Targeting the Tumor Immune Microenvironment Could Become a Potential Therapeutic Modality for Invasive Pituitary Adenoma.

Objective: The study aimed to explore the relationship between the invasiveness and immune cell infiltration in pituitary adenoma (PA) and provide the basis for immuno-targeting therapies.Methods: One hundred three patients who underwent surgery at a single institution were retrospectively identified. The infiltration of macrophages and T lymphocytes was quantitatively assessed in PA.Results: The number of CD68+ macrophages was positively correlated with Knosp (P=0.003) and MMP-9 grades (P=0.00). The infiltration of CD163+ macrophages differed among Knosp (P=0.022) and MMP-9 grades (P=0.04). CD8+ tumor-infiltrating lymphocytes (TILs) were also positively associated with Knosp (P=0.002) and MMP-9 grades (P=0.01). Interestingly, MGMT expression was positively correlated with MMP-9 staining extent (P=0.000). The quantity of CD8+ TILs (P= 0.016), CD68+ macrophages (P=0.000) and CD163+ macrophages (P=0.043) were negatively associated with MGMT expression levels. The number of CD68+ macrophages in the PD-L1 negative group was significantly more than in the PD-L1 positive group (P=0.01). The rate of PD-L1 positivity had positive correlations with the Ki-67 index (P=0.046) and p53 expression (P =0.029). Conclusion: Targeted therapy for macrophages and CD8+ TILs could be a helpful treatment in the future for invasive PA. Temozolomide (TMZ) may have better effects on the treatment of PA infiltrating more immune cells. Anti-PD-L1 therapy may better respond to PA with higher Ki-67, p53 expression and more infiltrating CD68+ macrophages. Multiple treatment modalities, especially combined immunotherapy, or immunotherapy with TMZ combination, could become a novel therapeutic strategy for invasive PA.

IM-2 Possibility of immunotherapy for the glioblastoma patients with O6-methyl-guanine DNA methyltransferase (MGMT) expression or promoter unmethylated

Background: MGMT is a DNA repair protein that removes the cytotoxic O6-methylguanine (O6MG) DNA lesions generated by TMZ; thereby, MGMT expression is mechanistically linked to TMZ resistance. However, thus far, there is no effective treatment for these patients with MGMT promoter unmethylated. Therefore, a new treatment for GBM patients with MGMT expression is urgently needed. To this end, we examined the tumor microenvironment in GBM with or without MGMT expression. Methods: Based on The Cancer Genome Atlas (TCGA) primary GBM cohort, the tumor-infiltrating lymphocytes (TILs) expression level was calculated using the CIBERSORTx algorithms and the single-sample Gene Set Enrichment Analysis (ssGSEA) method. Furthermore, the differential expression gene analysis was conducted and pathway analysis was performed using Ingenuity Pathway Analysis (IPA). The results were validated using the GBM cohort from the Chinese Glioma Genome Atlas (CGGA) database. In addition, TILs were isolated from 13 surgically removed primary GBM tumors in our institution. Their responses to autologous tumors were evaluated by IFNγ ELISA. Results: T cells CD8 score by CIBERSORTx was significantly higher in the MGMT-high tumor. Similarly, ssGSEA scores for activated CD8 T cell, Macrophage, activated B cell, and Type 1 T helper cell were significantly higher in the MGMT-high tumor. Conversely, T cells CD4 naive was significantly higher in the MGMT-low tumor. Consistently, tumor-reactive TILs were detected in the MGMT-high tumor. Pathway analysis showed that Rictor was highly enriched in the MGMT-high tumor and Rictor inhibited lymphocyte activation. Coclusion: In this study, we demonstrated that macrophage was highly activated in the MGMT-high tumors. Thus, CSF-1R inhibitor can be combined with immunotherapy in these MGMT-high tumors to enhance anti-tumor immune responses. In addition, TMZ + mTOR2 inhibitors + PD-1 inhibitors may be effective against the MGMT-L group.

DDRE-29. EGFR INHIBITION DOWNREGULATES MGMT AND SENSITIZES GBM CELLS TO TMZ

Glioblastoma (GBM) is a highly malignant type of adult brain tumor with a poor prognosis. Temozolomide (TMZ), a DNA alkylating agent, has been widely used as an effective first-line chemotherapeutic agent for the treatment of GBM patients. The efficacy of TMZ in GBM depends on the absence of the DNA repair protein MGMT which reverses the DNA damage induced by TMZ. The MGMT promoter is hypermethylated in about 45% of GBMs, resulting in lack of MGMT expression and increased responsiveness to TMZ. TMZ is less effective in MGMT unmethylated GBMs. We propose that EGFR inhibition downregulates MGMT and sensitizes glioma cells to TMZ and a combination of pretreatment with erlotinib followed by TMZ could be a useful therapeutic approach in MGMT expressing GBMs. As our experimental model, we used multiple MGMT unmethylated lines from the Mayo Clinic patient derived xenografts (PDXs) panel. Our data demonstrate that exposure of cells to erlotinib for 48h results in downregulation of MGMT at the mRNA and protein level. Additionally, EGFR inhibition activates the AP-1 transcription factor, and overexpression of AP-1 components Fos and Jun results in decreased MGMT expression in TMZ resistant PDXs, suggesting that AP-1 acts as a transcriptional repressor of MGMT. We further identified that the mice implanted with TMZ resistant PDXs pretreated with afatinib followed by TMZ treatment survived longer compared to those treated with TMZ alone. Thus, the use of EGFR inhibition may enhance the sensitivity of MGMT unmethylated GBMs to TMZ.

647 Possibility of immunotherapy for the glioblastoma patients with O6-methyl-guanine DNA methyltransferase (MGMT) expression or promoter unmethylated

BackgroundIt has been widely accepted that O6-methyl-guanine DNA methyltransferase (MGMT) promoter methylation in glioblastoma is associated with a benefit from temozolomide (TMZ) treatment. MGMT is a DNA repair protein that removes the cytotoxic O6-methylguanine (O6MG) DNA lesions generated by TMZ; thereby, MGMT expression is mechanistically linked to TMZ resistance. However, thus far, there is no effective treatment for these patients with MGMT promoter unmethylated. Therefore, a new treatment for GBM patients with MGMT expression is urgently needed.1 2 To this end, we examined the tumor microenvironment in GBM with or without MGMT expression.MethodsBased on The Cancer Genome Atlas (TCGA) primary GBM cohort, the tumor-infiltrating lymphocyte expression level was calculated using the CIBERSORTx algorithms and the single-sample Gene Set Enrichment Analysis (ssGSEA) method. Furthermore, the differential expression gene analysis was conducted and pathway analysis was performed using Ingenuity Pathway Analysis (IPA). The results were validated using the GBM cohort from the Chinese Glioma Genome Atlas (CGGA) database. In addition, tumor-infiltrating lymphocytes (TILs) were isolated from 13 surgically removed primary GBM tumors in our institution. Their responses to autologous tumors were evaluated by IFNγ ELISA.ResultsT cells CD8 score by CIBERSORTx was significantly higher in the MGMT-high tumor. Similarly, ssGSEA scores for activated CD8 T cell, Macrophage, activated B cell, and Type 1 T helper cell were significantly higher in the MGMT-high tumor. Conversely, T cells CD4 naive was significantly higher in the MGMT-low tumor. These results indicate that more immune cell infiltration is associated with MGMT-high tumors. Consistently, tumor-reactive TILs were detected in the MGMT-high tumor. Pathway analysis showed that oxidative phosphorylation (OXPHOS) was highly enriched in the MGMT-high tumor.There were many CD8 T cells and tumor-reactive T cells in the MGMT-high tumors. However, it has been reported that anti-PD-1/PD-L1 monotherapy was not effective in glioblastoma. In this study, we demonstrated that OXPHOS was highly activated in the MGMT-high tumors. Thus, metabolic therapy can be combined with immunotherapy in these MGMT-high tumors to enhance anti-tumor immune responses.ConclusionsAlthough MGMT-high tumors are resistant to TMZ, the existence of immune cell infiltration in the tumor microenvironment of MGMT-high tumors suggest the potential of immunotherapy in these patients.ReferencesStupp R, Hegi ME, Mason WP, van den Bent MJ, Taphoorn MJB, Janzer RC, Ludwin SK, Allgeier A, Fisher B, Belanger K, Hau P, Brandes AA, Gijtenbeek J, Marosi C, Vecht CJ, Mokhtari K, Wesseling P, Villa S, Eisenhauer E, Gorlia T, Weller M, Lacombe D, Cairncross Jy, Mirimanoff R-O, European Organisation for Research and Treatment of Cancer Brain Tumour and Radiation Oncology Groups; National Cancer Institute of Canada Clinical Trials Group. Lancet Oncol 2009;10(5):459–66.Wick W, Weller M, van den Bent M, Sanson M, Weiler M, von Deimling A, Plass C, Hegi M, Platten M, Reifenberger G. Nat Rev Neurol 2014;;10(7):372–85.Ethics ApprovalG3545-(26)ConsentWritten informed consent was obtained from the patient for publication of this abstract and any accompanying images. A copy of the written consent is available for review by the Editor of this journal.

Clinical Efficacy of Temozolomide and Its Predictors in Aggressive Pituitary Tumors and Pituitary Carcinomas: A Systematic Review and Meta-Analysis

Background: A growing number of evidences suggest that TMZ applications can generate impressive benefits for APT and PC patients. However, the definite role of TMZ for individuals remains unclarified due to the variation between studies. And the predictive factors to alter its efficacy remain debatable.Objective: To evaluate the long-term effectiveness and safety profile of TMZ in the treatment of pituitary malignancies, and delineate the predictors during its clinical employment.Results: A literature retrieval was conducted from online databases for studies published up to December 31, 2020. Twenty one studies involving 429 patients were identified. TMZ exhibited 41% radiological overall response rate (rORR). The biochemical response rate was determinate in 53% of the functioning subset. Two-year and 4-year survival rate were 79 and 61%, respectively. TMZ prolonged the median PFS and OS as 20.18 and 40.24 months. TMZ-related adverse events occurred in 19% of patients. Regarding predictors of TMZ response, rORR was dramatically improved in patients with low/intermediate MGMT expression than those with high-MGMT (>50%) (p < 0.001). The benefit of TMZ varied according to functioning subtype of patients, with greater antitumor activities in functioning subgroups and fewer activities in non-functioning sets (p < 0.001). Notably, the concomitant therapy of radiotherapy and TMZ significantly increased the rORR (p = 0.007).Conclusion: TMZ elicits clinical benefits with moderate adverse events in APT and PC patients. MGMT expression and clinical subtype of secreting function might be vital predictors of TMZ efficacy. In the future, the combination of radiotherapy with TMZ may further improve the clinical outcomes than TMZ monotherapy.

MGMT Immunohistochemical Expression in Colorectal Carcinoma and its Correlation with Tumor Progression

Introduction: There is an urgent need to identify predictive features and markers for colorectal carcinoma (CRC) progression and treatment. This study aimed to assess O6-methylguanine DNA methyltransferase (MGMT) expression in CRC and correlate with the clinico-pathological aspects of the tumor, also to evaluate the relationship between different histopathologic parameters and tumor progression. Material and Methods: The study was carried on 70 colectomy using formalin fixed paraffin embedded tumor tissue. Immunohistochemistry was used to detect MGMT expression, and clinico-pathologic aspects as well as Tumor budding, type of desmoplastic reaction, inflammatory lymphocytic milieu, pattern of invasive front and necrosis were assessed Then correlated with MGMT expression and tumor progression, using parametric and nonparametric statistical methods. Results: MGMT Loss of expression was detected in 42.9% of CRC cases. MGMT expression status was significantly correlated with tumor stage and metastatic status (p<0.05), while it was not correlated with other clinic-pathologic features, (p>0.05). Desmoplastic reaction (DR), tumor budding, stromal tumor infiltrating lymphocytes (TIL-S) and necrosis were correlated with tumor stage (p<0.05). DR correlated with tumor budding (p<0.05). Both types of TIL and Crohn’s-like lymphoid reaction (CLR) showed a mutual correlation (p<0.05). Conclusion: MGMT high expression and histopathologic parameters as DR, tumor budding, inflammatory lymphocytic milieu and necrosis could be correlated with CRC progression.

Alkylating agent rechallenge in metastatic pancreatic neuroendocrine tumours

A rechallenge is common after initial efficacy of alkylating-based chemotherapy (ALK) in pancreatic neuroendocrine tumors (PanNET). High MGMT expression seems associated with lower response to ALK. We aimed to evaluate the efficacy and toxicity of ALK rechallenge in PanNET. Secondly, to assess the evolution of MGMT expression under ALK. All consecutive patients with advanced PanNETs who received initial ALK (achieving tumor control) followed by a pause > 3 months, then an ALK rechallenge (ALK2) upon progression were retrospectively studied (Cohort A). Primary endpoint was progression-free survival under ALK2 (PFS2). The MGMT expression was retrospectively assessed by immunohistochemistry (H-score) in consecutive PanNET surgically resected following ALK (Cohort B). We foud that cohort A included 62 patients (median Ki67 8%), for which ALK1 followed by pause achieved an objective response rate of 55%, and a PFS1 of 23.7 months (95% IC, 19.8-27.6). ALK2 achieved no objective response, and stability in 62% of patients. The median PFS2 was 9.2 months (IC 95% 7.1-11.3). At multivariable analysis, a hormonal syndrome (p=0.032) and a pause longer than 12 months (p=0.041) were associated with a longer PFS2. In the cohort B (17 patients), the median MGMT H-score increased from 45 (IQR 18-105) before ALK, to 100 (IQR 56-180) after ALK (p=0.003). We conclude that after initial efficacy of ALK treatment, a pause followed by ALK rechallenge might be appropriate to prolong tumor control, improve quality of life and limit long-term adverse events. Increased MGMT expression under ALK might explain low efficacy of ALK rechallenge.