Additional analyses of MYC regulation of ATAD2.

Oligonucleotides used in this Study

Over 1.4 million new cases of prostate cancer (PCa) and 375,304 deaths were recorded in 2020. Piper guineense Schum and Thonn is used to treat various diseases including cancer. Three triterpenoid saponins namely guineenosides (Guin) A, B C were recently isolated from this plant. This study marks the initial documentation of the anticancer potential of these compounds. Anti-cell growth (MTT), proliferation (CCK-8), clonogenic assays were evaluated. Furthermore, the most potent compound (Guin-B) was investigated for its potential to induce apoptosis, inhibit cell migration/invasion as well as cell adhesion. Some key proteins in cell proliferation and migration were evaluated. Out of the 3 saponins isolated from P. guineense, Guin-B exhibited significant and concentration-dependent inhibition of PC3 (IC50 =10 µg/mL or 4.82 × 10− 6 µM) and LNCaP (IC50 = 8 µg/mL or 3.86 × 10− 6 µM) cell growth and proliferation as well as a potent inhibition (p < 0.001) of PC3 cell clone formation. Guin-B induced a potent (near 50

Additional analyses related to Figure 4

BackgroundAs prostate cancer (PCa) remains one of the leading causes of cancer-related death in men, it is important to develop effective therapeutic approaches for the treatment of metastatic PCa and to improve the accuracy of prognosis for aggressive tumors. Non-POU domain-containing octamer-binding (NONO) is involved in RNA transport and almost all steps of gene regulation, and aberrant expression is associated with tumorigenesis in various tumor entities.MethodsImmunohistochemical analysis of the expression of NONO was performed in a cohort of 405 patient samples, including 54 benign prostate tissues, 250 primary prostate tumors, and 101 metastatic tissue samples. To assess the role of NONO in PCa cells, siRNA-mediated knockdown of NONO in the metastatic PCa cell lines PC3 and DU145, followed by a series of functional assays including proliferation, transwell, western blotting, and real-time quantitative PCR, was used.ResultsThe current study showed that the nuclear expression of NONO increases during the progression of PCa and is highest in distant metastases. NONO regulates cell proliferation by increasing the expression of cell cycle-related genes. In addition, NONO modulates migration and invasion in PCa cells.ConclusionOverall, these results suggest inhibition of NONO may be a promising approach for the treatment of metastatic PCa.

Metastasis accounts for the overwhelming majority of cancer deaths. In prostate cancer and many other solid tumors, progression to metastasis is associated with drastically reduced survival outcomes, yet the mechanisms behind this progression remain largely unknown. ATPase family AAA domain containing 2 (ATAD2) is an epigenetic reader of acetylated histones that is overexpressed in multiple cancer types and usually associated with poor patient outcomes. However, the functional role of ATAD2 in cancer progression and metastasis has been relatively understudied. Here, we employ genetically engineered mouse models of prostate cancer bone metastasis, as well as multiple independent human cohorts, to show that ATAD2 is highly enriched in bone metastasis compared with primary tumors and significantly associated with the development of metastasis. We show that ATAD2 expression is associated with MYC pathway activation in patient datasets and that, at least in a subset of tumors, MYC and ATAD2 can regulate each other's expression. Using functional studies on mouse bone metastatic cell lines and innovative organ-on-a-chip bone invasion assays, we establish a functional role for ATAD2 inhibition in reducing prostate cancer metastasis and growth in bone.Implications: Our study highlights ATAD2 as a driver of prostate cancer progression and metastasis and suggests it may constitute a promising novel therapeutic target.

Metastatic prostate cancer (mPCa) is a widespread disease with high mortality. Unraveling molecular mechanisms of disease progression is of utmost importance. The microenvironment in visceral organs and the skeletal system is of particular interest as a harbinger of metastatic spread. Therefore, we performed a comprehensive transcriptomic analysis of prostate cancer lung metastases with a special focus on differentially expressed genes attributable to the microenvironment. Digital gene expression analysis using the NanoString nCounter analysis system was performed on formalin-fixed, paraffin-embedded (FFPE) tissue from prostate cancer (PCa) lung metastases (n = 24). Data were compared to gene expression data from primary PCa and PCa bone metastases. Bioinformatic analysis was performed using several publicly available tools. In comparison to prostate cancer bone metastases, 209 genes were significantly upregulated, and 100 genes were significantly downregulated in prostate cancer lung metastases. Among the up-regulated genes, the top 10 genes with the most significant P-value were HLA-DPB1, PTPRC, ITGB7, C3, CCL21, CCL5, ITGAM, SERPINA1, MFAP4, ARAP2 and among the down-regulated genes, the top 10 genes with the most significant P-value were FOXC2, TWIST1, CDK14, CHAD, IBSP, EPN3, VIT, HAPLN1, SLC44A4, TBX1. In PCa lung metastases genes associated with immunogenic responses were upregulated while genes associated with epithelial-mesenchymal transition were down-regulated. We also showed that CXCR3/CXCL10 axis plays a significant role in prostate cancer lung metastases in comparison to bone metastases. In this study, we comprehensively explored transcriptomic alterations in PCa lung metastases in comparison to primary PCa and PCa bone metastases. In PCa lung metastases genes associated with immunogenic responses are upregulated while genes associated with epithelial-mesenchymal transition are down-regulated. This points to a more immunogenic phenotype of PCa lung metastases thus potentially making patients more susceptible to immunotherapeutic approaches.

Rationale: Current therapies for metastatic osseous disease frequently fail to provide a durable treatment response. To date, there are only limited therapeutic options for metastatic prostate cancer, the mechanisms that drive the survival of metastasis-initiating cells are poorly characterized, and reliable prognostic markers are missing. A high aldehyde dehydrogenase (ALDH) activity has been long considered a marker of cancer stem cells (CSC). Our study characterized a differential role of ALDH1A1 and ALDH1A3 genes as regulators of prostate cancer progression and metastatic growth. Methods: By genetic silencing of ALDH1A1 and ALDH1A3 in vitro, in xenografted zebrafish and murine models, and by comparative immunohistochemical analyses of benign, primary tumor, and metastatic specimens from patients with prostate cancer, we demonstrated that ALDH1A1 and ALDH1A3 maintain the CSC phenotype and radioresistance and regulate bone metastasis-initiating cells. We have validated ALDH1A1 and ALDH1A3 as potential biomarkers of clinical outcomes in the independent cohorts of patients with PCa. Furthermore, by RNAseq, chromatin immunoprecipitation (ChIP), and biostatistics analyses, we suggested the molecular mechanisms explaining the role of ALDH1A1 in PCa progression. Results: We found that aldehyde dehydrogenase protein ALDH1A1 positively regulates tumor cell survival in circulation, extravasation, and metastatic dissemination, whereas ALDH1A3 plays the opposite role. ALDH1A1 and ALDH1A3 are differentially expressed in metastatic tumors of patients with prostate cancer, and their expression levels oppositely correlate with clinical outcomes. Prostate cancer progression is associated with the increasing interplay of ALDH1A1 with androgen receptor (AR) and retinoid receptor (RAR) transcriptional programs. Polo-like kinase 3 (PLK3) was identified as a transcriptional target oppositely regulated by ALDH1A1 and ALDH1A3 genes in RAR and AR-dependent manner. PLK3 contributes to the control of prostate cancer cell proliferation, migration, DNA repair, and radioresistance. ALDH1A1 gain in prostate cancer bone metastases is associated with high PLK3 expression. Conclusion: This report provides the first evidence that ALDH1A1 and PLK3 could serve as biomarkers to predict metastatic dissemination and radiotherapy resistance in patients with prostate cancer and could be potential therapeutic targets to eliminate metastasis-initiating and radioresistant tumor cell populations.

BACKGROUND/AIM:Metastatic prostate cancer (mPCa) results in high morbidity and mortality. Visceral metastases in particular are associated with a shortened survival. Our aim was to unravel the molecular mechanisms that underly pulmonary spread in mPCa. MATERIALS AND METHODS:We performed a comprehensive transcriptomic analysis of PCa lung metastases, followed by functional validation of candidate genes. Digital gene expression analysis utilizing the NanoString technology was performed on mRNA extracted from formalin-fixed, paraffin-embedded (FFPE) tissue from PCa lung metastases. The gene expression data from primary PCa and PCa lung metastases were compared, and several publicly available bioinformatic analysis tools were used to annotate and validate the data. RESULTS:In PCa lung metastases, 234 genes were considerably up-regulated, and 78 genes were significantly down-regulated when compared to primary PCa. Carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6) was identified as suitable candidate gene for further functional validation. CEACAM6 as a cell adhesion molecule has been implicated in promoting metastatic disease in several solid tumors, such as colorectal or gastric cancer. We showed that siRNA knockdown of CEACAM6 in PC-3 and LNCaP cells resulted in decreased cell viability and migration as well as enhanced apoptosis. Comprehensive transcriptomic analyses identified several genes of interest that might promote metastatic spread to the lung. CONCLUSION:Functional validation revealed that CEACAM6 might play an important role in fostering metastatic spread to the lung of PCa patients via enhancing proliferation, migration and suppressing apoptosis in PC-3 and LNCaP cells. CEACAM6 might pose an attractive therapeutic target to prevent metastatic disease.

BACKGROUND:Prostate cancer is one of the most common cancers in men in Europe. Several classes of agents can be considered for the treatment of metastatic prostate carcinoma, and their use is supported by extensive guidelines. In the treatment of metastatic castration-resistant prostate cancer (mCRPC), it is currently unclear which sequence of systemic therapies is most effective. Currently approved system therapies in the castration-resistant setting generally include hormone-manipulating agents, taxane-based chemotherapies, radioactive agents, or inhibitiors of DNA repair mechanisms. This study aims to summarize real world data of mCRPC therapy. METHODS:Retrospectively, 90 mCRPC patients undergoing treatment at the University Hospital Schleswig-Holstein, Lübeck Campus between February 2006 and March 2020 were identified. The patient data were analyzed for their treatment sequence and disease progression. Due to the inclusion period, the mCRPC therapy sequences studied were limited to: Abiraterone, Cabazitaxel, Docetaxel, Enzalutamide, Lutetium-177-PSMA and Radium-223. The analysis includes the therapy sequences and their duration, clinical information of the respective cohort, overall and cancer-specific survival (OS/CSS) as well as time to second-line therapy in relation to the respective first-line therapy. RESULTS:Approximately two-thirds of patients underwent a true therapy sequence (at least two of the drugs listed above), with this proportion halving by the third line.The majority of patients received the sequence (first/second line) abiraterone/docetaxel (n=13), followed by docetaxel/abiraterone (n=12) and abiraterone/enzalutamid (n=10) and docetaxel/docetaxel (n=8).Within the different docetaxel sequences, first-line (mean 4.7 months ± SD 3.1; median 4.0) and rechallenge (mean 5.3 months ± SD 5.9; median 3.0) therapy durations were the longest. The subjective side effect rate of docetaxel was lower in the second line, so that a better tolerability can be assumed here.The abiraterone/docetaxel sequence was used mainly in patients with metachronous metastases. Among the different sequences of abiraterone, first-line (mean 10.8 months ± SD 10.2; median 9.0) and second-line (mean 10.6 months ± SD 9.0; median 7.0) therapy durations were the longest.The sequence abiraterone/enzalutamide was prescribed mainly to older patients with synchronous metastases. Among the different enzalutamide sequences first-line (mean 9.6 months ± SD 7.1; median 7.0) and rechallenge (mean 11.0 ± SD 0.0; median 11.0) therapy durations were the longest.In contrast, the sequence docetaxel/docetaxel was used mainly in younger patients with a high initial PSA.The evaluation shows a trend that both abiraterone and enzalutamide can account for a survival advantage in the first line. CONCLUSION:Ultimately, an optimal treatment sequence cannot be confidently derived from these data.However, it was found that only a small proportion of patients underwent fourth- or even fifth-line treatment at all. Thus, the focus on first- and second-line in this study seems reasonable. It could be shown in a trend that docetaxel as first-line therapy seems to be disadvantegous regarding OS as well as CSS when compared to abiraterone or enzalutamide. However, due to the small number of patients in this study, a clear significance cannot be derived. Moreover, the subjectively better tolerability of docetaxel in the second-line setting could provide an impetus for treatment planning in multimorbid elderly patients in the future. The sequence abiraterone/docetaxel may offer a beneficial option for initial mCRPC therapy.

Hintergrund Das Prostatakarzinom zählt zu den häufigsten Krebserkrankungen bei Männern in Europa. Für die Behandlung des metastasierten Prostatakarzinoms kommen verschiedene Substanzklassen in Betracht, wobei die Verwendung durch umfangreiche Leitlinien gestützt wird. In der Therapie des metastasierten kastrationsresistenten Prostatakarzinoms (mCRPC) ist derzeitig unklar, welche Reihenfolge (Sequenz) der applizierten Systemtherapien am zielführendsten ist. Derzeitig zugelassene Systemtherapien in der kastrationsresistenten Situation umfassen im Allgemeinen hormonmanipulierende Präparate, Taxan-basierte Chemotherapien, radioaktive Substanzen oder Inhibitoren von DNA-Reparaturmechanismen. Diese Studie soll die Anwendungsrealität der mCRPC-Therapie zusammenfassen. Methoden Retrospektiv konnten 90 mCRPC-Patienten ermittelt werden, welche sich zwischen Februar 2006 und März 2020 im Universitätsklinikum Schleswig-Holstein, Campus Lübeck in Behandlung befanden. Die Patientendaten wurden daraufhin auf ihre Therapiesequenz und ihren Krankheitsverlauf untersucht. Die untersuchten mCRPC-Therapiesequenzen beschränken sich aufgrund der Einschlussperiode auf: Abirateron, Cabazitaxel, Docetaxel, Enzalutamid, Lutetium-177-PSMA und Radium-223. Die Auswertung beinhaltet die Therapiesequenzen und ihre -dauer, klinische Informationen der jeweiligen Kohorte, das allgemeine und krebsspezifische Überleben (OS/CSS) sowie die Zeit bis zur Zweitlinientherapie in Relation zur jeweiligen Erstlinientherapie. Ergebnisse Etwa zwei Drittel der Patienten wurden einer echten Therapiesequenz (mind. zwei der oben aufgeführten Medikamente) unterzogen, wobei sich dieser Anteil bereits in der Drittlinie halbierte. Die Mehrheit der Patienten erhielt dabei die Sequenz (Erst-/Zweitlinie) Abirateron/Docetaxel (n=13), gefolgt von Docetaxel/Abirateron (n=12) sowie Abirateron/Enzalutamid (n=10) und Docetaxel/Docetaxel (n=8). Innerhalb der verschiedenen Docetaxel-Sequenzen war die Therapiedauer der Erstlinie (Mittelwert 4,7 Monate ± SD 3,1; Median 4,0) sowie die der Rechallenge (Mittelwert 5,3 Monate ± SD 5,9; Median 3,0) am längsten. Die subjektive Nebenwirkungsrate von Docetaxel war in der in der Zweitlinie deutlich geringer, sodass hierbei von einer besseren Verträglichkeit ausgegangen werden kann. Die Sequenz Abirateron/Docetaxel fand vor allem bei Patienten mit metachroner Metastasierung Anwendung. In den verschiedenen Sequenzen von Abirateron war die Therapiedauer der Erstlinie (Mittelwert 10,8 Monate ± SD 10,2; Median 9,0) sowie die der Zweitlinie (Mittelwert 10,6 Monate ± SD 9,0; Median 7,0) am längsten. Die Sequenz Abirateron/Enzalutamid wurde vor allem älteren Patienten mit synchroner Metastasierung verabreicht. In den verschiedenen Enzalutamid-Sequenzen war die Therapiedauer der Erstlinie (Mittelwert 9,6 Monate ± SD 7,1; Median 7,0) und die der Rechallenge (Mittelwert 11,0 ± SD 0,0; Median 11,0) am längsten. Die Sequenz Docetaxel/Docetaxel fand dagegen vor allem bei jüngeren Patienten mit einem hohen initialen PSA-Wert Anwendung. Die Auswertung zeigt in einem Trend auf, dass sowohl Abirateron als auch Enzalutamid in der Erstlinie einen Überlebensvorteil ausmachen könnten. Schlussfolgerung Letztlich lässt sich eine optimale Therapiesequenz anhand dieser Daten nicht sicher ableiten. Es zeigte sich jedoch, dass nur ein geringer Anteil an Patienten überhaupt einer Viert- oder sogar Fünftlinie unterzogen wurde. Somit scheint der in dieser Studie gesetzte Fokus auf die Erst- und Zweitlinie sinnvoll. Es konnte in einem Trend gezeigt werden, dass Docetaxel als Erstlinientherapie einen eher nachteiligen Einfluss auf das OS sowie CSS zu nehmen scheint im Vergleich zu Abirateron oder Enzalutamid. Aufgrund der geringen Patientenanzahl dieser Studie ist eine deutliche Signifikanz hierbei jedoch nicht abzuleiten. Die subjektiv bessere Verträglichkeit von Docetaxel in der Zweitlinie könnte darüber hinaus zukünftig einen Impuls für die Therapieplanung multimorbider älterer Patienten geben. Die Sequenz Abirateron/Docetaxel bietet daher möglicherweise einen guten Einstieg in die mCRPC-Therapie.

Urothelial bladder cancer (UC) has a wide tumor biological spectrum with challenging prognostic stratification and relevant therapy-associated morbidity. Most molecular classifications relate only indirectly to the therapeutically relevant protein level. We improve the pre-analytics of clinical samples for proteome analyses and characterize a cohort of 434 samples with 242 tumors and 192 paired normal mucosae covering the full range of UC. We evaluate sample-wise tumor specificity and rank biomarkers by target relevance. We identify robust proteomic subtypes with prognostic information independent from histopathological groups. In silico drug prediction suggests efficacy of several compounds hitherto not in clinical use. Both in silico and in vitro data indicate predictive value of the proteomic clusters for these drugs. We underline that proteomics is relevant for personalized oncology and provide abundance and tumor specificity data for a large part of the UC proteome ( www.cancerproteins.org ).

ETHNOPHARMACOLOGICAL RELEVANCE:Commiphora kerstingii Engl is a tree which is 20-30 m in height and commonly called "ararrabi" in Hausa. It is found in the Sahelian region (Cameroon, Chad, and Nigeria) where it is utilized for the treatment of several ailments including cancer. AIM OF THE STUDY:This study was aimed at investigating the chemical constituents and cytotoxic effect of extracts and isolates from the stem barks and leaves of C. kerstingii. MATERIALS AND METHODS:Using classical chromatography technique coupled with spectroscopic analysis and literature information, ten (10) compounds were isolated from C. kerstingii stem barks and leaves, out of which two [kerstingilactone (3) and kerstinginone (10)] were new. To evaluate their potential cytotoxic effect, the impact on cell viability, growth, and proliferation was assessed using MTT and CCK-8 assays. Cell death mechanisms were analyzed via flow cytometry, and Western blotting was utilized to examine the expression of specific regulatory proteins. Furthermore, anti-metastatic properties were investigated through assays on cell migration, adhesion, and chemotaxis. RESULTS:Among the tested compounds, 2 (Masticadienonic Acid) and 10 (kerstinginone) exhibited significant dose-dependent inhibition of PC3 and LNCaP cell growth. Compound 2 displayed optimal inhibitory effects within a concentration range of 10-40 μg/mL, while compound 10 demonstrated potent growth inhibition at concentrations of 2.5-10 μg/mL. Both compounds suppressed cell proliferation and the formation of clones. Specifically, compound 2 induced apoptosis solely in the androgen-sensitive LNCaP prostate cancer cells, whereas compound 10 induced a stronger and concentration-dependent apoptotic response in both PC3 and LNCaP cells, resulting in approximately 50-70% apoptotic cells. It also induced potent cell migration/invasion arrest at concentrations ranging from 2.5 to 5 μg/mL and increased cell adhesion to the extracellular matrix. CONCLUSION:Kerstinginone exhibits potent cytotoxicity and apoptosis-inducing activity, making it a promising lead for discovering a new anticancer drug.

Background The immunological composition of the tumor microenvironment has a decisive influence on the biological course of cancer and is therefore of profound clinical relevance. In this study, we analyzed the cooperative effects of integrin β4 (ITGB4) on tumor cells and E-/P-selectin on endothelial cells within the tumor stroma for regulating tumor growth by shaping the local and systemic immune environment. Methods We used several preclinical mouse models for different solid human cancer types (xenograft and syngeneic) to explore the role of ITGB4 (shRNA-mediated knockdown in tumor cells) and E-/P-selectins (knockout in mice) for tumor growth; effects on apoptosis, proliferation and intratumoral signaling pathways were determined by histological and biochemical methods and 3D in vitro experiments; changes in the intratumoral and systemic immune cell composition were determined by flow cytometry and immunohistochemistry; chemokine levels and their attracting potential were measured by ELISA and 3D invasion assays. Results We observed a very robust synergism between ITGB4 and E-/P-selectin for the regulation of tumor growth, accompanied by an increased recruitment of CD11b + Gr-1 Hi cells with low granularity (i.e., myeloid-derived suppressor cells, MDSCs) specifically into ITGB4-depleted tumors. ITGB4-depleted tumors undergo apoptosis and actively attract MDSCs, well-known to promote tumor growth in several cancers, via increased secretion of different chemokines. MDSC trafficking into tumors crucially depends on E-/P-selectin expression. Analyses of clinical samples confirmed an inverse relationship between ITGB4 expression in tumors and number of tumor-infiltrating leukocytes. Conclusions These findings suggest a distinct vulnerability of ITGB4 Lo tumors for MDSC-directed immunotherapies. Graphical Abstract

S1.(a) Distribution of tumor vs. normal lung tissue in K, KE, KP and KPE mice at 4 weeks post tumor induction (top). Distribution of murine lung adenocarcinoma grades (G1, G2, G3) in corresponding K, KE, KP and KPE mice at 4 weeks post tumor induction (bottom). (b) Anti-Ercc1 immunostaining of lung sections of KE mice (n = 2) at 4 weeks post tumor induction (mouse KE.1 shown at two magnifications, mouse KE.2 shown at two magnifications) (top). Anti-Ercc1 immunostaining of lung sections of KE mice (n = 2) at 12 weeks post tumor induction (mouse KE.3 shown at two magnifications, mouse KE.4 shown at two magnifications) (bottom). (c) Anti-Ercc1 immunostaining of lung sections of KPE mice (n = 5) at 4 weeks post tumor induction (shown at different magnifications) and of KPE mice (n = 5) at 12 weeks post tumor induction (shown at different magnifications).

Ablative radiotherapy is a highly efficient treatment modality for patients with metastatic prostate cancer (PCa). However, a subset of patients does not respond. Currently, this subgroup with bad prognosis cannot be identified before disease progression. We hypothesize that markers indicative of radioresistance, stemness and/or bone tropism may have a prognostic potential to identify patients profiting from metastases‐directed radiotherapy. Therefore, circulating tumor cells (CTCs) were analyzed in patients with metastatic PCa (n = 24) during radiotherapy with CellSearch, multicolor flow cytometry and imaging cytometry. Analysis of copy‐number alteration indicates a polyclonal CTC population that changes after radiotherapy. CTCs were found in 8 out of 24 patients (33.3%) and were associated with a shorter time to biochemical progression after radiotherapy. Whereas the total CTC count dropped after radiotherapy, a chemokine receptor CXCR4‐expressing subpopulation representing 28.6% of the total CTC population remained stable up to 3 months. At once, we observed higher chemokine CCL2 plasma concentrations and proinflammatory monocytes. Additional functional analyses demonstrated key roles of CXCR4 and CCL2 for cellular radiosensitivity, tumorigenicity and stem‐like potential in vitro and in vivo. Moreover, a high CXCR4 and CCL2 expression was found in bone metastasis biopsies of PCa patients. In summary, panCK + CXCR4 + CTCs may have a prognostic potential in patients with metastatic PCa treated with metastasis‐directed radiotherapy.

Prostate cancer (PCa) Lung metastases are rarely resected, therefore PCa lung metastases are insufficiently molecularly characterized. We recently identified carcinoembryonic antigen cell adhesion molecule 6 (CEACAM6) as a potential driver of pulmonary metastatic spread. Here, we show the biological significance of CEACAM6 in PCa cell proliferation, apoptosis and migration. CEACAM6 was silenced by siRNA in PC3 cells. Functional assessment of apoptosis, cell viability, proliferation and migration were performed in siRNA-CEACAM6 PC3 cells. Non-treated and scrambled (scr) RNA PC3 cells were used as control. Following a specific knockdown of CEACAM6 in PC3 cells, the expression of CEACAM6 protein was significantly decreased in comparison to controls. Cell viability and cell counts decreased in CEACAM6 silent PC3 cells. In contrast, caspase-3 activity was highly elevated in siRNA-CEACAM6 PC3. Furthermore, by performing a cell scratch assay, the migration ratio in siRNA-CEACAM6 PC3 cells were significantly diminished compared with the control group after 48 hours of post transfection incubation. CEACAM6 as a cell adhesion molecule has been implicated in promoting metastatic disease in several solid tumours such as colorectal or gastric cancer. We could show that silencing of CEACAM6 has a significant functional effect on PCa cells. CEACAM6 might play an important role in fostering metastatic spread to the lung of PCa patients via enhancing proliferation and suppressing apoptosis. CEACAM6 might therefore pose an attractive therapeutic target to prevent metastatic disease.

S4. (a) Representative immunostaining of Pt-(GpG) adducts in KP and KPE cell lines after cisplatin exposure (left). Quantification of Pt-(GpG) adduct staining in KP (n = 5) and KPE (n = 6) cells after cisplatin exposure (right). P values were calculated using two-tailed t-test. (b) Clonogenic survival assay of mock-treated KP and cisplatin-treated KP cells, mock-treated KPE and cisplatin-treated KPE cells, mock-treated KPE post cis and cisplatin-treated KPE post cis cells, mock-treated KPE post cis clone 6 (*6) and cisplatin-treated KPE post cis clone 6 (*6). (c) Detailed representation of Fig. 4c.