Homologous Recombination Deficiency Across Subtypes of Primary Breast Cancer

Purpose - Homologous recombination deficiency (HRD) is highly prevalent in triple-negative breast cancer (TNBC) and associated with response to PARP inhibition (PARPi). Here, we studied the prevalence of HRD in non-TNBC to assess the potential for PARPi in a wider group of patients with breast cancer. Methods - HRD status was established using targeted gene panel sequencing (360 genes) and BRCA1 methylation analysis of pretreatment biopsies from 201 patients with primary breast cancer in the phase II PETREMAC trial (ClinicalTrials.gov identifier: NCT02624973). HRD was defined as mutations in BRCA1, BRCA2, BRIP1, BARD1, or PALB2 and/or promoter methylation of BRCA1 (strict definition; HRD-S). In secondary analyses, a wider definition (HRD-W) was used, examining mutations in 20 additional genes. Furthermore, tumor BRCAness (multiplex ligation-dependent probe amplification), PAM50 subtyping, RAD51 nuclear foci to test functional HRD, tumor-infiltrating lymphocyte (TIL), and PD-L1 analyses were performed. Results - HRD-S was present in 5% of non-TNBC cases (n = 9 of 169), contrasting 47% of the TNBC tumors (n = 15 of 32). HRD-W was observed in 23% of non-TNBC (n = 39 of 169) and 59% of TNBC cases (n = 19 of 32). Of 58 non-TNBC and 30 TNBC biopsies examined for RAD51 foci, 4 of 4 (100%) non-TNBC and 13 of 14 (93%) TNBC cases classified as HRD-S had RAD51 low scores. In contrast, 4 of 17 (24%) non-TNBC and 15 of 19 (79%) TNBC biopsies classified as HRD-W exhibited RAD51 low scores. Of nine non-TNBC tumors with HRD-S status, only one had a basal-like PAM50 signature. There was a high concordance between HRD-S and either BRCAness, high TIL density, or high PD-L1 expression (each P Conclusion - The prevalence of HRD in non-TNBC suggests that therapy targeting HRD should be evaluated in a wider breast cancer patient population. Strict HRD criteria should be implemented to increase diagnostic precision with respect to functional HRD

Impact of snus use in teenage boys on tobacco use in young adulthood; a cohort from the HUNT Study Norway

Background As smoking rates decreased, the use of Swedish snus (smokeless tobacco) concordantly increased in Norway. The role of snus as possible contributor to the reduction of smoking has been widely discussed. Our aim was to quantitate transitions in snus use, smoking and dual use of snus and cigarettes in a young male population. Methods This prospective cohort study includes 1346 boys participating in the Nord-Trøndelag Health Study in Young-HUNT1 1995–97, age 13–19 and in HUNT3 2006–08, age 23–30. Participants reported on tobacco use at both points of time. Models with binominal regression were applied to examine relative risks (RRs), of adolescent ever snus users, dual users or smokers (reference: never tobacco use), to be current snus only users, smokers (including dual users), or tobacco free in adulthood. Results Current tobacco use in this male cohort increased from 27% in adolescence to 49% in adulthood, increasing more for snus only use and dual use than for smoking only. The adjusted RR (95% CI) of becoming a smoker as young adult, was 2.2 (CI 1.7–2.7) for adolescent snus users, 3.6 (CI 3.0–4.3) for adolescent dual users, and 2.7 (CI 2.2–3.3) for adolescent smokers. RR to become snus only users as adults was 3.1 (2.5–3.9) for adolescent dual users, 2.8 (2.2–3.4) for adolescent snus users and 1.5 (1.0–2.2) for adolescent smokers. The adjusted RR for the transition from adolescent tobacco use to no tobacco use in adulthood was similar for snus users and smokers with RR 0.5 (CI 0.4–0.7), but considerably lower for dual users with RR 0.2 (CI 0.2–0.3). Conclusions The use of snus, with or without concurrent smoking, carried a high risk of adult smoking as well as adult snus only use. Dual use seemed to promote the opportunity to become snus only users in adulthood, but made it also more difficult to quit. The benefit of snus use for harm reduction is not evident in our cohort, as the combination of smoking and dual use resulted in high smoking rates among the young adults

Impact of snus use in teenage boys on tobacco use in young adulthood; a cohort from the HUNT Study Norway

Background As smoking rates decreased, the use of Swedish snus (smokeless tobacco) concordantly increased in Norway. The role of snus as possible contributor to the reduction of smoking has been widely discussed. Our aim was to quantitate transitions in snus use, smoking and dual use of snus and cigarettes in a young male population. Methods This prospective cohort study includes 1346 boys participating in the Nord-Trøndelag Health Study in Young-HUNT1 1995–97, age 13–19 and in HUNT3 2006–08, age 23–30. Participants reported on tobacco use at both points of time. Models with binominal regression were applied to examine relative risks (RRs), of adolescent ever snus users, dual users or smokers (reference: never tobacco use), to be current snus only users, smokers (including dual users), or tobacco free in adulthood. Results Current tobacco use in this male cohort increased from 27% in adolescence to 49% in adulthood, increasing more for snus only use and dual use than for smoking only. The adjusted RR (95% CI) of becoming a smoker as young adult, was 2.2 (CI 1.7–2.7) for adolescent snus users, 3.6 (CI 3.0–4.3) for adolescent dual users, and 2.7 (CI 2.2–3.3) for adolescent smokers. RR to become snus only users as adults was 3.1 (2.5–3.9) for adolescent dual users, 2.8 (2.2–3.4) for adolescent snus users and 1.5 (1.0–2.2) for adolescent smokers. The adjusted RR for the transition from adolescent tobacco use to no tobacco use in adulthood was similar for snus users and smokers with RR 0.5 (CI 0.4–0.7), but considerably lower for dual users with RR 0.2 (CI 0.2–0.3). Conclusions The use of snus, with or without concurrent smoking, carried a high risk of adult smoking as well as adult snus only use. Dual use seemed to promote the opportunity to become snus only users in adulthood, but made it also more difficult to quit. The benefit of snus use for harm reduction is not evident in our cohort, as the combination of smoking and dual use resulted in high smoking rates among the young adults

Toll-like receptor profiling of seven trophoblast cell lines warrants caution for translation to primary trophoblasts

Introduction Excessive placental inflammation is associated with pregnancy complications. Toll-like receptors (TLRs) are sensors for danger signals from infections and damaged tissue and initiate inflammation. Trophoblasts in the placenta broadly express TLRs. Trophoblast cell lines are used as surrogates for primary trophoblasts for in vitro studies, but the inflammatory translatability of trophoblast cell lines warrants examination. We aimed to assess TLR1-10 gene expression and activation in seven trophoblast cell lines and compare this to primary trophoblasts. Methods The five choriocarcinoma trophoblast cell lines BeWo, JAR, JEG-3, AC1M-32 and ACH-3P, and the two SV40 transfected trophoblast cell lines HTR-8/SVneo and SGHPL-5 were included and compared to primary first trimester trophoblasts (n = 6). TLR1-10 gene expression was analyzed by RT-qPCR. Cells were stimulated by specific TLR1-9 ligands for 24 h and cytokine release was measured by a 10-plex immunoassay. Results All choriocarcinoma cell lines demonstrated broad TLR gene expression, but lacked functional cytokine response to TLR ligand activation. In contrast, SV40 transfected cell lines showed restricted TLR gene expression, but SGHPL-5 cells displayed significantly increased levels of interleukin (IL)-6, IL-8, IL-12 and vascular endothelial growth factor A after TLR3 and/or TLR4 activation (P < 0.01), while TLR2 activation increased IL-6 and IL-8 levels (P < 0.05). HTR8/SVneo cells responded to TLR3 activation by increased IL-6 and interferon (IFN)-γ (P < 0.05). The SGHPL-5 TLR profile most closely resembled primary trophoblast. Discussion The characterized trophoblast cell line TLR profiles serve as a reference and warrant caution when selecting trophoblast cell lines as in vitro models for immune responses in primary trophoblasts

Clonal evolution in primary breast cancers under sequential epirubicin and docetaxel monotherapy

Background Subclonal evolution during primary breast cancer treatment is largely unexplored. We aimed to assess the dynamic changes in subclonal composition of treatment-naïve breast cancers during neoadjuvant chemotherapy. Methods We performed whole exome sequencing of tumor biopsies collected before, at therapy switch, and after treatment with sequential epirubicin and docetaxel monotherapy in 51 out of 109 patients with primary breast cancer, who were included in a prospectively registered, neoadjuvant single-arm phase II trial. Results There was a profound and differential redistribution of subclones during epirubicin and docetaxel treatment, regardless of therapy response. While truncal mutations and main subclones persisted, smaller subclones frequently appeared or disappeared. Reassessment of raw data, beyond formal mutation calling, indicated that the majority of subclones seemingly appearing during treatment were in fact present in pretreatment breast cancers, below conventional detection limits. Likewise, subclones which seemingly disappeared were still present, below detection limits, in most cases where tumor tissue remained. Tumor mutational burden (TMB) dropped during neoadjuvant therapy, and copy number analysis demonstrated specific genomic regions to be systematically lost or gained for each of the two chemotherapeutics. Conclusions Sequential epirubicin and docetaxel monotherapy caused profound redistribution of smaller subclones in primary breast cancer, while early truncal mutations and major subclones generally persisted through treatment. Trial registration ClinicalTrials.gov, NCT00496795 , registered on July 4, 2007

DHA induces ER stress and growth arrest in human colon cancer cells: associations with cholesterol and calcium homeostasis *s⃞

Polyunsaturated fatty acids (PUFAs) are normal constituents of the diet, but have properties different from other fatty acids (e.g., through generation of signaling molecules). N-3 PUFAs reduce cancer cell growth, but no unified mechanism has been identified. We show that docosahexaenoic acid (DHA; 22:6 n-3) causes extensive changes in gene expression patterns at mRNA level in the colon cancer cell line SW620. Early changes include unfolded protein response (UPR) and increased levels of phosphorylated eIF2α as verified at protein level. The latter is considered a hallmark of endoplasmic reticulum (ER) stress and is abundantly present already after 3 h. It may coordinate many of the downstream changes observed, including signaling pathways for cell cycle arrest/apoptosis, calcium homeostasis, cholesterol metabolism, ubiquitination, and proteasomal degradation. Also, eicosapentaenoic acid (EPA), but not oleic acid (OA), induced key mediators of ER stress and UPR at protein level. Accumulation of esterified cholesterol was not compensated for by increased total levels of cholesterol, and mRNAs for cholesterol biosynthesis as well as de novo synthesis of cholesterol were reduced. These results suggest that cytotoxic effects of DHA are associated with signaling pathways involving lipid metabolism and ER stress