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The definition of the gene expression profile of genes encoding Ion Channels and Transporters (ICT-GEP) represents a novel and attracting aspect in cancer. We determined the ICT-GEP of Follicular Lymphoma (FL), and compared it with that of the more aggressive Diffuse Large B Cell Lymphoma (DLBCL). cDNA microarray data were collected both from patients enrolled for this study, and from public datasets. In FL the ICT-GEP indicated the overexpression of both the K+ channel encoding gene KCNN4, and SLC2A1, which encodes the Glut1 glucose transporter. SLC2A1 turned out to represent the hub of a functional network, connecting channels and transporters in FL. Relapsed FL patients were characterised by 38 differentially expressed ICT genes, among which ATP9A, SLC2A1 and KCNN4 were under-expressed, indicating a down-regulation of both excitability and glycolysis. A completely different profile of K+ channel encoding genes emerged in DLBCL accompanied by the over-expression of the fatty acid transporter-encoding gene SLC27A1 as well as of the metabolism regulator NCoR1. This indicates a change in excitability and a shift towards an oxidative metabolism in DLBCL. Overall, the ICT-GEP may contribute to identifying novel lymphoma biomarkers related to excitability and metabolic pathways, with particular relevance for drug resistant, relapsed FL.

Summary Relapsed/refractory myeloma has a poor outcome because of multi-drug resistance, patient low-performance status and toxicity of conventional chemotherapy. To improve results, standard chemotherapeutics and drugs targeting the microenvironment are applied at the same time. Bortezomib, by inhibiting proteasome function, may enhance chemosensitivity to other drugs and overcome drug-resistance. Notably, doxorubicin and bortezomib may reciprocally increase their efficacy. Thus, to improve outcome whilst minimizing therapy-related toxicity, liposomal doxorubicin was added to a bortezomib-based combination. From January 2004, relapsed/refractory myeloma patients referred to our Institution received bortezomib 1·0 mg/m2 i.v. twice weekly for 2 weeks in a 28-d cycle for up to six cycles, oral dexamethasone 24 mg with the standard scheduling and thalidomide 100 mg continuously (VTD). From January 2005, liposomal doxorubicin, 50 mg/m2 (30 mg/m2 for patients older than 75 years), was added on day 4 of each cycle [VTD plus Myocet (MyVTD)]. In total, 70 patients were treated: 28 received VTD and 42 MyVTD. Baseline demographic and clinical characteristics were similar between the two groups. Toxicity was manageable although more pronounced with MyVTD. The overall response rate (81% vs. 50%, P = 0·009), time to progression (19 vs. 11 months, P = 0·01) and progression-free survival (15 vs. 8 months, P = 0·001) were significantly higher with MyVTD regimen, suggesting an improved quality of response.

Introduction: Standard chemotherapy represented by the R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, prednisone) regimen is successful in about 60% of patients (pts) with diffuse large B-cell lymphoma (DLBCL). Pts who do not benefit from this treatment, due to the development of tumor drug resistance, have a very poor prognosis. Currently, knowledge on reasons of treatment related failures in DLBCL are scanty and predictive biomarker of response are largely unknown. We hypothesized that polymorphisms of gene involved in the pharmacokinetics and pharmacodynamics of drugs included in R-CHOP regimen may play a role in predicting the outcome in DLBCL pts.Thus, we designed a multicentre prospective pharmacogenetic trial aimed at identifying gene polymorphisms potentially predictive of drug efficacy/resistance in DLBCL pts treated with R-CHOP. An interim analysis on the first 80 enrolled ptswas planned and has been performed. Methods: The study included chemonaive DLBCL pts at various stages of disease candidate to an R-CHOP standard treatment. The Ethical Committee of each participating centre approved the pharmacogenetic protocol, and all pts signed a written informed consent. According to the aims of this interim analysis, the impact of single nucleotide polymorphisms (SNPs) on R-CHOP efficacy was evaluated by objective response (OR) rate at the end of treatment. The efficacy of R-CHOP was evaluated according to the Cheson criteria by performing standard hematochemical and instrumental (TC and FDFG-PET) tests and defining complete remission (CR), partial remission (PR), non response or progressive disease (PD). Genomic DNA wasextracted from peripheral blood of 80 pts. Twentysingle nucleotide polymorphisms (SNPs) from18candidate genes (ABCB1, ABCC1, ABCC2, ABCG2, CYBA, CYP2C9, FCGR2A, GSTP1, IL2, MLH1, NCF4, NQO1, NQO2, RAC2, TNF, TOP2A, TP53, TUBB)involved in pharmacokinetics and pharmacodynamics of R-CHOP (www.pharmgkb.org) have been analysed by a genotyping array based on Affimetrix methodology. Univariate analysis was performed to evaluate associations between polymorphisms and clinical/pathological characteristics or OR (Fisher exact test). Multivariate logistic regression analysis was performed to estimate adjusted odds ratios along with the corresponding 95% confidence intervals for the polymorphisms and OR. Results: Median age was 63 years. There were 37 men and 43 women. 47.5 % of pts were in stage I-II,52.5 % of pts in stage III-IV. 27.5% of ptshad bulky disease, 43.8 % of pts had involvement of extranodal site. 47.5% of pts had pathological LDH value. According to the revised IPI, 15 % pts were in the low risk group, 58.7 % in the intermediate risk group, and 26.3 % in the high risk group.Overall, 468 courses of R-CHOP had been administered (mean: 5.85 courses, range: 4-6). 81% of pts had CR to R-CHOP whereas the remaining showed PR (14%) or PD(5%). No statistically significant correlation was found between OR and clinical characteristics of pts.However, stage III-IV pts showed a worst OR than stage I-II pts (77% vs 87% of CR, respectively); pts with bulky disease had worst OR than non-bulky disease pts(73% vs 84.5% of CR, respectively); ptswith R-IPI 3-5 a worst OR than pts with R-IPI 0-2 (71.5% vs 85% of CR, respectively). Univariate and multivariateanalysis identified TOPOII rs13695as a predictor of OR (p=0.042). Pts with CT or TT genotypesshowed worst OR than CC wild-type homozygous pts (odds ratio 3.070, CI95% 1.113-13.457). Also, a statistical trend toward significance was observed for MLH1 rs1800734 polymorphism (p=0.062): ptswith homozygous genotype for the mutant allele showed a better OR than wild-type and heterozygous pt genotypes. Conclusions: No significant relationship between clinical/pathological characteristics and OR was observed. Our preliminary data show that SNPs affecting a gene involved in doxorubicin pharmacodynamics, i.e. the drug target TOPOII, as well asone of the major components of DNA mismatch repair, i.e. MLH1 gene,may predict response in DLBCL pts treated with R-CHOP. These preliminary results from the interim analysis are promising and warrant completion of pt accrual to reach the planned number of cases at the end of our study. Acknowledgments This work was supported by a grant from the Associazione Giacomo Onlus, Castiglioncello (LI), Italy to E.M. and Cassa di Risparmio di Firenze, Firenze, Italy to S.N. Disclosures No relevant conflicts of interest to declare.

Bortezomib (Velcade®) is effective in MM as single agent. A strong rationale supports the association of bortezomib with anthracyclines. Bortezomib may increase the activity of anthracyclines by inhibiting maturation of P-gp, suppressing DNA repair and inducing phosphorylation and cleavage of Bcl-2. Anthracyclines down-regulate the bortezomib anti-apoptotic effect. In vitro studies demonstrated that liposome encapsulated doxorubicin can overcome MDR-1 overexpression. It has a longer half-life, lower cardiac toxicity and comparable efficacy. Thus, we added non-pegylate liposomal doxorubicin (Myocet®) to the low dose VTD regimen (Ciolli et al. Leukemia & Lymphoma2006;47(1): 171–173). Aims of the study were to evaluate the feasibility of treatment and stem cells harvest and transplantation, to compare ORR and TTP in respect to LD-VTD. The study was performed in accordance with the Declaration of Helsinki. Eligibility criteria: pts primary refractory (PR) or relapsed and refractory (R/R), with a LVEF more than 45% and a measurable disease. Therapy: bortezomib 1.0 mg/m2 i.v. bolus days 1, 4, 8 and 11 of a 28-d cycle, Myocet 50 mg/m2 i.v over 60 minutes on day 4 (1 hour after bortezomib), oral dexamethasone 24 mg on the day of and the day following each bortezomib dose. Thalidomide 100 mg/d if non controindicated. All toxicities were defined according to NCI criteria. Response was evaluated after each cycle. Pts with progressive disease (PD) were removed from the study, the others continued until best response for a maximum of 4 cycles. Time to response was from the date of the first administration of bortezomib to the first evidence of response. From June 2005, 28 pts, median age 68 years, entered the study: 5 stage IIA, 20 IIIA and 3 IIIB. 16 (57%) were the PR pts and 12(43%) those R/R. 8(29%) had a β2microglobulin >4 mg/L, 7 a PS >2 and 8 a pre-existing peripheral neuropathy (PN) grade 2. Median time from diagnosis was 4 years and a median of 4 (range 2–6) were the prior therapy lines. All pts had previously received thalidomide plus dexamethasone; 8 had previously received bortezomib. 13 (46%) did not receive thalidomide due to a pre-existing neurological toxicity. Haematological toxicity was registered 14–17 days from the start of therapy and lasted for a maximum of 4 days: grade 2 neutropenia in 3 pts, grade 4 thrombocytopenia in 5. A patient had a grade 4 gastric haemorrhage. 3 pts had pneumonia, 2 HZ infection and 1 a DVT. Other adverse advents of grade >1 were fatigue (50%), nausea (60%), diarrhoea (15%), alopecia (10%). None progression of PN was observed. No patient experienced clinical evidence of cardiac toxicity. At July 31th 2006,. all pts were valuable for response: 3 PD, 4 SD, 21 responders: 8 CR, 2 nCR, 7 PR,4 MR (ORR 75%). Median time to best response was 1 month (range 1– 2). 3 pts successfully harvested the PBSC and 1 was transplanted. After a median follow-up of 9 months, median TTP and OS had not been reached, 23 pts were alive and 5 (2 PD and 3 responders) had died. Myocet did not add toxicity to LD-VTD. PBSC has been successfully performed in eligible pts. More pts and an adequate follow-up are needed to draw any definitive conclusion about TTP and OS, however, this regimen appears feasible and effective with an increased ORR compared to our previous LD-VTD experience (75% vs 53%).