upregulation of several endogenous repeat elements (Alu, LTR). Single-cell RNA sequencing showed ZNF638 clustering in neural progenitor-like and oligodendrocyte-like cells, with increased retroelement expression in low ZNF638 cells. Tumors with lower ZNF638 expression showed increased CD8+ populations. Bulk RNA sequencing deconvolution revealed that ZNF638 is linked to reduced CD8 and DC infiltration in gliomas (RCD8=-0.202,RDC=-0.198;pCD8=8.79E-06,pDC=1.34E-05). In immunocompetent mice, ZNF638 KO and PD-L1 inhibition significantly improved survival (mOS >50 days, p<0.0001) and reduced tumor volume (p<0.001). Analysis of cohorts from multiple cancer types treated with anti-PD1 therapy demonstrated that ZNF638 expression predicts therapeutic response and overall survival. Our findings suggest that epigenetic reprogramming through HUSH inhibition may potentiate immunotherapy for GBM.

TMIC-75. ICAM-1 PROMOTES AN IMMUNOSUPPRESSIVE LANDSCAPE IN GLIOBLASTOMAS BY REGULATING THE PHENOTYPIC POLARIZATION STATE OF TUMOR ASSOCIATED MACROPHAGES

Abstract Glioblastoma (GBM) is a fatal adult CNS tumor, with a median overall survival of 12-15 months. Intercellular adhesion molecule 1 (ICAM-1) is a cell adhesion molecule whose expression is significantly greater in the mesenchymal transcriptional subtype of GBMs, whereby it is prognostic. We have previously shown that when ICAM-1 is knocked out in the tumor microenvironment (TME) of immune-competent GBM-bearing mice, these mice showed prolonged overall survival and reduced tumor volume. It is yet to be determined how ICAM-1 expression affects infiltration and functional polarization of tumor-associated macrophages (TAMs) and other immune cells in GBM, and if we can suppress the infiltration of M2-like TAMs into the TME by inhibiting ICAM-1. We used a mouse glioma model with and without knockout (KO) of the ICAM-1 gene and intracranially injected GL261 cells. ICAM-1 KO mice survived longer and had reduced tumor volume relative to wildtype, with survival and volume differences being rescued upon ICAM-1 bone marrow reconstitution. Upon endpoint, we performed cytometry by time-of-flight, and found that ICAM-1 KO tumors harbored a reduction in helper T cells, Tregs, and cytotoxic T cells, but an increased proportion in double-negative T cells relative to ICAM-1 WT tumors. ICAM-1 KO tumors also demonstrated an increased proportion of M1-like TAMs relative to M2-like. These results were validated upon administration of ISIS-3082 to tumor models, an antisense oligonucleotide for which a human-specific form has been fast-track approved for other diseases, that targets ICAM-1 transiently in peripheral immune cells prior to TME infiltration. These findings suggest that ICAM-1 expression influences immune cell infiltration, potentially modulates the functional state of TAMs towards a M1-like state, and that ICAM-1 may be targeted in TAMs to ‘reeducate’ them before entering the TME. This may be translated into clinical practice and combined with standards-of-care to render GBMs more sensitive to treatment.

CSIG-04. ANTI-TUMOR RESPONSE OF ONCOMAGNETIC MONOTHERAPY IN GLIOBLASTOMA

Abstract BACKGROUND Glioblastoma (GBM) is the most lethal primary malignant brain tumor with a median survival of 15–20 months. Concurrent temozolomide (TMZ) chemotherapy and radiation (XRT) remain the current standard of care (SOC) treatment for newly diagnosed GBM. The addition of tumor treating fields (TTFs) improves median survival for newly diagnosed GBM from 16 to 20.9 months. This modest improvement underscores the urgent need to identify a new treatment modality as a safe and effective therapy. PURPOSE The noninvasive Oncomagnetic device developed in our laboratory selectively kills glioblastoma (GBM) in vitro. We studied the cellular and molecular effects of Oncomagnetic monotherapy (OMT) on GBM cell lines and investigated immune response in a syngeneic mouse model. RESULTS OMT significantly reduces cell proliferation and cell survival in vitro. OMT induces persistent ROS primarily by increasing superoxide level in cancer cells and reduces mitochondrial-membrane potential in GBM cells. It also induces DNA damage and arrests cells in G1-phase of the cell cycle. Whole-body Oncomagnetic stimulation caused a substantial retardation of tumor growth and reduction of the contrast-enhanced tumor volume in 9.4T MRI scans. OMT showed significant increase in overall survival in treated mice. We also observed upregulated immune response indicated by RNA sequencing data obtained from OMT-treated GBM cells as well as in treated GBM mouse tumor sections by imaging mass cytometry analysis. The tSNE analysis of various immune clusters suggests significant increase in immune response by activating CD4+, CD8+, murine-macrophages and M1-macrophages located at tumor site in the treated group. CONCLUSION The obtained data indicate significant anti-tumor response by inhibiting cancer signaling pathways and an increased immune response. Our findings suggest unique mechanism of action for OMT stimulation and provide a strong rationale for standalone OMT for GBM.

SURG-42. THE T1/T2 RATIO IS ASSOCIATED WITH RESECTABILITY IN PATIENTS WITH IDH-MUTANT ASTROCYTOMAS CNS WHO GRADES 2 AND 3

Abstract PURPOSE IDH-mutant astrocytomas CNS WHO grade 2 and 3 show heterogenous appearance on magnetic resonance imaging (MRI). In the pre-molecular era, the discrepancy between T1 hypointense and T2 hyperintense tumor volume in absolute values has been proposed as a marker for diffuse tumor growth. We set out to investigate if a ratio of T1 to T2 tumor volume (T1/T2 ratio) is associated with resectability and overall survival in patients with IDH-mutant astrocytomas. METHODS Patient data from two centers (Center A: Sahlgrenska University Hospital, Gothenburg, Sweden; Center B: LMU University Hospital, Munich, Germany) were collected retrospectively. Inclusion criteria were as follows: pre- and postoperative MRI scans available for volumetric analysis (I), diagnosis of an IDH-mutant astrocytoma between 2003-2021 (II), and tumor resection at initial diagnosis (III). Tumor volumes were manually segmented. The T1/T2 ratio was calculated and correlated with extent of resection, residual T2 tumor volume, and overall survival. RESULTS The study comprised 134 patients with 65 patients included from Center A and 69 patients from Center B. The median overall survival was 134 months and did not differ between the cohorts (p=0.29). Overall, the median T1/T2 ratio was 0.79 (range 0.15-1.0). Tumors displaying a T1/T2 ratio of 0.33 or lower showed significantly larger residual tumor volumes postoperatively (median 17.9 cm3 versus 4.6 cm3, p=0.03). The median extent of resection in these patients was 65% versus 90% (p=0.03). The ratio itself did not correlate with overall survival. In multivariable analyses, larger postoperative tumor volumes were associated with shorter survival times (HR 1.02, 95% CI 1.01-1.03, p<0.01). CONCLUSION The T1/T2 ratio might be a good indicator for diffuse tumor growth on MRI and is associated with resectability in patients with IDH-mutant astrocytoma. This ratio might aid to identify patients in which an oncologically relevant tumor volume reduction cannot be safely achieved.

EXTH-65. EXPLORING ANTIHISTAMINES TO TREAT GLIOBLASTOMA

Abstract BACKGROUND Glioblastoma (GBM) is an aggressive and lethal primary malignant brain tumor with a 5-year survival of 6.8%. Interestingly, people with allergies, atopy, or asthma are less likely to develop GBM. A recent study recognized that high histamine receptor 1 (HRH1) expression is inversely related to overall and progression-free survival. METHODS We used a bioinformatics tool, iLINCS, which, after analyzing 99 GBM and 38 healthy samples, identified 36 drugs that can reverse GBM signatures. Based on the concordance score and blood-brain barrier permeability, we selected brompheniramine (Brom), a first-generation HRH1 antagonist, as a potential candidate. HRH1 expression analysis was done on human and mouse GBM cell lines by immunoblotting. The effect of Brom alone or in combination with temozolomide (TMZ) was investigated on proliferation, DNA damage and apoptosis in human (U118 and U251) and three syngeneic (EGFRvIII, p16-/- & GFAP Cre, PTEN+/-, p53R172H+/-, EGFRvIII & GFAP Cre, PTEN-/-, p53R172H-/- & GFAP Cre) cell lines. GBM orthograft mouse models were used to test the efficacy of combination therapy. RESULTS HRH1 is highly expressed in human and syngeneic GBM cells. This expression is further increased in TMZ-resistant cells. Interestingly, the addition of histamine, induced TMZ resistance in GBM cells. TMZ with Brom decreases proliferation in mouse and human GBM cells, and Brom is non-toxic to non-transformed cells. Combination treatment increases DNA damage and decreases the DNA repair mechanism, possibly by downregulating the expression of O6methylguanine-DNA methyltransferase (MGMT). In the GBM orthograft mouse model, the combination therapy reduced tumor volume and significantly increased survival compared to single-agent treatment. CONCLUSIONS Brom is non-toxic; Brom, in combination with TMZ, enhanced the anti-tumor efficacy of TMZ both in in vitro and in vivo GBM models. Further studies in spontaneous mouse models are planned.

CSIG-07. JAK1/STAT1/3 COOPERATES WITH THE CIC-FUSIONS TO DRIVE CIC-REARRANGED SARCOMAS

Abstract CIC-rearranged sarcoma (CRS) is an rare disease driven by a specific fusion protein involving the CIC gene. Occurrence in the brain is 3% in all CRS patients. The native CIC protein is a transcriptional repressor of the (RTK)/Ras/ERK signaling pathway, which is one of the most tumorigenic pathways in cancer. The most common rearrangement is with the double homeobox 4 (DUX4) transcription factor (CIC-DUX4), and others, such as CIC-NUTM1 fusions, have been identified in a subset of pediatric primitive neuroectodermal tumors. However, the molecular mechanisms by which CIC-fusions drive CRS remain unknown. Preliminary data shows that CIC-DUX4/NUTM1 fusions activate JAK and its downstream effector STAT1/3. We hypothesize that the JAK/STAT1/3 signal transduction pathway cooperates with CIC-fusions to induce the expression of oncogenic transcription factors ETV1/4/5 and drive these sarcomas. We show high levels of JAK1/STAT1/3 activation in patient-derived CRS cell lines (NCC-SCC-89/C) as compared to other sarcoma cell lines without the fusion. JAK1 inhibition using Solicitinib and Ruxolitinib reduced phosphorylation of STAT1/3 as well as protein and mRNA expression of ETV1/4/5, promoter activity of ETV5, cell proliferation and tumorgenicity. Interestingly DUX4 is involved in histone acetylation and STAT1 has been shown to activate p300/CBP complex which lead us to evaluating the role of STAT1 in the gene activation CRS. We elucidate mechanistically that the fusion proteins increase binding of STAT1/3 at the promoters of ETV 1/4/5. Importantly, JAK1 inhibition effectively reduces histone acetylation induced by CIC-fusions at the promoters of ETV1/4/5. To evaluate the pre-clinical effect of targeting the JAK1/STAT1/3 pathway, A CRS cell line (NCC-SCC-89/C) were grafted into NSG mice. Ruxolitinib treatment significantly reduced tumor volume, STAT activation, and ETV1/4/5. In conclusion, we show that the JAK1/STAT1/3 pathway plays a critical role in cooperating with CIC-fusions to drive CRS, providing insight into potential therapeutic avenues for these aggressive tumors.

IMMU-05. TNFR2 IS A NOVEL MARKER OF EXHAUSTION AND TNFR2 BLOCKADE IMPROVES SUBCUTANEOUS TUMOR CONTROL

Abstract Checkpoint inhibitors have been successful in various tumors. However, these treatments have failed in glioblastoma. Approaches harnessing the immune response are hindered by multiple factors, including T cell exhaustion. TOX is particularly important for the transcriptional and epigenetic reprogramming of exhausted T cells. While it is known that NFAT is upstream of TOX, our understanding of the upregulation of TOX remains incomplete. We hypothesized that tumor necrosis factor (TNF) could be involved in the upregulation of TOX as TNF can lead to the translocation of NFAT into the nucleus. We observed significant upregulation of the anti-inflammatory TNF receptor type II (TNFR2) in tumor-infiltrating T cells. This local upregulation mimics expression patterns of canonical exhaustion markers. TNFR2 expression is correlated with markers of exhaustion, including PD1, TIM3, and TOX. Furthermore, TNFR2 knock out (KO) CD8 T cells have significantly lower TOX expression, without the concomitant decrease of TIM3. Whereas previous studies have linked TIM3 and TOX expression, these data suggest that TIM3 and TOX are regulated independently. We utilized bulk RNA-sequencing to assess transcriptional regulation of WT and KO T cells and detected a significant reduction of T cell exhaustion and immune checkpoint pathways in TNFR2 KO T cells. Various exhaustion-related transcription factors and coinhibitory markers were significantly reduced. In contrast, a significant increase in AP1 transcription factors, commonly associated with T cell effector functions, was detected. Given this reduced exhaustion profile, we subsequently investigated the influence of TNFR2 on tumor burden. TNFR2 KO mice had significantly lower tumor burdens following subcutaneous tumor challenges. We subsequently treated mice with a TNFR2 antagonist. While the antagonist alone was not sufficient to improve tumor control, combination with anti-PD1 significantly reduced tumor volumes. These data provide evidence for a novel marker of exhaustion that could result in a unique therapeutic strategy.

SURG-39. LONG-TERM RESULTS OF INTERVENTION FOR GROWING INCIDENTAL MENINGIOMAS OR CONTINUED ACTIVE SURVEILLANCE, A PROSPECTIVE COHORT STUDY

Abstract Incidental meningiomas are common findings. They represent a clinically challenging cohort, without a detailed consensus on their management. The risk of tumor progression and that of intervention must be considered. Long-term prospective data on incidental meningiomas are sparse. We aimed to evaluate the long-term effect of Gamma Knife Radiosurgery (GKRS), surgical resection and continued observation for growing incidental meningiomas. A prospective database of 62 patients (70 tumors) commenced under active surveillance was established in 2009. Radiological and clinical data was obtained until sept 2023. The results of long-term observation (group 1) and intervention with GKRS (group 2) or surgery (group 3) at progression was analyzed. Due to growth, 41 (58.6%) tumors were treated. The mean growth rate was higher prior to GKRS (2.1 cm 3 /y) and surgery (0.4 cm 3 /y) than during long-term active surveillance (0.009 cm 3 /y) (p < 0.001). The meningiomas became in mean 31.3% and 99.1% smaller after GKRS and resection, respectively. In comparison, tumors under long-term surveillance increased in mean 27.2% (p < 0.001). According to the RANO response criteria, the complete, partial, and minimal response versus stable disease rates were higher following intervention, the overall progressive disease rates were similar (Group 1; 20.7%, group 2: 11.4% and group 3: 16.7%) (p = 0.232). Treatment versus no-treatment did not affect overall survival (Group 1; 10.3 y, group 2: 11.8 y and group 3: 13.5 y (p = 0.264). No symptom development was registered in group 1. In group 2, 2.9% experienced transient (grade 2) adverse events, and in group 3, 50% suffered permanent (grade 4) adverse events according to the Common Terminology Criteria for Adverse Events (CTCEAv5). Intervention reduces tumor volume; however, the clinical significance remains uncertain and side effects are not negligible. Active surveillance is safe, saves > 40% of patients from unnecessary interventions and does not compromise future treatments. The optimal timing of intervention should be further explored.

BIOM-74. IDENTIFICATION OF EMP2 AS A TARGETABLE BIOMARKER IN GLIOBLASTOMA TUMORS RESISTANT TO ANTI-ANGIOGENIC THERAPIES

Abstract BACKGROUND The role of Epithelial Membrane Protein-2 (EMP2) in mediating resistance to current anti-angiogenic drugs in the treatment of glioblastoma (GBM) is an intriguing hypothesis that may provide insights into improving the limited survival benefits observed in current therapies. Investigating the interaction between EMP2 and anti-angiogenic drugs such as bevacizumab or Aflibercept could lead to a better understanding of the mechanisms underlying resistance, potentially opening up new avenues for more effective treatment strategies. OBJECTIVE To use EMP2 as a targetable biomarker for anti-VEGF resistance and test combination therapy with anti-VEGF (Aflibercept or bevacizumab), and a novel anti-EMP2 mAb in SB28 and GL261 GBM mouse models. METHODS Tumor tissue was collected prospectively from patients undergoing surgery for suspected glioblastoma and stained for EMP2 and HIF-2a using standard immunohistochemistry. Patients had banked tumor tissue both before and after bevacizumab treatment with 3 months of clinical and radiologic follow-up available. To establish syngeneic models, 10-week-old mice were subcutaneously implanted with murine SB28 or GL261 GBM cells. RESULTS Immunohistochemistry conducted on pre- and post-bevacizumab-treated patient samples revealed an increase in EMP2 and HIF-2a protein expression post-treatment. This increase in EMP2 expression correlated with reduced survival time. Similarly, the in vivo models showed a significant upregulation in both markers in the Aflibercept-treated cohort compared to the control. Treatment with either polyclonal sera to VEGF or Aflibercept failed to reduce tumor load or provide a therapeutic benefit. However, combination therapy with Aflibercept and anti-EMP2 mAb was found to significantly reduce tumor volume and weight in both SB28 and GL261 tumors. These findings highlight the potential effectiveness of combination therapies targeting EMP2 and VEGF in reducing tumor load in GBM. CONCLUSION Our results suggest that EMP2 can be a promising new therapeutic target that can be combined with anti-VEGF treatment to potentially increase overall survival in GBM patients.

BIOM-37. EXPLORING L-TYPE AMINO ACID TRANSPORTER 1 (LAT1) AS A DIAGNOSTIC MARKER AND THERAPEUTIC TARGET IN MALIGNANT PERIPHERAL NERVE SHEATH TUMORS (MPNST)

Abstract OBJECTIVES Malignant peripheral nerve sheath tumors (MPNST) are aggressive soft tissue sarcomas that frequently arise from benign plexiform neurofibromas (PN). Approximately half of MPNST are associated with Neurofibromatosis type 1 (NF1). Despite partial elucidation of the molecular mechanisms underlying MPNST, diagnosis remains challenging, and prognosis is poor. Previously, we found that MPNST cells have elevated βIII-spectrin, a cytoskeletal protein involved in cell morphology and regulation of transporters, including L-type Amino Acid Transporter 1 (LAT1). LAT1 facilitates transport of large neutral amino acids and is upregulated in other cancer types. Our objective is to investigate LAT1 as a diagnostic marker and therapeutic target for MPNST. METHODS Gene and protein expression were determined by qPCR and WES protein analysis, respectively. The effect of shRNA knockdown of LAT1 in MPNST cells was assessed using IncuCyte proliferation assays as well as multiple mouse models. Tumor uptake of [18F] FDOPA, primarily transported by LAT1, in MPNST PDX tumors was assessed using PET/CT imaging. RESULTS LAT1 was expressed in MPNST (n=14) vs. benign PN tissues (n=6). Additionally, in human and murine MPNST cells, shRNA-mediated knockdown of LAT1 suppressed proliferation of MPNST cells in vitro and tumor growth in vivo. In a subcutaneous tumor growth model, intratumor injection of shLAT1 lentivirus reduced tumor volume to approximately 50% of control. Furthermore, uptake of [18F] FDOPA by LAT1 in MPNST PDX tumors was detected by PET/CT imaging. WU-356 PDX tissues exhibited higher LAT1 expression as well as increased uptake of [18F] FDOPA in tumors. Conversely, PET imaging showed significantly lower uptake of [18F] FDOPA in LAT1 knockdown tumors (p<0.05). CONCLUSIONS These results suggest that LAT1 promotes the growth and survival of MPNST cells. In addition, these findings demonstrate the potential use of LAT1 and the radiotracer [18F] FDOPA as a diagnostic biomarker in the management of MPNST.

Sitravatinib in patients with solid tumors selected by molecular alterations: results from a Phase Ib study.

Aim: We report clinical activity and safety of sitravatinib in patients with advanced cancer from basket cohorts with specific molecular alterations, in a Phase Ib study.Materials & methods: Patients with advanced solid tumors harboring amplification, mutation, or rearrangement of MET, AXL, RET, NTRK, DDR2, KDR, PDGFRA, KIT or CBL received sitravatinib once daily. Primary end point was confirmed objective response rate (ORR).Results: In total, 113 patients were enrolled following a median of 3 (range 1-18) prior systemic regimens. Altered RET (n = 31), CBL (n = 31) and MET (n = 17) were most frequent cohorts. Overall, 68.9% had reduced tumor volume and most (61.5%) had a best objective response of stable disease. ORR was highest in patients with RET-rearranged non-small cell lung cancer (21.1%) but did not differ significantly from the null hypothesis (ORR ≤15%; p = 0.316). Median progression-free survival and overall survival (5.7 and 24.2 months, respectively) were also longest in the RET-rearranged non-small cell lung cancer cohort. Diarrhea (61.1%), fatigue (50.4%) and hypertension (46.9%) were the most frequent treatment-emergent adverse events. Most treatment-emergent adverse events were mild-to-moderate in severity. The study closed before the planned number of patients were enrolled in all cohorts.Conclusion: Sitravatinib had a manageable safety profile with modest signals of clinical activity in patients with molecularly selected solid tumors.Clinical trial registration: www.clinicaltrials.gov identifier is NCT02219711.

A comprehensive and systematic review on Curcumin as a promising candidate for the inhibition of melanoma growth: From pre-clinical evidence to molecular mechanisms of action

BackgroundMelanoma, a highly malignant skin tumor, can develop systemic metastases during the early stage. Several studies of melanoma animal models indicate that curcumin, a natural plant extract, inhibits melanoma growth through various mechanisms. To evaluate the relationships among different experimental conditions, curcumin itself, its derivatives, and special formulations, it is necessary to conduct a systematic review and meta-analysis. PurposeThis meta-analysis aims to evaluate the potential of Curcumin as a drug for inhibiting the growth of melanoma and to determine the optimal dosage range and treatment duration for Curcumin administration. MethodsA systematic search of studies published from inception to December 2023 was conducted across six databases (PubMed, Web of Science, Embase, China National Knowledge Infrastructure, Wanfang Data, and VIP). Methodological quality was assessed using SYRCLE's RoB tool. Study heterogeneity was assessed using Cochran's Q test and I2 statistics. Publication bias risk was evaluated using a funnel plot. All analyses were performed using R (version 4.3.3). Additionally, three-dimensional effect analysis and machine learning techniques were utilized to determine the optimal dosage range and treatment duration for Curcumin administration. ResultsForty studies involving 989 animals were included. The results demonstrated that, relative to the control group, administration of Curcumin resulted in a significant reduction in tumor volume. [SMD=−3.44; 95 % CI (−4.25, −2.63); P<0.01; I2 = 79 %] and tumor weight [SMD=−1.93; 95 % CI (−2.41, −1.45); P<0.01; I2 = 75 %]. Additionally, Curcumin demonstrated a significant capacity to decrease the number of lung tumor nodules and microangiogenesis, as well as to extend survival time, in animal models. The results from three-dimensional effect analysis and machine learning emphasize that the optimal dosage range for Curcumin is 25–50 mg/kg, with an intervention duration of 10–20 days. ConclusionCurcumin can inhibit the growth of melanoma, and the dose-response relationship is not linear. However, further large-scale animal and clinical studies are required to confirm these conclusions.

4‑Methoxydalbergione inhibits the tumorigenesis and metastasis of lung cancer through promoting ferroptosis via the DNMT1/system Xc‑/GPX4 pathway.

Lung cancer is responsible for the highest number of tumor‑related deaths worldwide. A flavonoid extracted from the heartwood of Dalbergia sissoo Roxb., 4‑methoxydalbergione (4‑MD), exhibits potent anticancer activity in multiple malignancies; however, the potential anticancer activity of 4‑MD in lung cancer has not yet been elucidated. In the present study, A549 cells were treated with increasing concentrations of 4‑MD, and cell viability was assessed using a Cell Counting Kit‑8 assay. In addition, colony formation, 5‑ethynyl‑2'‑deoxyuridine, wound healing and Transwell assays were conducted to evaluate cell proliferation, migration and invasion, respectively. Cell morphology was observed using transmission electron microscopy, and ferroptosis was determined using thiobarbituric acid reactive substance, lipid reactive oxygen species (ROS) and iron assays. Moreover, molecular docking was used to verify the potential interaction between 4‑MD and DNA methyltransferase 1 (DNMT1). Tumor‑bearing mice were established and treated with 10 or 30 mg/kg 4‑MD, and tumor volume and weight were recorded. Immunohistochemistry and Prussian blue staining were conducted to examine Ki‑67 expression and iron deposition in tumor tissues, and protein expression was further explored using western blot analysis. The results of the present study revealed that 4‑MD significantly inhibited cell proliferation, migration, invasion and epithelial‑mesenchymal transition in a concentration‑dependent manner. Notably, 4‑MD induced ferroptosis via increased lipid peroxidation, lipid ROS and Fe2+ levels. In addition, it was revealed that 4‑MD can directly bind to DNMT1 to inhibit expression, and inhibit solute carrier family 7 member 11 (SLC7A11; also known as cystine‑glutamate antiporter) and glutathione peroxidase 4 expression. Following DNMT1 overexpression, the observed antitumor activity and ferroptosis‑promoting effects of 4‑MD were partially reversed. Furthermore, 4‑MD significantly inhibited tumor growth in vivo, and reduced tumor volume and weight. In addition, Ki‑67 expression was reduced while iron deposition was increased in the tumor tissues of mice following treatment with 4‑MD. In conclusion, 4‑MD may exhibit anticancer activity through the promotion of DNMT1‑mediated cell ferroptosis. Thus, 4‑MD may have potential as a novel therapeutic agent in the treatment of lung cancer.

Nanotechnology‐Based Systems for Enhancing the Efficacy of Sonodynamic Therapy in Cancer Treatment

AbstractSonodynamic therapy (SDT) is a minimally invasive cancer therapeutic modality that utilizes low‐intensity ultrasound to activate sensitizers for the production of cytotoxic reactive oxygen species (ROS) and the site‐specific ablation of tumors. Compared to conventional cancer treatments, SDT has no adverse side effects, has been shown to radically reduce tumor volume, and has the potential to raise antitumor immune response. Ever‐increasing studies have demonstrated that nanotechnology can significantly enhance the efficacy of SDT against certain cancers and improve its therapeutic effects. Nanosystems are developed that can improve the intratumoral delivery and cellular uptake of sensitizers, can reduce intrinsic cancer defenses against ROS, can act synergistically for the production of ROS and can tackle cancer immune tolerance. In this review, the various strategies to enhance SDT performance with the assistance of nanotechnology, and the outstanding research outcomes, are discussed.

The Inhibitory Effect and Mechanism of the Histidine-Rich Peptide rAj-HRP from Apostichopus japonicus on Human Colon Cancer HCT116 Cells

Colon cancer is a common and lethal malignancy, ranking second in global cancer-related mortality, highlighting the urgent need for novel targeted therapies. The sea cucumber (Apostichopus japonicus) is a marine organism known for its medicinal properties. After conducting a bioinformatics analysis of the cDNA library of Apostichopus japonicus, we found and cloned a cDNA sequence encoding histidine-rich peptides, and the recombinant peptide was named rAj-HRP. Human histidine-rich peptides are known for their anti-cancer properties, raising questions as to whether rAj-HRP might exhibit similar effects. To investigate whether rAj-HRP can inhibit colon cancer, we used human colon cancer HCT116 cells as a model and studied the tumor suppressive activity in vitro and in vivo. The results showed that rAj-HRP inhibited HCT116 cell proliferation, migration, and adhesion to extracellular matrix (ECM) proteins in vitro. It also disrupted the cytoskeleton and induced apoptosis in these cells. In vivo, rAj-HRP significantly inhibited the growth of HCT116 tumors in BALB/c mice, reducing tumor volume and weight without affecting the body weight of the tumor-bearing mice. Western blot analysis showed that rAj-HRP inhibited HCT116 cell proliferation and induced apoptosis by upregulating BAX and promoting PARP zymogen degradation. Additionally, rAj-HRP inhibited HCT116 cell adhesion and migration by reducing MMP2 levels. Further research showed that rAj-HRP downregulated EGFR expression in HCT116 cells and inhibited key downstream molecules, including AKT, P-AKT, PLCγ, P38 MAPK, and c-Jun. In conclusion, rAj-HRP exhibits significant inhibitory effects on HCT116 cells in both in vitro and in vivo, primarily through the EGFR and apoptosis pathways. These findings suggest that rAj-HRP has the potential as a novel targeted therapy for colon cancer.

G9a/DNMT1 co-targeting inhibits non-small cell lung cancer growth and reprograms tumor cells to respond to cancer-drugs through SCARA5 and AOX1

The treatment of non-small cell lung cancer (NSCLC) patients has significantly improved with recent therapeutic strategies; however, many patients still do not benefit from them. As a result, new treatment approaches are urgently needed. In this study, we evaluated the antitumor efficacy of co-targeting G9a and DNMT1 enzymes and its potential as a cancer drug sensitizer. We observed co-expression and overexpression of G9a and DNMT1 in NSCLC, which were associated with poor prognosis. Co-targeting G9a/DNMT1 with the drug CM-272 reduced proliferation and induced cell death in a panel of human and murine NSCLC cell lines. Additionally, the transcriptomes of these cells were reprogrammed to become highly responsive to chemotherapy (cisplatin), targeted therapy (trametinib), and epigenetic therapy (vorinostat). In vivo, CM-272 reduced tumor volume in human and murine cell-derived cancer models, and this effect was synergistically enhanced by cisplatin. The expression of SCARA5 and AOX1 was induced by CM-272, and both proteins were found to be essential for the antiproliferative response, as gene silencing decreased cytotoxicity. Furthermore, the expression of SCARA5 and AOX1 was positively correlated with each other and inversely correlated with G9a and DNMT1 expression in NSCLC patients. SCARA5 and AOX1 DNA promoters were hypermethylated in NSCLC, and SCARA5 methylation was identified as an epigenetic biomarker in tumors and liquid biopsies from NSCLC patients. Thus, we demonstrate that co-targeting G9a/DNMT1 is a promising strategy to enhance the efficacy of cancer drugs, and SCARA5 methylation could serve as a non-invasive biomarker to monitor tumor progression.