Spontaneous Tumors Research Articles

STING agonists and PI3Kγ inhibitor co-loaded ferric ion-punicalagin networks for comprehensive cancer therapy

Nanoparticles-based drug delivery system has been a promising approach for the treatment of colorectal cancer (CRC), which can be combined with chemotherapy, targeted therapy and immunotherapy to improve the treatment of CRC. 2′3’ cyclic guanosine monophosphate-adenosine monophosphate (cGAMP) is an agonist of the STING signaling pathway activating antitumor immunity. IPI-549 is a small-molecule inhibitor for phosphatidylinositol 3-kinase γ (PI3Kγ), which can induce M1 macrophages polarization to provide pro-inflammatory microenvironment to suppress tumors. Here, we developed a ferric ion-punicalagin network (Fe-PU), which can be not only used as an inducer of ferroptosis, but also serve as a carrier to load cGAMP and IPI-549 to obtain nanohybrid (Fe-PU/CD-IPI). In order to improve the delivery effect and targeted ability to CRC, a cyclic arginine-glycine-aspartic acid peptide linked-bovine serum albumin were utilized to modify Fe-PU/CD-IPI to prepare nanohybrid Fe-PU/CD-IPI@cBSA. The therapeutic effect of various nanohybrids were validated in the mice with spontaneous tumor in the colorectal area and tumor-bearing mice, which lead to the increase of ferroptosis, the activation of STING signaling pathway, and the repolarization of macrophages. Collectively, the cGAMP and IPI-549 co-loaded nanohybrids effectively reshaped the tumor immune microenvironment, and exhibited prominent treatment effect of anti-colorectal cancer in vitro, patient-derived organoids, and in vivo.

Development of Vincristine‐Loaded Nano Graphene Oxides for Dual Therapies Against Canine Anaplastic Carcinoma Cells Obtained From a Dog with Spontaneous Mammary Tumor

AbstractAmong pet species, mammary tumors are most commonly encountered in dogs. In the traditional treatment of mammary tumors in dogs; surgical operation, chemotherapy, immunotherapy, cryotherapy, radiotherapy and homeopathy are used. Herein, we reported vincristine loaded nano graphene oxide (Vin@nGO) for simultaneous photothermal therapy (PTT) and chemotherapy under near infrared laser (NIR, 808 nm), whereas GO was used both as a platform for vincristine and as a photothermal (PT) agent, and vincristine was used as a chemotherapeutic agent. Spontaneous mammary tumor masses of a dog were removed and primary tumor cells were obtained and propagated by tissue culture from tumor tissue samples under appropriate conditions. Then, these cells were incubated with Vin@nGO and exposed to NIR laser. NIR laser irradiation at 808 nm was converted to heat by nGO, then the target cancer cells were destroyed by both hyperthermia and chemotherapy. MTS analysis was performed to test the effectiveness of this dual therapy on cell proliferation and a statistically significant (p < 0.05) decrease in cell proliferation/viability was found (90% reduction compared to the control group (p < 0.0001)). Our study data also shed light on usability of Vin@nGO in the in vivo model for future work.

Abstract C037: Chemotherapy causes de novo induction of invasive breast cancer cells

Abstract Metastasis is the leading cause of breast cancer (BC) death, and tumor cells must migrate and invade to metastasize. BC cells that express the actin regulatory protein MenaINV have an enhanced ability to migrate and intravasate within the primary tumor, and extravasate at secondary sites. Though chemotherapy improves patient survival, treatment with paclitaxel, a chemotherapy used for BC, leads to upregulation of MenaINV and an increase in metastasis in mice. MenaINV expression can be induced in BC cells through Notch1 signaling with macrophages, which are often recruited to tumors in response to chemotherapy. MenaINV-expressing cells are also resistant to paclitaxel, begging the question of whether paclitaxel increases MenaINV by de novo induction or by selectively killing non-MenaINV-expressing cells. We hypothesized that paclitaxel causes de novo MenaINV induction by increasing macrophage-tumor cell Notch signaling. Understanding this pro-metastatic effect of chemotherapy is crucial to refining treatment strategies. MMTV-PyMT mice bearing spontaneous mammary tumors were used as a model of BC. Mice were treated with paclitaxel or vehicle every 5 days for a total of 3 treatments. To study the role of macrophages in the paclitaxel-mediated MenaINV increase, in addition to paclitaxel, one cohort of mice was co-administered the macrophage-depleting agent clodronate or vehicle. To study the role of Notch signaling, a second cohort of mice was co-administered the Notch inhibitor DAPT or vehicle. Immunofluorescence staining was used to evaluate tumors for: %MenaINV+ area, proportion of apoptotic cells, macrophage density, and macrophage expression of Notch ligands Jag1 and Jag2. Paclitaxel-treated tumors expressed significantly more MenaINV even when tumor cell death did not increase, indicating that chemotherapy increases MenaINV expression by induction. Both macrophages and Notch signaling were needed for this effect to occur. Interestingly, paclitaxel did not increase macrophage infiltration or macrophage expression of Notch ligands, suggesting that paclitaxel may increase MenaINV by acting on tumor cells to promote Notch signaling. Paclitaxel causes de novo MenaINV induction that is macrophage- and Notch-dependent. Studies aiming to further elucidate the mechanism behind paclitaxel-mediated MenaINV induction are ongoing. These results lay the groundwork for novel microenvironment-based therapies to alleviate the pro-metastatic effects of chemotherapy in BC. Citation Format: Madeline Friedman-DeLuca, George S. Karagiannis, Camille L. Duran, Luis Rivera Sanchez, Prachiben P. Patel, Suryansh Shukla, Nicole D. Barth, Ved P. Sharma, Michael Papanicolaou, John S. Condeelis, Maja H. Oktay, David Entenberg. Chemotherapy causes de novo induction of invasive breast cancer cells [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr C037.

Abstract A004: E2F5 conditional knockout in mammary epithelium drives organotropic metastasis in breast cancer

Abstract Breast cancer ranks as the most commonly diagnosed cancer in women with 90% of associated deaths caused by distant metastasis. The metastatic lesions in brain, lung, liver, bone and lymph nodes in human breast cancer are not arbitrary, but rather are a complex mechanism of extrinsic and intrinsic regulatory factors controlling organ-specific colonization. Specialized mechanisms driving bone, brain and lung cancer have been described. However, the studies of liver and lymph node metastatic drivers are limited despite the prevalence of metastases in these organs. A significant limitation is the lack of genetically engineered mouse models that develop mammary tumors that spontaneously metastasize to liver and lymph nodes. Our previous research identified the role for E2F5 in both mammary gland development and breast cancer. Loss of E2F5 resulted in tumorigenesis after a latency of one year and 70% of mice showed metastatic lesions in lung, lymph node and liver. Transplantation of these spontaneous tumors into FVB MMTV Cre mice recapitulated the metastatic pattern observed in the primary tumors. We propose that E2F5 regulates drivers of metastasis and potentiates organ tropism to lymph nodes and the liver. To address this hypothesis, we conducted serial transplantations of tumors originating from the lymph nodes and liver back into the mammary gland. This enrichment study led to a significant increase in organ-specific metastasis, with 60% and 77% of penetrance observed in two distinct mouse lineages with liver (LVM) and lymph node tropism (LNM). Immunohistochemical studies confirmed a complete epithelial-mesenchymal transition (EMT) phenotype in LVM that is conserved in the cell lines generated. LNM showed a partial EMT with a more epithelial phenotype in cell lines. We observed the presence of an immune enriched environment in primary and metastatic tumors in both lineages. Bulk-RNA sequencing studies uncovered the presence of unique organotropic pathways associated with liver and lymph node metastasis. We observed enriched pathways from the primary tumor as major component of our organotropic drivers with a clear distinction between liver and lymph node enriched lineages. Additionally, a gradual acquisition of distant organ-specific markers and signaling pathways suggests that phenotypic mimicry and tumor plasticity are key for successful events of colonization to liver and lymph nodes in our model. With these results, we unravel potential mechanisms responsible for the organotropic metastases in breast cancer under the absence of the transcription factor E2F5. Citation Format: Jesus A. Garcia Lerena, Briana To, Jing-Ru Jhan, Eran Andrechek. E2F5 conditional knockout in mammary epithelium drives organotropic metastasis in breast cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr A004.

IMMU-36. MULTI-OMIC CHARACTERIZATION OF A SYNGENEIC MOUSE CANCER STEM CELL MODEL OF GBM FOR PRECLINICAL APPLICATIONS

Abstract Glioblastoma (GBM) is resistant to many therapies including immunotherapies. There is an urgent need to explore the role of the microenvironment in resistance to therapy with tailored pre-clinical models. Using multi-omic approaches we describe a syngeneic cancer stem cell mouse model of GBM, with a spontaneous amplification of Igf2. We investigate whether Igf2 influences tumour and microenvironment cells to promote immunosuppression in GBM. Three cell lines were previously established from spontaneous brain tumours in C57Bl/6 Trp53+/-/Nf1+/- mice and maintained under neural stem cell culture conditions. Whole genome sequencing (WGS) was used to determine single nucleotide, copy number, and structural variation, revealing loss of both copies of Trp53 and Nf1 in all three cell lines. However only one cell line, mBT0309, developed tumors when orthotopically allografted. Analysis of genomic alterations and transcriptomics revealed that mBT309 exhibits amplification of the Igf2 loci and overexpression was confirmed at the RNA and protein level. Spatial and single cell transcriptomics showed that Igf2 is overexpressed in mBT0309 allografted tumours and correlated with specific transcriptomic programs. A high-parameter Imaging Mass Cytometry (IMCTM) panel was used for spatial proteomic analysis, to monitor the development of tumours in a time-course experiment. In vitro growth characteristics and stem marker expression in both transcriptomic and spatial IMC data suggest that high levels of Igf2 may regulate GBM stemness features. Analysis of human GBM datasets revealed that Igf2 amplified tumours have reduced CXCL11 expression. These findings suggest that high levels of Igf2 may influence immunosuppression by reducing T-cell recruitment. This syngeneic immunocompetent GBM stem cell model harbouring Igf2 amplification adds to the suite of models to study the GBM microenvironment and its role in immunosuppression and resistance to therapy.

Comparative analysis of canine and human HtrA2 to delineate its role in apoptosis and cancer.

Therapeutically, targeting the pro- and anti-apoptotic proteins has been one of the major approaches behind devising strategies to combat associated diseases. Human high-temperature requirement serine protease A2 (hHtrA2), which induces apoptosis through both caspase-dependent and independent pathways is implicated in several diseases including cancer, ischemic heart diseases, and neurodegeneration, thus making it a promising target molecule. In the recent past, the canine model has gained prominence in the understanding of human pathophysiology that was otherwise limited to the rodent system. Moreover, canine models in cancer research provide an opportunity to study spontaneous tumors as their size, lifespan, and environmental exposure are significantly closer to that of humans compared with laboratory rodents. Therefore, using HtrA2 as a model protein, comparative analysis has been done to revisit the hypothesis that canines might be excellent models for cancer research. We have performed evolutionary phylogenetic analyses that confirm a close relationship between canine and human HtrA2s. Molecular modeling demonstrates structural similarities including orientation of the catalytic triad residues, followed by in silico docking and molecular dynamics simulation studies that identify the potential interacting partners for canine HtrA2 (cHtrA2). In vitro biophysical and protease studies depict similarities in interaction with their respective substrates as well as transient transfection of cHtrA2 in mammalian cell culture shows induction of apoptosis. This work, therefore, promises to open a new avenue in cancer research through the study of spontaneous cancer model systems in canines.

Nerve Sheath Tumor in a Cynomolgus Macaque (Macaca fascicularis) and Diagnostic Differentiation from Amelanotic Melanoma

AbstractThere are few reports describing spontaneous neoplasms in cynomolgus macaques, despite the frequent use of this species in laboratory research. This report describes cytologic, histologic, and immunohistochemical findings of a cutaneous to subcutaneous nerve sheath tumor located within the haired skin of the abdomen of a 2.5-y-old, intact, female, captive Mauritius cynomolgus macaque. The nerve sheath tumor was well demarcated, partially encapsulated, densely cellular, and extended from the subcutis to the most superficial dermis, abutting the epidermis. Neoplastic cells formed intersecting streams and had a high mitotic count (18 per 2.37 mm2). Due to the substantial morphologic overlap of this neoplasm with amelanotic melanoma, particularly the close association with the epidermis, immunohistochemistry was required for definitive diagnosis. Neoplastic cells were immunoreactive to vimentin, S-100, SOX10, laminin, collagen IV, and CD56, and negative for melan-A, tyrosinase, MITF, and HMB45. This immunohistochemical profile is diagnostic for nerve sheath tumor based on human and canine criteria and rules out amelanotic melanoma. Despite incomplete excision, the nerve sheath tumor had not grossly recurred after 1 mo, at which point the animal was euthanized for unrelated reasons. This report underscores the importance of using an immunohistochemical panel in cases of cutaneous and subcutaneous spindle cell neoplasms, as there is substantial morphologic and immunohistochemical overlap between nerve sheath tumors and melanocytic neoplasms due to their shared neuroectodermal origin. To our knowledge, this is the first report of a nerve sheath tumor in a cynomolgus macaque, and one of the few reports of spontaneous neoplasia in this species.

Response of Spontaneous Oral Tumors in Canine Cancer Patients Treated with Stereotactic Body Radiation Therapy (SBRT).

To describe outcome and toxicity for dogs with oral tumors, specifically oral malignant melanoma (OMM), squamous cell carcinoma (SCC), and soft tissue sarcoma (STS) after stereotactic body radiation therapy (SBRT). A single institution retrospective study was conducted. Outcomes were analyzed using Kaplan-Meier analysis and Cox proportional hazard analysis. Treatment responses at different time points were evaluated with Pearson's Chi-squared test to identify prognostic factors. Acute and late toxicities were recorded according to VRTOG criteria and were analyzed to identify risk factors. Adverse events other than acute and late toxicities were recorded. A total of 98 patients met the inclusion criteria (OMM n = 37; SCC n = 18; STS n = 43). The SBRT prescription was 1-6 fractions, with a total dose range of 12-40 Gy. Local progression-free survival (PFS) for OMM, SCC, and STS was 187, 253, and 161 days, respectively. Overall PFS was 152 days and median survival time (MST) was 270 days, with no statistical difference between tumor types. The presence of lymph node metastasis and the use of elective nodal irradiation (ENI) were associated with shorted PFS and MST. Severe acute toxicities to organs at risk affected 10/85 (11.8%) of patients. Osteoradionecrosis and oronasal fistula formation occurred in 23/81 (28.4%) of patients and was significantly associated with tumor type (SCC, P = 0.006). BRT can be offered as a treatment option for oral tumors in dogs. Toxicities were common and warrant risk factor considerations and adjustments to current SBRT protocols.

Combining ERAP1 silencing and entinostat therapy to overcome resistance to cancer immunotherapy in neuroblastoma

BackgroundCheckpoint immunotherapy unleashes tumor control by T cells, but it is undermined in non-immunogenic tumors, e.g. with low MHC class I expression and low neoantigen burden, such as neuroblastoma (NB). Endoplasmic reticulum aminopeptidase 1 (ERAP1) is an enzyme that trims peptides before loading on MHC class I molecules. Inhibition of ERAP1 results in the generation of new antigens able of inducing potent anti-tumor immune responses. Here, we identify a novel non-toxic combinatorial strategy based on genetic inhibition of ERAP1 and administration of the HDAC inhibitor (HDACi) entinostat that increase the immunogenicity of NB, making it responsive to PD-1 therapy.MethodsCRISPR/Cas9-mediated gene editing was used to knockout (KO) the ERAP1 gene in 9464D NB cells derived from spontaneous tumors of TH-MYCN transgenic mice. The expression of MHC class I and PD-L1 was evaluated by flow cytometry (FC). The immunopeptidome of these cells was studied by mass spectrometry. Cocultures of splenocytes derived from 9464D bearing mice and tumor cells allowed the assessment of the effect of ERAP1 inhibition on the secretion of inflammatory cytokines and activation and migration of immune cells towards ERAP1 KO cells by FC. Tumor cell killing was evaluated by Caspase 3/7 assay and flow cytometry analysis. The effect of ERAP1 inhibition on the immune content of tumors was analyzed by FC, immunohistochemistry and multiple immunofluorescence.ResultsWe found that inhibition of ERAP1 makes 9464D cells more susceptible to immune cell-mediated killing by increasing both the recall and activation of CD4+ and CD8+ T cells and NK cells. Treatment with entinostat induces the expression of MHC class I and PD-L1 molecules in 9464D both in vitro and in vivo. This results in pronounced changes in the immunopeptidome induced by ERAP1 inhibition, but also restrains the growth of ERAP1 KO tumors in vivo by remodelling the tumor-infiltrating T-cell compartment. Interestingly, the absence of ERAP1 in combination with entinostat and PD-1 blockade overcomes resistance to PD-1 immunotherapy and increases host survival.ConclusionsThese findings demonstrate that ERAP1 inhibition combined with HDACi entinostat treatment and PD-1 blockade remodels the immune landscape of a non-immunogenic tumor such as NB, making it responsive to checkpoint immunotherapy.

Burned-out testicular cancer - case report

Introduction and Purpose Germ cell tumors of the testis are the most common malignant tumors in young men aged 15-40, although their incidence in the male population is about 1-1.5%. 90-98% of all testicular tumors originate from germ cells. There has been an increase in the incidence of testicular tumors in recent years, suggesting the influence of both genetic and environmental factors. Testicular cancer, which accounts for about 5% of all urological cancers and mainly affects young men. Most of these cancers originate from germ cells, and cases of spontaneous tumor regression are extremely rare. Aim of the study We present the case of a 36-year-old man who was found to have a left testicular tumor with negative tumor markers during physical examination and testicular ultrasound. The tumor was surgically removed, and the patient was discharged home. Histopathological examination showed that the tumor had completely necrosed, and a CT scan showed no metastasis. Results and Future directions The phenomenon of testicular tumor “burnout” has been described since 1927, but it was only in 2016 that the WHO recognized it as a separate disease entity. Despite the rarity of this phenomenon, more and more cases are being reported in the literature, allowing for a better understanding of the underlying mechanisms. It is important to remember that the tumor can give a variable clinical picture and its symptoms are not necessarily related to the location of the tumor, but to its metastasis. Physical and ultrasound examinations are key to making a correct diagnosis. Histologic features such as scar formation, intravesicular calcification, lymphoplasmocytic infiltration and testicular atrophy can help diagnose germ cell tumor. The findings of this case demonstrate the need for diagnostic vigilance, especially in patients with equivocal tumor marker results but with clinical signs of testicular tumor.

PD-L1 expression in testicular germ cell tumors undergoing spontaneous regression.

Spontaneous regression of testicular germ cell tumors is a well-known phenomenon; however, the precise mechanisms of spontaneous regression are still unknown. Our study aimed to investigate programmed death-ligand 1 (PD-L1) expression in spontaneously regressed testicular germ cell tumors, exploring the link between the immune response and spontaneous regression. From a sample of 356 testicular germ cell tumors, we singled out 5 completely regressed and 6 partially regressed tumors. In four out of six cases with partial regression, a residual seminoma component was found, while in the remaining two cases, an embryonal carcinoma component was found. Comparisons were made with 20 pure seminomas and 20 mixed germ cell tumors (MGCTs). A semiquantitative immunohistochemical analysis of PD-L1 expression in tumor cells and intra/peritumoral lymphocytes was performed. There was no PD-L1 expression in tumors with complete regression. All partially regressed tumors showed expression in intra/peritumoral lymphocytes within the tumor remnants. Expression was significantly more frequent in pure seminomas compared to MGCTs (P = 0.004). A positive correlation was demonstrated between the seminoma component and the proportion of PD-L1 positive lymphocytes, with a Kendall's Tau-b coefficient of 0.626 (P < 0.001). Tumor cells showed PD-L1 expression in three MGCTs within the embryonal carcinoma component. Our results support an immunological mechanism of spontaneous tumor regression, with the strongest potential in testicular tumors containing seminoma components. However, further research is necessary to determine the role of PD-L1 ligand more precisely in the microenvironment of spontaneously regressed tumors.

Intramuscular myxoma: unusual observation of spontaneous tumor size shrinkage.

Soft tissue tumors, whether benign or malignant, may grow over time or remain stable, but they usually do not spontaneously decrease in size. However, there are exceptions, such as inflammatory conditions, desmoid tumors, or benign cysts. Intramuscular myxomas are benign soft tissue tumors that typically present as a solitary, slow-growing, painless mass. They are generally treated by surgical resection, after which recurrence is rare. Here, we present a brief series of three unusual cases of intramuscular myxomas that spontaneously decreased in size. They were located in the cervical region, the right lower extremity, and the paravertebral lumbar region. Imaging findings and percutaneous biopsies confirmed the diagnosis in all cases. Follow-up imaging showed a spontaneous reduction in lesion volume over time, far exceeding the amount of tissue sample removed during biopsy. This unusual observation of spontaneous shrinkage may call into question the subsequent therapeutic approach to these lesions.

A temporal (phospho-)proteomic dataset of neurotrophic receptor tyrosine kinase signalling in neuroblastoma

Neurotrophic receptor tyrosine kinases (TrkA, TrkB, TrkC), despite their homology, contribute to the clinical heterogeneity of the childhood cancer neuroblastoma. TrkA expression is associated with low-stage disease and is often seen with spontaneous tumour regression. Conversely, TrkB is present in unfavourable neuroblastomas that often harbour amplification of the MYCN oncogene. The role of TrkC is less clearly defined, although some studies suggest its association with a favourable outcome. Understanding the differences in activity of Trk receptors that drive divergent clinical phenotypes as well as the influence of MYCN amplification on downstream Trk receptor signalling remains poorly understood. Here, we present a comprehensive label-free mass spectrometry-based total proteomics and phosphoproteomics dataset (432 raw files with FragPipe search outputs; available on PRIDE with accession number PXD054441) where we identified and quantified 4,907 proteins, 16,744 phosphosites and 5,084 phosphoproteins, derived from NGF/BDNF/NT-3 treated TrkA/B/C-overexpressing neuroblastoma cells with differential MYCN status. Analysing our dataset offers valuable insights into TrkA/B/C receptor signalling in neuroblastoma and its modulation by MYCN status; and holds potential for advancing therapeutic strategies in this challenging childhood cancer.

8123 Identification Of Glutamine Metabolism As A Druggable Therapeutic Vulnerability For Adrenocortical Carcinoma

Abstract Disclosure: V. Chortis: None. C. Yao: None. C. Ribeiro: None. L. Kelley: None. L. Nagano: None. M. Berber: None. S. Raveenthiraraj: None. P. Vendramini: None. K. Kiseljak-Vassiliades: None. D.L. Carlone: None. M.C. Haigis: None. K.S. Borges: None. D.T. Breault: None. Dysregulation of cellular metabolism is a hallmark feature of cancer that can be targeted therapeutically but remains underexplored in adrenocortical carcinoma (ACC), despite the urgent need for improved treatments. We employed a recently developed transgenic mouse model of ACC (BPCre), driven by the two most common mutations in human ACC (β-catenin gain-of-function and p53 loss-of-function), aiming to characterize in vivo ACC metabolism at a tissue level and to identify metabolic vulnerabilities. We employed metabolomics (liquid chromatography-mass spectrometry) and transcriptomics (bulk RNA-seq) to compare tissue metabolism in spontaneous BPCre tumors with control adrenals. Polar metabolites were analyzed using a library that provides a representative cross-section of major carbon and nitrogen-handling pathways. We identified widespread dysregulation of tumor metabolism (64/175 metabolites significantly dysregulated, FDR&amp;lt;0.05), with the most significantly dysregulated pathways including purine and pyrimidine metabolism, proline and glutamate metabolism, glycolysis, and the pentose phosphate pathway (FDR&amp;lt;0.05). Orthogonal analysis by RNA-seq revealed congruent metabolic pathway alterations. Comparison of the most significantly dysregulated pathways between BPCre mouse tumors and human ACC using publicly accessible human transcriptomic datasets (TCGA and GTEx) revealed extensive overlap. Given the prominent dysregulation of several glutamine (Gln)-dependent metabolic pathways in mouse and human ACC, we hypothesized that Gln catabolism is likely to represent a targetable metabolic vulnerability in ACC. Pharmacological targeting of Gln metabolism using the Gln antagonist 6-Diazo-5-Oxo-L-Norleucine (L-DON) induced marked cytotoxicity in two cell lines derived from BPCre tumors and three human ACC cell lines (NCI-H295R, CU-ACC1-2), as assessed by cell viability and clonogenic assays. Remarkably, this effect was rescued only by nucleoside supplementation, suggesting that Gln’s contribution to de novo purine and pyrimidine biosynthesis is the critical metabolic dependency in ACC cells. Treatment with the L-DON pro-drug JHU-083, in vivo, led to marked growth inhibition of subcutaneous mouse ACC tumor implants in both immunocompetent and immunodeficient mice. As expected, tissue metabolomics of JHU-083-treated tumors confirmed extensive depletion of purine metabolites. Tumor immunophenotyping by flow cytometry revealed significantly increased infiltration with Natural Killer cells, likely potentiating the anti-tumor effect of glutamine antagonism in immunocompetent mice. JHU-083 was also effective against human NCI-H295R xenografts, in vivo. This work reveals important new insights into ACC metabolism and provides pre-clinical evidence supporting Gln antagonism as a new metabolic treatment approach for ACC. Presentation: 6/2/2024

The role of cytoreductive nephrectomy in the era of immune checkpoint inhibitors: A review of current evidence and ongoing trials.

Renal cell carcinoma (RCC) was diagnosed in over 400 000 individuals globally in 2020, making it a significant global health concern. The incidence of RCC varies by region and overall mortality rates have been declining. This decline is attributed in part to advancements in early cancer detection through imaging and the development of more effective systemic therapies. Cytoreductive nephrectomy (CN) was adopted as a standard treatment for metastatic RCC (mRCC) based on clinical experience and early clinical trials. However, the treatment landscape has shifted with the introduction of tyrosine kinase inhibitors (TKI) in 2007 and, more recently, immune checkpoint inhibitors (ICIs). Dual ICI therapy and combinations of ICIs with TKIs are collectively referred to as immuno-combination therapies and have become standard first-line treatments. This review examines the evolving role of CN in the era of immuno-combination therapies, with a focus on patient selection and the timing of surgery. The immunogenic nature of RCC, characterized by spontaneous tumor regression and immune cell infiltration, suggests a potential benefit from combining CN with ICI therapy to enhance treatment outcomes. This is supported by several clinical studies that reported improved outcomes; however, these were limited by their retrospective nature. Ongoing clinical trials, such as NORDIC-SUN, PROBE, and SEVURO-CN, are expected to provide critical insights into the role of CN in the ICI era. Their findings will ultimately guide future clinical decision-making and further refine treatment strategies for mRCC.

Comparative Analysis of Metastatic Thyroid Carcinoma versus Ectopic Thyroid Carcinoma in Lateral Neck Masses without Identifiable Primary Thyroid Carcinoma

Background/Objectives: Thyroid carcinoma, presenting as a lateral neck mass without an identifiable primary tumor within the thyroid, poses a diagnostic challenge. This comparative analysis aimed to explore the differences between metastatic thyroid carcinoma and ectopic thyroid carcinoma, as both present with a lateral neck mass without evidence of primary thyroid carcinoma. Methods: Searches were conducted for studies on thyroid carcinoma in the lateral neck without evidence of primary thyroid carcinoma. A total of 39 patients were identified from 32 reported studies. Results: Metastatic and ectopic thyroid carcinomas were found in 11 and 28 patients, respectively. Metastatic thyroid carcinoma is characterized by evidence of spontaneous primary tumor regression within the thyroid and commonly associated with multiple lymph node metastases in central and lateral neck compartments. Ectopic thyroid carcinoma is more commonly diagnosed in younger patients and is frequently identified in branchial cleft cysts. The coexistence of normal thyroid tissue adjacent to the ectopic thyroid carcinoma was confirmed, and patients with ectopic thyroid carcinoma exhibited significantly higher rates of second-stage thyroidectomy or neck dissection. When complete surgical excision was considered adequate, excision alone was chosen for patients with ectopic thyroid carcinoma. Conclusions: Identifying these differences is valuable for the differential diagnosis and development of treatment strategies for metastatic and ectopic thyroid carcinomas in lateral neck masses without evidence of primary thyroid tumor.

Letrozole-Based Near-Infrared Dynamic Imaging Targeting Ductal-Vascular RhoJ From Pancreatic Intraepithelial Neoplasia to Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) relies heavily on neoangiogenesis for its progression, making early detection crucial. Here, LTZi-MHI148 (Letrozole inhibitor bonding with MHI-148 dye), a near-infrared (NIR) fluorescent agent is developed, to target RhoJ (Ras Homolog Family Member J), a protein expressed in neonatal vasculature, for both imaging and therapy of early PDAC. This agent is synthesized by conjugating Letrozole with MHI-148, exhibiting excellent NIR characteristics and photostability. In vitro studies showed that LTZi-MHI148 selectively accumulated within pancreatic cancer cells through Organic Anion Transporting Polypeptide (OATP) transporters and bound to cytoplasmic RhoJ. In vivo, the probe effectively targeted neoangiogenesis and Pancreatic Intraepithelial Neoplasias (PanINs) in various PDAC models, including the orthotopic, ectopic, spontaneous, and tamoxifen-induced tumors. Notably, LTZi-MHI148 detected preneoplastic PanIN lesions with Overexpressed RhoJ and active neoangiogenesis in both spontaneous and tamoxifen-induced PDAC murine models. Longitudinal imaging studies revealed that RhoJ-targeted neoangiogenesis tracks lesion progression, highlighting LTZi-MHI148's utility in monitoring disease progression. Furthermore, multiple LTZi-MHI148 administrations attenuated PanINs to PDAC progression, suggesting its potential as a therapeutic intervention. These findings underscore the translational potential of LTZi-MHI148 for the early detection and targeted therapy of PDAC, utilizing NIR-I/II imaging to monitor RhoJ overexpression in precancerous ductal neoplasia associated with neoangiogenesis.

The Comparative Oncology of Canine Malignant Melanoma in Targeted Therapy: A Systematic Review of In Vitro Experiments and Animal Model Reports

Canine malignant melanoma (CMM) is highly aggressive and mostly located in the oral cavity. CMM is the predominant type of canine oral malignancy and shows striking homologies with human mucosal melanoma. In comparative oncology, canine oral melanomas (COMs), as spontaneous tumor models, have the potential to acquire a unique value as a translational model of rare human melanoma subtypes. This review aims to provide a comprehensive summary of targeted therapies for canine malignant melanoma and to enrich the field of comparative oncology. Following the PRISMA guidelines, a comprehensive literature search was conducted across databases for studies from 1976 to April 2024. Studies were selected based on their relevance to targeted treatments. A total of 30 studies met the inclusion criteria. Based on the treatment approaches, the studies were further categorized into immunotherapies, small molecule signaling inhibitors, indirect kinase inhibitors, and other alternative strategies. Some treatments have been shown to result in stable disease or partial response, accounting for 29% (monoclonal antibody) and 76.5% (micro-RNA therapies) in clinical trials. Moreover, in vitro experiments of small molecule inhibitors, including cell signaling inhibitors and indirect kinase inhibitors, have shown the potential to be an effective treatment option for the development of therapeutic strategies in canine malignant melanoma. The observed response in in vitro experiments of CMM (particularly the oral and certain cutaneous subtypes) to drugs used in the treatment of human melanoma underlines the resemblance to human melanoma, therefore supporting the notion that CMM may be a valuable model for understanding rare human melanoma subtypes and exploring potential therapeutic avenues in preclinical trials. Finally, this literature review serves as a valuable resource for the development of therapeutic strategies for CMM and highlights the potential for translating these findings to human cancer treatment.

Abstract A044: Persistence of fetal splanchnic gene signature defines a tumor-restraining fibroblast subtype in pancreatic cancer

Abstract The pancreas is composed of the epithelial and mesenchymal cells. While mesenchymal fibroblasts are a minor component of the normal pancreas, fibroblast population expands drastically during tumorigenesis. In pancreatic ductal adenocarcinoma (PDAC), cancer associated fibroblasts (CAFs) play critical and complex roles in the tumor microenvironment. This study sought to define the origin, heterogeneity and function of pancreatic cancer associated fibroblasts. Recently we performed a series of lineage tracing studies in genetically engineered mouse models. This identified the splanchnic mesenchyme (a tissue layer adjacent to the developing fetal pancreatic epithelium) as the fetal origin of the adult pancreatic resident fibroblasts and pancreatic CAFs. Single cell transcriptomic analysis indicated persistent and dynamic gene expressions along the pancreatic mesenchymal trajectory during development, homeostasis, precancer and cancer. Intriguingly, we found that two splanchnic transcription factors, GATA6 and FOXF1, are expressed in only subsets of adult pancreatic fibroblasts in temporally and spatially distinct patterns. Similar patterns were observed in both mouse models and human patient samples. To determine the role of GATA6 in fibroblasts during tumorigenesis, we constructed a dual DNA recombinase mouse genetic model. DNA recombinase FlpO directs expression of an oncogene Kras (G12D mutation) and loss of a tumor suppressor p53 in pancreatic epithelial cells to induce spontaneous tumor formation in the pancreas, and DNA recombinase Cre deletes Gata6 specifically in fibroblasts. This fibroblast specific Gata6 deletion resulted in altered number and gene expression of CAFs as well as increased tumor burden in the pancreas. This suggests a non-cell autonomous function of GATA6 within CAFs to restrain pancreatic cancer progression. In summary, this study delineated a continuous cell trajectory of the mesenchymal lineage in the pancreas across different life stages. Additionally, this study provides evidence that persistent and selective gene expressions along the mesenchymal trajectory contributes to pancreatic CAF heterogeneity and such persistence may constitute an inherent host defense mechanism to restrain pancreatic cancer. The enhancement of this mechanism could be explored further for therapeutic benefits. Citation Format: Lu Han, Tom Walter, Joseph Beaudet, Caroline Everett, Gustavo Leone, Michael Ostrowski. Persistence of fetal splanchnic gene signature defines a tumor-restraining fibroblast subtype in pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A044.

Molecular Imaging of p53 in Mouse Models of Cancer Using a Radiolabeled Antibody TAT Conjugate with SPECT.

Mutations of p53 protein occur in over half of all cancers, with profound effects on tumor biology. We present the first-to our knowledge-method for noninvasive visualization of p53 in tumor tissue invivo, using SPECT, in 3 different models of cancer. Methods: Anti-p53 monoclonal antibodies were conjugated to the cell-penetrating transactivator of transcription (TAT) peptide and a metal ion chelator and then radiolabeled with 111In to allow SPECT imaging. 111In-anti-p53-TAT conjugates were retained longer in cells overexpressing p53-specific than non-p53-specific 111In-mIgG (mouse IgG from murine plasma)-TAT controls, but not in null p53 cells. Results: In vivo SPECT imaging showed enhanced uptake of 111In-anti-p53-TAT, versus 111In-mIgG-TAT, in high-expression p53R175H and medium-expression wild-type p53 but not in null p53 tumor xenografts. The results were confirmed in mice bearing genetically engineered KPC mouse-derived pancreatic ductal adenocarcinoma tumors. Imaging with 111In-anti-p53-TAT was possible in KPC mice bearing spontaneous p53R172H pancreatic ductal adenocarcinoma tumors. Conclusion: We demonstrate the feasibility of noninvasive invivo molecular imaging of p53 in tumor tissue using a radiolabeled TAT-modified monoclonal antibody.