Metastatic Burden Research Articles

Abstract A102: ROR2/Wnt5a signaling regulates directional cell migration and early tumor cell invasion in ovarian cancer

Abstract Adhesion to and clearance of the mesothelial monolayer are key events in the early metastatic seeding of ovarian cancer. To identify mediators of ovarian cancer cell adhesion and invasion, we developed and conducted a cellular, high-throughput genetic screen using a human kinome siRNA library. Through this screen, we identified ROR2 as a driver of ovarian cancer adhesion and invasion. ROR2 is a receptor tyrosine kinase that interacts with Wnt5a ligand to activate non-canonical Wnt signaling and has been previously shown to be upregulated in ovarian cancer tissue. However, no prior study has evaluated the mechanistic role of ROR2 in ovarian cancer. Using in vitro adhesion and mesothelial clearance assays, we show that ROR2 promotes ovarian cancer cell adhesion and clearance of the mesothelial monolayer, key early steps in ovarian cancer metastasis. Further, we show that depletion of ROR2 reduces metastatic tumor burden in a syngeneic mouse model of ovarian cancer. Additionally, using a microfluidic device capable of producing stable fluid gradients, we demonstrate that ROR2 expression in ovarian cancer cells serves to drive directed cell migration of tumor cells, preferentially to areas of high Wnt5a ligand. As Wnt5a has been previously shown to be produced and upregulated in the mesothelial cells of the ovarian cancer microenvironment, these findings represent a novel role for ROR2 to promote the homing of ovarian tumor cells to the mesothelial lined metastatic niche by using Wnt5a as a chemotactic cue. These findings support the role of ROR2 as a critical factor contributing to the steps of ovarian cancer metastasis. Therapeutic targeting of the ROR2/Wnt5a signaling axis could provide a means of improving treatment for patients with advanced ovarian cancer. Citation Format: Whitney R. Grither, Breanna Baker, Vasilios A. Morikis, Katherine C. Fuh, Gregoery D Longmore. ROR2/Wnt5a signaling regulates directional cell migration and early tumor cell invasion in ovarian cancer [abstract]. In: Proceedings of the AACR Special Conference on Ovarian Cancer; 2023 Oct 5-7; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_2):Abstract nr A102.

PROM2 overexpression induces metastatic potential through epithelial-to-mesenchymal transition and ferroptosis resistance in human cancers.

Despite considerable therapeutic advances in the last 20 years, metastatic cancers remain a major cause of death. We previously identified prominin-2 (PROM2) as a biomarker predictive of distant metastases and decreased survival, thus providing a promising bio-target. In this translational study, we set out to decipher the biological roles of PROM2 during the metastatic process and resistance to cell death, in particular for metastatic melanoma. Methods and results: We demonstrated that PROM2 overexpression was closely linked to an increased metastatic potential through the increase of epithelial-to-mesenchymal transition (EMT) marker expression and ferroptosis resistance. This was also found in renal cell carcinoma and triple negative breast cancer patient-derived xenograft models. Using an oligonucleotide anti-sense anti-PROM2, we efficaciously decreased PROM2 expression and prevented metastases in melanoma xenografts. We also demonstrated that PROM2 was implicated in an aggravation loop, contributing to increase the metastatic burden both in murine metastatic models and in patients with metastatic melanoma. The metastatic burden is closely linked to PROM2 expression through the expression of EMT markers and ferroptosis cell death resistance in a deterioration loop. Our results open the way for further studies using PROM2 as a bio-target in resort situations in human metastatic melanoma and also in other cancer types.

The value of the improved percutaneous and intravenous contrast-enhanced ultrasound diagnostic classification in sentinel lymph nodes of breast cancer.

Metastatic burden of sentinel lymph node (SLN) in breast cancer patient is the basis for the decision to choose SLN biopsy or axillary lymph node dissection (ALND). However, the diagnostic performance of the previous percutaneous contrast-enhanced ultrasound (P-CEUS) and intravenous contrast-enhanced ultrasound (IV-CEUS) pattern were not satisfied. This study aimed to establish new classification based on structural characteristics for P-CEUS and IV-CEUS of SLN in breast cancer and evaluate the diagnostic efficacy. This retrospective study included consecutive breast cancer patients who had not received neoadjuvant therapy in the First Affiliated Hospital of Sun Yat-sen University between June 2019 and December 2021. Conventional ultrasound, P-CEUS and IV-CEUS were performed. The new classification methods for P-CEUS and IV-CEUS of SLN were established based on structural characteristics of SLN. Pathology was considered as the gold standard, the diagnostic efficacy of P-CEUS, IV-CEUS and combined contrast-enhanced ultrasound in SLNs was analyzed. The detection rate of SLN by P-CEUS in 368 patients was 95.42%. The P-CEUS pattern of SLNs was divided into six types. The IV-CEUS sequence was divided into three types. The IV-CEUS mode was divided into four types. Among the 438 SLNs detected by P-CEUS, 105 (23.97%) were malignant and 333 (76.03%) were benign. Among the previously classified P-CEUS, P-CEUS, IV-CEUS and combined contrast-enhanced ultrasound, the latter had the highest diagnostic efficacy (P<0.05), with sensitivity, specificity, positive predictive value, negative predictive value, accuracy and area under curve (AUC) of 81.90% (86/105), 97.30% (324/333), 90.53% (86/95), 94.46% (324/343), 93.61% (410/438) and 0.896 (0.864-0.923), respectively. The new classification of the P-CEUS and IV-CEUS features of SLNs was performed based on structural characteristics of lymph nodes. Compared with the previously classified P-CEUS, the new classification method has higher diagnostic performance. The combination of new classified P-CEUS and IV-CEUS is helpful to further improve the diagnostic performance of SLNs.

Poor patient outcome correlates with active engulfment of cytokeratin positive CTCs within cancer-associated monocyte population in lung cancer

High rates of mortality in non-small cell lung cancer lung cancer is due to inherent and acquired resistance to systemic therapies and subsequent metastatic burden. Metastasis is supported by suppression of the immune system at secondary organs and within the circulation. Modulation of the immune system is now being exploited as a therapeutic target with immune checkpoint inhibitors. The tracking of therapeutic efficacy in a real-time can be achieved with liquid biopsy, and evaluation of circulating tumour cells and the associated immune cells. A stable liquid biopsy biomarker for non-small cell lung cancer lung cancer has yet to be approved for clinical use. We performed a cross-sectional single-site study, and collected liquid biopsies from patients diagnosed with early, locally advanced, or metastatic lung cancer, undergoing surgery, or systemic therapy (chemotherapy/checkpoint inhibitors). Evaluation of overall circulating tumour cell counts, or cluster counts did not correlate with patient outcome. Interestingly, the numbers of Pan cytokeratin positive circulating tumour cells engulfed by tumour associated monocytes correlated strongly with patient outcome independent of circulating tumour cell counts and the use of checkpoint inhibitors. We suggest that Pan cytokeratin staining within monocytes is an important indicator of tumour-associated inflammation post-therapy and an effective biomarker with strong prognostic capability for patient outcome.

Oleic acid-PPARγ-FABP4 loop fuels cholangiocarcinoma colonization in lymph node metastases microenvironment.

Lymph node metastasis is a significant risk factor for patients with cholangiocarcinoma, but the mechanisms underlying cholangiocarcinoma colonization in the lymph node microenvironment remain unclear. We aimed to determine whether metabolic reprogramming fueled the adaptation and remodeling of cholangiocarcinoma cells to the lymph node microenvironment. Here, we applied single-cell RNA sequencing of primary tumor lesions and paired lymph node metastases from patients with cholangiocarcinoma and revealed significantly reduced intertumor heterogeneity and syntropic lipid metabolic reprogramming of cholangiocarcinoma after metastasis to lymph nodes, which was verified by pan-cancer single-cell RNA sequencing analysis, highlighting the essential role of lipid metabolism in tumor colonization in lymph nodes. Metabolomics and in vivo CRISPR/Cas9 screening identified PPARγ as a crucial regulator in fueling cholangiocarcinoma colonization in lymph nodes through the oleic acid-PPARγ-fatty acid-binding protein 4 positive feedback loop by upregulating fatty acid uptake and oxidation. Patient-derived organoids and animal models have demonstrated that blocking this loop impairs cholangiocarcinoma proliferation and colonization in the lymph node microenvironment and is superior to systemic inhibition of fatty acid oxidation. PPARγ-regulated fatty acid metabolic reprogramming in cholangiocarcinoma also contributes to the immune-suppressive niche in lymph node metastases by producing kynurenine and was found to be associated with tumor relapse, immune-suppressive lymph node microenvironment, and poor immune checkpoint blockade response. Our results reveal the role of the oleic acid-PPARγ-fatty acid-binding protein 4 loop in fueling cholangiocarcinoma colonization in lymph nodes and demonstrate that PPARγ-regulated lipid metabolic reprogramming is a promising therapeutic target for relieving cholangiocarcinoma lymph node metastasis burden and reducing further progression.

Impact of Complete Surgical Resection of Metastatic Lesions in Patients with Advanced Renal Cell Carcinoma in the Era of Tyrosine Kinase Inhibitors and Immune Checkpoint Inhibitors.

Complete metastasectomy (CM) in metastatic renal cell carcinoma (mRCC) has demonstrated efficacy in the cytokine era, but its effectiveness in the era of tyrosine kinase inhibitors (TKIs) and immune checkpoint inhibitors (ICIs) remains unclear. A multi-institutional database included clinicopathological data of 367 patients with mRCC. Patients were divided into two groups: the CM group and the non-CM group. These two groups were compared before and after propensity score matching (PSM). Cox proportional hazard models were used to detect factors associated with disease-free survival (DFS) and overall survival (OS) from mRCC diagnosis. The CM group showed a significant association with longer overall survival compared to the non-CM group in the PSM-unadjusted cohorts (p < 0.001, hazard ratio 0.49, 95% confidence interval 0.35-0.69), but no superiority was noted in the adjusted cohorts. The median DFS after CM was 24 months, with no significant differences based on relapse timing. Notably, the international metastatic RCC database consortium risk categories and metastatic burden were associated with DFS. This study supports the potential of CM in mRCC management during the TKI/ICI era, although limitations including sample size and selection bias need to be considered.

Comparison of ex vivo bioluminescence imaging, Alu-qPCR and histology for the quantification of spontaneous lung and bone metastases in subcutaneous xenograft mouse models

Bioluminescence imaging (BLI) is a non-invasive state-of-the-art-method for longitudinal tracking of tumor cells in mice. The technique is commonly used to determine bone metastatic burden in vivo and also suitable ex vivo to detect even smallest bone micro-metastases in spontaneous metastasis xenograft models. However, it is unclear to which extent ex vivo BLI correlates with alternative methods for metastasis quantification. Here, we compared ex vivo BLI, human DNA-based Alu-qPCR, and histology for the quantification of bone vs. lung metastases, which are amongst the most common sites of metastasis in prostate cancer (PCa) patients and spontaneous PCa xenograft models. Data from 93 immunodeficient mice were considered, each of which were subcutaneously injected with luciferase/RGB-labeled human PCa PC-3 cells. The primary tumors were resected at ~ 0.75 cm³ and mice were sacrificed ~ 3 weeks after surgery and immediately examined by ex vivo BLI. Afterwards, the right lungs and hind limbs with the higher BLI signal (BLIHi bone) were processed for histology, whereas the left lung lobes and hind limbs with the lower BLI signal (BLILo bone) were prepared for Alu-qPCR. Our data demonstrate remarkable differences in the correlation coefficients of the different methods for lung metastasis detection (r ~ 0.8) vs. bone metastasis detection (r ~ 0.4). However, the BLI values of the BLIHi and BLILo bones correlated very strongly (r ~ 0.9), indicating that the method per se was reliable under identical limitations; the overall level of metastasis to contralateral bones was astonishingly similar. Instead, the level of lung metastasis only weakly to moderately correlated with the level of bone metastasis formation. Summarized, we observed a considerable discrepancy between ex vivo BLI and histology/Alu-qPCR in the quantification of bone metastases, which was not observed in the case of lung metastases. Future studies using ex vivo BLI for bone metastasis quantification should combine multiple methods to accurately determine metastatic load in bone samples.

Genomic discordances and heterogeneous mutational burden, PD-L1 expression and immune infiltrates of non-small cell lung cancer metastasis

AimsTo investigate the genomic discordances and heterogeneous mutational burden, PD-L1 expression and immune cell (IC) infiltrates of non-small cell lung cancer (NSCLC) metastasis.MethodsSurgical samples from 41 cases of NSCLC with...

Predicting abiraterone efficacy in advanced prostate cancer: Insights from marker of proliferation Ki-67.

176 Background: The marker of proliferation Ki-67 (KI67) is a well-known biomarker reflecting cell proliferation activity and was reported to be associated with the treatment efficacy of chemotherapy in different tumors. However, it is unknown whether KI67 also has a role in predicting the efficacy of the next-generation hormone therapies, e.g., abiraterone, for patients with prostate cancer (PCa). Methods: Clinicopathological data of 144 men with metastatic PCa who received abiraterone therapy were retrospectively collected. The KI67 positivity was examined by immunohistochemistry (IHC) using the prostate biopsy specimen. The predictive value of KI67 and other factors on the therapeutic efficacy of abiraterone treatment was explored. Kaplan-Meier curve and COX regression analysis were used for survival analysis. The endpoints were PSA progression-free survival (PSA-PFS) and radiographic progression-free survival (rPFS) according to the PCWG3 criteria. Results: Among the 144 included patients, 82 (56.9%) and 62 (43.1%), respectively, received abiraterone therapy at metastatic hormone-sensitive (mHSPC) and castration-resistant PCa (mCRPC) stage. The median KI67 positivity was 20% for the total cohort (interquartile range: 10%-30%). When taken as a continuous variable, KI67 positivity was adversely related to both PSA-PFS (HR, 95%CI: 1.02, 1.01-1.03, P=0.001) and rPFS (HR, 95%CI: 1.02, 1.02-1.03, P=0.001) of the abiraterone therapy, while was not associated with PSA response. The results of multivariate COX regression analysis implied that, after adjusting by mHSPC/mCRPC stage, ISUP grading, pain score, visceral metastasis, and metastatic burden, KI67 was still an independent predictor of PSA-PFS (HR, 95%CI: 1.02, 1.01-1.03, P=0.001) and rPFS (HR, 95%CI: 1.02, 1.01-1.03, P=0.008) of the abiraterone therapy. Moreover, when patients were divided into two groups with high and low KI67, using cutoffs of 10%, 20%, and 30%, KI67 remained a significant predictor of both PSA-PFS (HR: 1.67, 1.91, 2.29; P value: 0.052, 0.004, &lt;0.001) and rPFS (HR: 1.54, 1.83, 1.88; P value: 0.119, 0.008, 0.008) of the abiraterone treatment. Subgroup analysis based on whether patients received abiraterone at mHSPC or mCRPC stage suggested that KI67 30% (median PSA-PFS: 11.4- vs. 27.6-Mo, P&lt;0.001; median rPFS: 16.6- vs. 33.6-Mo, P=0.001) was the optimal cutoff to maximize patient prognostic differentiation for the mHSPC cohort, while KI67 20% (median PSA-PFS: 6.9- vs. 14.9-Mo, P=0.033; median rPFS: 10.1- vs. 16.5-Mo, P=0.030) was the optimal cutoff for the mCRPC cohort. Conclusions: Our study revealed that KI67 positivity in prostate biopsy specimens was a robust predictor of abiraterone efficacy for patients with advanced PCa. Further validation using datasets from other centers is needed to strengthen the findings of our work.

Abstract PR03: CSF-1R antibody targeting therapy with combined metronomic chemotherapy and immune checkpoint blockade enhance a B and T cell response to attenuate metastatic triple negative breast cancer

Abstract Background: Increased macrophage infiltration and elevated levels of Epithelial to Mesenchymal Transition (EMT) gene signatures are associated with a poor outcome following neo-adjuvant chemotherapy in patients with triple negative breast cancer (TNBC). Further, metastatic disease with overall the poorest survival rates has a cold tumor immune microenvironment (TIME), especially in lung and liver metastases. Interestingly, an abundance of macrophages are observed at these metastatic sites. We have extensively characterized multiple preclinical syngeneic Trp53-/- tumor models that have extensive Tumor Associated Macrophage (TAM) infiltration. These tumor models transcriptionally resemble “claudin-low”, “basal-like” and “luminal-like” breast cancers. From these models the “claudin-low” closely phenocopy the high EMT/TAM subtype observed in patients. Objective: Accordingly, we asked if SNDX-ms6352 a novel specific, high affinity monoclonal antibody targeting CSF-1R in combination with Chemotherapy (CTX) and immune checkpoint blockade might provide an alternative approach for treating established lung and liver metastases. Methods: Single cells from primary T12 mammary tumors were introduced either via tail vein injection to obtain lung metastases or portal vein (PV) injection for liver metastases. Mice were injected with BrdU and multiple lung and liver tissues are harvested randomly to identify established metastases. The mice were randomized into treatment groups and administered four weekly treatments of IgG, CTX, SNDX-ms6352, CTX+SNDX-ms6352 or CTX+SNDX-ms6352+aPD1. Tissues were harvested and fixed overnight in 4% PFA then placed in 70% EtOH for paraffin embedding and stained for H&amp;E, Immunohistochemistry (IHC), Immunofluorescence (IF) and Imaging Mass Cytometry (IMC). Results: Following four weekly treatments of IgG, SNDX-ms6352 and CTX+/-SNDX-ms6352 we observed a decrease in lung metastatic burden and an increase in overall survival as compared to IgG, CTX and SNDX-ms6352 alone. Interestingly we also observed increased CD8 T-cell infiltration, but only in the combination treated mice 28 and 56 days post treatment. However, lung metastases recurred in all of the mice several weeks later. Data from IMC showed intra-tumoral heterogeneity with increased PD-L1 and Ki67. Short term treated lung and liver metastases with the addition of aPD1 demonstrated increased CD20+ B and CD8+ T cell expansion and cleared most of the lung and liver metastatic burden. These results suggest that targeting macrophages enhances the immunostimulatory effect of metronomic CTX with immune checkpoint blockade in treating not only primary tumors, but also established lung and liver metastases via the activation of the tumor immune microenvironment. Supported by R01CA148761-12 and American Cancer Society – AutoZone Breast Cancer Postdoctoral Fellowship, PF-22-163-01-MM grants. Citation Format: Diego A Pedroza, Qian Zhu, Weigou Wu, Paul Porter, Xiang H-F Zhang, Jeffrey M Rosen. CSF-1R antibody targeting therapy with combined metronomic chemotherapy and immune checkpoint blockade enhance a B and T cell response to attenuate metastatic triple negative breast cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Breast Cancer Research; 2023 Oct 19-22; San Diego, California. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_1):Abstract nr PR03.

The prognostic value of ductal adenocarcinoma of the prostate in patients with advanced prostate cancer treated with abiraterone acetate.

167 Background: Previous studies have indicated that ductal adenocarcinoma of the prostate (DA) is associated with adverse prognosis in patients with localized prostate cancer (PCa). However, the clinical significance of DA in advanced PCa remains largely ambiguous. This study endeavors to investigate the relationship between the existence of DA in prostate biopsy specimens and treatment outcomes among patients with advanced PCa receiving abiraterone acetate (AA) therapy. Methods: We retrospectively analyzed data from 440 patients with advanced PCa who received AA at either the metastatic hormone-sensitive (mHSPC, N=123) or castration-resistant PCa (mCRPC, N=317) stage. The presence of DA and its proportion was evaluated based on prostate biopsy specimens. Kaplan-Meier curves and COX regression were used to evaluate the predictive significance of DA on AA efficacy, including PSA response, PSA progression-free survival (PSA-PFS), and radiographic progression-free survival (rPFS). Results: In aggregate, DA was detected in 35/440 (8.0%) patients, with 13/440 (3.0%) and 22/440 (5.0%) men harboring DA&lt;5% and ≥5%, respectively, in the total tumors. The overall PSA response rate in the entire cohort was 302/440 (68.6%), and it was comparable for people with and without DA, with both groups having a response rate of 68.6%. Of note, compared with patients without DA, men with DA have significantly longer median PSA-PFS (mPSA-PFS, 12.1- vs. 25.0-Mo, P=0.005) and median rPFS (mrPFS, 19.8 vs. 38.2-Mo, P=0.034). Subgroup analysis based on DA proportion further revealed that in comparison with PCa without DA, DA≥5% was an indicator of favorable PSA-PFS (HR, 95%CI: 0.28, 0.14-0.56, p&lt;0.001) and rPFS (HR, 95%CI: 0.35, 0.18-0.72, p=0.004), while DA&lt;5% was not. In multivariate COX regression analysis, after adjusting by clinicopathological factors, including mCRPC/mHSPC stage, ISUP grading, metastatic burden, pain score and pretreatment hemoglobin, alkaline phosphatase, and lactate dehydrogenase, DA≥5% was still independently associated with better therapeutic efficacy of AA treatment (PSA-PFS: HR, 95%CI: 0.36, 0.17-0.74, p&lt;0.001; rPFS: HR, 95%CI: 0.47, 0.23-0.96, p=0.040). Conclusions: A DA proportion ≥5% in the tumor is related to improved treatment efficacy for individuals with advanced PCa receiving AA. Thus, in the pathological diagnosis of PCa, it is necessary to routinely report the presence and proportion of DA in order to aid with treatment decision-making.

Optimal sequential therapy for metastatic castration-resistant prostate cancer after androgen receptor pathway inhibitors in the up-front era.

168 Background: The optimal sequential therapy for metastatic castration-resistant prostate cancer (mCRPC) following the use of up-front androgen receptor pathway inhibitors (ARPIs) for metastatic hormone-sensitive prostate cancer (mHSPC) is still unclear. Methods: A total of 220 patients who received systemic therapy for de novo mHSPC at Nagoya University Hospital and its affiliated institutions between 2014 and 2022 were included in the study. Patients who received up-front therapy with ARPI as first-line therapy for mHSPC, followed by ARPI as primary therapy for mCRPC were defined as the AA group, followed by docetaxel (DOC) as primary therapy for mCRPC were defined as the AD group. On the other hand, the vAA group was defined as patients who received androgen deprivation therapy alone or combined androgen blockade as “vintage” first-line therapy for mHSPC, followed by two consecutive ARPIs as sequential therapy for mCRPC, and the vAD group was defined as patients who received ARPI followed by DOC as sequential therapy for mCRPC. Patient characteristics and progression-free survival (PFS) of the second agent after the first ARPI were compared in each group. Results: We identified 14 patients in the AA group, 16 in the AD group, 84 in the vAA group, and 59 in the vAD group. There were no differences in patient background such as initial PSA, Gleason score, and metastatic burden between the groups, while patient age was lower in the AD and vAD groups compared to the AA and vAA groups. The PFS of the second agent in the AA group was significantly shorter than that in the vAA group (median, 6.8 vs. 1.9 months; hazard ratio [HR], 0.25; 95% confidence interval [CI], 0.25-0.87; p=0.017). In contrast, there was no significant difference in PFS of the second agent in the vAD and AD groups (median, 7.0 vs. 4.0 months; HR, 0.63; 95%CI, 0.29-1.34; p=0.235). Conclusions: The effect of each agent after ARPI therapy in the UP-FRONT era may be poorer than that in the VINTAGE era. In the setting after up-front ARPI, sequential therapy with DOC may be more appropriate than sequential therapy with ARPI.

Role of Bone Scan Index (BSI) in the Prognosis and Treatment Efficacy in Castration-Sensitive Prostate Cancer Patients with Bone Metastasis

Bone is the most common metastatic site in prostate cancer (PCa). Although the extent of disease (EOD) grade is used for evaluating burden of bone metastasis, the accuracy of bone metastasis classification needs improvement. Bone scan index (BSI) was developed as a quantitative tool to enhance the interpretability and clinical relevance of the bone scan. This study aimed to explore the role of BSI using BONENAVI® software in determining the prognosis and treatment efficacy in castration-sensitive PCa (mCSPC) patients with bone metastasis. We retrospectively reviewed 61 mCSPC patients with bone metastasis who had received primary androgen deprivation therapy (PADT) at our institution. All patients received PADT with luteinizing hormone-releasing hormone agonist or surgical castration accompanied by first-generation antiandrogen, bicalutamide. Bone scans were performed with ⁹⁹[m]Tc-MDP. BSI (%) was divided into two groups (＜1.0 and ≧1.0), and BSI response rates(change at 0 months to after 6 months) were determined using thresholds of 45% decline. Castration-resistant prostate cancer (CRPC) -free survival (CRPC-FS) and Overall survival (OS) rates were analyzed using the Kaplan-Meier method. The median follow-up was 41. 9 months. Overall, 16 patients (26. 2%) died. Multivariate analysis on pretreatment factors revealed that hemoglobin (P＝0.03) and BSI (P＝0.04) were independent prognostic factors for OS. The 5-year OS rates in patients with low BSI and high BSI were 84.6% and 39.2%, respectively (P＝0.02). In 40 patients who had a bone scan before and after PADT, OS rates in patients with a good response (≧45%) were significantly higher than those with a poor response (＜45%) (P＝0.001). Nadir PSA titers within 6 months after the start of treatment (P＝0.005), Hb (P＝0.003), and BSI change (P＝0.014) were independent prognostic factors for OS. In mCSPC patients with bone metastases, BSI at diagnosis was an important predictor of CRPC progression and OS as a pre-treatment factor, and BSI change rate and PSA nadir as post-treatment factors.

A real-world observational study characterizing patients (pts) with metastatic castration-sensitive prostate cancer (mCSPC) treated with or without androgen receptor pathway inhibitors (ARPIs).

59 Background: mCSPC treatment is rapidly evolving with androgen deprivation therapy (ADT) plus ARPIs viewed as the standard of care for most pts. Data are needed to understand the real-world characteristics and treatment patterns of mCSPC (PC) pts in this dynamic field. This study aimed to define clinical features of mCSPC pts, estimate the proportion of pts receiving ARPIs, and evaluate clinical and demographic features associated with ARPI use. Methods: This retrospective cohort study used data from electronic health records and administrative databases in the province of Alberta, Canada. All newly diagnosed metastatic PC pts between 2017-2020 were identified. Pts were considered to have mCSPC if they initiated ADT within 30 days prior to, or at any time after diagnosis. ARPI exposure was defined as receiving ARPI within 180 days of initiating ADT. ARPI-naïve pts may have received ADT alone or other non-ARPI therapy (i.e., docetaxel) within 180 days of initiating ADT. Multivariable logistic regression was used to evaluate pt characteristics related to the receipt of ARPI. Overall survival (OS) was defined as the date of diagnosis to death from any cause or last known contact. Results: Of the 976 mCSPC pts identified, 33.5% received an ARPI. In ARPI exposed pts, the median time from ADT to ARPI start was 7.9 weeks and median time on ARPI was 13.1 months (mos). In multivariable analyses, ARPI use was associated with younger pt age, more recent diagnosis, fewer comorbidities, a higher metastatic burden, treatment centre, referral to a medical oncologist, and prior local therapy (all p&lt;0.05). The median time to next therapy was 24.6 mos (95%CI=19.5-33.0) vs. 18.5 mos (95%CI+16.9-21.1), and median OS was 38.5 mos (95%CI=32.8-NA) vs. 34.2 mos (95%CI=33.3-38.8) for APRI exposed vs. ARPI-naive pts, respectively (p=0.03). Conclusions: This study identified factors associated with ARPI use in mCSPC, potentially allowing targeted efforts to improve ARPI uptake for pts in whom use is low. These findings are important as they reflect outcomes seen in real world pts.

The effect of radiotherapy on the course of disease in prostatic cancer with high metastatic burden.

112 Background: Many retrospective studies suggest that local RT for the prostate in the metastatic setting improves overall survival, however the question of whether metastatic burden is an independent factor influencing the results of localized treatment was not directly addressed. Moreover, results are conflicting with the outcomes from prospective trials on the subject. Thus overall, there is a limited, conflicting and insufficient data on the effect of radiotherapy on prostate cancer patients with a significant metastatic burden. Here we show that RT treatment in prostate cancer patients with a significant metastatic burden (four or more bone metastases with one or more outside the vertebral bodies or pelvis or a visceral metastases) is beneficial and extends overall survival while reducing symptoms. Methods: We conducted a retrospective study examining 100 high metastatic burden prostate cancer patients treated in Hadassah Ein Karem Medical Center between the years of 2004-2022. Data was collected from the medical records of the hospital. Variables that were recorded include basic demographics, time and durations of hospitalization, cancer staging, treatment course, RT course, imaging after RT, vital status, clinical symptoms and PSA levels. Tumors could be of any grade (Gleason score 6–10) and T or N stage (T1-T4; N0-N1; M1). Prostate RT included external EBRT to the prostate with/ without pelvis. Radiotherapy dosage between 30-60 gy using conventional fractionation. Results: Overall Survival was higher in the RT group when compared to NRT group (P &lt; 0.046). Progression free survival (PFS) was not significantly higher in the RT group as compared to the NRT group (P &lt; 0.153). However, a trend towards better PFS was noted. Biochemical progression free survival was higher in the RT group when compared to NRT group (P &lt; 0.033). Time without symptom worsening as defined by time since treatment beginning until worsening of a symptom or appearance of a new symptom reported by the patient included urinary incontinence, urinary urgency, and increased frequency, need for catheter or urinary obstruction. Time without symptom worsening was higher in the RT group as compared to NRT group (P &lt; 0.017). The amount of hospitalization wasn’t significantly higher in the RT group when compared to NRT group (P &lt; 0.460). The duration of hospitalization was significantly higher in the NRT group when compared to the RT group (P &lt; 0.038). Conclusions: Here we show for the first time an increase of overall survival specifically in the high metastatic burden group when adding the RT to a standard treatment of choice (ADT with or without chemotherapy). We also demonstrate an improvement in QOL of our patients.

Abstract A018: Dihydroceramide desaturase 1 (DES1) promotes anchorage-independent survival and metastasis of HER2-positive and basal breast cancer through regulation of caspase 14

Abstract Despite improvements in therapies, metastasis still leads to high mortality rates in breast cancer (BC). Consequently, discovering new therapies that are effective at targeting metastatic disease remains a critical need, particularly for the aggressive HER2+ and basal BC subtypes. The ability of cancer cells to survive in the absence of an extracellular matrix – termed anchorage-independent survival (AIS) – is central to the metastatic cascade and has emerged as a target biology of interest for reducing metastatic burden. Previously, we identified the SL enzyme dihydroceramide desaturase 1 (DES1) as a critical node for AIS of HER2+ BC cells. Here, we have extended these data to show that targeting DES1 through pharmacological inhibitors (4HPR), siRNA, or CRISPR-cas9 knockout was able to overcome AIS in basal BC cells as well as HER2+ BC cells that, functionally, led to a reduction of in vitro colony formation in soft agar. Extending these findings in vivo, we found that DES1 knockout led to a significant reduction in lung metastasis of both HER2+ and basal BC cells in murine models of experimental metastasis. Moreover, treatment of PyMT mice with the DES1 inhibitor 4HPR was able to reduce metastatic burden. To define downstream effectors of DES1 required for AIS, we performed a microarray analysis of control and DES1 knockout cells in suspension, leading us to identify caspase 14 as a downstream effector of DES1 in HER2+ and basal BC. Consistent with these findings, siRNA knockdown of caspase 14 led to a striking decrease in cell viability in colony formation assays. Finally, analysis of publicly available data found that high levels of DES1 and Caspase 14 were associated with poor clinical outcomes in both HER2+ and basal BC. Taken together, these data establish DES1 as a mediator of BC metastasis, likely through promoting AIS, and define caspase 14 as a target of DES1-driven signaling. Furthermore, results suggest that both DES1 and caspase 14 expression may be useful as biomarkers of metastasis-prone tumors. Citation Format: Deanna M Peperno, Danielle Lambadis, Victoria Alvarado, Andrew E Resnick, Christopher J Clarke. Dihydroceramide desaturase 1 (DES1) promotes anchorage-independent survival and metastasis of HER2-positive and basal breast cancer through regulation of caspase 14 [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Breast Cancer Research; 2023 Oct 19-22; San Diego, California. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_1):Abstract nr A018.

Abstract A051: Mitochondrial metabolism and fission as dynamic drivers of breast cancer metastasis

Abstract Approximately 30% of women with early-stage breast cancer will develop metastatic disease, making efforts to define drivers of metastasis critical. Our group found that metastatic breast cancer cells express an oxidative phosphorylation (OXPHOS) gene signature. Further, inhibiting OXPHOS reduced metastatic seeding, thus revealing the functional role of mitochondrial metabolism in breast cancer metastasis. It is understood that mitochondria are highly dynamic organelles that are regulated by a balance of fission and fusion proteins; however, it remains poorly understood how mitochondrial metabolism and their dynamics promote cancer spreading. To interrogate mitochondrial heterogeneity within primary tumors, we sorted tumor cells from patient derived xenograft (HCI010) and murine (4T1) breast tumors stratified by low or high uptake of tetramethylrhodamine methyl ester (TMRM, a dye that accumulates in active mitochondria undergoing OXPHOS). Low or high cell fractions were re-injected intravenously in new cohorts of mice. We found greater metastatic burden in the lungs of mice injected with the TMRM high fraction. Furthermore, we uncovered that the TMRM high cells had greater mitochondria abundance than TMRM low cells. We hypothesized the differences in mitochondrial content we observed in primary tumor cells is caused by shifts in cellular expression of mitochondria fission and fusion proteins. Consistent with this hypothesis, we found that TMRM high, metastatic tumor cells had greater expression of mitochondria fission gene, DRP1, and lower expression of fusion gene, MFN2. Collectively, our results revealed heterogeneity in tumor cell mitochondrial content within a primary tumor that appears to influence the fitness of a cell to metastasize. Lastly, our data supports a role for mitochondrial fission in breast cancer metastasis. Future directions for our studies will include genetic and pharmacological modulation of mitochondrial fission and fusion to better understand why it influences metastasis and at what point during the metastatic cascade. Citation Format: Hannah Savage, Dennis Ma, Devon Lawson. Mitochondrial metabolism and fission as dynamic drivers of breast cancer metastasis [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Breast Cancer Research; 2023 Oct 19-22; San Diego, California. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_1):Abstract nr A051.

PD-1 inhibitor plus oncolytic vaccinia virus is a safe and effective treatment option for metastatic renal cell carcinoma

BackgroundAlthough a combination of immune checkpoint inhibitors (ICIs) is recommended as the first line treatment option for metastatic renal cell carcinoma (mRCC), several immune-related adverse events (irAEs) occur, especially hepatitis. We explored the therapeutic benefits and safety profile of combining oncolytic vaccinia virus, JX-594, with a programmed cell death protein-1 (PD-1) inhibitor.MethodsWe used early-stage and advanced-stage orthotopic murine mRCC models developed by our group. PD-1 inhibitor monotherapy or a PD-1 inhibitor combined with either JX-594 or a cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) inhibitor were systemically injected through the peritoneum. An immunofluorescence analysis was performed to analyze the tumor immune microenvironment (TIME). irAEs were assessed in terms of hepatitis.ResultsIn the early-stage mRCC model mice, the combination of JX-594 and a PD-1 inhibitor significantly decreased the primary tumor size and number of lung nodules, compared with the ICI combination, but the JX-594 and PD-1 inhibitor combination and ICI combination did not differ significantly in the advanced-stage mRCC model mice. The JX-594 and PD-1 inhibitor combination induced tumor-suppressing TIME changes in both the early- and advanced-stage mRCC models. Furthermore, mice treated with the ICI combination had significantly greater hepatic injuries than those treated with the JX-594 and PD-1 inhibitor combination which was evaluated in early-stage mRCC model.ConclusionsThe JX-594 and PD-1 inhibitor combination effectively reduced primary tumors and the metastatic burden, similar to ICI combination therapy, through dynamic remodeling of the TIME. Furthermore, hepatitis was significantly decreased in the JX-594 and PD-1 inhibitor combination group, suggesting the potential benefit of that combination for reducing ICI-induced toxicity.

A telomere-targeting drug depletes cancer initiating cells and promotes anti-tumor immunity in small cell lung cancer

There are few effective treatments for small cell lung cancer (SCLC) underscoring the need for innovative therapeutic approaches. This study focuses on exploiting telomerase, a critical SCLC dependency as a therapeutic target. A prominent characteristic of SCLC is their reliance on telomerase activity, a key enzyme essential for their continuous proliferation. Here we utilize a nucleoside analog, 6-Thio-2’-deoxyguanosine (6TdG) currently in phase II clinical trials, that is preferentially incorporated by telomerase into telomeres leading to telomere dysfunction. Using preclinical mouse and human derived models we find low intermittent doses of 6TdG inhibit tumor growth and reduce metastatic burden. Anti-tumor efficacy correlates with a reduction in a subpopulation of cancer initiating like cells (CICs) identified by their expression of L1CAM/CD133 and highest telomerase activity. 6TdG treatment also leads to activation of innate and adaptive anti-tumor responses. Mechanistically, 6TdG depletes CICs and induces type-I interferon signaling leading to tumor immune visibility by activating tumor cell STING signaling. We also observe increased sensitivity to irradiation after 6TdG treatment in both syngeneic and humanized SCLC xenograft models both of which are dependent on the presence of host immune cells. This study underscores the immune-enhancing and metastasis-reducing effects of 6TdG, employing a range of complementary in vitro and in vivo SCLC preclinical models providing a potential therapeutic approach to SCLC.

Subgroup analyses of the phase 3 CAIRO3 trial: Association of number and total volume of metastases on survival in patients with metastatic colorectal cancer based on a deep-learning automatic segmentation model.

130 Background: Maintenance treatment with capecitabine and bevacizumab after capecitabine, oxaliplatin, and bevacizumab therapy in metastatic colorectal cancer (mCRC) is shown to be effective with observation as an acceptable alternative. Metastatic burden might aid in shared decision-making and the aim of this exploratory analysis is to assess the prognostic and predictive value of number and total volume of metastases in mCRC patients included in the CAIRO3 study. Methods: Patients in the CAIRO3 study with liver and/or lung metastases were included. Every individual liver and lung metastasis was counted and volumes were assessed using deep-learning automatic segmentation software (SAS analytical platform and Philips IntelliSpace). Lymph node metastases were counted manually. Each scan was supervised by a dedicated radiologist. Kaplan-Meier analyses were used to estimate progression-free survival (PFS) and overall survival (OS) using log-rank testing for statistical comparison. Hazard ratios (HR) were estimated using Cox regression models. Results: A total of 156 patients were included with 154 OS events, in which 8198 metastases were identified: 5179 were liver metastases, 2835 lung metastases, and 184 lymph node metastases. Patients had a mean age of 63 years and 67% were male. At randomization, the median number of metastases was 9 and mean total metastatic volume was 116 cm3. Median OS for patients with ≤10 versus &gt;10 metastases was 22.0 months (95%CI: 19.3-29.3) and 14.3 months (95% CI: 11.6-20.9, p-value &lt;0.01), with similar differences for low/high tumor volume. In multivariable analysis number and total volume were independent prognostic markers for PFS and OS, respectively, after adjustment for age, sex, WHO performance score, treatment arm, serum lactate dehydrogenase (LDH) level, site of primary tumor, time to metastases, and treatment response. Benefit of maintenance therapy was most pronounced in patients with ≤10 compared to &gt;10 metastases with a PFS HR of 0.24 (0.12-0.46) compared to 0.52 (0.28-0.96) and OS HR of 0.75 (0.42-1.34) compared to 1.17 (0.66-2.08), demonstrating potential predictive value. Conclusions: Metastatic burden is associated with a poor prognosis in patients with mCRC. Furthermore, patients with limited metastatic disease (1-10 metastases) benefit most from maintenance treatment and this could potentially be a predictive biomarker. [Table: see text]