Skeletal Tumor Burden Research Articles

Impact of somatic DNA repair mutations on bone-related efficacy endpoints in patients with metastatic castration-resistant prostate cancer (mCRPC): A retrospective analysis.

214 Background: Up to 80% of mCRPC have bone metastases and about 25% of metastatic prostate cancer harbor mutations in DNA damage repair defects (DDR). DDR gene mutations are a negative prognostic factor, being associated with short metastasis-free survival and cancer-specific survival. We analyzed the impact of somatic mutations in DDR genes on bone-related efficacy endpoints in mCRPC. Methods: We retrospectively analyzed the mutational status of mCRPC patients per FoundationOne analysis in tissue biopsy or, when it was not possible, in liquid biopsy (performed at the onset of mCRPC). We evaluated the impact of DDR gene mutations on bone-related efficacy endpoints at the time of mCRPC diagnoses, by dividing patients in two main groups (mutated DDR, specifically, mutations in at least one of BRCA1/2, ATM, FANCA, ATR, CHEK2, RAD51, PALB2, and CDK12 genes: group A; wild-type DDR: group B). We investigated differences between the two groups in terms of time from bone metastases onset to death, skeletal metastatic tumor burden (sites and number of lesions), skeletal-related events (SRE) incidence, and time to first on-study SRE. Results: Of the 131 patients with mCRPC enrolled, 115 had bone metastases. Clinical data distribution, according to molecular profile, is presented. There is no difference in terms of sites of metastases and time from bone metastases onset to death between the 2 groups. Mutated DDR status was associated with bone metastases volume (p = 0.0325), but did not affect SRE incidence and time to SRE onset. Conclusions: In our analysis, mutated DDR status was associated with higher bone metastases volume, although a not detrimental effect on the other bone-related efficacy endpoints was observed. [Table: see text]

Automated quantification of PET/CT skeletal tumor burden in prostate cancer using artificial intelligence: The PET index

PurposeConsistent assessment of bone metastases is crucial for patient management and clinical trials in prostate cancer (PCa). We aimed to develop a fully automated convolutional neural network (CNN)-based model for calculating PET/CT skeletal tumor burden in patients with PCa.MethodsA total of 168 patients from three centers were divided into training, validation, and test groups. Manual annotations of skeletal lesions in [18F]fluoride PET/CT scans were used to train a CNN. The AI model was evaluated in 26 patients and compared to segmentations by physicians and to a SUV 15 threshold. PET index representing the percentage of skeletal volume taken up by lesions was estimated.ResultsThere was no case in which all readers agreed on prevalence of lesions that the AI model failed to detect. PET index by the AI model correlated moderately strong to physician PET index (mean r = 0.69). Threshold PET index correlated fairly with physician PET index (mean r = 0.49). The sensitivity for lesion detection was 65–76% for AI, 68–91% for physicians, and 44–51% for threshold depending on which physician was considered reference.ConclusionIt was possible to develop an AI-based model for automated assessment of PET/CT skeletal tumor burden. The model’s performance was superior to using a threshold and provides fully automated calculation of whole-body skeletal tumor burden. It could be further developed to apply to different radiotracers. Objective scan evaluation is a first step toward developing a PET/CT imaging biomarker for PCa skeletal metastases.

Parathyroid Hormone-Related Protein Inhibition Blocks Triple-Negative Breast Cancer Expansion in Bone Through Epithelial to Mesenchymal Transition Reversal.

ABSTRACTParathyroid hormone‐related protein (PTHrP) plays a major role in skeletal metastasis but its action mechanism has not been fully defined. We previously demonstrated the crucial importance of PTHrP in promoting mammary tumor initiation, growth, and metastasis in a mouse model with a mammary epithelium‐targeted Pthlh gene ablation. We demonstrate here a novel mechanism for bone invasion involving PTHrP induction of epithelial to mesenchymal transition (EMT) and cancer stem cells (CSCs) regulation. Clustered regularly interspaced short palindromic repeats (CRISPR)‐mediated Pthlh gene ablation was used to study EMT markers, phenotype, and invasiveness in two triple‐negative breast cancer (TNBC) cell types (established MDA‐MB‐231 and patient‐derived PT‐TNBC cells). In vitro, Pthlh ablation in TNBC cells reduced EMT markers, mammosphere‐forming ability, and CD44high/CD24low cells ratio. In vivo, cells were injected intratibially into athymic nude mice, and therapeutic treatment with our anti‐PTHrP blocking antibody was started 2 weeks after skeletal tumors were established. In vivo, compared to control, lytic bone lesion from Pthlh ‐ablated cells decreased significantly over 2 weeks by 27% for MDA‐MB‐231 and by 75% for PT‐TNBC‐injected mice (p < 0.001). Micro‐CT (μCT) analyses also showed that antibody therapy reduced bone lytic volume loss by 52% and 48% for non‐ablated MDA‐MB‐231 and PT‐TNBC, respectively (p < 0.05). Antibody therapy reduced skeletal tumor burden by 45% and 87% for non‐ablated MDA‐MB‐231 and PT‐TNBC, respectively (p < 0.002) and caused a significant decrease of CSC/EMT markers ALDH1, vimentin, and Slug, and an increase in E‐cadherin in bone lesions. We conclude that PTHrP is a targetable EMT molecular driver and suggest that its pharmacological blockade can provide a potential therapeutic approach against established TNBC‐derived skeletal lesions. © 2021 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

The multifaceted roles of the chemokines CCL2 and CXCL12 in osteophilic metastatic cancers.

Breast and prostate cancers have a great propensity to metastasize to long bones. The development of bone metastases is life-threatening, incurable, and drastically reduces patients' quality of life. The chemokines CCL2 and CXCL12 and their respective receptors, CCR2 and CXCR4, are central instigators involved in all stages leading to cancer cell dissemination and secondary tumor formation in distant target organs. They orchestrate tumor cell survival, growth and migration, tumor invasion and angiogenesis, and the formation of micrometastases in the bone marrow. The bone niche is of particular importance in metastasis formation, as it expresses high levels of CCL2 and CXCL12, which attract tumor cells and contribute to malignancy. The limited number of available effective treatment strategies highlights the need to better understand the pathophysiology of bone metastases and reduce the skeletal tumor burden in patients diagnosed with metastatic bone disease. This review focuses on the involvement of the CCL2/CCR2 and CXCL12/CXCR4 chemokine axes in the formation and development of bone metastases, as well as on therapeutic perspectives aimed at targeting these chemokine-receptor pairs.

Inhibitory Effects of Cathepsin K Inhibitor (ODN-MK-0822) on the Paracrine Pro-Osteoclast Factors of Breast Cancer Cells.

Breast cancer (BC) produces bone resorptive cytokines and growth factors that accelerate the development of osteoclasts (OCs), leading to osteolytic bone metastases. In the Long-term Odanacatib Fracture Trial (LOFT), the skeletal-metastasized breast cancer subjects who received odanacatib (ODN) had a delayed tumour progression and skeletal tumour burden as a result of anti-resorptive effects through inhibition of cathepsin K (CTSK). In this study, we explored the effect of ODN, a CTSK inhibitor, on the paracrine pro-osteoclast activity of breast cancer cells. An immunohistochemistry study was performed to demonstrate CTSK and PTHrP expression in the samples of primary breast carcinoma. Expression of CTSK mRNA and protein was confirmed by the reverse transcription PCR and western blotting analysis in two human breast cancer cell lines, MDA-MB-231 and MCF-7 BC cell lines. Cells were incubated with sub-lethal amounts of ODN, and their conditioned supernatants were assessed for their capacity to differentiate PBMCs of healthy donors into osteoclast and its interference on bone-resorbing activities. We also measured the mRNA levels of major pro-osteoclast (pro-OC) factors in ODN-treated breast cancer cells and their secreted levels by semi-quantitative reverse transcription PCR and protein expression by immunoblotting. Different staining intensity was observed in samples containing PTHrP and CTSK in various histological grades of breast carcinoma. A significant positive relationship was found between CTSK expression and histological grade of BC and presence or absence of distant metastasis. The present study results also indicate that ODN has no effects on OCs number, however, ODN decreases the mRNA expression of secreted pro-OC factors such as PTHrP, CXCR-4, and TNF-α. Immunoblot indicates that ODN treatment decreased the protein expression of CTSK, IL-6, and IL-1β, and thus lowered protein levels paralleled the defective phosphorylation of NF-κB. Moreover, there was a significant reduction in the level of growth factors such as IGF-1, PDGF, and TGFβ expression at transcriptional level after ODN treatment as compared to control. ODN has shown to prevent osteolytic metastasis by interacting with the NF-κB pathway, inhibiting bone resorptive cytokines and growth factors. This effect can also be taken into account the delayed development of metastatic bone disease found in the long-term odanacatib fracture trial (LOFT) study.

Abstract 5026: Efficacy of anti-PD-1, IDO inhibitor, chemotherapy and bone-targeting agent on tumor growth in a syngeneic bone metastasis model of triple-negative breast cancer

Abstract Triple-negative breast cancer (TNBC) is an aggressive cancer with treatment options limited to standard-of-care chemotherapy. TNBC patients develop metastases in high incidence, and almost all patients have bone metastases at end-stage disease. Bone metastases are mainly osteolytic, causing severe bone loss that can be prevented by bone-targeting agents. Currently, there is no efficient cure for bone metastases. Activation of the patient’s own immune system by immunotherapies is widely studied in various cancers, but their efficacy on bone metastases is not well established. In this study, we aimed to evaluate the efficacy of immunotherapies in comparison to standard-of-care compounds in a preclinical TNBC bone metastasis model. Mouse 4T1-GFP TNBC cells were inoculated intracardially to immunocompetent female Balb/c mice to model bone metastasis. Treatment of single agents of chemotherapy (cyclophosphamide, 100 mg/kg), bone-targeting agent (zoledronic acid, 0.1 mg/kg), and immunotherapies programmed cell death 1 antibody (mouse anti-PD-1, 10 mg/kg) and indoleamine-pyrrole 2,3-dioxygenase (IDO) inhibitor (epacadostat, 100 mg/kg), and the combination of anti-PD-1 and epacadostat were started on the day following the cancer cell inoculations. The study was terminated about 2 weeks after the inoculations. Tumor burden was evaluated at sacrifice by GFP imaging in skeleton and soft tissues, and tumor-induced bone loss by X-ray imaging. Tumor infiltrating lymphocytes (TILs) were evaluated by immunohistochemical stainings of CD3+, CD4+ and CD8+ T cells. Cyclophosphamide decreased skeletal tumor burden and the area of tumor-induced osteolytic bone lesions. It also decreased the number of soft tissue metastases in ovaries and adrenal glands. Zoledronic acid decreased osteolytic lesion area but had no effect on tumor burden. Anti-PD-1, epacadostat and their combination had no effect on tumor burden, on the development of osteolytic lesions, or on the number of soft tissue metastases. Low number or no TILs were observed in the tumors growing in bone obtained from vehicle-treated mice. Immunotherapies did not prevent tumor growth in bone or tumor-induced bone changes in the syngeneic TNBC model. This is probably a consequence of the low number of TILs in the tumors growing in bone. Further studies are needed to evaluate the effects of treatment combinations and to increase the responsiveness to immunotherapies especially in bone metastases. Citation Format: Tiina Emilia Kähkönen, Mari I. Suominen, Jenni H. Mäki-Jouppila, Jussi M. Halleen, Jenni Bernoulli. Efficacy of anti-PD-1, IDO inhibitor, chemotherapy and bone-targeting agent on tumor growth in a syngeneic bone metastasis model of triple-negative breast cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5026.

A novel artificial intelligence biomarker: Validation of the automated bone scan index (aBSI) in veterans with metastatic prostate cancer.

e17507 Background: Prostate cancer commonly metastasizes to the bone and is associated with reduced survival, pathologic fractures and bone pain. The assessment of bone lesions is made with the technetium Tc99m(99mTc) bone scan, which relies on the subjective interpretation of radiologists and has a wide interobserver variability. There is an unmet need for a more objective and quantifiable measurement tool. Progenics Pharmaceuticals has introduced an automated bone scan index (aBSI), which employs artificial intelligence to quantify skeletal tumor burden. The automated bone scan index has been prospectively validated and is reproducible in large Phase III studies. The aBSI was validated by our study in the Veteran population at the West LA VA Medical Center. Methods: The first positive technetium 99 Tc99m bone scans of veterans diagnosed with metastatic, castration-sensitive prostate cancer were evaluated. Since 2011, a total of 107 evaluable patient bone scans were studied (n = 107). Patients with visceral metastases were excluded to evaluate only those with skeletal metastases. An automated bone scan index (aBSI) was generated for each scan using the Progenics Pharmaceuticals’ artificial intelligence platform. Multivariate analysis of aBSI with overall survival, prostate cancer specific survival, time from diagnosis to first positive bone scan, age at diagnosis, ethnicity, and Gleason score was assessed. Results: The study demonstrated a wide range of aBSI values (Range 0-16.84). Values calculated above the Median aBSI value (1.0) were prognostic for Overall Survival (p = 0.0009) and Prostate Cancer-Specific Survival (p = 0.0011). Patients in the highest quartile of aBSI values (range 5.2-16.84) showed a statistically significant Prostate Cancer-Specific Mortality (p = 0.0300) when compared to the lowest two quartiles (Range 0-1.07). The time from diagnosis to the first positive Tc99m bone scan statistically correlated with aBSI values (p = 0.0016). Multivariate analysis using Cox regression was utilized in the final statistical analysis of prostate cancer-specific mortality and overall survival. Conclusions: The automated Bone Scan Index provides a quantifiable and validated artificial intelligence biomarker to address an unmet need among metastatic prostate cancer patients. This tool was validated among Veterans, a pertinent population that is commonly affected by metastatic prostate cancer.

JZL184, A Monoacylglycerol Lipase Inhibitor, Induces Bone Loss in a Multiple Myeloma Model of Immunocompetent Mice

Multiple myeloma (MM) patients develop osteolysis characterised by excessive osteoclastic bone destruction and lack of osteoblast bone formation. Pharmacological manipulation of monoacylglycerol lipase (MAGL), an enzyme responsible for the degradation of the endocannabinoid 2-arachidonoyl glycerol (2-AG), reduced skeletal tumour burden and osteolysis associated with osteosarcoma and advanced breast and prostate cancers. MM and hematopoietic, immune and bone marrow cells express high levels of type 2 cannabinoid receptor and osteoblasts secrete 2-AG. However, the effects of MAGL manipulation on MM have not been investigated. Here, we report that treatment of pre-osteoclasts with non-cytotoxic concentrations of JZL184, a verified MAGL inhibitor, enhanced MM- and RANKL-induced osteoclast formation and size in vitro. Exposure of osteoblasts to JZL184 in the presence of MM cell-derived factors reduced osteoblast growth but had no effect on the ability of these cells to mature or form bone nodules. In vivo, administration of JZL184 induced a modest, yet significant, bone loss at both trabecular and cortical compartments of long bones of immunocompetent mice inoculated with the syngeneic 5TGM1-GFP MM cells. Notably, JZL184 failed to inhibit the in vitro growth of a panel of mouse and human MM cell lines, or reduce tumour burden in mice. Thus, MAGL inhibitors such as JZL184 can exacerbate MM-induced bone loss.

Abstract B144: Lack of immunotherapy efficacy in a syngeneic bone metastasis model of triple-negative breast cancer

Abstract Background: Triple-negative breast cancer (TNBC) is an aggressive cancer with treatment options limited to standard-of-care chemotherapy. TNBC patients develop metastases in high incidence, and almost all patients have bone metastases at end-stage disease. Bone metastases are mainly osteolytic, causing severe bone loss that can be prevented by bone-targeted agents. Currently, there is no efficient cure for bone metastases. Activation of the patient’s own immune system by immunotherapies is widely studied in various cancers, but their efficacy on bone metastases is not well established. In this study, we aimed to evaluate the efficacy of immunotherapies in comparison to standard-of-care compounds in a preclinical TNBC bone metastasis model. Materials and methods: Mouse 4T1-GFP TNBC cells were inoculated intracardially to immunocompetent female Balb/c mice to model bone metastasis. Treatment of single agents of chemotherapy (cyclophosphamide, 100 mg/kg), bone-targeting agent (zoledronic acid, 0.1 mg/kg), and immunotherapies programmed cell death 1 antibody (mouse anti-PD-1, 10 mg/kg) and indoleamine-pyrrole 2,3-dioxygenase (IDO) inhibitor (epacadostat, 100 mg/kg), and the combination of anti-PD-1 and epacadostat were started on the day following the cancer cell inoculations. The study was terminated 12-13 days after the inoculations. Tumor burden was evaluated by GFP imaging and tumor-induced bone loss by X-ray imaging at sacrifice. Tumor infiltrating lymphocytes (TILs) were evaluated by immunohistochemical stainings of CD3+, CD4+ and CD8+ T cells. Results: Cyclophosphamide decreased skeletal tumor burden and the area of tumor-induced osteolytic bone lesions. Zoledronic acid decreased the area of osteolytic lesions but had no effect on skeletal tumor burden. Anti-PD-1, epacadostat and their combination had no effect on skeletal tumor burden or in the development of osteolytic lesions. Low number or no TILs were observed in the tumors growing in bone obtained from vehicle-treated mice. Conclusions: Immunotherapies did not prevent tumor growth in bone or tumor-induced bone changes in the syngeneic TNBC model. This is probably caused by the low number of TILs in the tumors growing in bone. Further studies are needed to evaluate the effects of treatment combinations and to increase the responsiveness to immunotherapies especially in bone metastases. Citation Format: Tiina E Kähkönen, Mari I Suominen, Jenni HE Mäki-Jouppila, Jussi M Halleen, Jenni Bernoulli. Lack of immunotherapy efficacy in a syngeneic bone metastasis model of triple-negative breast cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr B144. doi:10.1158/1535-7163.TARG-19-B144

Abstract 2007: Validation of a preclinical model for bone metastatic triple negative breast cancer

Abstract Metastatic breast cancer (MBC) is the most advanced stage of breast cancer, diagnosed as stage IV. Typically, MBC metastases occur in the bones, liver, brain, and lungs. Bone metastases cause significant morbidity and mortality in late-stage breast cancer patients. At present, there is no efficient curative or preventive treatment for bone metastases. Therefore, establishment and validation of well characterized in vivo models of breast cancer bone metastasis is of utmost importance. The aim of this study was to validate an intracardiac model of bone MBC utilizing GFP tagged triple negative (estrogen receptor; ER, progesterone receptor; PR and human epidermal growth factor receptor 2; HER2 negative) human breast cancer cells, which could be used to study efficacy of new potential treatments. Female athymic nude mice (Hsd: Athymic Nude-Foxn1nu) were used in this study. Bone seeking MDA-MB-231(SA)-GFP human breast cancer cells were inoculated into the left cardiac ventricle of 4-6 weeks old female mice. Allocation to treatment groups (n=15) was performed by stratification procedure based on animal weight prior to the beginning of the study. To validate the model, the current standard-of-care (SOC) treatment (Paclitaxel) was administered between study days 13-17 at the therapeutic dose. Serum bone biomarker TRACP5b was measured before cancer cell inoculation and at sacrifice. Planar X-ray of hind limbs and ex vivo fluorescence imaging of the whole body were performed at sacrifice. The observed tumor take rate was 100%. 33% of mice in the vehicle group were cachectic, but mice treated with SOC exhibited only mild and temporary weight loss. Tumors were observed mainly in skeletal sites but also in soft tissues. The SOC treatment decreased both total and skeletal tumor burden. Based on X-ray images, tumor induced osteolytic bone lesions were smaller in the SOC treated group compared to vehicle though no differences were observed in total bone lesion area. Serum TRACP5b was increased in the vehicle group as a result of osteolytic lesions, and decreased with the SOC treatment. In conclusion, an intracardiac model of MBC was successfully validated with the SOC treatment paclitaxel that efficiently inhibited disease progression. Therefore, this intracardiac model provides a promising tool for testing new treatments against bone and soft tissue MBC in vivo. Citation Format: Justyna Zdrojewska, Mari I. Suominen, Katja M. Fagerlund, Jussi M. Halleen, Jenni Bernoulli, Johann Zimmermann. Validation of a preclinical model for bone metastatic triple negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2007.

Loss of the Vitamin D Receptor in Human Breast Cancer Cells Promotes Epithelial to Mesenchymal Cell Transition and Skeletal Colonization.

Expression of the vitamin D receptor (VDR) is thought to be associated with neoplastic progression. However, the role of the VDR in breast cancer metastasis to bone and the molecular mechanisms underlying this process are unknown. Employing a rodent model (female Balb/c nu/nu mice) of systemic metastasis, we here demonstrate that knockdown of the VDR strongly increases the metastatic potential of MDA-MB-231 human breast cancer cells to bone, resulting in significantly greater skeletal tumor burden. Ablation of VDR expression promotes cancer cell mobility (migration) and invasiveness, thereby facilitating skeletal colonization. Mechanistically, these changes in tumor cell behavior are attributable to shifts in the expression of proteins involved in cell adhesion, proliferation, and cytoskeletal organization, patterns characteristic for epithelial-to-mesenchymal cell transition (EMT). In keeping with these experimental findings, analyses of human breast cancer specimens corroborated the association between VDR expression, EMT-typical changes in protein expression patterns, and clinical prognosis. Loss of the VDR in human breast cancer cells marks a critical point in oncogenesis by inducing EMT, promoting the dissemination of cancer cells, and facilitating the formation of tumor colonies in bone. © 2019 American Society for Bone and Mineral Research.

Deep learning for segmentation of 49 selected bones in CT scans: First step in automated PET/CT-based 3D quantification of skeletal metastases

PurposeThe aim of this study was to develop a deep learning-based method for segmentation of bones in CT scans and test its accuracy compared to manual delineation, as a first step in the creation of an automated PET/CT-based method for quantifying skeletal tumour burden. MethodsConvolutional neural networks (CNNs) were trained to segment 49 bones using manual segmentations from 100 CT scans. After training, the CNN-based segmentation method was tested on 46 patients with prostate cancer, who had undergone 18F-choline-PET/CT and 18F-NaF PET/CT less than three weeks apart. Bone volumes were calculated from the segmentations. The network’s performance was compared with manual segmentations of five bones made by an experienced physician. Accuracy of the spatial overlap between automated CNN-based and manual segmentations of these five bones was assessed using the Sørensen-Dice index (SDI). Reproducibility was evaluated applying the Bland-Altman method. ResultsThe median (SD) volumes of the five selected bones were by CNN and manual segmentation: Th7 41 (3.8) and 36 (5.1), L3 76 (13) and 75 (9.2), sacrum 284 (40) and 283 (26), 7th rib 33 (3.9) and 31 (4.8), sternum 80 (11) and 72 (9.2), respectively. Median SDIs were 0.86 (Th7), 0.85 (L3), 0.88 (sacrum), 0.84 (7th rib) and 0.83 (sternum). The intraobserver volume difference was less with CNN-based than manual approach: Th7 2% and 14%, L3 7% and 8%, sacrum 1% and 3%, 7th rib 1% and 6%, sternum 3% and 5%, respectively. The average volume difference measured as ratio volume difference/mean volume between the two CNN-based segmentations was 5–6% for the vertebral column and ribs and ≤3% for other bones. ConclusionThe new deep learning-based method for automated segmentation of bones in CT scans provided highly accurate bone volumes in a fast and automated way and, thus, appears to be a valuable first step in the development of a clinical useful processing procedure providing reliable skeletal segmentation as a key part of quantification of skeletal metastases.

Is interim 18F-fluoride PET/CT a predictor of outcomes after radium-223 therapy?

ObjectiveTo determine whether an interim 18F-fluoride positron-emission tomography/computed tomography (PET/CT) study performed after the third cycle of radium-223 dichloride (223RaCl2) therapy is able to identify patients that will not respond to treatment.Materials and MethodsWe retrospectively reviewed 34 histologically confirmed cases of hormone-refractory prostate cancer with bone metastasis in patients submitted to 223RaCl2 therapy. All of the patients underwent baseline and interim 18F-fluoride PET/CT studies. The interim study was performed immediately prior to the fourth cycle of 223RaCl2. The skeletal tumor burden-expressed as the total lesion fluoride uptake above a maximum standardized uptake value of 10 (TLF10)-was calculated for the baseline and the interim studies. The percent change in TLF10 between the baseline and interim studies (%TFL10) was calculated as follows: %TFL10 = interim TLF10 - baseline TLF10 / baseline TLF10. End points were overall survival, progression-free survival, and skeletal-related events.ResultsThe mean age of the patients was 72.4 ± 10.2 years (range, 43.3-88.8 years). The %TLF10 was not able to predict overall survival (p = 0.6320; hazard ratio [HR] = 0.753; 95% confidence interval [CI]: 0.236-2.401), progression-free survival (p = 0.5908; HR = 1.248; 95% CI: 0.557-2.797) nor time to a bone event (p = 0.5114; HR = 1.588; 95% CI: 0.399-6.312).ConclusionThe skeletal tumor burden on an interim 18F-fluoride PET/CT, performed after three cycles of 223RaCl2, is not able to predict overall survival, progression-free survival, or time to bone event, and should not be performed to monitor response at this time.

MiRNA-30 Family Members Inhibit Breast Cancer Invasion, Osteomimicry, and Bone Destruction by Directly Targeting Multiple Bone Metastasis-Associated Genes.

miRNAs are master regulators of gene expression that play key roles in cancer metastasis. During bone metastasis, metastatic tumor cells must rewire their biology and express genes that are normally expressed by bone cells (a process called osteomimicry), which endow tumor cells with full competence for outgrowth in the bone marrow. Here, we establish miR-30 family members miR-30a, miR-30b, miR-30c, miR-30d, and miR-30e as suppressors of breast cancer bone metastasis that regulate multiple pathways, including osteomimicry. Low expression of miR-30 in primary tumors from patients with breast cancer were associated with poor relapse-free survival. In addition, estrogen receptor (ER)-negative/progesterone receptor (PR)-negative breast cancer cells expressed lower miR-30 levels than their ER/PR-positive counterparts. Overexpression of miR-30 in ER/PR-negative breast cancer cells resulted in the reduction of bone metastasis burden in vivoIn vitro, miR-30 did not affect tumor cell proliferation, but did inhibit tumor cell invasion. Furthermore, overexpression of miR-30 restored bone homeostasis by reversing the effects of tumor cell-conditioned medium on osteoclastogenesis and osteoblastogenesis. A number of genes associated with osteoclastogenesis stimulation (IL8, IL11), osteoblastogenesis inhibition (DKK-1), tumor cell osteomimicry (RUNX2, CDH11), and invasiveness (CTGF, ITGA5, ITGB3) were identified as targets for repression by miR-30. Among these genes, silencing CDH11 or ITGA5 in ER-/PR-negative breast cancer cells recapitulated inhibitory effects of miR-30 on skeletal tumor burden in vivo Overall, our findings provide evidence that miR-30 family members employ multiple mechanisms to impede breast cancer bone metastasis and may represent attractive targets for therapeutic intervention.Significance: These findings suggest miR-30 family members may serve as an effective means to therapeutically attenuate metastasis in triple-negative breast cancer. Cancer Res; 78(18); 5259-73. ©2018 AACR.

Validation of the Semiautomatic Quantification of 18F-Fluoride PET/CT Whole-Body Skeletal Tumor Burden.

Our purpose was to validate a semiautomatic quantification of the skeletal tumor burden on 18F-fluoride PET/CT using manual quantification as a reference. Methods: We quantified 51 18F-fluoride PET/CT examinations performed on female breast cancer patients. Clinical information (age; time of disease presentation; presence of visceral metastases; and time to death, progression, or a bone event) was recorded. The total volume of 18F-fluoride-avid skeletal metastases and the total activity of 18F-fluoride-avid metastases were calculated manually and semiautomatically. Results: Manual and semiautomatic metrics correlated strongly (P < 0.0001; 95% confidence interval, 0.9300-0.9769). On multivariable analysis, the semiautomatic measures of total activity for 18F-fluoride-avid metastasis correlated significantly with overall survival (P = 0.0001) and progression-free survival (P = 0.0006). Approximate times for calculating skeletal tumor burden (semiautomatic vs. manual) were, respectively, 30 s versus 321 s in patients with fewer than 5 metastases, 120 s versus 640 s in patients with 5-10 metastases, and 240 s versus 1207s in patients with more than 10 metastases. Conclusion: Semiautomatic quantification of whole-body 18F-fluoride PET/CT skeletal tumor burden can replace manual quantification in breast cancer patients and is a strong independent biomarker of prognosis.

Assessment of Skeletal Tumor Load in Metastasized Castration-Resistant Prostate Cancer Patients: A Review of Available Methods and an Overview on Future Perspectives.

Metastasized castration-resistant prostate cancer (mCRPC), is the most advanced form of prostate neoplasia, where massive spread to the skeletal tissue is frequent. Patients with this condition are benefiting from an increasing number of treatment options. However, assessing tumor response in patients with multiple localizations might be challenging. For this reason, many computational approaches have been developed in the last decades to quantify the skeletal tumor burden and treatment response. In this review, we analyzed the progressive development and diffusion of such approaches. A computerized literature search of the PubMed/Medline was conducted, including articles between January 2008 and March 2018. The search was expanded by manually reviewing the reference list of the chosen articles. Thirty-five studies were identified. The number of eligible studies greatly increased over time. Studies could be categorized in the following categories: automated analysis of 2D scans, SUV-based thresholding, hybrid CT- and SUV-based thresholding, and MRI-based thresholding. All methods are discussed in detail. Automated analysis of bone tumor burden in mCRPC is a growing field of research; when choosing the appropriate method of analysis, it is important to consider the possible advantages as well as the limitations thoroughly.

TRAF2 in osteotropic breast cancer cells enhances skeletal tumour growth and promotes osteolysis

NFκB plays an important role in inflammation and bone remodelling. Tumour necrosis factor receptor associated factor 2 (TRAF2), a key component of NFκB signalling, has been identified as an oncogene, but its role in the regulation of breast cancer osteolytic metastasis remains unknown. Here, we report that stable overexpression of TRAF2 in parental and osteotropic sub-clones of human MDA-MB-231 (MDA-231) breast cancer cells increased cell growth and motility in vitro, whereas TRAF2 knockdown was inhibitory. In vivo, TRAF2 overexpression in the parental MDA-231-P cells enhanced tumour growth after orthotopic injection into the mammary fat pad of mice but failed to promote the metastasis of these cells to bone. In contrast, overexpression of TRAF2 in osteotropic MDA-231-BT cells increased skeletal tumour growth, enhanced osteoclast formation and worsened osteolytic bone loss after intra-tibial injection in mice. Mechanistic and functional studies in osteotropic MDA-231-BT and osteoclasts revealed that upregulation of TRAF2 increased the ability of osteotropic MDA-231-BT cells to migrate and to enhance osteoclastogenesis by a mechanism dependent, at least in part, on NFκB activation. Thus, the TRAF2/NFκB axis is implicated in the regulation of skeletal tumour burden and osteolysis associated with advanced breast cancer.

223Ra Therapy of Advanced Metastatic Castration-Resistant Prostate Cancer: Quantitative Assessment of Skeletal Tumor Burden for Prognostication of Clinical Outcome and Hematologic Toxicity.

The aim of this study was to investigate the prognostic value of the quantitative assessment of skeletal tumor burden on bone scintigraphy (Bone Scan Index [BSI]) in patients who have advanced metastatic castration-resistant prostate cancer (mCRPC) and are receiving 223RaCl2 We hypothesized that the BSI can serve as a prognostic biomarker of overall survival (OS) and hematologic toxicity and as a tool for response assessment in patients with mCRPC treated with 223RaCl2Methods: This study was a retrospective investigation of a Danish cohort of mCRPC patients who received 223RaCl2 therapy between March 2014 and October 2015 and for whom baseline bone scintigraphy was available. Bone scintigraphy studies were reviewed and graded according to the extent of disease. Furthermore, an automated BSI (EXINI BoneBSI) was obtained for baseline scintigraphy studies and follow-up scans after 3 cycles as well as at the end of therapy. Clinical outcomes were OS and occurrence of hematologic toxicity of grades 2-5. Associations between the BSI and clinical outcomes were investigated in multivariate regression models including the visual assessment of bone scintigraphy and other relevant covariates. Results: A total of 88 patients were included. The median number of completed 223RaCl2 cycles was 4, and 27 patients (31%) completed 6 cycles. The BSI was significantly associated with OS in the multivariate analysis; the median OS for patients with a BSI of greater than 5 was 8.2 mo, and the median OS for patients with a BSI of less than or equal to 5 was 15.0 mo (hazard ratio, 2.65 [95% confidence interval, 1.5-4.71]; P = 0.001). Likewise, the baseline BSI was prognostic for the occurrence of hematologic toxicity; patients with a BSI of greater than 5 had an odds ratio of 3.02 (95% confidence interval, 1.2-7.8; P = 0.02) for toxicity. The BSI declined during therapy in 44% of the patients who completed 3 cycles of 223RaCl2 (n = 52) and in 84% of the patients after the end of therapy (n = 32). There was no significant association between a change in the BSI during therapy and OS. Conclusion: The BSI is a promising biomarker for prognostication of OS and hematologic toxicity in late-stage mCRPC patients receiving 223RaCl2 Further prospective studies are needed to evaluate the potential of the BSI for response assessment in 223RaCl2 therapy.

Repeatability of quantitative parameters of 18F-fluoride PET/CT and biochemical tumour and specific bone remodelling markers in prostate cancer bone metastases

Purpose18F-fluoride PET/CT exhibits high sensitivity to delineate and measure the extent of bone metastatic disease in patients with prostate cancer. 18F-fluoride PET/CT could potentially replace traditional bone scintigraphy in clinical routine and trials. However, more studies are needed to assess repeatability and biological uptake variation. The aim of this study was to perform test-retest analysis of quantitative PET-derived parameters and blood/serum bone turnover markers at the same time point.Ten patients with prostate cancer and verified bone metastases were prospectively included. All underwent two serial 18F-fluoride PET/CT at 1 h post-injection. Up to five dominant index lesions and whole-body 18F-fluoride skeletal tumour burden were recorded per patient. Lesion-based PET parameters were SUVmax, SUVmean and functional tumour volume applying a VOI with 50% threshold (FTV50%). The total skeletal tumour burden, total lesion 18F-fluoride (TLF), was calculated using a threshold of SUV of ≥15. Blood/serum biochemical bone turnover markers obtained at the time of each PET were PSA, ALP, S-osteocalcin, S-beta-CTx, 1CTP and BAP.ResultsA total of 47 index lesions and a range of 2–122 bone metastases per patient were evaluated. Median time between 18F-fluoride PET/CT was 7 days (range 6–8 days). Repeatability coefficients were for SUVmax 26%, SUVmean 24%, FTV50% for index lesions 23% and total skeletal tumour burden (TLF) 35%. Biochemical bone marker repeatability coefficients were for PSA 19%, ALP 23%, S-osteocalcin 18%, S-beta-CTx 22%, 1CTP 18% and BAP 23%.ConclusionsQuantitative 18F-fluoride uptake and simultaneous biochemical bone markers measurements are reproducible for prostate cancer metastases and show similar magnitude in test-retest variation.

18F-Fluoride PET/CT tumor burden quantification predicts survival in breast cancer.

PurposeIn bone-metastatic breast cancer patients, there are no current imaging biomarkers to identify which patients have worst prognosis. The purpose of our study was to investigate if skeletal tumor burden determined by 18F-Fluoride PET/CT correlates with clinical outcomes and may help define prognosis throughout the course of the disease.ResultsBone metastases were present in 49 patients. On multivariable analysis, skeletal tumor burden was significantly and independently associated with overall survival (p < 0.0001) and progression free-survival (p < 0.0001). The simple presence of bone metastases was associated with time to bone event (p = 0.0448).Materials and MethodsWe quantified the skeletal tumor burden on 18F-Fluoride PET/CT images of 107 female breast cancer patients (40 for primary staging and the remainder for restaging after therapy). Clinical parameters, primary tumor characteristics and skeletal tumor burden were correlated to overall survival, progression free-survival and time to bone event. The median follow-up time was 19.5 months.Conclusions18F-Fluoride PET/CT skeletal tumor burden is a strong independent prognostic imaging biomarker in breast cancer patients.