Germline Genetic Factors Affecting Interferon Alpha Therapy in Polycythemia Vera

Introduction: Interferon alpha (IFNa) based therapies can induce sustained hematological and molecular responses (HR/MR) in Polycythemia Vera (PV) and other Myeloproliferative Neoplasms (MPN), however, not all patients respond sufficiently. While previous studies suggest that disease driving somatic mutations and genomic aberrations do not predict response to IFNa, the role of common germline polymorphisms remains elusive. We addressed the effect of germline genetic factors on PV therapy with Ropeginterferon alfa-2b (Ropeg), a novel monopegylated IFNa. We performed genome-wide association studies (GWAS) as an unbiased approach, and additionally evaluated a previously reported influence of polymorphisms at the IFNL4 locus in a large PV patient cohort. Methods: Genomic DNA was isolated from whole blood of PV patients (n=115) on Ropeg therapy, who provided consent within the PROUD-PV (NCT01949805) and CONTI-PV (NCT02218047) study programs. Patients were genotyped for >900k tagging single nucleotide polymorphisms (SNPs) across the genome on the Affymetrix SNP 6.0 array platform. Additional IFNL4 SNPs were typed using an amplicon-based next generation sequencing approach. Association analyses were performed using the PLINK toolset. JAK2-V617F mutant allele burden was quantified using a qPCR-based assay (ipsogen MutaQuant, Qiagen). MR was measured based on changes in mutant allele burden upon therapy as defined by the European LeukemiaNet (ELN) criteria. Results: To test for potential associations between germline genetic variation and response to IFNa therapy, we performed GWAS for MR data upon 12 months (M), 18M, 24M and 30M and 36M follow-up on Ropeg treatment. Genome-wide tagging SNPs were tested both for association in a case-control setup applying chi-square statistics as well as for quantitative trait association, using peripheral blood JAK2-V617F mutational burden changes from baseline as continuous variables. After Bonferroni correction for multiple testing, none of those analyses revealed a statistically significant association, suggesting the absence of strong germline predisposition factors for MR. Germline variation at the interferon-lambda (IFNL) locus was previously reported to strongly influence viral clearance during IFNa therapy of Hepatitis C (reviewed in Wack et al., Nat Immunol, 2015). While the same variants were recently reported to also affect HR during IFNa treatment in an MPN patient cohort (Lindgren et al., EJH, 2018), their potential impact on MR as surrogate marker for the size of the malignant MPN clone has not yet been evaluated. Upon testing for MR in our PV cohort, in a case-control setup we observed a statistically significant association only upon 36M follow-up (p=0.02; OR=2.51; 95%CI=[1.14-5.62]; Figure 1a). Notably, testing for change of JAK2-V617F mutational burden under Ropeg treatment as quantitative trait, we found the association to be present at formal statistical significance at all stages during follow-up (Figure 1a). For the Hepatitis C association, it is now widely acknowledged that a diplotype of two coding variants covers most of the causality, where rs368234815_TT disrupts the open reading frame (no IFNL4) while rs117648444_G results in the impaired IFNL4-S70 in contrast to the fully functional IFNL4-P70, the latter paradoxically impacting negatively on viral clearance (Terczynska-Dyla et al., Nat Commun, 2014). Similarly, in our PV cohort genotyping and phasing of this diplotype revealed a significant influence on MR in carriers of the fully functional IFNL4-P70 (p=0.01; Figure 1b; shown are patients at 36M follow-up (n=70)). Conclusions: The absence of a strong germline predisposition factor for Ropeg treatment response in our cohort implies that any PV patient may be eligible for Ropeg therapy independently of their genetic makeup. While the IFNL4 diplotype is of potential use for patient stratification, it remains to be investigated whether increase of treatment duration and/or dose adjustments can overcome the adverse effect of functional IFNL4 on Ropeg therapy. Longitudinal monitoring of JAK2-V617F mutational burden under Ropeg treatment in conjunction with determination of the IFNL4 germline genetic status may allow for optimizing patient management. Disclosures Krejcy: AOP Orphan Pharmaceuticals AG: Employment. Klade:AOP Orphan Pharmaceuticals AG: Employment. Zoerer:AOP Orphan Pharmaceuticals AG: Employment. Gisslinger:Janssen-Cilag: Honoraria; Roche Austria GmbH: Consultancy; Novartis Pharma GmbH: Consultancy, Honoraria, Research Funding; Myelopro GmbH: Consultancy; Celgene GmbH: Honoraria; Pharma Essentia: Other: Personal fees; AOP Orphan Pharmaceuticals: Consultancy, Honoraria, Research Funding. Kralovics:Novartis: Honoraria; AOP Orphan Pharmaceuticals AG: Honoraria, Other: Advisory board; Pharma Essentia: Honoraria; Qiagen: Honoraria; MyeloPro Diagnostics and Research: Equity Ownership.

Identifying Cytokine Biomarkers of Response to Pegylated-Interferon Therapy in Polycythemia Vera and Essential Thrombocythemia: Correlative Analysis from the MPN-RC 111/112 Trials

Background: Pegylated Interferon-alpha (PEG-IFNa) improves hematologic parameters and reduces mutant allele fraction in patients with myeloproliferative neoplasms (MPNs); however, widespread clinical use is limited by frequent discontinuation due to side effects. Identifying biomarkers of response to PEG-IFNa therapy might identify patient subsets most likely to derive benefit from this therapy. Serum pro-inflammatory cytokine levels are increased in MPN patients and associated with adverse clinical outcomes. The effect of PEG-IFNa therapy on cytokine levels in MPN patients and correlation with clinical response has not been elucidated. We hypothesized that identification of cytokine signatures could predict for clinical/molecular response to PEG-IFNa in MPNs. Methods: Pre- and post-treatment serum samples were collected from available patients enrolled on the Myeloproliferative Disorders Research Consortium (MPD-RC) 111/112 trials-multicenter studies assessing efficacy of PEG-IFNa in patients with high-risk polycythemia vera/essential thrombocythemia either up front or in the setting of hydroxyurea failure/intolerance. Serum was prepared using the Millipore MAP Human 41-Cytokine Magnet Bead System (Millipore; Cat:HCYTOMAG-60K) and run on the Luminex FLEXMAP 3D platform. Data was analyzed using Luminex xPonent 4.3 software. Correlative mutational data on 156 genes implicated in myeloid malignancies was also available for all patients. Age-matched serum from healthy individuals (N=6) was obtained from AllCellsTM for use as wild-type (WT) controls. Baseline cytokine samples were compared between groups of interest by use of Wilcoxon rank-sum test. Change from baseline in cytokines for each group were compared to the null hypothesis of zero change. P values <0.05 were considered statistically significant. Results: Pre- and post-treatment serum samples were available on 29 MPN patients from the MPN-RC 111/112 cohorts. 21 patients were classified as complete responders (CRs) vs. 8 non-responders (NRs) using European LeukemiaNet criteria. Baseline cytokine levels were generally increased in both CRs and NRs compared to WT, including significant increases in Eotaxin (p=0.023), IL3 (p=0.048), IP10 (p=0.04), and TNFa (p=0.006) (Table 1), as well as a non-significant trend increase in IL8 (p=0.076). Notably, other cytokines were decreased at baseline in comparison to WT, including EGF (p<0.001), as well as MDC (p=0.040) and PDGFaabb (p=0.02), in line with previous studies. Compared to CRs, NRs displayed significantly lower levels of GRO (p=0.020), MDC (p=0.003), and PDGFaabb (p=0.026) at baseline along with elevated levels of IP10 (p=0.028) suggesting pre-treatment levels of these cytokines might predict response to therapy. Comparison of pre- and post- treatment samples revealed a general increase in median cytokine levels in CRs, with significant changes in GM-CSF (p=0.012), IP10 (p<0.001), MCP1 (p=0.015) and IFNa itself (p=0.010). Notably, median cytokine levels in NRs post-treatment, including IFNa, were relatively unchanged from baseline despite on-going PEG-IFNa therapy. Additional pre-treatment serum samples, as well as correlative analysis of cytokine changes with JAK-STAT driver mutant allele fraction and mutational profiling are underway and will be presented at the meeting. Conclusions: Preliminary findings suggest individual cytokines are affected by PEG-IFNa in MPNs. Furthermore, baseline levels of GRO, MDC, PDGFaabb, and IP10 correlate with response to PEG-IFNa. These data may identify MPN patients most likely to respond to PEG-IFNa therapy and those for whom this therapy may be most appropriate. Further analysis with a larger cohort of treatment-naïve patients is underway to validate these findings and identify additional molecular and cytokine signatures of response to PEG-IFNa therapy. Disclosures Yacoub: Agios: Speakers Bureau; Hylapharm: Equity Ownership; Dynavax: Equity Ownership; Cara: Equity Ownership; Ardelyx: Equity Ownership; Incyte: Consultancy, Honoraria, Speakers Bureau; Seattle Genetics: Honoraria, Speakers Bureau; Novartis: Consultancy, Speakers Bureau. Mascarenhas:Incyte: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Research Funding; Roche: Consultancy, Research Funding; Merck: Research Funding; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; CTI Biopharma: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Research Funding; Promedior: Research Funding; Merus: Research Funding; Pharmaessentia: Consultancy, Membership on an entity's Board of Directors or advisory committees. Levine:Lilly: Honoraria; Qiagen: Membership on an entity's Board of Directors or advisory committees; Imago Biosciences: Membership on an entity's Board of Directors or advisory committees; Loxo: Membership on an entity's Board of Directors or advisory committees; Prelude Therapeutics: Research Funding; Isoplexis: Membership on an entity's Board of Directors or advisory committees; Roche: Consultancy, Research Funding; Gilead: Consultancy; Amgen: Honoraria; C4 Therapeutics: Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Research Funding; Novartis: Consultancy. Hoffman:Merus: Research Funding. Rampal:Constellation, Incyte, and Stemline Therapeutics: Research Funding; Agios, Apexx, Blueprint Medicines, Celgene, Constellation, and Jazz: Consultancy.

Generalized Molluscum Contagiosum Successfully Treated with Interferon-Alpha in a Patient with Folliculotropic Mycosis Fungoides

We present the case of a 50-year-old patient with folliculotropic mycosis fungoides (FMF) unresponsive to retinoids and the chemotherapeutic regimens CHOP, gemcitabine, and brentuximab-vedotin. During immunosuppressive therapy, the patient developed extensive progressive molluscum contagiosum. The mollusca did not respond to topical imiquimod but showed a swift complete response to interferon-alpha 2a (IFNa). Recently, the patient started with alemtuzumab as induction therapy for an allogenic stem cell transplantation and simultaneously continued IFNa therapy.

Impact of the Molecular Profile of Malignant Clones on the Response to Interferon Alpha (IFNa) Therapy in JAK2V617F-Negative Essential Thrombocythemia (ET)

Background: The majority of ET patients (pts) without JAK2 or MPL mutations present somatic mutations in the calreticulin gene (CALR). However, a series of mutations in other genes involved in the epigenome, the splicing machinery or leukemic transformation have been described in ET, but contrary to MF, their impact has not been clearly assessed. We have previously shown that IFNa may considerably reduce the JAK2-mutated clones, and we also observed that TET2 mutated clones are resistant to IFNa therapy in JAK2+TET2+ pts with polycythemia vera. As little is known about the IFNa response of clones harboring mutations other than JAK2V617F in ET, we took advantage of a cohort of pts without JAK2 mutation but CALRmutated and treated with IFNa (according to international and local guidelines) to assess the dynamics of the different MPN clones during therapy. Aims: 1) To determine, using a Next Generation Sequencing (NGS) approach, the molecular pattern of mutations in genes previously shown to have a prognostic impact in MPNs in a series of CALR-mutated ET pts; 2) To study the evolution of these mutational patterns during IFNa therapy. Methods: JAK2 and MPL-negative ET pts followed in our department fulfilling the following criteria were included in the study: presence of a CALR mutation; availability of at least 3 sequential blood samples including one taken before IFNa; IFNa therapy for at least 3 months; informed consent for molecular analysis. Total DNA was extracted from blood samples (Qiagen blood DNA mini kits) for molecular analyses. Mutations in TET2, ASXL1, EZH2, SRSF2, IDH1, IDH2, SH2B3 and CSF3R were searched through a NGS approach on a MiSeq instrument using a TruSeq custom amplicon approach (Illumina). CALRgene mutation detection was done by direct Sanger sequencing of exon 9, and mutant allele burden (%CALR) was estimated by fragment analysis (GeneMapper software , Life technologies) with a sensitivity of about 1%. Both sequencing and fragment analysis were performed on a 3500xL DX Genetic Analyser (Life technologies). Molecular response was defined as complete (CMR) when CALR mutation was no longer detectable, partial (PMR) when %CALR was decreased by >50%, minor (mMR) when %CALR was decreased by 25 to 49%, and non responders if %CALR was reduced by less than 25%. Results: Among 238 ET pts without JAK2 or MPL mutations, we identified 52 pts (22%) treated with peg-IFNa-2a, of whom 24 fulfilled the inclusion criteria. Median age was 52 years (range 31-68), 67% were women, median follow up was 12 years (range : 1.5 – 29) and median IFN treatment duration was 30 months (range: 12 – 102). 23/24 (96%) pts achieved complete or partial hematological response to IFNa. CALR mutations (present in all patients, quantifiable in all but 1 because of 1bp difference between mutated and WT) were of type 1 in 10 (42%), type 2 in 8 (33%) and of neither type in 6 (25%) pts, respectively. In addition to CALR, a second mutation was found in 8 pts (33%) by NGS, affecting ASXL1 (n=2), TET2 (n=2), IDH2 (n=2), CSF3R (n=1) and SH2B3 (n=1) genes. Comparison of sequential samples showed that the %CALR decreased from a median of 43% (range: 8-57) to 26% (range: 0-49) in the last sample (p= 0.018). In details, %CALR decreased in 13 (57%) pts, including 1 CMR, 7 PMR and 5 mMR. Interestingly, molecular response to IFNa of CALR+clones appeared poorer in pts with additional mutations (vs. pts with CALR mutation alone): 50% were non responders (vs. 40%), %CALR even increased during follow up in 25% (vs. 7%), and the median decrease in %CALR was 32% (vs. 45%). Dynamics of clones harboring additional mutated genes showed that, in contrast to CALR, mutant allele burden did not vary in 6/8 pts. However, an IDH2 mutation decreased from 9% to 1% in 1 pt, and a TET2 mutation increased from 1% to 21% in another (while %CALR remained unchanged in both pts, at 44% and 19% respectively). Of note, after 12 years of median follow-up, no emergence of new mutation was observed in any patient. Conclusion: In this cohort of JAK2V617F-negative ET, 96% of pts achieved hematological response with IFNa therapy. However, molecular profiling suggests that the mutational pattern found in malignant clones modulates the molecular response to IFNa: 1) Existence of more than 1 mutation induces poorer molecular response; 2) Clones with CALR mutation alone are sensitive to IFNa; 3) Clones harboring mutations in genes other than CALR seem less responsive to IFNa therapy. Disclosures Off Label Use: Interferon alpha was used off-label in selected ET patients according to local and international guidelines (Barbui et al., J Clin Oncol. 2011 Feb 20;29(6):761-70).

Clinical Features, Treatment Outcome, and Predictive Biomarkers In Adult T-Cell Leukemia/Lymphoma: University Of Miami Experience

Introduction Adult T-cell leukemia/lymphoma (ATLL) is a rare aggressive malignancy with a poor prognosis caused by HTLV-1. Miami is proximal to the Caribbean where HTLV-I is endemic, and we encounter a relatively high number of ATLL cases. Herein, we have performed the largest single institution retrospective analysis of ATLL patients (pts) to date in the U.S. We studied ATLL patient characteristics, treatment patterns, and disease outcome. In addition, we investigated the expression of IRF-4/MUM-1 in available specimens. Previously, our group demonstrated an association between lack of IRF-4/MUM-1 and response to AZT-interferon-alpha (AZT/IFNa) therapy in a small ATLL cohort. IRF-4/MUM-1 is a putative NF-kB target gene that encodes a transcription factor. IRF-4/MUM-1 expression has been associated with interferon resistance in preclinical studies, and is a poor prognostic marker in some lymphomas. One of our objectives is evaluate and validate IRF-4/MUM-1 as a potential biomarker for treatment selection and outcome in ATLL. Methods We analyzed 125 pts diagnosed with ATLL in UM/JMH between 1987 and 2013. We evaluated MUM-1 protein expression using immunohistochemistry (IHC) on either tissue sections, or cytospins prepared from CD4+-enriched peripheral blood leukemic specimens using 30% nuclear staining as a cut-off positive value, or by western blot (WB) analysis in some cases where fresh or DMSO preserved ATLL cells were not available. Kaplan-Meier survival curves, log-rank test where used for survival analysis. Mann-Whitney's U test was used to compare non-normally distributed continuous variables. Pearson's chi-squared or Fischer's exact tests were used to compare categorical variables. Results ATLL pts were 45% male and 55% female with a median age of 51 (17-91). The great majority of pts were Afro-Caribbean (82%), followed by U.S. African American (12%) and South American (6%). A total of 109 pts have been analyzed for treatment response so far, including 51 acute, 50 lymphomatous (L), 6 chronic (5 unfavorable), and 2 smoldering types. The median overall survival (OS) for acute and L was 6 and 10 months respectively, and not reached for chronic and smoldering types (figure 1). Fifty-six pts (34 acute, 14 L, 5 unfavorable chronic, 1 chronic, and 2 smoldering) were treated with high-dose AZT/IFNa as first line therapy. The complete and overall response rates (CR and ORR) after AZT/IFNa for acute/unfavorable chronic (A/UC) vs. L types were 25% vs. 0.7%, and 54% vs. 21% respectively. Seventy-seven pts received chemotherapy at some point during their treatment. The CR rate and ORR for A/UC vs. L-type pts treated with chemotherapy-based regimens were 40 % vs. 21%, and 70% vs. 77% respectively. However, we observed a significantly longer median progression-free survival (PFS) and sustained responses in pts with A/UC ATLL who achieved a CR with AZT/IFNa (168.1 wks), as compared to chemotherapy (61.1 wks), which translated into an overall survival benefit. Next, in order to determine whether IRF-4/MUM-1 predicted response to AZT/IFNa, 66 ATLL cases were analyzed by IHC and/or WB. The results showed that 38.5% of A/UC were IRF-4/MUM-1+ as compared to 82.1% in the L type (<P.0001). Evaluable pts for AZT/IFNa response demonstrated that 55% of A/UC MUM-1(-) cases had a CR as compared to 0% in MUM-1+ pts (P=.009). In the L group, AZT/IFNa responses were minimal and were mainly limited to stable disease. Subgroup analysis showed that median OS for MUM-1(-) vs. MUM-1+ in A/UC subtype was 38.4 wks vs. 27.6 wks (P=0.275) (figure 2), respectively. In the L subtype, median OS for MUM-1(-) vs. MUM-1+ was 25.7 wks vs. 60.3 wks (P=0.02) (figure 3), respectively. Finally, the median PFS in AZT/IFNa-treated A/UC pts favored the MUM-1(-) as compared to MUM-1+ cases. Conclusion Our data demonstrate that AZT/IFNa therapy is beneficial in leukemic type (A/UC) lacking IRF-4/MUM-1 expression, while L type is generally resistant to this treatment. On the other hand, IRF-4/MUM-1 expression is associated with a favorable outcome in L subtype. We have identified IRF-4/MUM-1 expression as a predictive marker that could be used in deciding upfront therapy (i.e. AZT/IFNa vs. standard chemotherapy) in leukemic ATLL subtypes. Our study findings must be confirmed and validated in a larger ATLL cohort. Disclosures: No relevant conflicts of interest to declare.

Analysis Of Molecular Responses and Chromosomal Aberrations In Patients With Polycythemia Vera Treated With Peg-Proline-Interferon Alpha-2b (AOP2014/P1101) In The Peginvera Study

Polycythemia vera (PV) is one of the most common type of BCR-ABL negative myeloproliferative neoplasms. It is characterized by elevated erythrocyte mass, variable presence of thrombocytosis and leukocytosis, predisposition to secondary myelofibrosis, thrombosis, bleeding and leukemic transformation. Most patients carry the JAK2-V617F mutation (up to 95%) or the less common JAK2 exon 12 mutations (around 3%). The current treatments include phlebotomy, low-dose aspirin, hydroxyurea, interferon alpha (IFNa) and bone marrow transplantation. However, PV is still lacking a curative treatment, with the exception of cases with successful bone marrow transplantation (in the spent phase) and few reports of complete clinical and molecular remission using IFNa. Measuring mutant JAK2 burden offers an opportunity to evaluate efficacy of therapy on the molecular level. We have previously reported sound clinical and molecular responses of PV patients treated with a new, once every 14 days formulation of peg-proline-IFNa-2b (AOP2014/P1101) in the Phase I/II clinical study PEGINVERA. The rational of the cytogenetic evaluation in this study is to investigate if chromosomal aberrations have an influence on the clinical course of PV patients, treated with AOP2014/P1101. It might be that cytogenetically complex patients have lower response rates to IFNa therapy. Furthermore, cytogenetic lesions may serve as additional markers to evaluate the response in parallel to JAK2 mutational burden analysis. Results of JAK2 mutational burden as well as high-resolution SNP array-based cytogenetic analysis, in 45 patients, treated with AOP2014/P1101, are presented here. Genome-wide human SNP 6.0 Affymetrix arrays were performed for the baseline sample (at the start of IFNa treatment) and latest follow-up sample. Mutant JAK2-V617F burden was determined by allele specific-PCR and quantitative PCR. For JAK2 exon 12 mutations a fragment analysis-based assay was used. Molecular response, defined by at least 10% decrease in mutant JAK2 burden, was observed in 73% of patients. The median follow-up time of patients was 500 days. The median follow-up time of molecular responding patients was 633 days, for partial responders 959 days and for non-responding patients 168 days. At least one chromosomal aberration was present in 69% of patients, of which chromosome 9p uniparental disomies (9pUPDs) were the most prevalent ones. Molecular non-responding patients did not have recurrent specific cytogenetic lesions or more chromosomal aberrations than responding patients. Molecular responses analyzed by JAK2 mutational burden correlated well with cytogenetic changes. A complete cytogenetic remission with around 3% residual JAK2-V617F burden could be achieved in 3 patients, all showing chromosome 9p UPDs at baseline. Interestingly, one patient had in addition to the 9p UPD a chromosome 14q UPD and another one trisomies of chromosome 8 and 9. This indicates that IFNa therapy is not restricted to mutant JAK2 clones but is also able to target other aberrant clones with common genetic changes found in MPN. Cytogenetic lesions found in follow-up samples that were not detected at baseline may indicate clonal evolution during IFNa therapy. These emerging clones might be responsible for acquisition of IFNa resistance and/or acceleration of disease progression. We found 3 such patients in our study, 1 with molecular response that acquired a small clone with a deletion of chromosome Y, 1 with partial molecular response where a single gene deletion on chromosome 10p (USP6NL) was detected and 1 that had no molecular response which showed a single gene gain on chromosome 3q (FXR1) as well as a single gene deletion on chromosome 7p (NXPH1). Additional follow-up samples will be necessary to assess the impact of the clonal evolution in these patients. Not only acquired somatic changes (including large chromosomal aberrations and point mutations) but also germline variants might influence IFNa response or resistance. Since we did not observe any difference in cytogentic lesions between molecular responding and non-responding patients, we suspect that in some cases germline variants are likely to influence the outcome of IFNa therapy. Further characterization of AOP2014/P1101 treated patients and later follow-up samples will help to better understand the clonal evolution and molecular responses during long-term IFNa treatment. Disclosures: Them: AOP Orphan Pharmaceuticals AG: Research Funding. Gisslinger:AOP Orphan Pharmaceuticals AG: Research Funding. Buxhofer-Ausch:AOP Orphan Pharmaceuticals AG: Research Funding. Greil:AOP Orphan Pharmaceuticals AG: Research Funding. Thaler:AOP Orphan Pharmaceuticals AG: Research Funding. Schloegl:AOP Orphan Pharmaceuticals AG: Research Funding. Gastl:AOP Orphan Pharmaceuticals AG: Research Funding. Berg:AOP Orphan Pharmaceuticals AG: Research Funding. Bagienski:AOP Orphan Pharmaceuticals AG: Research Funding. Zahriychuk:AOP Orphan Pharmaceuticals AG: Employment. Klade:AOP Orphan Pharmaceuticals AG: Employment. Kralovics:AOP Orphan Pharmaceuticals AG: Research Funding.

Predictive utility of serum cytokine levels in patients with metastatic renal cell carcinoma (MRCC) treated with interferon alfa (IFNa)

14503 Background: IFNa may prolong survival in MRCC patients (pts) due to stimulation of cell-mediated immunity. We hypothesized that IFNa exerts an anti-tumor effect by upregulation of Th1 cytokines and that patients (pts) with elevated serum levels of Th1 cytokines either at baseline (BL) or after treatment with IFNa would have a superior clinical outcome. Methods: Cytokine profiling was performed on 104 pts with MRCC treated in a randomized phase III trial with IFNa 0.5 million units (MU) subcutaneously (SC) twice daily or 5 MU SC daily. Serum samples were collected at BL (n = 104) and after 8 weeks of IFNa therapy (C1) (n = 89). Cytokine concentrations were determined using a 16-plex immunoassay. The linear mixed-effect model was fit to assess the change of cytokine levels from BL to C1. Cox proportional hazards model was fit to evaluate the effect of BL cytokine levels or change of cytokine levels from BL to C1 on the risk of death. Results: Of 16 cytokines evaluated (IL-1b, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12 p40, IL-13, IL-15, IL-17, IFNa, IFNg, GM-CSF, TNFa, VEGF), lower BL TNFa, IL-8 (Th1), and IL-13 (Th2) levels were associated with clinical benefit (major response or progression-free status at 6 months) (p = 0.01 and 0.03, respectively). By multivariate analysis, only extremely low or high levels of IFNa (p = 0.02) and IL-12 (p = 0.002) at BL were associated with an increased risk of death. IFNa therapy after C1 produced higher levels of several Th1 cytokines (IL-8, IL-12 p40, IL-15) (p < .001) and lower levels of Th2 cytokines (IL-4, IL-13). Unexpectedly, there were significantly lower levels of TNFa and GM-CSF (Th1) and higher levels of IL-10 (Th2) with IFNa. Only an increase of IL-2 levels from BL to C1 (RR 1.45; p = 0.05) correlated with an increased risk of death. Conclusions: Lower BL serum levels of TNFa, IL-8, and IL-13 were associated with clinical benefit to IFNa. Although IFNa therapy favored a shift towards a Th1 response, this effect alone did not correlate with clinical outcome. No significant financial relationships to disclose.

Serum Islet Cell Autoantibodies During Interferon α Treatment in Patients With HCV-Genotype 4 Chronic Hepatitis

Chronic hepatitis C virus (HCV) infection is a leading cause of end-stage liver disease worldwide and HCV genotype 4 (HCV4) is predominant in African and Middle Eastern countries. It is well established that interferon-α (IFNa) treatment for HCV may trigger serum autoantibodies against pancreatic islet cells (ICA) in a subgroup of patients. Available data on the incidence of ICA during IFNa therapy for chronic HCV4 infection are not conclusive. We investigated the appearance of ICA in 40 naïve Egyptian patients (38 males, 32 ± 6 years) with histologically defined chronic HCV4 infection undergoing IFNa treatment at a dose of 9-million U/week for 24 weeks. Serum samples were collected at baseline and following IFNa therapy and ICA were detected using indirect immunofluorescence. Baseline evaluation indicated that 2/40 (5%) patients had detectable serum ICA. After the completion of the treatment scheme, 12/38 (32%) previously ICA negative patients became ICA positive; however, no patient developed impaired glucose tolerance (IGT) or diabetes during follow-up. In conclusion, we submit that IFNa treatment for chronic hepatitis C (CHC) may induce serum ICA in one-third of Egyptian patients with HCV4. These autoantibodies, however, do not lead to alterations in glucose metabolism.

Eicosapentaenoic Acid (EPA) Attenuates the Anemia Due to Ribavirin/Interferon a Treatment in Patients with Chronic Hepatitis C.

Ribavirin(RBV)is a water soluble synthetic guanosine analog that exerts antiviral activity against DNA and RNA viruses after intracellular phosphorylation. Current studies indicate that combination therapy with RBV and Interferon a (IFNa) is associated with higher rates of sustained virological and biochemical response compared to IFN a monotherapy for chronic hepatitis type C (CHC). Hemolytic anemia is the major side effect of the therapy, with 67% of treated patients developing anemia. The exact mechanism for RBV-related anemia is unknown, but it is thought to be decreasing deformability of eythrocytes resulting from accumulation of phosphorylated RBV in erythrocytes. Highly purified eicosapentaenoic acid (EPA) is widely used for the treatment of hyperlipidemia and atherosclerosis in Japan. We have investigated that EPA has a beneficial effect in patients with RBV-related anemia (Int J Mol Med11:729–732, 2003). Our aim of this study is to establish the safer treatment of RBV/IFNa using EPA for CHC patients and elucidate the mechanism of EPA action for improving anemia. [Patients and methods] Thirty three patients with CHC were randomized to RBV/IFNa therapy (800mg/6MU/day) with or without EPA (1800mg/day) for 12 weeks (14:+EPA; 19:-EPA). Erythrocyte filterability (Nickel mesh filtration method; Jpn J Physiol53: 481-486, 2003) was measured before and after 12 weeks of treatment in 8+EPA and 9 -EPA patients. Statistical analysis was performed by Wilcoxon and ANOVA test. [Result] As shown in Fig. 1, hemoglobin (Hb) levels decreased in both groups by the treatment. However, decrease of Hb levels significantly attenuated by the addition of EPA at 4 and 8 weeks after initiation of the treatment. This effect continued for 12 weeks. Initial mean erythrocyte filterability (%) at study entry was 65.3% and 68.1% in the + and -EPA group, respectively. After 12 weeks treatment, improvement of erythrocyte filterability occurred in 6/8 patients in the +EPA group. In contrast, 7/9 patients resulted in the decrease of erythrocyte filterability in the -EPA group. As shown in Fig 2, the mean erythrocyte filterability (%) for the +EPA group was 71.0%, against 51.4% in the -EPA group (P=0.01) after 12 weeks. [Discussion and Conclusion] EPA significantly attenuated the anemic effect of RBV/IFNa therapy for CHC patients. Erythrocyte filterability decreased in the -EPA group. On the contrary, decrease of erythrocyte filterability was not found in the +EPA group. Therefore, EPA improved the anemic side effect of RBV/IFNa via the stabilizing action of the membrane by amelioration of erythrocyte filterability. This study would be useful for clinical application.