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Purpose: A disturbed myeloid lineage development with abnormally abundant neutrophils and impaired dendritic cell (DC) differentiation may contribute to tumor immune escape. We investigated the effect of sunitinib, a tyrosine kinase inhibitor of fms-like tyrosine kinase-3, KIT, and vascular endothelial growth factor receptors, on myeloid differentiation in renal cell cancer (RCC) patients.Experimental Design: Twenty-six advanced RCC patients were treated with sunitinib in a 4-week on/2-week off schedule. Enumeration and extensive phenotyping of myeloid subsets in the blood was done at baseline and at weeks 4 and 6 of the first treatment cycle. Baseline patient data were compared with sex- and age-matched healthy donor data.Results: Baseline frequencies of DC subsets were lower in RCC patients than in healthy donors. After 4 weeks of sunitinib treatment, a generalized decrease in myeloid frequencies was observed. Whereas neutrophils and monocytes, which were both abnormally high at baseline, remained low during the 2-week off period, DC rates recovered, resulting in a normalized myeloid lineage distribution. Subsequent to sunitinib treatment, an increase to high levels of myeloid DC (MDC) subset frequencies relative to other myeloid subsets, was specifically observed in patients experiencing tumor regression. Moreover, high CD1c/BDCA-1+ MDC frequencies were predictive for tumor regression and improved progression-free survival.Conclusion: The sunitinib-induced myeloid lineage redistribution observed in advanced RCC patients is consistent with an improved immune status. Immunologic recovery may contribute to clinical efficacy as suggested by the finding of highly increased MDC frequencies relative to other myeloid subsets in patients with tumor regression.

Purpose: The survival of patients with clear cell metastatic renal cell carcinoma (cc-mRCC) has improved substantially since the introduction of tyrosine kinase inhibitors (TKI). With the fact that TKIs interact with immune responses, we investigated whether polymorphisms of genes involved in immune checkpoints are related to the clinical outcome of cc-mRCC patients treated with sunitinib as first TKI.Experimental Design: Twenty-seven single-nucleotide polymorphisms (SNP) in CD274 (PD-L1), PDCD1 (PD-1), and CTLA-4 were tested for a possible association with progression-free survival (PFS) and overall survival (OS) in a discovery cohort of 550 sunitinib-treated cc-mRCC patients. SNPs with a significant association (P < 0.05) were tested in an independent validation cohort of 138 sunitinib-treated cc-mRCC patients. Finally, data of the discovery and validation cohort were pooled for meta-analysis.Results: CTLA-4 rs231775 and CD274 rs7866740 showed significant associations with OS in the discovery cohort after correction for age, gender, and Heng prognostic risk group [HR, 0.84; 95% confidence interval (CI), 0.72–0.98; P = 0.028, and HR, 0.73; 95% CI, 0.54–0.99; P = 0.047, respectively]. In the validation cohort, the associations of both SNPs with OS did not meet the significance threshold of P < 0.05. After meta-analysis, CTLA-4 rs231775 showed a significant association with OS (HR, 0.83; 95% CI, 0.72–0.95; P = 0.008). Patients with the GG genotype had longer OS (35.1 months) compared with patients with an AG (30.3 months) or AA genotype (24.3 months). No significant associations with PFS were found.Conclusions: The G-allele of rs231775 in the CTLA-4 gene is associated with an improved OS in sunitinib-treated cc-mRCC patients and could potentially be used as a prognostic biomarker. Clin Cancer Res; 24(10); 2350–6. ©2018 AACR.

Supplementary Figure S1 from Sunitinib-Induced Myeloid Lineage Redistribution in Renal Cell Cancer Patients: CD1c+ Dendritic Cell Frequency Predicts Progression-Free Survival

Supplementary Table S1 Selected tagging SNPs Supplementary Figure S1 Kaplan-Meier plot for overall survival by genotype of CTLA-4 rs231775 in discovery cohort (A), validation cohort (B) and combined cohort (C). Supplementary Figure S2 The association of CTLA-4 rs231775 with CTLA-4 gene expression in multiple tissues (A) and in testis tissue (B, p=1.0Ã-10-7) using data from the Genotype-Tissue Expression (GTEx) dataset. Supplementary Figure S3 The survival curve shows the association of CTLA-4 gene expression with overall survival in patients with clear cell renal cell carcinoma (p=0.00255) through OncoLnc, which links The Cancer Genome Atlas (TCGA) survival data to mRNA expression levels (23). (low: lower 50 percentile, high: upper 50 percentile).

Purpose: We examined whether pretreatment levels of angiogenesis- or hypoxia-related proteins and their changes after one cycle of first-line bevacizumab-based therapy were associated with response, PFS, or OS in patients with metastatic breast cancer.Experimental Design: We included 181 patients enrolled in the phase II ATX trial evaluating first-line paclitaxel and bevacizumab without or with capecitabine (NTR1348). Plasma samples were analyzed for VEGF-A, soluble VEGFR2 (sVEGFR2), angiopoietin 2 (ANG2), soluble TIE2 (sTIE2), IL6, IL8, and carbonic anhydrase 9 (CA9). Baseline serum CA15-3 was documented. HR was adjusted for confounding factors. Where appropriate, an optimal cut-off value defining a high and a low group was determined with Martingale residuals.Results: At baseline, multiple proteins were significantly associated with PFS (ANG2, IL6, IL8, CA9, CA15-3) and OS (ANG2, sTIE2, IL6, IL8, CA9, CA15-3). After one cycle, VEGF-A, ANG2, sTIE2, and IL8 significantly decreased, while sVEGFR2 and CA9 significantly increased. The relative change in sVEGFR2 (P = 0.01) and IL8 (P = 0.001) was associated with response. Defining optimal cut-off, patients with a high CA9 rise (>2.9%) had better PFS (HR 0.45) and OS (HR 0.54) than those with low/no rise.Conclusions: Multiple angiogenesis- or hypoxia-related proteins were prognostic for PFS and OS. Molecular agents targeting these proteins might be beneficial in patients with high levels. Changes in IL8 or sVEGFR2 levels at second cycle appear predictive for response. Changes in CA9 levels during bevacizumab-based therapy for prediction of PFS and OS merit further study. Clin Cancer Res; 22(7); 1611–20. ©2016 AACR.

Supplementary Table 1. Clinical outcome of original trial population and biomarker population Supplementary Table 2. Baseline plasma protein levels vs baseline patient and tumor characteristics Supplementary Table 3. Spearman Rho correlation coefficients among protein levels at baseline Supplementary Table 4. Plasma protein changes vs baseline patient and tumor characteristics Supplementary Table 5. Change in levels of proteins per treatment arm Supplementary Table 6. Spearman Rho correlation coefficients among protein changes on C2D1 from baseline Supplementary Figure 1A. A plot of Martingale residual vs log-transformed CA9 change for death Supplementary Figure 1B. Estimation of an optimal cut-off point for CA9 change Supplementary Figure 1C. Changes in CA9 levels in the subgroups of a high rise and a low/no rise

Purpose Everolimus plus exemestane (EVE/EXE) is a registered treatment option for ER-positive, HER2-negative (ER +/HER2-) metastatic breast cancer (MBC), but resistance mechanisms limit efficacy. We aimed to find markers that might help select patients with a higher chance on benefit from EVE/EXE. Methods Immunohistochemistry (IHC) of PTEN, p-AKT(Thr308), p-AKT(Ser473), p-4EBP1, p-p70S6K, p-S6RP(Ser240/244), p-ERK1/2 and p-S6RP (Ser235/236) was performed on primary tumour tissue and on biopsies immediately taken from ER +/HER2- MBC patients before the start of standard EVE/EXE (Eudract 2013-004120-11). Unsupervised hierarchical clustering was executed to create heatmaps to distinguish subgroups of preferentially activated and less-activated PI3K/MAPK proteins. Uni- and multivariate Cox models were used for associations with PFS. Results Primary tumour tissue from 145 patients was retrieved. Median PFS was 5.4 months. Patients without (neo)adjuvant therapy (p = 0.03) or bone only disease (p = 0.04) had longer PFS on EVE/EXE. In primary tumours, neither single proteins nor PI3K/MAPK-associated heatmap subgroups were significantly associated with PFS. In 21 patients a non-osseous biopsy obtained before dosing was useful for continuous scoring, which demonstrated upregulation of several proteins as compared to readings in corresponding primary tumour tissues. These comparisons revealed that increased expression of p-4EBP1 was significantly associated with worse PFS (multivariate HR 3.69, p = 0.05). Conclusions IHC of single proteins or heatmap subgroups of the differentially activated PI3K/MAPK pathways was not able to discriminate patients on EVE/EXE with poor or better PFS. Upregulation of p-4EBP1 in pre-treatment biopsies as compared to levels in primary tumours pointed towards shorter PFS.

We determined whether progression-free survival (PFS) in metastatic breast cancer (MBC) patients receiving everolimus plus exemestane (EVE/EXE) varies depending on circulating tumour DNA (ctDNA) characteristics. Baseline plasma cell-free DNA (cfDNA) from 164 postmenopausal women with ER-positive, HER2-negative MBC refractory to a non-steroidal aromatase inhibitor and treated with standard EVE/EXE (Everolimus Biomarker Study, Eudract 2013-004120-11) was characterised for 10 relevant breast cancer genes by next generation sequencing with molecular barcoding. ctDNA molecule numbers, number of mutations and specific variants were related with PFS and overall survival (OS). Missense hotspot mutations in cfDNA were detected in 125 patients. The median of 54 ctDNA molecules per mL plasma distinguished patients with high and low/no ctDNA load. Patients with low/no ctDNA load (N=102) showed longer median PFS of 5.7 months (P=0.006) and OS of 124.8 months (P=0.008) than patients with high ctDNA load (N=62) (4.4 months and 107.7 months, respectively) in multivariate analyses. Patients with

Purpose Correct identification of tumour receptor status is important for treatment decisions in breast cancer. [18F]FES PET and [18F]FDHT PET allow non-invasive assessment of the oestrogen (ER) and androgen receptor (AR) status of individual lesions within a patient. Despite standardised analysis techniques, interobserver variability can significantly affect the interpretation of PET results and thus clinical applicability. The purpose of this study was to determine visual and quantitative interobserver variability of [18F]FES PET and [18F]FDHT PET interpretation in patients with metastatic breast cancer. Methods In this prospective, two-centre study, patients with ER-positive metastatic breast cancer underwent both [18F]FES and [18F]FDHT PET/CT. In total, 120 lesions were identified in 10 patients with either conventional imaging (bone scan or lesions > 1 cm on high-resolution CT, n = 69) or only with [18F]FES and [18F]FDHT PET (n = 51). All lesions were scored visually and quantitatively by two independent observers. A visually PET-positive lesion was defined as uptake above background. For quantification, we used standardised uptake values (SUV): SUVmax, SUVpeak and SUVmean. Results Visual analysis showed an absolute positive and negative interobserver agreement for [18F]FES PET of 84% and 83%, respectively (kappa = 0.67, 95% CI 0.48–0.87), and 49% and 74% for [18F]FDHT PET, respectively (kappa = 0.23, 95% CI − 0.04–0.49). Intraclass correlation coefficients (ICC) for quantification of SUVmax, SUVpeak and SUVmean were 0.98 (95% CI 0.96–0.98), 0.97 (95% CI 0.96–0.98) and 0.89 (95% CI 0.83–0.92) for [18F]FES, and 0.78 (95% CI 0.66–0.85), 0.76 (95% CI 0.63–0.84) and 0.75 (95% CI 0.62–0.84) for [18F]FDHT, respectively. Conclusion Visual and quantitative evaluation of [18F]FES PET showed high interobserver agreement. These results support the use of [18F]FES PET in clinical practice. In contrast, visual agreement for [18F]FDHT PET was relatively low due to low tumour-background ratios, but quantitative agreement was good. This underscores the relevance of quantitative analysis of [18F]FDHT PET in breast cancer. Trial registration ClinicalTrials.gov, NCT01988324. Registered 20 November 2013, https://clinicaltrials.gov/ct2/show/NCT01988324?term=FDHT+PET&draw=1&rank=2.

The phosphatidylinositol-3-kinase (PI3K) and mitogen-activated protein kinase(MAPK) pathways are frequently activated in breast cancer. We recently demonstrated the importance of analyzing multiple proteins as read-out for pathway activation in ER+/HER2- breast cancer, since single proteins are known to provide insufficient information. Here, we determined pathway activation in other primary breast cancer intrinsic subtypes derived from postmenopausal patients. Tumor blocks were recollected and immunohistochemistry was performed using antibodies against PTEN, p-AKT(Thr308), p-AKT(Ser473), p-p70S6K, p-4EBP1, p-S6RP(Ser235/236) and p-ERK1/2, followed by unsupervised hierarchical clustering. In 32 ER+/HER2+, 37 ER-/HER2+ and 74 triple-negative breast cancer patients, subgroups were identified with preferentially activated (A) and preferentially not activated (N) proteins. These subgroups likely reflect tumors with differences in biological behavior as well as treatment outcome.

Background IGF-1R is overexpressed in a substantial number of breast cancer cases. When phosphorylated, IGF-1R activates the PI3K and MAPK pathways. In preclinical studies, IGF-1R overexpression has been described to be associated with tamoxifen resistance. Expression of activated IGF-1R has not been established as a marker for endocrine resistance in the clinic. We tested the value of p-IGF-1R to predict adjuvant tamoxifen benefit in IGF-1R-positive tumors from ER+ breast cancer patients and its possible association with PI3K/MAPK activation. We also carried out cell line experiments to illustrate a potential causal link between IGF-1R activation status, PI3K/MAPK pathway activation and tamoxifen resistance. Methods Primary tumor blocks from 563 ER+ (stage I-III) postmenopausal patients previously randomized between tamoxifen (1 to 3 years) vs. no adjuvant therapy (IKA trial 1982-1993; Beelen et al, Breast Cancer Res, 2014) were recollected. Immunohistochemistry scoring on tissue microarrays included PTEN, p-IGF-1R, p-AKT(Thr308), p-AKT(Ser473) and p-p70S6K(Thr389) by cytoplasmic intensity (0-3), of p-4EBP1(Ser65) and p-ERK1/2(Thr202/204) by percentage of tumor cells with positive nuclear staining and of p-S6RP(Ser235/236) by percentage of tumor cells with positive membranous staining. Informative data on p-IGF-1R staining was available for 364 IGF-1R-positive tumors. Multivariate Cox models including standard prognostic factors were used to assess hazard ratios (HR) for recurrence-free interval for tamoxifen treatment, p-IGF-1R status (negative vs. positive) and their interaction. MCF-7 and T47D cell lines were used to validate the clinical findings. IncuCyte cell proliferation experiments were performed with various IGF-1R-related growth stimulating and inhibiting conditions. Western blots were carried out on cells under various growth conditions to analyze whether activation of IGF-1R would be associated with PI3K/MAPK pathway activation. Results Patients having tumors without p-IGF-1R expression derived significant benefit from tamoxifen (HR 0.4290, 95% CI 0.2356 - 0.7813; p = 0.00566), while those having tumors with p-IGF-1R expression had no benefit (HR 0.8051, 95% CI 0.2643 – 2.453; p = 0.70). The p for interaction was not significant (p = 0.32106). Tumors positive for p-IGF-1R had more expression of (phospho)proteins downstream of the PI3K/MAPK pathways. These results were supported by Western blots from the cell lines examined under various growth conditions. In both cell lines, linsitinib (a dual IGF-1R and insulin receptor inhibitor) was able to block IGF-1R signaling, preventing activation of the PI3K and MAPK pathways and abrogating cell proliferation in the presence of tamoxifen. Conclusions Postmenopausal breast cancer patients with p-IGF-1R-positive tumors appear to derive no benefit from adjuvant tamoxifen. Tumors with p-IGF-1R expression were associated with PI3K/MAPK pathway activation. In breast cancer cell lines with activated IGF-1R signaling, addition of linsitinib to endocrine therapy can restore sensitivity. This combination might be an interesting treatment option for tamoxifen-resistant patients. Citation Format: Kruger DT, Alexi X, Opdam M, Schuurman KG, Sanders J, van der Noort V, Boven E, Zwart W, Linn SC. Phosphorylation of insulin-like growth factor-1 receptor (IGF-1R) is associated with tamoxifen resistance by activating the PI3K/MAPK pathway [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr P4-08-06.

Background Several (activated) proteins of the phosphatidylinositol-3-kinase (PI3K) and/or mitogen-activated protein kinase (MAPK) pathway have been analyzed for an association with adjuvant tamoxifen benefit in estrogen receptor-positive (ER+), HER2-negative (HER2-) breast cancer. Currently, no single protein has yielded convincing results allowing clinical implementation. Combining activated proteins could provide more robust predictive potential. We used unsupervised hierarchical clustering of 7 activated proteins downstream in these pathways, discriminated by expression profile, to analyze adjuvant tamoxifen benefit in ER+/HER2- postmenopausal breast cancer patients. Further, we developed a classification tool based on the generated heatmap groups by pathway activation status to categorize future patients into sensitive for or resistant to adjuvant tamoxifen. Methods Primary tumor blocks from 489 ER+/HER2- (stage I-III) postmenopausal patients previously randomized between tamoxifen (1 to 3 years) vs no adjuvant therapy (IKA trial 1982-1993; Beelen et al, Breast Cancer Res, 2014) were used for immunohistochemistry on tissue microarrays. Median follow-up of the patients without a recurrence event was 8.5 years. Scoring of PTEN, p-AKT(Thr308), p-AKT(Ser473) and p-p70S6K(Thr389) was by cytoplasmic intensity (0-3), p-4EBP1(Ser65) and pERK1/2(Thr202/204) by percentage of tumor cells with positive nuclear staining and p-S6RP(Ser235/236) by the percentage of tumor cells with positive membranous staining. Unsupervised hierarchical clustering was performed on tissue from 293 patients with informative data on all 7 proteins. Cox models were used to assess tamoxifen benefit (defined as the Hazard ratio (HR) of tamoxifen treatment vs no adjuvant therapy for recurrence-free interval) and to compare this benefit between subgroups generated by clustering as well as generated by the classification tool. Cox models included covariates (age, histological grade and subtype, tumor size, PR status) and were stratified for lymph node status. Results Two distinct groups were identified based on hierarchical clustering: one with preferentially activated pathway markers (A) and one with relatively no activation (N). Patients in group N derived significant benefit from tamoxifen (multivariable HR 0.28, 95% CI 0.14 − 0.56, p = 0.000324), while patients from group A had no benefit (multivariable HR 1.57, 95% CI 0.44 – 5.56, p = 0.482), p for interaction 0.018. Contradictory, patients in group A had a better prognosis compared to those in group N (multivariable HR 0.17, 95% CI 0.04 − 0.7, p = 0.014). The classification tool was also successful in differentiating ER+/HER2- patients with or without tamoxifen benefit (p for interaction = 0.016). Conclusions Hierarchical clustering of PI3K/MAPK pathway proteins is able to differentiate patients with adjuvant tamoxifen benefit and patients who would potentially need alternative treatment, for instance with inhibitors of the PI3K or MAPK pathway. The classification tool distinguishing patients with and without tamoxifen benefit should be validated in an independent cohort of postmenopausal patients with primary ER+/HER2- breast cancer. Citation Format: Kruger DT, Beelen K, Opdam M, Sanders J, van der Noort V, Boven E, Linn SC. Clustering of activated proteins of the PI3K and MAPK pathways distinguishes ER+/HER2- primary breast cancer patients with preferential tamoxifen benefit [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr PD4-12.

In addition to the well-known estrogen receptor (ER) and human epidermal growth factor receptor 2, the androgen receptor (AR) is also a potential drug target in breast cancer treatment. Whole-body imaging can provide information across lesions within a patient. ER expression in tumor lesions can be visualized by F-18-fluoroestradiol (F-18-FES) PET, and AR expression has been visualized in prostate cancer patients with F-18-fluorodihydrotestosterone (F-18-FDHT) PET. Our aim was to assess the concordance between F-18-FDHT and F-18-FES PET and tumor AR and ER expression measured immunohistochemically in patients with metastatic breast cancer. Methods: Patients with ER-positive metastatic breast cancer were eligible for the study, irrespective of tumor AR status. The concordance of F-18-FDHT and F-18-FES uptake on PET with immunohistochemical expression of AR and ER in biopsies of corresponding metastases was analyzed. Patients underwent F-18-FDHT PET and F-18-FES PET. A metastasis was biopsied within 8 wk of the PET procedures. Tumor samples with more than 10% and 1% nuclear tumor cell staining were considered, respectively, AR-and ER-positive. Correlations between PET uptake and semiquantitative immunohistochemical scoring (percentage positive cells x intensity) were calculated. The optimum threshold of SUV to discriminate positive and negative lesions for both AR and ER was determined by receiver-operating-characteristic analysis. Results: In the 13 evaluable patients, correlation (R-2) between semiquantitative AR expression and F-18-FDHT uptake was 0.47 (P = 0.01) and between semiquantitative ER expression and F-18-FES uptake 0.78 (P = 0.01). The optimal cutoff for AR-positive lesions was an SUVmax of 1.94 for F-18-FDHT PET, yielding a sensitivity of 91% and a specificity of 100%; the optimal cutoff was an SUVmax of 1.54 for F-18-FES PET, resulting in a sensitivity and specificity of 100% for ER. Conclusion: F-18-FDHT and F-18-FES uptake correlate well with AR and ER expression levels in representative biopsies. These results show the potential use of whole-body imaging for receptor status assessment, particularly in view of biopsy-associated sampling errors and heterogeneous receptor expression in breast cancer metastases.

The identification of biomarkers able to predict clinical benefit from vascular endothelial growth factor receptor (VEGFR) tyrosine kinase inhibitors is urgently needed. Recently, Maitland and colleagues described an association between KDR-rs34231037 and soluble VEGFR2 levels as well as pazopanib pharmacodynamics. We investigated in a well-characterized series of metastatic clear cell renal cell carcinoma patients whether rs34231037 could influence sunitinib response. Clinical data and DNA were available from an international series of 276 patients. KDR-rs34231037 association with sunitinib response, clinical benefit, and progression-free survival was analyzed using logistic and Cox regression analyses. We found that G-allele carriers were over-represented among patients with clinical benefit during sunitinib treatment compared with those refractory to the treatment (odds ratio: 3.78; 95% confidence interval: 1.02-14.06; P=0.047, multivariable analysis). In conclusion, rs34231037 variant carriers seemed to have better sunitinib response than wild-type patients. Moreover, the association with tumor size reduction suggests that this single nucleotide polymorphism might also identify patients with successful tumor downsizing under anti-VEGFR therapy.

Preclinical studies indicate that activated IGF‐1R can drive endocrine resistance in ER‐positive (ER+) breast cancer, but its clinical relevance is unknown. We studied the effect of IGF‐1R signaling on tamoxifen benefit in patients and we searched for approaches to overcome IGF‐1R‐mediated tamoxifen failure in cell lines. Primary tumor blocks from postmenopausal ER+ breast cancer patients randomized between adjuvant tamoxifen versus nil were recollected. Immunohistochemistry for IGF‐1R, p‐IGF‐1R/InsR, p‐ERα(Ser118), p‐ERα(Ser167) and PI3K/MAPK pathway proteins was performed. Multivariate Cox models were employed to assess tamoxifen efficacy. The association between p‐IGF‐1R/InsR and PI3K/MAPK pathway activation in MCF‐7 and T47D cells was analyzed with Western blots. Cell proliferation experiments were performed under various growth‐stimulating and ‐inhibiting conditions. Patients with ER+, IGF‐1R‐positive breast cancer without p‐IGF‐1R/InsR staining (n = 242) had tamoxifen benefit (HR 0.41, p = 0.0038), while the results for p‐IGF‐1R/InsR‐positive patients (n = 125) were not significant (HR 0.95, p = 0.3). High p‐ERα(Ser118) or p‐ERα(Ser167) expression was associated with less tamoxifen benefit. In MCF‐7 cells, IGF‐1R stimulation increased phosphorylation of PI3K/MAPK proteins and ERα(Ser167) regardless of IGF‐1R overexpression. This could be abrogated by the dual IGF‐1R/InsR inhibitor linsitinib, but not by the IGF‐IR‐selective antibody 1H7. In MCF‐7 and T47D cells, stimulation of the IGF‐1R/InsR pathway resulted in cell proliferation regardless of tamoxifen. Abrogation of cell growth was regained by addition of linsitinib. In conclusion, p‐IGF‐1R/InsR positivity in ER+ breast cancer is associated with reduced benefit from adjuvant tamoxifen in postmenopausal patients. In cell lines, stimulation rather than overexpression of IGF‐1R is driving tamoxifen resistance to be abrogated by linsitinib., What's new? In breast cancer, the growth factor receptor IGF‐1R is frequently overexpressed, boosting cell proliferation and transformation. Prior work has shown that IGF‐1R pathway activation promotes treatment resistance in cell lines. Here, the authors evaluated whether the pathway contributes to tamoxifen resistance in patients. In patients with ER+ breast cancer, patients whose cancers tested negative for activated IGF‐R1 benefited from adjuvant tamoxifen, while those with cancers tested positive showed little improvement. Next, they showed that treating breast cancer cells with linsitinib can thwart IGF‐1R signaling and restore tamoxifen's efficacy. Patients with activated IGF‐1R pathways may therefore do better with tamoxifen combined with linsitinib than tamoxifen alone.

ContextStudies indicate adverse effects of breast cancer (BC) and cancer treatment on cognitive function.ObjectivesTo investigate the effects of systemic treatment on cognitive performance in BC patients.MethodsParticipants were BC patients scheduled to receive systemic treatment (BC + SYST; n = 31), or no systemic treatment (BC; n = 24) and no-cancer (NC) controls (n = 33). Neuropsychological examinations were used to study cognitive performance on 18 tests grouped into eight cognitive domains, before adjuvant treatment (T1) and six months after chemotherapy (T2), or at similar intervals. We also assessed health-related quality of life, anxiety and depression, mood, stress, and cognitive problems. Analysis of variance was used to assess group differences of cognitive performance and multivariate normative comparison to classify impairment, comparing scores of each participant against the distribution of the scores of NC controls.ResultsOf BC + SYST, 16% were cognitively impaired at T2, compared to 4% in BC and 6% in NC. Although not significant, we observed moderate effect sizes for worse performance in the BC + SYST group compared to NC (Flanker congruent [effect size {ES} = 0.44] and stimulus incongruent [ES = 0.44]) and compared to BC (Controlled Oral Word Association Test [ES = 0.47], digit span [ES = 0.41], and Hopkins Verbal Learning Test immediate [ES = 0.71] and delayed recall [ES = 0.65]). Cognitively impaired patients had a significantly lower estimated premorbid intelligence, worse physical and social functioning, and more distress at T2 compared to unimpaired patients.ConclusionOur findings indicate that cognitive impairment after systemic treatment occurs in a subset of BC patients. The predictive value of demographic and psychosocial factors in cognitive impairment should be further investigated in a larger sample of impaired patients.

Purpose: The survival of patients with clear cell metastatic renal cell carcinoma (cc-mRCC) has improved substantially since the introduction of tyrosine kinase inhibitors (TKI). With the fact that TKIs interact with immune responses, we investigated whether polymorphisms of genes involved in immune checkpoints are related to the clinical outcome of cc-mRCC patients treated with sunitinib as first TKI. Experimental Design: Twenty-seven single-nucleotide polymorphisms (SNP) in CD274 (PD-L1), PDCD1 (PD-1), and CTLA-4 were tested for a possible association with progression-free survival (PFS) and overall survival (OS) in a discovery cohort of 550 sunitinib-treated cc-mRCC patients. SNPs with a significant association (P < 0.05) were tested in an independent validation cohort of 138 sunitinib-treated cc-mRCC patients. Finally, data of the discovery and validation cohort were pooled for meta-analysis. Results: CTLA-4 rs231775 and CD274 rs7866740 showed significant associations with OS in the discovery cohort after correction for age, gender, and Heng prognostic risk group [HR, 0.84; 95% confidence interval (CI), 0.72–0.98; P = 0.028, and HR, 0.73; 95% CI, 0.54–0.99; P = 0.047, respectively]. In the validation cohort, the associations of both SNPs with OS did not meet the significance threshold of P < 0.05. After meta-analysis, CTLA-4 rs231775 showed a significant association with OS (HR, 0.83; 95% CI, 0.72–0.95; P = 0.008). Patients with the GG genotype had longer OS (35.1 months) compared with patients with an AG (30.3 months) or AA genotype (24.3 months). No significant associations with PFS were found. Conclusions: The G-allele of rs231775 in the CTLA-4 gene is associated with an improved OS in sunitinib-treated cc-mRCC patients and could potentially be used as a prognostic biomarker. Clin Cancer Res; 24(10); 2350–6. ©2018 AACR.

Background: The phosphatidylinositol-3-kinase (PI3K) and/or mitogen-activated protein kinase (MAPK) pathways are frequently activated in breast cancer which can result in antioestrogen resistance. Single protein markers failed to be introduced into clinical practice. We, therefore, aimed to find a better read-out of activation of the PI3K and MAPK pathways in ER+/HER2− breast cancer. Assessment of seven PI3K/MAPK proteins might better reflect pathway activation and distinguish patients without adjuvant tamoxifen benefit. Methods: Tumour blocks were recollected from 293 primary postmenopausal ER+/HER2− breast cancer patients randomised between tamoxifen and no adjuvant therapy. PTEN, p-AKT(Thr308), p-AKT(Ser473), p-p70S6K, p-4EBP1, p-ERK1/2 and p-S6RP expression was assessed by immunohistochemistry followed by unsupervised hierarchical clustering. The primary endpoint was recurrence-free interval. Multivariate Cox models were used to assess tamoxifen benefit. A classification tool was developed based on protein expression profile. Results: Subgroups were identified with preferentially activated (A) and preferentially not activated (N) proteins. Patients in group N derived significant benefit from tamoxifen (multivariate hazard ratio (HR) = 0.23, p = 0.000101), while patients from group A did not (multivariate HR = 1.37, p = 0.64), p for interaction 0.020. Our generated classification tool confirmed these results (p for interaction 0.024). Conclusions: Hierarchical clustering of seven PI3K/MAPK proteins reflects pathway activation and can guide treatment decisions in primary ER+/HER2− postmenopausal breast cancer patients.

Purpose We evaluated the effectiveness of a low-intensity, home-based physical activity program (Onco-Move) and a moderate- to high-intensity, combined supervised resistance and aerobic exercise program (OnTrack) versus usual care (UC) in maintaining or enhancing physical fitness, minimizing fatigue, enhancing health-related quality of life, and optimizing chemotherapy completion rates in patients undergoing adjuvant chemotherapy for breast cancer. Patients and Methods We randomly assigned patients who were scheduled to undergo adjuvant chemotherapy (N = 230) to Onco-Move, OnTrack, or UC. Performance-based and self-reported outcomes were assessed before random assignment, at the end of chemotherapy, and at the 6-month follow-up. We used generalized estimating equations to compare the groups over time. Results Onco-Move and OnTrack resulted in less decline in cardiorespiratory fitness (P < .001), better physical functioning (P ≤ .001), less nausea and vomiting (P = .029 and .031, respectively) and less pain (P = .003 and .011, respectively) compared with UC. OnTrack also resulted in better outcomes for muscle strength (P = .002) and physical fatigue (P < .001). At the 6-month follow-up, most outcomes returned to baseline levels for all three groups. A smaller percentage of participants in OnTrack required chemotherapy dose adjustments than those in the UC or Onco-Move groups (P = .002). Both intervention groups returned earlier (P = .012), as well as for more hours per week (P = .014), to work than the control group. Conclusion A supervised, moderate- to high-intensity, combined resistance and aerobic exercise program is most effective for patients with breast cancer undergoing adjuvant chemotherapy. A home-based, low-intensity physical activity program represents a viable alternative for women who are unable or unwilling to follow the higher intensity program.

Background The phase II ATX trial aimed at evaluating safety and efficacy of first-line AT or ATX for HER2-negative LR/MBC (NTR1348; BOOG 2006-06). Plasma samples were collected for investigation of circulating proteins involved in angiogenesis and their possible association with therapy outcome. We here report the prognostic value of plasma VEGF-A, soluble (s)VEGFR-2, ANG2, sTIE2, IL6, IL8 and CA9 at baseline (C1D1) and their changes after cycle 1 (C2D1). Methods 312 patients were randomized 1:1 to AT (T 90 mg/m2 d1, 8, 15 & A 10 mg/kg d1, 15 q4w x 6 cycles → A 15 mg/kg d1 q3w for next cycles) or ATX (T 90 mg/m2 d1, 8, A 15 mg/kg d1 & X 825 mg/m2 bid d1–14 q3w x 8 cycles → A & X at same dose q3w for next cycles). Plasma proteins were measured by immunoassays (R&D Systems and MSD). The association of (continuous) protein levels on C1D1 and their changes on C2D1 with response, PFS and OS were evaluated by Mann-Whitney U test and univariate Cox regression analysis. Results The biomarker cohort (n=181) and trial cohort had similar baseline characteristics and clinical outcome. After a median follow-up of 46 months, there were 178 (98%) PFS events and 152 (84%) deaths. On C1D1, levels of ANG2 and sTIE2 were significantly higher in patients with hormone-receptor positive disease compared to those with triple-negative disease (p=.025 and p=.001, respectively). A high level of VEGF-A was noted in patients with visceral metastases (p=.028) and those with an increasing number of metastatic sites (p=.017). High levels of ANG2, IL6, IL8 and CA9 were significantly associated with poor PFS and OS (Table 1). Protein levels were not associated with response. Table 1. Association of circulating proteins at baseline with PFS and OSPFS HR (95%CI)PFS p-valueOS HR (95%CI)OS p-valueANG22.3 (1.0-4.9).043.1 (1.4-7.0).007IL61.6 (1.1-2.3).021.9 (1.3-2.8).001IL81.9 (1.3-2.8).0012.8 (1.8-4.3) On C2D1, levels of all proteins, except for IL6, had significantly changed. Whereas VEGF-A, ANG2, sTIE2 and IL8 decreased significantly, sVEGFR2 and CA9 showed a significant increase. The median relative change of both IL8 and sVEGFR2 was significantly different between patients having CR/PR vs. SD/PD (for IL8: -19.4% vs. 22.3%, p=.001 and for sVEGFR2: 6.5 vs. 2.1%, p=.01). A large relative increase in CA9 level was associated with better PFS (HR = 0.36, 95% CI, 0.19 – 0.68, p=.002) and OS (HR = 0.50, 95% CI, 0.27 – 0.94, p=.03). All patients had very low levels of free VEGF-A on C2D1 (median 8 pg/ml). Conclusions In patients with HER2-negative LR/MBC receiving first-line bevacizumab-containing chemotherapy, high baseline plasma levels of ANG2, IL6, IL8 and CA9 indicate a high risk for poor PFS and OS. These proteins might be useful for stratification according to prognosis. Moreover, relative decrease in IL8 and increase in sVEGFR2 on C2D1 are associated with response, whereas a large relative increase in CA9 is associated with better PFS and OS. Changes in these proteins after one cycle might be early indicators of efficacy of bevacizumab combined with chemotherapy. Financial support from Roche Netherlands. Citation Format: Siu W Lam, Nienke M Nota, Steffen M de Groot, Agnes Jager, Monique MEM Bos, Sabine C Linn, Joan van den Bosch, Hans J Braun, Ankie MT van der Velden, Maartje Los, Johanneke EA Portielje, Judith R Kroep, Aafke H Honkoop, Carolien H Smorenburg, Bea Tanis, Johanna MGH van Riel, Jetske M Meerum Terwogt, Marien O den Boer, Joep Douma, Frank Jeurissen, Johan Berends, Harm van Tinteren, Epie Boven. Plasma biomarker analysis in patients with HER2-negative locally recurrent or metastatic breast cancer (LR/MBC) treated with first-line bevacizumab (A) and paclitaxel (T) without or with capecitabine (X) [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P3-06-09.

Purpose Erlotinib is an orally administered tyrosine kinase inhibitor used for treatment of non-small cell lung cancer. Understanding actual use of medication is essential for optimizing treatment conditions. Methods In this multicentre prospective observational study, patients starting erlotinib treatment were followed for 4 months. Adherence was assessed using a medication event monitoring system (MEMS). Area under the curve (AUC) was determined after 1, 2 and 4 months. Before start and at monthly intervals, patients filled out questionnaires about attitude towards medication and disease, quality of life, symptoms and use in daily practice. Results Sixty-two patients (median age 63.5 years, 53 % male) were included of whom 15 were still on treatment after 4 months. MEMS data of 55 patients revealed a mean adherence of 96.8 +/- 4.0 %. Over one-third of patients had an adherence rate < 95 %. At 1 month, 21 % of patients did not always correctly take erlotinib without food. Associated risk factors were older age, suboptimal adherence, ocular symptoms and stomatitis (all p < 0.05). After 1 month of treatment, fatigue (91 %) and rash (86 %) were the most common symptoms reported. AUCss of erlotinib was higher in patients with rash and patients with moderate-severe anorexia (both p < 0.05). Conclusion Though adherence to erlotinib treatment is generally high, non-adherence might be an issue in a considerable number of patients. To support optimal erlotinib intake, clinicians need to take adequate measures to ameliorate symptoms and to address adherence and correct intake without food. Especially older patients and those who experience stomatitis may need extra attention.

Background: In our exploratory studies, we associated single nucleotide polymorphisms (SNPs) in candidate genes with the efficacy and toxicities of sunitinib in metastatic renal cell carcinoma (mRCC). Objective: To see whether previously reported associations of SNPs with sunitinib-induced toxicities and efficacy in mRCC can be confirmed in a large cohort of patients. Design, setting, and participants: The mRCC patients treated with sunitinib and a DNA sample available were pooled from three exploratory studies conducted in the United States, Spain, and the Netherlands. A total of 22 SNPs and 6 haplotypes in 10 candidate genes related to the pharmacokinetics and pharmacodynamics of sunitinib were selected for association testing. Outcome measurements and statistical analysis: SNPs and haplotypes were tested for associations with toxicity, dose reductions, progression-free survival (PFS), overall survival (OS), and best objective response. Results and limitations: A total of 333 patients were included. We confirmed 2 of the 22 previously reported SNP associations. The presence of CYP3A5*1 was associated with dose reductions (odds ratio: 2.0; 95% confidence interval [CI], 1.0-4.0, p = 0.039). The presence of CGT in the ABCB1 haplotype was associated with an increased PFS (hazard ratio: 1.9; 95% CI, 1.3-2.6; p < 0.001) and remained significant after Bonferroni correction. These associations are consistent with prior observations. Conclusions: The confirmation of previously reported associations between polymorphisms in CYP3A5 and ABCB1 with sunitinib toxicity and efficacy, respectively, indicates that genotyping of these genetic variants will be useful for guiding sunitinib treatment. A prospective validation study is needed to confirm our findings on ABCB1 and CYP3A5 genetic polymorphisms. Patient summary: We confirmed that variants in genes involved in processing sunitinib through the body have an effect on sunitinib treatment outcome. These findings confirm the potential of testing for these genetic variants to improve individual patient care for patients with metastatic renal cell carcinoma treated with sunitinib. (C) 2015 European Association of Urology. Published by Elsevier B.V. All rights reserved.

We examined whether genetic polymorphisms (SNPs) in the capecitabine activation pathway and CDA enzymatic activity were associated with prognosis, benefit from capecitabine-containing treatment or capecitabine-related toxicities. The study population comprised 188 metastatic breast cancer patients of the ATX trial (EudraCT 2006-006058-83) randomized for first-line paclitaxel and bevacizumab with (ATX) or without capecitabine (AT). Cumulative capecitabine dose until grade ≥2 hand-foot syndrome or until first dose reduction were toxicity endpoints. We genotyped CDA c.-451C. >. T (rs532545), CDA c.-33delC (rs3215400) and CES2 c.-806C. >. G (rs11075646). CDA activity in baseline serum was measured with a spectrophotometric assay and values were analyzed using a median cut-off or as continuous variable. CDA c.-33delC was prognostic for overall survival (OS) independent of hormone receptor status. For the predictive analysis, progression-free survival benefit from ATX over AT was observed in patients with a CDA c.-33del/del or del/insC genotype, a CDA c.-451CC or CT genotype, and a CES2 c.-806CC genotype compared with their counterparts. There was a higher response rate for ATX over AT in patients with a CDA c.-451CT or TT genotype. Patients with high CDA enzymatic activity had more benefit from capecitabine, while this was marginally observed in the CDA low group. Toxicity endpoints were not associated with any candidate markers. In conclusion, CDA c.-33delC was associated with OS. Since particular SNPs in CDA and CES2 were associated with benefit from the addition of capecitabine to AT, their predictive value should be explored in a higher number of patients.

// Xiaoyan Liu 1, 2 , Jesse J. Swen 1 , Epie Boven 3 , Daniel Castellano 4, 5 , Hans Gelderblom 6 , Ron H.J. Mathijssen 7 , Cristina Rodriguez-Antona 8, 9 , Jesus Garcia-Donas 5, 10 , Brian I. Rini 11 , Henk-Jan Guchelaar 1 1 Department of Clinical Pharmacy and Toxicology, Leiden University Medical Center, Leiden, The Netherlands 2 Institute of Clinical Pharmacology, Qilu Hospital of Shandong University, Jinan, China 3 Department of Medical Oncology, VU University Medical Center, Amsterdam, The Netherlands 4 Oncology Department, Hospital Universitario 12 de Octubre, Madrid, Spain 5 Spanish Oncology Genitourinary Group (SOGUG), Madrid, Spain 6 Department of Medical Oncology, Leiden University Medical Center, Leiden, The Netherlands 7 Department of Medical Oncology, Erasmus MC Cancer Institute, Rotterdam, The Netherlands 8 Hereditary Endocrine Cancer Group, Spanish National Cancer Research Centre (CNIO), Madrid, Spain 9 ISCIII Center for Biomedical Research on Rare Diseases (CIBERER), Madrid, Spain 10 Oncology Unit, Clara Campal Comprehensive Cancer Center, Madrid, Spain 11 Department of Solid Tumor Oncology, Cleveland Clinic Taussig Cancer Institute (CCF), Cleveland, Ohio, USA Correspondence to: Henk-Jan Guchelaar, email: H.J.Guchelaar@lumc.nl Keywords: sunitinib, metastatic renal cell carcinoma, VEGFR1 , validation study, meta-analysis Received: August 20, 2016 Accepted: November 07, 2016 Published: November 25, 2016 ABSTRACT VEGFR1 rs9582036 and rs9554320 were previously reported the association with sunitinib progression-free survival (PFS) and overall survival (OS) in patients with metastatic renal cell carcinoma (mRCC). Hereafter, the association of both single nucleotide polymorphisms (SNPs) with PFS/OS was confirmed in two independent mRCC cohorts. The aim of the current study was to validate the associations of both SNPs with sunitinib outcome in three independent well-characterized cohorts (SUTOX, CCF and SOGUG) including 286 sunitinib-treated mRCC patients, as well as to perform a meta-analysis of current and published data combined. We found that rs9582036 and rs9554320 showed a significant association with sunitinib PFS in the CCF cohort (HR: 0.254, 95%CI: 0.092-0.703; P=0.008 and HR: 0.430, 95%CI: 0.200-0.927; P=0.031, respectively). Patients with the variant genotype of rs9582036 and rs9554320 had a shorter median PFS. No significant association of both SNPs with sunitinib PFS or OS was detected in either the SUTOX or SOGUG cohort. After the combination of all available data into a meta-analysis, the association of both SNPs with sunitinib PFS or OS did not achieve the threshold for statistical significance. Our findings suggest that, although VEGFR1 rs9582036 and rs9554320 are involved in sunitinib therapy outcome, its clinical use as biomarkers for prediction of sunitinib outcome in mRCC patients is limited, due to inconsistent findings when analyzing all existing studies together.

Breast cancer is traditionally considered as a heterogeneous disease. Molecular profiling of breast cancer by gene expression studies has provided us an important tool to discriminate a number of subtypes. These breast cancer subtypes have been shown to be associated with clinical outcome and treatment response. In order to elucidate the functional consequences of altered gene expressions related to each breast cancer subtype, proteomic technologies can provide further insight by identifying quantitative differences at the protein level. In recent years, proteomic technologies have matured to an extent that they can provide proteome-wide expressions in different clinical materials. This technology can be applied for the identification of proteins or protein profiles to further refine breast cancer subtypes or for discovery of novel protein biomarkers pointing towards metastatic potential or therapy resistance in a specific subtype. In this review, we summarize the current state of knowledge of proteomic research on molecular breast cancer classification and discuss important aspects of the potential usefulness of proteomics for discovery of breast cancer-associated protein biomarkers in the clinic.

Colorectal cancer (CRC) is a major health problem. Biomarkers associated with molecular changes in cancer cells can aid early detection, diagnosis, prognosis, therapy selection, and disease monitoring. Tumor tissue secretomes are a rich source of candidate biomarkers. To identify CRC protein biomarkers, secretomes of four pairs of human CRC tissue and patient-matched normal colon tissue samples, and secretomes of five CRC cell lines were analyzed by GeLC-MS/MS. Subsequent data analysis was based on label-free spectral counting, Ingenuity Pathway Analysis, Secretome/SignalP, STRING and Cytoscape, resulting in 2703 protein identifications in the tissue secretomes, of which 409 proteins were significantly more present in CRC samples than in controls. Biomarker selection of 76 candidates was based on consistent and abundant over-representation in cancer- compared to control-secretomes, and presumed neoplastic origin. Overlap analysis with previously obtained datasets revealed 21 biomarkers suited for early detection of CRC. Immunohistochemistry confirmed overexpression in CRC of one candidate marker (MCM5). In conclusion, a human reference dataset of 76 candidate biomarkers was identified for which we illustrate that combination with existing pre-clinical datasets allows pre-selection of biomarkers for blood- or stool-based assays to support clinical management of CRC. Further dedicated validation studies are required to demonstrate their clinical applicability.Tissue secretome proteomes are a rich source of candidate biomarkers. Several secretome proteome datasets have been obtained from pre-clinical in vitro and in vivo colorectal cancer (CRC) model systems, yielding promising CRC biomarkers obtained under well-defined experimentally controlled conditions. However, which of these biomarker proteins are actually secreted by human CRC samples was not known. To our knowledge, this is the first study that directly compares secretome proteomes from clinically relevant human CRC tissues to patient-matched normal colon tissues. We identified 76 human CRC protein biomarkers that may facilitate blood-based or stool-based assay development to support clinical management of CRC. Overlap analysis with datasets from well-defined pre-clinical studies helps to determine what clinical application suits these human CRC biomarkers best, i.e. early detection, diagnosis, prognosis, therapy selection, and/or disease monitoring of CRC. This is demonstrated for a CRC mouse model dataset, revealing 21 human CRC biomarkers suited for early detection of CRC.

Capecitabine is an oral prodrug of 5-fluorouracil (5-FU) and approved for treatment of various malignancies. Hereditary genetic variants may affect a drug's pharmacokinetics or pharmacodynamics and account for differences in treatment response and adverse events among patients. In this review we present the current knowledge on genetic variants, commonly single-nucleotide polymorphisms (SNPs), tested in cohorts of cancer patients and possibly useful for prediction of capecitabine efficacy or toxicity. Capecitabine is activated to 5-FU by CES, CDA and TYMP , of which SNPs in CDA and CES2 were found to be associated with efficacy and toxicity. In addition, variants in genes of the 5-FU metabolic pathway, including TYMS , MTHFR and DPYD also influenced capecitabine efficacy and toxicity. In particular, well-known SNPs in TYMS and DPYD as well as putative DPYD SNPs had an association with clinical outcome as well as adverse events. Inconsistent findings may be attributable to factors related to ethnic differences, sample size, study design, study endpoints, dosing schedule and the use of multiple agents. Of the SNPs described in this review, dose reduction of fluoropyrimidines based on the presence of DPYD variants * 2A (rs3918290), * 13 (rs55886062), −2846A>T (rs67376798) and −1236G>A/HapB3 (rs56038477) has already been recommended. Other variants merit further validation to establish their definite role in explanation of interindividual differences in the outcome of capecitabine-based therapy.

Background: The purpose of this study was to evaluate single-nucleotide polymorphisms (SNPs) in genes encoding key metabolising enzymes or involved in pharmacodynamics for possible associations with paclitaxel-induced peripheral neuropathy. Methods: The study population consists of 188 women from the multicenter, randomised, phase II ATX trial (BOOG2006-06; EudraCT number 2006-006058-83) that received paclitaxel and bevacizumab without or with capecitabine as first-line palliative therapy of HER2-negative metastatic breast cancer. Genotyping of CYP2C8*3 (c.416G>A), CYP3A4*22 (c.522-191C>T), TUBB2A (c.-101T>C), FGD4 (c.2044-236G>A) and EPHA5 (c.2895G>A) was performed by real-time PCR. Toxicity endpoints were cumulative dose (1) until first onset of grade ⩾1 peripheral neuropathy and (2) until first paclitaxel dose reduction from related toxicity (NCI-CTCAE version 3.0). SNPs were evaluated using the Kaplan–Meier method, the Gehan–Breslow–Wilcoxon test and the multivariate Cox regression analysis. Results: The rate of grade ⩾1 peripheral neuropathy was 67% (n=126). The rate of dose reduction was 46% (n=87). Age ⩾65 years was a risk factor for peripheral neuropathy (HR=1.87, P

Background Newly identified SNPs in genes encoding metabolizing enzymes (CYP2C8, CYP3A4) and drug target (TUBB2A) of paclitaxel have been associated with PNP1-3. A recent GWAS has found novel SNPs in EPHA5 and FGD4 possibly predictive for PNP4. Likewise, SNPs in genes (CDA, CES2) involved in capecitabine activation may play a role in HFS5. Here, we attempted to confirm these SNPs as predictive markers for PNP and HFS in patients (pts) receiving first-line paclitaxel (T) and bevacizumab (A) without or with capecitabine (X). Patients and methods In the phase II ATX trial (NTR1348;BOOG2006-06), 312 pts were randomized 1:1 to AT (T 90 mg/m2 d1, 8, 15 & A 10 mg/kg d1, 15 q4w x 6 cycles → A 15 mg/kg d1 q3w for next cycles) or ATX (T 90 mg/m2 d1, 8, A 15 mg/kg d1 & X 825 mg/m2 bid d1–14 q3w x 8 cycles → the same dose of A & X q3w for next cycles). Toxicity was graded by using NCI CTCAE v3 at each cycle. Germline DNA was isolated for genotyping CYP2C8*3 (1196A>G & 416G>A), CYP3A4*22 (522-191C>T), EPHA5 2895G>A, FGD4 2044-236G>A, CES2 823C>G, TUBB2A 101C>T, TUBB2A 112G>A, CDA 943insC and CDA 451C>T. Results of TUBB2A and CDA SNPs will be presented at the meeting. The association between SNPs and toxicity was analyzed for 1) maximum grade of treatment-related toxicity per patient by Pearson’s X2 or Fisher's exact test; 2) cumulative dose level of drugs until first grade ≥1 toxicity or first dose reduction by Kaplan-Meier curve and Gehan-Breslow test. Results 188 pts had SNPs genotyped; characteristics reflected the trial cohort. Table 1 shows the maximum grade of PNP. When grouped into grades 0–1 and 2–3, no differences in rates of PNP were noted among SNPs. Median cumulative dose of T until grade ≥1 PNP was 1,800 mg/m2 (95% CI 1,534–2,065). Carriers of the CYP2C8 416 A-allele had a significantly lower cumulative dose of T until grade ≥1 PNP compared to those with a homozygous G/G genotype (Gehan-Breslow p=0.011). Carriers of the FGD4 2044-236 A-allele had a significantly higher cumulative dose until first dose reduction of T compared to those with a homozygous G/G genotype (Gehan-Breslow p=0.037). Table 1. toxicity by maximum grade per patient Grade 0123Paclitaxel-treated patients (N=188)PNP, No. (%)62 (33)33 (36)41 (22)17 (9)Capecitabine-treated patients (N=93)HFS, No. (%)17 (18)21 (23)25 (27)30 (32) For ATX-treated pts, rates of HFS are shown in Table 1. With regard to maximum grade of HFS, there was a trend of a lower rate of grade 2–3 HFS for CES 823C/G compared to C/C genotype (33% vs 64%, Fisher’s exact p=0.059). The median cumulative dose of X until grade ≥1 HFS was 98.7 g/m2 (95%CI 82.1–115). CES 823C>G was not associated with the cumulative dose of X until grade ≥1 HFS or dose reduction. Conclusions Our results indicate that CYP2C8 416G>A and FGD4 2044-236G>A might be predictive markers for paclitaxel-induced neurotoxicity, whereas reported associations of other SNPs with toxicity could not be confirmed. 1Hertz et al. Ann Oncol 2013, 2De Graan et al. Clin Cancer Res 2013, 3Leandro-Garcia et al. Clin Cancer Res 2012, 4Baldwin et al. Clin Cancer Res 2012, 5Caronia et al. Clin Cancer Res 2011 Financial support from Roche Netherlands. Citation Format: Siu W Lam, Charlotte N Frederiks, Tahar van der Straaten, Steffen M de Groot, Agnes Jager, Monique MEM Bos, Sabine C Linn, Joan van den Bosch, Hans J Braun, Ankie MT van der Velden, Maartje Los, Jojanneke EA Portielje, Judith R Kroep, Aafke H Honkoop, Carolien H Smorenburg, Bea Tanis, Johanna MGH van Riel, Jetske M Meerum Terwogt, Marien O den Boer, Joep Douma, Frank Jeurissen, Johan Berends, Henk-Jan Guchelaar, Epie Boven. Genetic polymorphisms (SNPs) as predictive markers for paclitaxel-induced peripheral neuropathy (PNP) and capecitabine-induced hand-foot syndrome (HFS) in HER-2 negative metastatic breast cancer patients [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P6-08-09.

In contrast to various signatures that predict the prognosis of breast cancer patients, markers that predict chemotherapy response are still elusive. To detect such predictive biomarkers, we investigated early changes in protein expression using two mouse models for distinct breast cancer subtypes who have a differential knock-out status for the breast cancer 1, early onset (Brca1) gene. The proteome of cisplatin-sensitive BRCA1-deficient mammary tumors was compared with that of cisplatin-resistant mammary tumors resembling pleomorphic invasive lobular carcinoma. The analyses were performed 24 h after administration of the maximum tolerable dose of cisplatin. At this time point, drug-sensitive BRCA1-deficient tumors showed DNA damage, but cells were largely viable. By applying paired statistics and quantitative filtering, we identified highly discriminatory markers for the sensitive and resistant model. Proteins up-regulated in the sensitive model are involved in centrosome organization, chromosome condensation, homology-directed DNA repair, and nucleotide metabolism. Major discriminatory markers that were up-regulated in the resistant model were predominantly involved in fatty acid metabolism, such as fatty-acid synthase. Specific inhibition of fatty-acid synthase sensitized resistant cells to cisplatin. Our data suggest that exploring the functional link between the DNA damage response and cancer metabolism shortly after the initial treatment may be a useful strategy to predict the efficacy of cisplatin.

Background In a Dutch multicentre study, elderly (65 + year) metastatic breast cancer patients, eligible for first-line chemotherapy, were randomised between two types of single-agent chemotherapy. As accrual was slow, with 78 randomised patients between April 2007 and September 2011, we explored potential barriers in the accrual process and their consequences for characteristics of included patients. Methods We sent surveys on the reasons for non-inclusion to all coordinating investigators. We also examined inclusion in a concurrent, non-elderly breast cancer study of the trialists' group and analysed baseline geriatric characteristics of included patients. Results Investigators from fifteen participating centres returned the survey. Most commonly reported barriers to inclusion were: patient's refusal of chemotherapy ( n = 8) or of randomisation ( n = 9), impaired cognition ( n = 3) and insufficient cardiac function ( n = 2). Oncologists' preference for combination regimens over single-agent chemotherapy was reported twice. Twenty-eight potentially eligible patients, aged 65–71 years, were included in a concurrent, study investigating combination chemotherapy in fit non-elderly patients with metastatic breast cancer. However, baseline characteristics of the included patients showed that the OMEGA study succeeded in including frail and older patients, with a performance status of 2 in 22% of patients and 54% of patients aged 75 years or older. Conclusion Accrual in this study was mainly hampered by patient's refusal or preference for a particular type of treatment, and an overall condition considered as too fit or too frail for inclusion. Future trials in elderly metastatic breast cancer patients should focus on non-restrictive inclusion criteria as well as on education of physicians and elderly patients on the advantages of trial participation.

Purpose The purpose of our study was to evaluate the effect of cognitive behavioral therapy (CBT), physical exercise (PE), and of these two interventions combined (CBT/PE) on menopausal symptoms (primary outcome), body image, sexual functioning, psychological well-being, and health-related quality of life (secondary outcomes) in patients with breast cancer experiencing treatment-induced menopause. Patients and Methods Patients with breast cancer reporting treatment-induced menopausal symptoms (N = 422) were randomly assigned to CBT (n = 109), PE (n = 104), CBT/PE (n = 106), or to a waiting list control group (n = 103). Self-report questionnaires were completed at baseline, 12 weeks, and 6 months. Multilevel procedures were used to compare the intervention groups with the control group over time. Results Compared with the control group, the intervention groups had a significant decrease in levels of endocrine symptoms (Functional Assessment of Cancer Therapy–Endocrine Symptoms; P < .001; effect size, 0.31-0.52) and urinary symptoms (Bristol Female Lower Urinary Tract Symptoms Questionnaire; P = .002; effect size, 0.29-0.33), and they showed an improvement in physical functioning (36-Item Short Form Health Survey physical functioning subscale; P = .002; effect size, 0.37-0.46). The groups that included CBT also showed a significant decrease in the perceived burden of hot flashes and night sweats (problem rating scale of the Hot Flush Rating Scale; P < .001; effect size, 0.39-0.56) and an increase in sexual activity (Sexual Activity Questionnaire habit subscale; P = .027; effect size, 0.65). Most of these effects were observed at both the 12-week and 6-month follow-ups. Conclusion CBT and PE can have salutary effects on endocrine symptoms and, to a lesser degree, on sexuality and physical functioning of patients with breast cancer experiencing treatment-induced menopause. Future work is needed to improve the design and the planning of these interventions to improve program adherence.

Background: Only a small proportion of patients respond to anti-VEGF therapy, pressing the need for a reliable biomarker that can identify patients who will benefit. We studied the biological activity of anti-VEGF antibodies in patients' blood during anti-VEGF therapy by using the Ba/F3-VEGFR2 cell line, which is dependent on VEGF for its growth. Methods: Serum samples from 22 patients with cancer before and during treatment with bevacizumab were tested for their effect on proliferation of Ba/F3-VEGFR2 cells. Vascular endothelial growth factor as well as bevacizumab concentrations in serum samples from these patients were determined by enzyme linked immunosorbent assay (ELISA). Results: The hVEGF-driven cell proliferation was effectively blocked by bevacizumab (IC50 3.7 μg ml−1; 95% CI 1.7–8.3 μg ml−1). Cell proliferation was significantly reduced when patients' serum during treatment with bevacizumab was added (22–103% inhibition compared with pre-treatment). Although bevacizumab levels were not related, on-treatment serum VEGF levels were correlated with Ba/F3-VEGFR2 cell proliferation. Conclusions: We found that the neutralising effect of anti-VEGF antibody therapy on the biological activity of circulating VEGF can be accurately determined with a Ba/F3-VEGFR2 bioassay. The value of this bioassay to predict clinical benefit of anti-VEGF antibody therapy needs further clinical evaluation in a larger randomised cohort.

Breast cancer arising in female BRCA1 mutation carriers is characterized by an aggressive phenotype and early age of onset. We performed tandem mass spectrometry-based proteomics of secretomes and exosome-like extracellular vesicles from BRCA1-deficient and BRCA1-proficient murine breast tumor models to identify extracellular protein biomarkers, which can be used as an adjunct to current diagnostic modalities in patients with BRCA1-deficient breast cancer. We identified 2,107 proteins, of which 215 were highly enriched in the BRCA1-deficient secretome. We demonstrated that BRCA1-deficient secretome proteins could cluster most human BRCA1- and BRCA2-related breast carcinomas at the transcriptome level. Topoisomerase I (TOP1) and P-cadherin (CDH3) expression was investigated by immunohistochemistry on tissue microarrays of a large panel of 253 human breast carcinomas with and without BRCA1/2 mutations. We showed that expression of TOP1 and CDH3 was significantly increased in human BRCA1-related breast carcinomas relative to sporadic cases (p = 0.002 and p < 0.001, respectively). Multiple logistic regression showed that TOP1 (adjusted odds ratio [OR] 3.75; 95% confidence interval [95% CI], 1.85 - 7.71, p < 0.001) as well as CDH3 positivity (adjusted OR 2.45; 95% CI, 1.08 - 5.49, p = 0.032) were associated with BRCA1/2-related breast carcinomas after adjustment for triple-negative phenotype and age. In conclusion, proteome profiling of secretome using murine breast tumor models is a powerful strategy to identify non-invasive candidate biomarkers of BRCA1-deficient breast cancer. We demonstrate that TOP1 and CDH3 are closely associated to BRCA1-deficient breast cancer. These data merit further investigation for early detection of tumors arising in BRCA1 mutation carriers.

Purpose Earlier, the association of single nucleotide polymorphisms (SNPs) with toxicity and efficacy of sunitinib has been explored in patients with metastatic renal cell carcinoma (mRCC). Recently, additional SNPs have been suggested as potential biomarkers. We investigated these novel SNPs for association with sunitinib treatment outcome in mRCC patients. Methods In this exploratory study, we selected SNPs in genes CYP3A4, NR1I2, POR, IL8, IL13, IL4-R, HIF1A and MET that might possibly be associated with sunitinib treatment outcome. Each SNP was tested for association with progression-free survival (PFS) and overall survival (OS) by Cox-regression analysis and for clinical response and toxicity using logistic regression. Results We included 374 patients for toxicity analyses, of which 38 patients with non-clear cell renal cell cancer were excluded from efficacy analyses. The risk for hypertension was increased in the presence of the T allele in IL8 rs1126647 (OR = 1.69, 95 % CI = 1.07–2.67, P = 0.024). The T allele in IL13 rs1800925 was associated with an increase in the risk of leukopenia (OR = 6.76, 95 % CI = 1.35–33.9, P = 0.020) and increased prevalence of any toxicity > grade 2 (OR = 1.75, 95 % CI = 1.06–2.88, P = 0.028). No significant associations were found with PFS, OS or clinical response. Conclusions We show that polymorphisms in IL8 rs1126647 and IL13 rs1800925 are associated with sunitinib-induced toxicities. Validation in an independent cohort is required. Electronic supplementary material The online version of this article (doi:10.1007/s00228-015-1935-7) contains supplementary material, which is available to authorized users.

Item does not contain fulltext Hypertension is an important side effect of sunitinib treatment. In a retrospective study in 255 patients, single-nucleotide polymorphisms (SNPs) in vascular endothelial growth factor A (VEGFA), vascular endothelial growth factor receptor (VEGFR)-2, endothelin-1 (ET-1), and endothelium-derived nitric oxide synthase (eNOS) were multivariately tested against hypertension grades and changes in systolic blood pressure (SBP), diastolic BP (DBP), and mean arterial BP (MAP). Next, the association between hypertension and survival in patients with metastatic renal cell cancer (mRCC) was studied. Greater elevations in SBP and MAP were associated with the presence of a haplotype in VEGFA (P = 0.014 and P = 0.036, respectively). The tendency to develop grade 3 hypertension was associated with this haplotype and also with a SNP in eNOS (P = 0.031 and P = 0.045, respectively). In mRCC patients, sunitinib-induced hypertension was found to confer a survival benefit, with the mean overall survival being prolonged by 7.2 months (P = 0.035 and P = 0.026 for SBP and DBP elevations, respectively). Genetic polymorphisms in VEGFA and eNOS independently predict rise in BP and/or development of severe hypertension in sunitinib-treated patients. Grade 3 hypertension was found to be an independent factor for overall survival in patients with mRCC.

BACKGROUND: As a prelude to combination studies aimed at resistance reversal, this dose-escalation/dose-expansion study investigated the selective Src kinase inhibitor saracatinib (AZD0530) in combination with carboplatin and/or paclitaxel.METHODS: Patients with advanced solid tumours received saracatinib once-daily oral tablets in combination with either carboplatin AUC 5 every 3 weeks (q3w), paclitaxel 175 mgm(-2) q3w, paclitaxel 80mgm(-2) every 1 week (q1w), or carboplatin AUC 5 plus paclitaxel 175mgm(-2) q3w. The primary endpoint was safety/tolerability.RESULTS: A total of 116 patients received saracatinib 125 (N - 20), 175 (N - 44), 225 (N - 40), 250 (N - 9), or 300mg (N - 3). There were no clear dose-related trends within each chemotherapy regimen group in number or severity of adverse events (AEs). However, combining all groups, the occurrence of grade >= 3 asthenic AEs (all causality) was dose-related (125 mg, 10%; 175 mg, 20%; >= 225 mg, 33%), and grade >= 3 neutropenia occurred more commonly at doses >= 225 mg. There was no evidence that saracatinib affected exposure to carboplatin or paclitaxel, or vice versa. Objective responses were seen in 5 out of 44 patients (11%) receiving carboplatin plus paclitaxel q3w, and 5 out of 24 (21%) receiving paclitaxel q1w.CONCLUSION: Saracatinib doses up to 175 mg with paclitaxel with/without carboplatin showed acceptable toxicity in most patients, and are suitable for further trials.British Journal of Cancer (2012) 106, 1728-1734. doi:10.1038/bjc.2012.158 www.bjcancer.com Published online 24 April 2012 (C) 2012 Cancer Research UK