Tumor Growth Rate Research Articles

Abstract 3608: MLL1 regulates cytokine-driven cell migration and metastasis

Abstract Cell migration, particularly in three-dimensional (3D) environments, is a critical requirement for metastasis - the spread of cancer cells from a primary tumor to distant sites. Despite causing the vast majority of cancer-related deaths, targeting metastasis remains challenging clinically. Cancer cell migration can be triggered and maintained by elevated levels of cytokines, which are presumed to be predominantly secreted by immune cells in the tumor microenvironment. However, recent work has highlighted the importance and impact of cytokine production by cancer cells themselves. While the signaling cascades that regulate cell migration have been studied extensively, the epigenetic regulation of cell migration is still poorly understood. MLL1 is a histone methyltransferase, which along with its scaffold protein Menin, is integral to methylation at the histone 3 lysine 4 (H3K4) site. Here, we show that the MLL1-Menin interaction is essential for the migration of cancer cells and that the disruption of this interaction impairs cell migration and metastasis. MLL1 depletion reduces 3D cell migration in vitro and led to lesser metastatic burden and prolonged survival in vivo. Reduced lung metastatic burden was observed even after accounting for the difference in the growth rate of primary tumors. Mechanistically, MLL1-Menin interaction controls actin filament assembly via the IL-6/pSTAT3/Arp3 axis and acto-myosin contractility via the TGF-β1/Gli2/ROCK1/2/pMLC2 axis, which together regulate dynamic protrusion generation and 3D cell migration. MLL1-Menin inhibition also decreases cell proliferation by inhibiting mitosis and cell cycle-related pathways. MLL1 depletion decreases primary tumor growth, metastatic spread, and metastatic outgrowth in mouse models of triple-negative breast cancer (TNBC), thus improving survival. When comparing metastatic burden at a threshold tumor size, rather than at a timepoint, MLL1 depletion still lowered metastatic burden. Combining an MLL1-Menin inhibitor with Paclitaxel, a standard TNBC chemotherapeutic, abrogated tumor growth and metastasis in a syngeneic TNBC mouse model. Overall, our studies show that MLL1 is an epigenetic regulator of 3D cancer cell migration, which exerts its effects by controlling the production of the key cytokines IL-6 and TGF-β1. Our in vivo studies demonstrate the potential of targeting the MLL1-Menin interaction in metastasis prevention and its potential to be combined with currently administered chemotherapeutics. Citation Format: Praful R. Nair, Ludmila Danilova, Estibaliz Gómez-de-Mariscal, Dongjoo Kim, Rong Fan, Arrate Muñoz-Barrutia, Elana Fertig, Denis Wirtz. MLL1 regulates cytokine-driven cell migration and metastasis. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3608.

Abstract 5164: Liposomal Delivery of an Immunostimulatory CpG Induces Robust Antitumor Immunity and Long-Term Immune Memory by Reprogramming Tumor-Associated Macrophages

Abstract Background: The efficacy of current T cell-based immunotherapy is known to be limited by various factors that induce immune suppression in the tumor microenvironment (TME). Tumor-associated macrophages (TAMs) are representative TME-comprising immune suppressive cells and influence tumor growth and progression. TAMs are considered as an attractive target for cancer immunotherapy. However, current TAMs-targeting strategies are not sufficient to induce continuous antitumor responses. Here, we suggest a new strategy that can reprogram TAMs and further bridge innate-to-adaptive immunity by utilizing immunostimulatory CpG oligodeoxynucleotides (ODNs). In this study, we investigated the new role of TLR9 agonist CpG ODN as an immunotherapeutic agent and tested whether liposomal delivery of CpG could modulate the function of TAMs and lead to remodeling of the TME. Methods: A liposome-based nanomaterial encapsulating CpG ODN was constructed. A small lipid nanoparticle (SLNP) used as the CpG delivery platform in this study was made of a cationic cholesterol derivative and two biocompatible phospholipids. The CpG-encapsulating SLNP (SLNP@CpG) was prepared by a thin-film formation and rehydration method. In vitro studies were conducted to assess whether SLNP@CpG could reprogram macrophages. To assess antitumor therapeutic efficacy of the SLNP@CpG, two subcutaneous tumor models were established in mice and tumor growth and survival rates of mice were monitored. For tumor rechallenge experiment, mice showing complete tumor regression were rechallenged with same cancer cells and their splenocytes were analyzed to confirm the development of immune memory. To elucidate the immunological mechanism of SLNP@CpG-mediated antitumor efficacy, changes of immune cell subpopulations in treated tumors were analyzed by flow cytometry. Results: SLNP@CpG enhanced macrophage-mediated phagocytosis of cancer cells and tumor antigen cross-presentation, and skewed the polarization state of macrophages in vitro. Intratumorally injected SLNP@CpG exerted its therapeutic efficacy in an established E.G7-OVA tumor via uptake by TAMs. SLNP@CpG treatment significantly suppressed the E.G7 tumor growth and also considerably prolonged the survival of mice, with 83.3% of mice becoming tumor-free. Local administration of SLNP@CpG resisted E.G7 tumor rechallenge by inducing immunological memory and long-term antitumor immunity. Local administration of SLNP@CpG even exerted its antitumor efficacy in an aggressive B16-F10 melanoma by remodeling TME towards immune stimulation and tumor elimination. Conclusion: Liposomal delivery of CpG via local treatment reprogrammed TAMs by enhancing phagocytic activity and repolarizing M2 to M1 phenotype and also reshaped immunosuppressive TME, leading to antitumor immunity and long-term memory responses. Citation Format: Yujin Kim, Sangyong Jon. Liposomal Delivery of an Immunostimulatory CpG Induces Robust Antitumor Immunity and Long-Term Immune Memory by Reprogramming Tumor-Associated Macrophages. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5164.

Abstract 5165: In vivo therapeutic validation of NRT-YHD_001, a novel macrophage checkpoint inhibitor, in liver cancer

Abstract Previously, we demonstrated that recovery of Histone deacetylase 6 (HDAC6) elicited let-7i-5p suppression to de-repress the thrombospondin-1 (TSP1) expression, and thereby it occupied CD47 receptor to block CD47-SIRPα mediated anti-phagocytosis of macrophage in liver cancer. NRT-YHD_001 is a modified antisense miRNA of let-7i-5p for liver cancer treatment. NRT-YHD_001 was substituted with O-methoxyethyl and phosphothioate for stability, and N-acetylgalactosamine was attached for specific delivery to the liver. In in vitro phagocytosis assay, NRT-YHD_001 showed macrophage phagocytosis activity with similar or better results compared to non-modified antisense let-7i-5p (AS_let-7i-5p). In addition, it was confirmed that NRT-YHD_001 inhibited endogenous let-7i-5p of liver cancer cells and activated macrophage phagocytosis at 4°C or 25°C for up to 2 weeks. When NRT-YHD_001 activity was investigated in vivo after 1 week, 2 weeks, and 4 weeks after administration of single dose NRT-YHD_001 to the tail vein of a mouse, NRT-YHD_001 was not degraded, and let-7i-5p was inhibited while maintaining activity in liver tissue for 2 weeks after injection. To investigate the in vivo therapeutic effect, a mouse spontaneous (Ras-transgenic) liver cancer model was used and compared with the sorafenib-administered group. After administration of NRT-YHD_001 and sorafenib each week at the time when liver mass was detected, the tumor size and growth rate were compared. As a result, the NRT-YHD_001 administration group showed significant therapeutic effect compared to sorafenib. These results showed that NRT-YHD_001 had a strong anticancer effect by converting the macrophage of don't eat me into eat me signal in the tumor microenvironment of liver cancer, thereby removing or inhibiting liver cancer cells. Keywords: let-7i-5p, macrophage, phagocytosis, liver cancer, NRT-YHD_001 Citation Format: Suk Woo Nam, Sangyean Kim, Min Jeong Na, Eunbi Shin, Jin Woong Ha, Soyoung Jeon, Jeong Hwan Yoon. In vivo therapeutic validation of NRT-YHD_001, a novel macrophage checkpoint inhibitor, in liver cancer. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5165.

Abstract 2486: Dissecting the lineage specific roles of BCL-XL in breast cancer metastasis

Abstract B-cell lymphoma X-large (BCL-XL) is a member of the BCL-2 protein family and has been known as an anti-apoptotic protein. As a viable target for cancer therapy, we recently found that proteolysis-targeting chimeras (PROTAC, currently under phase I clinical investigation) against BCL-XL kill cancer cells as well as tumor-induced regulatory T cells (TI-Tregs). Interestingly, BCL-XL has been shown to promote breast cancer metastasis via a function independent of its anti-apoptotic function. We reason that BCL-XL-targeting PROTACs can be potentially used to treat metastatic breast cancer - by direct killing of circulating tumor cells at their vulnerable stages and/or by eliciting anti-cancer immunity via T-reg cell depletion. To our surprise, our pilot experiments using two syngeneic breast cancer models showed opposite results with one model showing reduced metastasis by BCL-XL PROTAC treatment and the other model showing increased metastasis. Genetic deletion of BCL-XL (KO) in those two cell lines reproduces the effect of PROTAC on metastasis, suggesting the on-targeting but contrasting function of BCL-XL in breast cancer metastasis. Our central hypothesis is that BCL-XL may play different roles in breast cancer metastasis depending on the genetics/proteomics of cellular cues from different cancer cells. To confirm the function of BCL-XL in different breast cancer cell lines, we generated BCL-XL KO lines from 4 other syngeneic mouse breast cancers. We confirmed that the majority of breast cancer cells depend on BCL-XL for metastasis, including 4T1, AT-3, E0771.lmb etc; whereas only the Py8119 model exhibits an opposite phenotype. For primary tumor growth, AT-3 and E0771.lmb cells grow bigger tumors in the WT group than in the BCL-XL KO group; and 4T1 and Py8119 cells exhibit similar tumor growth rate between WT and KO cells. For metastatic breast cancer therapy, we treated 4T1-tumor bearing mice with BCL-XL-targeting PROTAC that nearly significantly reduces metastatic spread in the lung without significant change in primary tumor growth. In addition, we found a systematic inhibition of TI-Tregs by PROTACs indicating a lineage-specific function of BCL-XL in breast cancer metastasis. We conclude that BCL-XL is a viable therapeutic target for treating breast cancer metastasis, but caution is required as BCL-XL inhibition may be detrimental for certain cancers. Our future effort will focus on the mechanisms explaining why BCL-XL inhibits metastasis in some cancer types, which can be used as an exclusion criterion for BCL-XL-targeting therapy. Citation Format: Madison E. Carelock, Mo Jiao, Weizhou Zhang. Dissecting the lineage specific roles of BCL-XL in breast cancer metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2486.

Abstract 5189: Comparisons of Rituximab and CD19xCD3 BiTE antitumor response across multiple humanized NSG strains engrafted with human hematopoietic stem cells

Abstract Humanized NSG mice engrafted with hematopoietic stem cells (HSCs) is a well-known in vivo platform for reconstitution of human immune system in a mouse host. In this study we tried to dissect the role of various human cytokines in the generation of the human immune system, expansion of human B cell lymphoma cells, and drug response to two anti-cancer therapies - Rituximab and CD19XCD3 BiTE (Bispecific T-cell Engager). To exclude donor variabilities, we first humanized five NSG-based, human cytokine-transgenic mouse strains (NSG, NSG-IL15, NSG-SGM3, NSG-SGM3-IL15, and NSG-FLT3L) with HSCs from one donor (0084). We found that tumor growth was enhanced in NSG-SGM3 mice when compared to NSG mice. In contrast, IL15 expression suppressed tumor growth in NSG-IL15 and NSG-SGM3-IL15 mice. Furthermore, NK cell frequencies in blood are higher in NSG-IL15 and NSG-SGM3-IL15 than in other strains. suggesting that NK cell-mediated cytotoxicity is involved in suppression of tumor cell growth in the strains expressing human IL-15. Rituximab treatment resulted in tumor growth inhibition and significant B cell depletions across NSG-IL15, NSG-SGM3, and NSG-SGM3-IL15 mice engrafted with 0084 PBMC. We also observed a donor variability of tumor growth rates in NSG-SGM3-IL15 mice engrafted with donors 0084, 0662A, and 0733. The tumor growth in NSG-SGM3-IL15 mice engrafted with 0084 PBMC was significantly slower than the one in mice engrafted with the other two donors (0084 vs. 0662A, P=0.002; 0084 vs. 0733, P=0.004 on day 21); interestingly, this strain of mice exhibited higher frequencies of CD8+ T cells and lower frequencies of NK cells. Next, we tested CD19XCD3 BiTE treatment in multiple NSG strains engrafted with HSCs from different donors. The BiTE administration inhibited Raji tumor growth and significantly depleted B cells in all tested strains as NSG-SGM3-IL15 showed highest frequencies of NK cells and CD8+ T cells. The combination of Rituximab and BiTE showed even better tumor inhibition efficacy. In conclusion, regardless of the donor and strain variations in tumor growth and human immune cell subset development, the administrations of rituximab or BiTE significantly and consistently inhibited tumor growth and depleted peripheral B cells across all mouse strains. These results indicate that utilizing a comprehensive humanized mouse platform could drive more impactful conclusions in terms of evaluating the anti-tumor efficacy of immune therapies. Citation Format: Won Yeong Kang, Guoxiang Yang, Ilian Radichev, Li-Chin Yao, Mingshan Cheng, James G. Keck. Comparisons of Rituximab and CD19xCD3 BiTE antitumor response across multiple humanized NSG strains engrafted with human hematopoietic stem cells. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5189.

Abstract 2741: Sexual dimorphism induces differential immunomodulatory outcomes from local nanoparticle and focused ultrasound therapy in murine model of head and neck cancer

Abstract Nanoparticles (NPs) and therapeutic devices are increasingly being investigated to re-engineer poorly immunogenic tumors into activated ′hot′ targets. Innate & adaptive immunological responses in females are known to be relatively superior to males. However, the development of NPs and therapeutic devices generally lacks attention to the impact of sex on therapeutic outcomes. Herein, we evaluated the role of sexual dimorphism in the immune-mediated remission of murine head & neck (MOC2) tumors following in-situ therapy with immunogenic cell death-enhancing calreticulin nanoparticle (CRT-NP) and focused ultrasound-based histotripsy (HT). CRT-NP was synthesized with DOTAP and cholesterol lipids using the thin film and extrusion methodology and loaded with a full-length clone CRT-expressing vector. CRT-NP was characterized for size and Ζ potential (ZP) using dynamic light scattering. To assess in vivo efficacy, male & female mice bearing MOC2 tumor xenografts were randomized into the following groups (n=6mice/group)¦ 1) Control, 2) CRT-NP, & 3) HT. Three intratumoral CRT-NP injections (20µg CRT-DNA/injection) were given 2 days apart 7-days post-inoculation. For comparing gender-mediated tumor immunomodulation, two local HT treatments were similarly administered in male & female MOC2 cohorts. Mice were sacrificed on day 20-post inoculation & the tumors, tumor-draining lymph nodes, spleen, & serum were processed for assessing inflammatory signatures using flow cytometry, & cytokine analysis. CRT-NPs demonstrated a hydrodynamic diameter of ≈260nm, ΖP of +17mv & a PDI ≤0.3. Compared to the untreated controls that showed a pronounced increase in tumor volumes, male & female mice treated with CRT-NP showed a ≈65% reduction in tumor growth post-treatment. HT treatment of female mice similarly suppressed the MOC2 tumor growth rates ≈64% vs. corresponding untreated control, however, it was ineffective in inducing remission of male mice tumors. The reduced HT efficacy in male mice correlated with increased infiltration of immunosuppressive NK+PD1+ cells & CD11b+PDL1+ macrophage in tumors. In contrast, the improved efficacy in female mice with both CRT and HT treatments associated with increased infiltration of CD4+ cells, CD8+ cells, NK cells, macrophage (CD11b+CD86+MHCII+), & a reduction in the M-MDSC population in the treated tumors. An additional correlation of local immune cell immunomodulation with systemic serum cytokine responses is currently in the works. CRT-NP was effective in both male & female mice. HT-induced tumor remission in females, but male mice demonstrate immunoresistance. These suggest that immunomodulatory therapies should account for sexual dimorphism as a variable. A higher accumulation of immunosuppressive cells in male TME can impact the anti-tumor immunity of immunomodulatory therapies in preclinical and clinical settings. Citation Format: Sri Vidhya Chandrasekar, Akansha Singh, Ashish Ranjan. Sexual dimorphism induces differential immunomodulatory outcomes from local nanoparticle and focused ultrasound therapy in murine model of head and neck cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2741.

Abstract 4780: Comparison of safety and efficacy outcomes with nonthermal plasma and tirapazamine in porcine skin and mouse melanoma models

Abstract A novel preclinical experimental therapy for various cancer types including melanoma is the combination of nonthermal plasma (NTP) and tirapazamine (TPZ) delivered in situ. This patented combination therapy delivers reactive oxygen species (ROS) and reactive nitrogen species (RNS) from the atmosphere that are activated and ionized with helium gas to create the state of matter known as plasma. The combination with the prodrug, TPZ, that only activates to become a ROS in hypoxic environments, shows synergistic targeting of cancer cells. In this study, we assayed the effects of the dual therapy on porcine skin, which is the closest model to represent human skin and is commonly used for testing drug therapy. The results demonstrated that porcine skin showed no significant cell killing with the therapy. We also compared the combination therapy in normal skin keratinocytes and 1205Lu cells derived from a melanoma metastasis to the lung. The cells were treated with NTP and TPZ and maintained in a hypoxic chamber. Four days following therapy the keratinocytes continued to grow in culture, whereas the melanoma cells expressing endogenous gap junctions, or overexpressed Cx43 gap junctions showed decreased viability. However, the melanoma cells with a dominant negative Cx43 mutant did not show this loss in viability. This confirms our previous report that gap junctions mediate the passage of derivatives of ROS to increase the extent of cell death. To determine the effects of NTP on regulating the lethal dose of TPZ, we performed IC50 curves for TPZ in the presence or absence of NTP in B16-F10 murine metastatic melanoma cells. We demonstrated a decrease in the IC50 from 28 μM without NTP to 17.4 μM with NTP. In further analysis, the effects of treating with NTP in the presence or absence of tissue culture media were compared. The results showed a highly significant increased efficacy of NTP in the absence of media. The combination therapy was more effective than each independent treatment. To further confirm the experimental analysis, we induced tumor formation with the B16-F10 cells in a syngeneic C57BL/6 mouse model. The tumors from mice that were untreated or treated with NTP or TPZ alone, showed a significant increase in tumor volume as compared to the tumors from mice receiving the combination treatment in situ. In comparison of the tumor growth rates, the control, NTP-, or TPZ-treated tumors had increased by approximately 350% by day 10, whereas the tumors in the mice that received the dual therapy only increased by an average of 70%. The results from these studies collectively implicate the combination of NTP and TPZ as a potential synergistic treatment for in situ melanomas that offer selectivity as compared to normal tissue. Citation Format: Matthew Yehl, Dominik Kucharski, Michelle Eubank, Xavier Ramadan, Ahmad Dianat, Bianca Pham, Timothy C. Hutcherson, Sandra Sexton, Shoshanna N. Zucker. Comparison of safety and efficacy outcomes with nonthermal plasma and tirapazamine in porcine skin and mouse melanoma models. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4780.

Abstract 2504: Antiarrhythmic drugs repositioning to counteract the activated phenotype of prostate cancer-associated fibroblasts

Abstract The interplay between cancer cells and adjacent stroma is fundamental to sustain carcinogenesis and promote the acquisition of additional malignant features. Cancer-associated fibroblasts (CAFs) have been shown to fuel prostate cancer (PCa) development and metastasis by mutually interacting with tumor cells. Comparative gene expression profile analysis of CAFs and normal fibroblasts, showed that CAFs are characterized by a positive enrichment of gene sets that regulate cytoskeleton remodeling and muscle contractility, including genes coding for calcium, potassium and sodium channels. Here, for the first time, we investigated the effects of cation channel inhibitors, currently used as antiarrhythmic drugs, on CAF-activated state and CAF-PCa cell interplay. Our results showed that antiarrhythmic drugs are able to interfere with crucial features of reactive prostate CAFs, as indicated by changes in cell morphology and modulation of specific fibroblast activation markers, such as α-SMA, fibroblast activation protein and collagen type I. In addition, although to a variable extent, antiarrhythmics reduced CAF motility and hindered CAF capability to remodel extracellular matrix, as assessed by 3D gel contraction and scratch assays. Interestingly, conditioned medium and co-culture experiments reveled that CAFs were able to promote cell growth and induce epithelial to mesenchymal transition (EMT) in PCa cells, as indicated by E-cadherin and β-catenin down-regulation and Snail and Vimentin up-regulation. Interestingly, antiarrhythmics partially reverted CAF-mediated protumoral effects, abrogating their ability to foster PCa cell proliferation and plasticity. Moreover, based on previous evidence indicating the involvement of matrix metalloproteinase (MMP) in CAF-induced EMT on PCa cells, we found that antiarrhythmics impaired the secretory phenotype of CAFs, as indicated by the reduction of MMP-2 releasing in the medium. Finally, gene expression analysis highlighted that antiarrhythmic-treated CAFs showed a gene expression pattern resembling that of normal fibroblasts. Consistent with our in vitro findings, in vivo experiments confirmed that intra-tumoral administration of medium form antiarrhythmic-treated CAF reduced tumor growth rate of PCa xenografts compared to medium form untreated CAFs. Overall, cation channels inhibition interferes with crucial features of prostate CAFs, affecting their activated state. Moreover, co-culture experiments indicate that antiarrhythmics are able to impair CAF-induced proliferative spur and EMT in PCa cells, and also induce tumor growth delay in vivo. Collectively, such data suggest new opportunities for the therapeutic repositioning of antiarrhythmic drugs with the aim of targeting CAFs and hence interrupting their supportive boost in PCa. Citation Format: Valentina Doldi, Monica Tortoreto, Stefano Percio, Paolo Gandellini, Nadia Zaffaroni. Antiarrhythmic drugs repositioning to counteract the activated phenotype of prostate cancer-associated fibroblasts [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2504.

Abstract 619: TP53 gain-of-function mutation modulates the immunosuppressive microenvironment in non-HPV associated oral squamous cell carcinoma

Abstract Background: TP53 is the most common mutated gene in head and neck cancer, including oral squamous cell carcinoma (OSCC), a lethal disease with limited therapeutic options. In OSCC, for example, TP53 R175H is a hot-spot, high-risk, functional mutation that inhibits the function of WT p53, however these correlations and functional roles of high-risk p53 mutations had not been experimentally addressed using syngeneic oral tumor models. To understand the functional role of TP53 mutations in modulating the tumor immune microenvironment (TIME) and response to immunotherapy, we used a syngeneic oral tumor model (ROC3 cell) derived from a genetically engineered mouse model expressing the p53 R172H (human, R175H). Methods: Orthotopic tongue injections were performed to (i) characterize tumor growth rate (ii) the TIME landscape by flow cytometry and opal multiplex immunofluorescence (mIF); and (iii) immunotherapy drug studies. We inactivated mutant p53 expression by shRNA and CRISPR Cas9, profiled gene expression changes by HTG EdgeSeq array, and validated these findings by qPCR, Western blot and immunochemistry. Tumor response was achieved by anti-PD1 and STING agonist therapies. Patients’ gene expression analyzed by CIBERSORT and TIME landscape performed by mIF. The spatial distribution of TIME was evaluated by Visiopharm software and UMAP. Results: We demonstrated that disruption of mutant p53R172H in ROC3 cells altered cytokine and chemokine expression levels and consequently affected the maturation and infiltration of different immune subsets, including cytotoxic CD8+ T cells, helper CD4+ T cells, Tregs, G-MDSCs, tumor-associated macrophages and neutrophils in ROC3 tumors. In addition, our spatial TIME analysis revealed that p53R172H mutation promoted the infiltration of Tregs, M2 macrophages, and exhausted CD8+ T cells, and excluded active CD8+ and CD11+ cells in both the tumor cores and on the tumor edges. Interestingly, it seems that p53R172H also regulates the spatial distribution of immunocyte populations, and their distribution between central and peripheral intratumoral locations. Moreover, we found that p53R172H mutation not only disrupted the phosphorylation of STING pathway which supported the recently reported finding, but also decreased the absolute amount of STING protein. Furthermore, we detected that p53-mutated tumor-infiltrating CD8+ and CD4+ T cells expressed programmed cell death protein 1 (PD-1), and that these tumors responded to anti-PD-1 and STING agonist therapy. Furthermore, Human OSCC samples also revealed associations between immune cell populations and the TP53R175H mutation, which paralleled the findings from our ROC3 tumor model. Conclusions: Our findings collectively demonstrate that the mutant TP53 GOF mutation modulates the tumor immune landscape and is associated with reduced overall survival in patients with OSCC. Citation Format: Yewen Shi, Xiaoyong Ren, Shaolong Cao, Tongxin Xie, Xi Chen, Chieko Michikawa, Abdullah A. Osman, Andrew G. Sikora, Jeffrey N. Myers, Roberto Rangel. TP53 gain-of-function mutation modulates the immunosuppressive microenvironment in non-HPV associated oral squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 619.

Baseline tumor growth rate highlights the heterogeneity of well differentiated gastroenteropancreatic neuroendocrine tumors and predicts for increases in Ki67 index over time.

Refined risk stratification for gastroenteropancreatic neuroendocrine tumors (GEP-NETs) has the potential to improve comparisons of study populations across clinical trials and facilitate drug development. Tumor growth rate (TGR) is a radiological metric with demonstrated prognostic value in well differentiated grade 1 and 2 (G1-2) GEP-NETs, but little is known about TGR in G3 NETs. In this retrospective study of 48 patients with advanced G1-3 GEP-NET, we calculated baseline TGR (TGR0 ) from radiological images of metastases acquired prior to first-line therapy and evaluated its association with disease characteristics and outcomes. The median pretreatment Ki67 proliferation index for G1-3 tumors combined was 5% (range = 0.1%-52%) and median TGR0 was 4.8%/month (m) (range = 0%-45.9%/m). TGR0 correlated with pretreatment Ki67 across G1-3 pooled and within G3 GEP-NET. Patients with higher TGR0 (>11.7%/m) tumors, which were primarily G3 pancreatic NETs, exhibited decreased time to first therapy (median, 2.2 vs. 5.3 months; p = .03) and shorter overall survival (median, 4.1 years vs. not reached; p = .003). Independent of therapies given, higher TGR0 GEP-NETs experienced a greater incidence of Ki67 increase (100 vs. 50%; p = .02) and greater magnitude of Ki67 change (median, 14.0 vs. 0.1%; p = .04) upon serial biopsy. Importantly, TGR0 , but not grade, predicted for future Ki67 increase in this series. Given the heterogeneity of well differentiated GEP-NETs, future clinical trials may benefit from stratification for TGR0 , particularly in G1-2 tumors, in which TGR0 does not correlate with Ki67. TGR0 has the potential to noninvasively identify patients with previously undiagnosed grade progression and those in whom more or less frequent monitoring may be appropriate. Additional research is needed to determine the prognostic and predictive value of TGR0 in larger and more homogeneously treated cohorts, and to ascertain if post-treatment TGR has value in previously treated patients starting a new line of therapy.

MP41-19 BILATERAL MULTIFOCAL RENAL ONCOCYTOMA: CLINICAL CHARACTERISTICS AND OUTCOMES OF MANAGEMENT

You have accessJournal of UrologyCME1 Apr 2023MP41-19 BILATERAL MULTIFOCAL RENAL ONCOCYTOMA: CLINICAL CHARACTERISTICS AND OUTCOMES OF MANAGEMENT Shiva Singh, Aditi Chaurasia, Nikhil Gopal, Noah Pinson, W. Marston Linehan, Ashkan Malayeri, and Mark Ball Shiva SinghShiva Singh More articles by this author , Aditi ChaurasiaAditi Chaurasia More articles by this author , Nikhil GopalNikhil Gopal More articles by this author , Noah PinsonNoah Pinson More articles by this author , W. Marston LinehanW. Marston Linehan More articles by this author , Ashkan MalayeriAshkan Malayeri More articles by this author , and Mark BallMark Ball More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003279.19AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Renal oncocytomas are benign neoplasms, accounting for 3-7% of renal tumors. A select number of patients can present with bilateral and/or multifocal oncocytomas and negative germline testing. There is limited data on the clinical course of this subset of patients and optimum treatment strategy remains unclear. This study seeks to evaluate the renal functional and oncological outcomes in patients with bilateral multifocal renal oncocytomas. METHODS: We retrospectively reviewed a cohort of 64 patients with histopathologically confirmed bilateral multifocal renal oncocytomas with negative genetic testing who underwent active surveillance, partial or radical nephrectomy at our institution. Demographics, oncologic outcomes, renal function change and tumor growth rate using 2-dimensional measurements on cross-sectional imaging were assessed. Renal function outcomes were assessed using serum creatinine levels within and between two groups (based on management strategy) with Wilcoxon signed ranked test and Mann-Whitney test respectively. RESULTS: A total of 64 patients (51 males, 13 females) with a median age of diagnosis of 64 years and mean follow-up of 5.7 years were studied. 19 patients underwent only active surveillance, and 45 patients underwent at least one surgical intervention during the study period. Median age of first procedure was 61.5 years. Interventions included 2 renal ablation, 52 partial, and 13 radical nephrectomies. The median growth rate was 0.21 cm/year (0.16-0.27 cm/year), with a new tumor diagnosed on imaging every five years. The median change in serum creatinine was 0.13 mg/dL on active surveillance and 0.12 mg/dL after surgery (p=0.6). No metastases were noted in this cohort throughout the study period. The median age of death was 82.4 years. CONCLUSIONS: Bilateral multifocal renal oncocytomas with negative genetic workup are benign tumors with an indolent course and are seen primarily in 6th decade of life. Considering a minimal difference in functional outcomes between active surveillance and surgical management, surgeons may consider the additional clinical characteristics of patients to design the individual management plan. Source of Funding: This research was supported, in part, by the Intramural Research Program of the National Institutes of Health Clinical Center. © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e564 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Shiva Singh More articles by this author Aditi Chaurasia More articles by this author Nikhil Gopal More articles by this author Noah Pinson More articles by this author W. Marston Linehan More articles by this author Ashkan Malayeri More articles by this author Mark Ball More articles by this author Expand All Advertisement PDF downloadLoading ...

Development's Inaugural Outstanding Paper Prize.

Development's Inaugural Outstanding Paper Prize.

MP41-18 CLINICAL MANIFESTATIONS OF BIRT-HOGG-DUBE SYNDROME IN A LARGE CLINICAL COHORT

You have accessJournal of UrologyCME1 Apr 2023MP41-18 CLINICAL MANIFESTATIONS OF BIRT-HOGG-DUBE SYNDROME IN A LARGE CLINICAL COHORT Brian Cortese, Raju Chelluri, David Ostrowski, Daniel Roberson, Phillip Pierorazio, and Katherine Nathanson Brian CorteseBrian Cortese More articles by this author , Raju ChelluriRaju Chelluri More articles by this author , David OstrowskiDavid Ostrowski More articles by this author , Daniel RobersonDaniel Roberson More articles by this author , Phillip PierorazioPhillip Pierorazio More articles by this author , and Katherine NathansonKatherine Nathanson More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003279.18AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Birt-Hogg-Dube (BHD) syndrome is an underdiagnosed autosomal dominant condition caused by folliculin (FLCN) gene mutations and is characterized by fibrofolliculomas, pulmonary cysts, and renal masses. Our study characterizes our center’s cohort of BHD patients to describe the range of presentations. METHODS: A single center, retrospective cohort study of patients with BHD syndrome was performed. The medical record was queried for clinical data corresponding to renal mass, pulmonary, and dermatologic findings. Results were reported as median and inter-quartile range (IQR). Germline mutation status is reported. RESULTS: Eighty-one BHD patients were identified. Median age of BHD diagnosis was 38 [28-57] years old. Ten (12.3%) patients had a total of 15 renal masses. Age at first tumor diagnosis was 58 [50.8-60.2] years with median lesion size being 1.5 [1-2.1] cm. Six (60%) tumor patients were managed non-operatively and 4 (40%) underwent resection. Surveilled tumor growth rate was 0 [-0.02 – 0] cm/yr during a median 2.3 [2.3-5.6] year follow-up. Twenty-six (32.1%) patients had characteristic skin findings at diagnosis with fibrofolliculomas being documented in 47 (58%) total patients at a median age of 46.5 [33-58] years. Twenty-eight (34.6%) patients had a history of pneumothorax at diagnosis with pneumothorax affecting 34 (42%) total patients. Notably, 9/47 (19.1%) patients with fibrofolliculomas and 4/34 (11.8%) patients with pneumothorax history had a renal mass. Four (40%) renal tumor patients had pneumothorax history. CONCLUSIONS: Renal tumors were found to affect 12.3% of BHD patients in this clinical cohort. BHD-associated renal masses were noted to mostly small, indolent with a static growth rate. Only 19% patients with fibrofolliculomas and 12% of patients with pneumothorax developed a renal mass. Of patients with a renal mass, 40% experienced a pneumothorax in their lifetime. Limitations of this study include its retrospective nature, small sample size of BHD patients with renal lesions, and low follow-up time. Source of Funding: None © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e563 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Brian Cortese More articles by this author Raju Chelluri More articles by this author David Ostrowski More articles by this author Daniel Roberson More articles by this author Phillip Pierorazio More articles by this author Katherine Nathanson More articles by this author Expand All Advertisement PDF downloadLoading ...

Renal cell carcinoma management: real-world practice and challenges at a national level.

Renal cell carcinoma (RCC) management has seen a revolution over the last decades. Six Lebanese oncologists discussed recent updates in RCC management and outlined the challenges and future directions in Lebanon. Sunitinib continues to be a first-line choice for metastatic RCC in Lebanon, except for intermediate- and poor-risk patients. Immunotherapy is not always accessible to patients or selected routinely as first-line therapy. More data are needed on the sequencing of immunotherapy and tyrosine kinase inhibitor treatments and on the use of immunotherapy beyond progression and/or after failure of immunotherapy in the first-line setting. For second-line management, the clinical experience with axitinib for low tumor growth rate and nivolumab after progression on tyrosine kinase inhibitors make those two agents the most widely used. Several challenges affect the Lebanese practice, limiting the accessibility and availability of the medications. Reimbursement remains the most critical challenge, especially with the socioeconomic crisis of October 2019.

Hyperprogressive disease during atezolizumab plus bevacizumab treatment in patients with advanced hepatocellular carcinoma from Japanese real-world practice

BackgroundHyperprogressive disease (HPD) is a phenomenon with greatly accelerated tumor growth and clinical deterioration rates compared to pre-therapy, in patients treated with immune checkpoint inhibitors (ICI). The aim of this study is to clarify the reality of HPD in patients with advanced hepatocellular carcinoma (HCC) who were treated with atezolizumab plus bevacizumab (Atez/Bev) using tumor dynamics.MethodsMedical records of consecutive patients with advanced HCC who were treated with Atez/Bev were retrospectively reviewed. HPD was defined as a more than two- or fourfold increase in tumor growth rate (TGR) or tumor growth kinetics rate (TGKR) before and after treatment. Overall survival (OS) and baseline characteristics with or without HPD were analyzed.ResultsA total of 85 patients were included in the analysis. When HPD was defined as a twofold of TGR or TGKR, 8 patients (8/85, 9.4%) had HPD and 11 had PD without HPD. A total of 5 patients (5/85, 5.9%) were diagnosed with HPD and 14 with PD without HPD when HPD was defined as a fourfold of TGR or TGKR. No significant difference was observed in the baseline characteristics between HPD and non-HPD.ConclusionThe prevalence of HPD in patients with advanced HCC treated with Atez/Bev was lower than those treated with nivolumab monotherapy. The HPD mechanism in ICI combined with antibodies targeting vascular endothelial growth factor (VEGF) remains to be elucidated.

Contemporary Management of Small Renal Masses by Urologic Oncologists: A 2022 Korean Renal Cancer Study Group Practice Pattern Survey

Purpose: Increased abdominal imaging brought about an explosive increase in the incidental detection of small renal masses (SRMs). In the absence of optimal guidelines for health screening, as well as subsequent diagnostic and therapeutic action plans, incidentally detected SRMs may likewise increasingly become a dilemma, especially in an aging society. In the current study, we aimed to describe the current practice patterns for incidentally detected SRMs among urologic oncologists and to identify key indicators in action plans for active surveillance.Materials and Methods: A survey containing 18 questions on SRM management patterns was designed. In June 2022, an online survey was sent to all 711 active members of the Korean Urological Oncology Society via email. After response collection, a consensus meeting of the Korean Renal Cancer Study Group, which 19 specialists attended, was held to analyze the results.Results: In total, 176 responses from participants practicing in an academic setting were obtained (24.8%, 176 of 711). Regarding the age of patients with SRMs, 42.6% (n=72) responded that they would recommend diagnostic evaluation and definitive treatment for anyone under 80 years of age as long as the patient was healthy. The most commonly used target indicators for surveillance termination were a tumor growth rate above a certain velocity (57.9%, n=102) and size increase above a certain diameter (36.9%, n=65). Renal mass biopsy was recommended in very select cases (<10% of all patients) by most respondents (53.4%, n=94), followed by “not using it at all” in 25.6% (n=45).Conclusions: We described the current practice patterns for incidentally detected SRMs among urologic oncologists and identified key indicators in action plans for active surveillance. This survey provided robust information, empowering physicians with a detailed knowledge of practice patterns and valuable insights on SRMs.

Impact of diabetes and chronic kidney disease on active surveillance outcomes for small renal masses: A cohort study.

The American Cancer Society estimates 79,000 individuals will be diagnosed with kidney cancer in 2022, most of which are initially found as small renal masses (SRMs). Proper management of SRM patients includes careful evaluation of risk factors such as medical comorbidities and renal function. To investigate the importance of these risk factors, we examined their effect on crossover to delayed intervention (DI) and overall survival (OS) in patients undergoing active surveillance (AS) for SRMs. This is an Institutional Review Board-approved retrospective analysis of AS patients presented at kidney tumor conferences with SRMs between 2007 and 2017. Univariable and multivariable logistic regression analyses were performed to determine how factors including estimated glomerular filtration rate (eGFR), diabetes, and chronic kidney disease are associated with DI and OS. A total of 111 cases were reviewed. In general, AS patients were elderly and had significant comorbidities. On univariate analysis, intervention was more likely to occur in patients with a younger age (P = 0.01), better kidney function (P = 0.01), and higher tumor growth rates (GRs) (P = 0.02). Higher eGFR was associated with better survival (P = 0.03), while higher tumor GRs (P = 0.014), greater Charlson Comorbidity Index (P = 0.01), and larger tumors (P = 0.01) were associated with worse OS. Of the comorbidities, diabetes was found to be an independent predictor of worse OS (P = 0.01). Patient-level factors - such as diabetes and eGFR - are associated with the rate of DI and OS among SRM patients. Consideration of these factors may facilitate better AS protocols and improve patient outcomes for those with SRMs.

B cells as modulators of HPV+ oropharyngeal cancer in a preclinical model.

Among the different immune cells present within tumors, B cells also infiltrate human papillomavirus-positive (HPV+) oropharyngeal tumors. However, the role of B cells during programmed death-1 (PD-1) blockade in HPV+ oropharyngeal cancer needs to be better defined. By using the preclinical mouse model for HPV+ oropharyngeal cancer (named mEER), we characterized B cells within tumors and determined their functional role in vivo during PD-1 blockade. We determined that treatment naïve tongue-implanted tumors, which we have previously demonstrated to be sensitive to PD-1 blockade, contained high infiltration of CD8+ T cells and low infiltration of B cells whereas flank-implanted tumors, which are resistant to PD-1 blockade, contain a higher frequency of B cells compared to T cells. Moreover, B cell-deficient mice (µMt) and B cell-depleted mice showed a slower tumor growth rate compared to wild-type (WT) mice, and B cell deficiency increased CD8+ T cell infiltration in tumors. When we compared tongue tumor-bearing mice treated with anti-PD-1, we observed that tumors that responded to the therapy contained more T cells and B cells than the ones that did not respond. However, µMt mice treated with PD-1 blockade showed similar tumor growth rates to WT mice. Our data suggest that in untreated mice, B cells have a more pro-tumorigenic phenotype potentially affecting T cell infiltration in the tumors. In contrast, B cells are dispensable for PD-1 blockade efficacy. Mechanistic studies are needed to identify novel targets to promote the anti-tumorigenic function and/or suppress the immunosuppressive function of B cells in HPV+ oropharyngeal cancer.

Cognitive issues in patients with IDH mutant gliomas: from neuroscience to clinical neuropsychology.

The understanding of cognitive symptoms in patients with IDH-Mutant gliomas (IDH-Mut) is rapidly developing. In this article, we summarize the neuroscientific knowledge base regarding the influence of IDH-Mut tumors and their treatment on cognition and provide guidance regarding the management of these symptoms in patients. We performed a review of peer reviewed publications relevant to IDH-Mut glioma and cognitive outcomes and provide an overview of the literature as well as a case example to clarify management strategies. At the time of presentation, patients with IDH-Mut gliomas have a favorable cognitive profile as compared with those with IDH-wild type (WT) tumors. The relatively low cognitive burden may reflect the slower growth rate of IDH-Mut tumors, which is less disruptive to both local and widespread neural networks. Human connectomic research using a variety of modalities has demonstrated relatively preserved network efficiency in patients with IDH-Mut gliomas as compared with IDH-WT tumors. Risk of cognitive decline from surgery can potentially be mitigated by careful integration of intra-operative mapping. Longer term cognitive risks of tumor treatment, including chemotherapy and radiation, are best managed by instituting neuropsychological assessment as part of the long-term care of patients with IDH-Mutant glioma. A specific timeline for such integrative care is provided. Given the relative recency of the IDH-mutation based classification of gliomas, as well as the long time course of this disease, a thoughtful and comprehensive strategy to studying patient outcomes and devising methods of cognitive risk reduction is required.

Potential anti-cancer effects of hibernating common carp (Cyprinus carpio) plasma on B16-F10 murine melanoma: In vitro and in vivo studies

Melanoma is the major type of skin cancer, which its treatment is still a challenge in the world. In recent years, interest in hibernation-based therapeutic approaches for various biomedical applications has been increased. Many studies indicated that some factors in the blood plasma of hibernating animals such as alpha-2-macroglobulin (A2M) cause anti-proliferative effects. Considering that, the present study was conducted to investigate the anti-cancer effects of hibernating common carp plasma (HCCP) on murine melanoma (B16-F10) in vitro and in vivo. The effect of HCCP on cell viability, migration, apoptosis rate, and cell cycle distribution of B16-F10 cells, tumor growth, and rate of survival were evaluated. To investigate the role of A2M in the anti-cancer effects of HCCP, the gene of interest and proteins in HCCP and non-hibernating common carp plasma (NHCCP) were evaluated by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) assay as well as sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and mass spectrometry analysis. Based on our findings, HCCP significantly decreased B16-F10 cell viability. Moreover, HCCP caused morphological alternations, inhibition of migration, induction of apoptosis, and significantly induced the cell cycle arrest at the G2/M phase. In addition, A2M level was significantly increased in HCCP compared with NHCCP. Taken together, our findings suggested that HCCP had the potential to be a promising novel therapeutic target for cancer treatment because of its anti-cancer properties.