Characterization of real-world treatment practices and outcomes among patients with chronic lymphocytic leukemia treated in a Finnish tertiary center.

Objectives: We conducted this retrospective study to characterize the change in chronic lymphocytic leukemia (CLL) treatment patterns between 2005 and 2019, to understand the treatment sequencing across the course of the disease, and to investigate how targeted agents and prognostic testing were implemented into the patient care. Methods: This study included adult patients with CLL treated at the Hospital District of Southwest Finland during the study period. Data were collected from the Turku University Hospital data lake. Results: In total, 122 and 60 patients received first- and second-line treatments for CLL, respectively. The shift from conventional chemoimmunotherapy to targeted treatments in recent years (2014-2019) was observed. The median overall survival times were not reached in patients treated with targeted agents compared to conventional standard treatments in first- and second-line settings and improved toward the end of the study period. Prognostic testing increased during the study follow-up and patients with unmutated immunoglobulin heavy-chain variable showed significantly poorer overall survival and time-to-next-treatment outcomes than patients with mutated immunoglobulin heavy-chain variable. Conclusions: This real-world study implicated added value of targeted chemo-free therapies as reported in randomized clinical trials, and highlighted the necessity of prognostic testing in order to improve treatment selection and patient outcomes.

Methyltransferase expression and tumor suppressor gene methylation in sporadic and familial colorectal cancer.

Molecular mechanisms underlying coordinated hypermethylation of multiple CpG islands in cancer remain unclear and studies of methyltransferase enzymes have arrived at conflicting results. We focused on DNMT1 and DNMT3B, DNA methyltransferases responsible for (de novo) methylation, and EZH2, histone (H3K27) methyltransferase, and examined their roles in tumor suppressor gene (TSG) methylation patterns we have previously established in sporadic and familial cancers. Our investigation comprised 165 tumors, stratified by tissue of origin (117 colorectal and 48 endometrial carcinomas) and sporadic vs. familial disease (57 sporadic vs. 60 familial, mainly Lynch syndrome, colorectal carcinomas). By immunohistochemical evaluation, EZH2 protein expression was associated with a TSG methylator phenotype. DNMT1, DNMT3B, and EZH2 were expressed at significantly higher levels in tumor vs. normal tissues. DNMT1 and EZH2 expression were positively correlated and higher in microsatellite-unstable vs. microsatellite-stable tumors, whether sporadic or hereditary. Ki-67 expression mirrored the same pattern. Promoter methylation of the methyltransferase genes themselves was addressed as a possible cause behind their altered expression. While DNMT1 or EZH2 did not show differential methylation between normal and tumor tissues, DNMT3B analysis corroborated the regulatory role of a distal promoter region. Our study shows that methyltransferase expression in cancer depends on the tissue of origin, microsatellite-instability status, cellular proliferation, and--in the case of DNMT3B--promoter methylation of the respective gene. Translation of methyltransferase expression into DNA methylation appears complex as suggested by the fact that except for EZH2, no clear association between methyltransferase protein expression and TSG methylation was observed.

LINE-1 hypomethylation in familial and sporadic cancer.

Increased and decreased methylation at specific sequences (hypermethylation and hypomethylation, respectively) is characteristic of tumor DNA compared to normal DNA and promotes carcinogenesis in multiple ways including genomic instability. Long interspersed element (LINE), an abundant class of retrotransposons, provides a surrogate marker for global hypomethylation. We developed methylation-specific multiplex ligation-dependent probe amplification assays to study LINE-1 methylation in cases of colorectal, gastric, and endometrial cancer (N = 276), stratified by patient category [sporadic; Lynch syndrome (LS); familial colorectal cancer type X (FCCX)] and microsatellite instability status. Within each patient group, LINE-1 showed lower methylation in tumor DNA relative to paired normal DNA and hypomethylation was statistically significant in most cases. Interestingly, normal colorectal mucosa samples from different patient groups displayed differences in LINE-1 methylation that mirrored differences between the respective tumor tissues, with a decreasing trend for LINE-1 methylation from patients with sporadic colorectal cancer to LS to FCCX. Despite the fact that the degree of LINE-1 methylation is generally tissue specific, normal colorectal mucosa, gastric mucosa, and endometrium from LS patients showed similar levels of LINE-1 methylation. Our results suggest that the degree of LINE-1 methylation may constitute a "field defect" that may predispose normal tissues for cancer development.

Epigenetic signatures of familial cancer are characteristic of tumor type and family category.

Tumor suppressor genes (TSG) may be inactivated by methylation of critical CpG sites in their promoter regions, providing targets for early detection and prevention. Although sporadic cancers, especially colorectal carcinoma (CRC), have been characterized for epigenetic changes extensively, such information in familial/hereditary cancer is limited. We studied 108 CRCs and 63 endometrial carcinomas (EC) occurring as part of hereditary nonpolyposis CRC, as separate familial site-specific entities or sporadically, for promoter methylation of 24 TSGs. Eleven genes in CRC and 6 in EC were methylated in at least 15% of tumors and together accounted for 89% and 82% of promoter methylation events in CRC and EC, respectively. Some genes (e.g., CDH13, APC, GSTP1, and TIMP3) showed frequent methylation in both cancers, whereas promoter methylation of ESR1, CHFR, and RARB was typical of CRC and that of RASSF1(A) characterized EC. Among CRCs, sets of genes with methylation characteristic of familial versus sporadic tumors appeared. A TSG methylator phenotype (methylation of at least 5 of 24 genes) occurred in 37% of CRC and 18% of EC (P = 0.013), and the presence versus absence of MLH1 methylation divided the tumors into high versus low methylation groups. In conclusion, inactivation of TSGs by promoter methylation followed patterns characteristic of tumor type (CRC versus EC) and family category and was strongly influenced by MLH1 promoter methylation status in all categories. Paired normal tissues or blood displayed negligible methylation arguing against a constitutional methylation abnormality in familial cases.