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Precision medicine can significantly improve outcomes for cancer patients, but implementation requires comprehensive characterization of tumor cells to identify therapeutically exploitable vulnerabilities. Here we describe somatic biallelic TET2 mutations in an elderly patient with acute myeloid leukemia (AML) that was chemoresistant to anthracycline and cytarabine (Ara-C), but acutely sensitive to 5'-azacitidine (5'-Aza) hypomethylating monotherapy resulting in long-term morphological remission. Given the role of TET2 as a regulator of genomic methylation, we hypothesized that mutant TET2 allele dosage affects response to 5'-Aza. Using an isogenic cell model system and an orthotopic mouse xenograft, we demonstrate that biallelic TET2 mutations confer sensitivity to 5'-Aza compared to cells with monoallelic mutation. Our data argue in favor of using hypomethylating agents for chemoresistant disease or as first line therapy in patients with biallelic TET2-mutated AML and demonstrate the importance of considering mutant allele dosage in the implementation of precision medicine for cancer patients.

Myelodysplastic syndromes are clonal disorders with morphological dysplasia, a variable degree of cytopenia and a risk of transformation to acute myeloid leukemia. Prognosis is very variable and is defined by blast count, cytopenia, cytogenetics and more recently by somatic mutations, with IPSS or revised IPSS score being the most widely used to assess disease risk. HSCT remains the only curative treatment to date, with high-risk patients obtaining the biggest benefit. However, NRM should be carefully assessed before indicating the transplant in this usually old population, where organ toxicity and comorbid conditions are to be considered. Multi-domain assessment tools, such as CGA (comprehensive geriatric assessment) and EBMT score, are useful in this context and might guide physician decisions regarding the transplant. Indeed, with the development of reduced intensity conditioning regimens, the number of patient candidates for an HSCT has increased. Regarding pre-transplant treatment, patients with a blast excess > 10% might be treated with HMAs or chemotherapy, although there are no randomized trials confirming the benefit of this approach, even when achieving a complete response. Concerning donor choice, matched sibling donors continue to be the first option, although matched unrelated donors, and more recently haploidentical donors, have proven to be valid options and should be offered in the absence of a related donor. Relapse remains the main cause of transplantation failure. MRD assessment and pre-emptive or prophylactic use of HMA or other targeted inhibitors with or without DLI are accepted strategies to reduce relapse risk, but the prognosis in this context remains dismal, and is the subject for several ongoing clinical protocols.

Acute myeloid leukemia is a heterogeneous disease defined by a large spectrum of genetic aberrations that are potential therapeutic targets. New targeted therapies have changed the landscape for a disease with poor outcomes. They are more effective than standard chemotherapy with a good safety profile. For “fit patients” in first-line, the combination of gemtuzumab ozogamicin or midostaurin with intensive chemotherapy or Vyxeos is now considered the “standard of care” for selected patients. On the other hand, for “unfit patients”, azacitidine-venetoclax has been consolidated as a frontline treatment, while other combinations with magrolimab or ivosidenib are in development. Nevertheless, global survival results, especially in relapsed or refractory patients, remain unfavorable. New immunotherapies or targeted therapies, such as Menin inhibitors or sabatolimab, represent an opportunity in this situation. Future directions will probably come from combinations of different targeted therapies (“triplets”) and maintenance strategies guided by measurable residual disease.

The value of measurable residual disease (MRD) in elderly patients with acute myeloid leukemia (AML) is inconsistent between those treated with intensive vs hypomethylating drugs, and unknown after semi-intensive therapy. We investigated the role of MRD in refining complete remission (CR) and treatment duration in the phase 3 FLUGAZA clinical trial, which randomized 283 elderly AML patients to induction and consolidation with fludarabine plus cytarabine (FLUGA) vs 5-azacitidine. After consolidation, patients continued treatment if MRD was =0.01% or stopped if MRD was

Clonal evolution in acute myeloid leukemia (AML) originates long before diagnosis and is a dynamic process that may affect survival. However, it remains uninvestigated during routine diagnostic workup. We hypothesized that the mutational status of bone marrow dysplastic cells and leukemic blasts, analyzed at the onset of AML using integrated multidimensional flow cytometry (MFC) immunophenotyping and sorting (FACS) with next-generation sequencing (NGS), could reconstruct leukemogenesis. Dysplastic cells were detected by MFC in 285 of 348 (82%) newly-diagnosed AML patients. Presence of dysplasia according to MFC and WHO criteria had no prognostic value in the elderly. NGS of dysplastic cells and blasts isolated at diagnosis identified three evolutionary patterns: stable (n=12/21), branching (n=4/21) and clonal evolution (n=5/21). In patients achieving complete response, integrated MFC and FACS with NGS showed persistent measurable residual disease (MRD) in phenotypically normal cell types, as well as the acquisition of genetic traits associated with treatment resistance. Furthermore, whole-exome sequencing of dysplastic and leukemic cells at diagnosis and of MRD uncovered different clonal involvement in dysplastic myelo-erythropoiesis, leukemic transformation and chemoresistance. Altogether, we showed that it is possible to reconstruct leukemogenesis in approximately 80% of newly diagnosed AML patients, using techniques other than single-cell multiomics. ACKNOWLEDGEMENTS: The authors acknowledge the patients, caregivers, and the biobank of the University of Navarra. This work was supported by grants from the Área de Oncología del Instituto de Salud Carlos III, Centro de Investigacion Biom ´ edica en ´ Red (CIBER-ONC) (CB16/12/00369, CB16/12/00233, CB16/12/ 00489, and CB16/12/00284), Instituto de Salud Carlos III/Subdireccion General de Investigaci ´ on Sanitaria (FIS numbers PI16/ ´ 01661, PI16/00517, and PI19/01518), and the Plan de Investigacion´ de la Universidad de Navarra (PIUNA 2014-18). This work was supported internationally by the Cancer Research UK, FCAECC, and AIRC under the Accelerator Award Program (EDITOR).

Cytomegalovirus (CMV) reactivation is an important cause of complications after hematopoietic stem cell transplantation (HSCT). Discrepancies between serologic and cellular CMV-specific immune response have been reported. This study evaluated the impact of lack of CMV-specific CD8

Precision medicine can significantly improve outcomes for cancer patients, but implementation requires comprehensive characterization of tumor cells to identify therapeutically exploitable vulnerabilities. Here we describe somatic biallelic TET2 mutation (focal deletion and nonsense mutation) in an elderly patient with acute myeloid leukemia (AML) that was chemoresistant to anthracycline and cytarabine, but acutely sensitive to 5’-azacitidine (5’-Aza) hypomethylating monotherapy, resulting in long-term morphological remission (overall survival (OS) 850 days). Given the role of TET2 as a regulator of genomic methylation, we hypothesized that mutant TET2 allele dosage affects response to 5’-Aza. Using an isogenic cell model system and an orthotopic mouse xenograft, we demonstrate that biallelic TET2 mutations confer sensitivity to 5’-Aza compared to cells with monoallelic mutation. We subsequently identified 29 additional patients from the Study Alliance Leukemia biobank with chromosome 4 abnormalities and identified two further patients with complex biallelic TET2 mutations, including one with trisomy 4, homozygosity across the long arm and an inactivating point mutation. We also screened patients recruited to the PETHEMA FLUGAZA phase 3 clinical trial and identified three patients with biallelic TET2 mutations, two of whom had responded very well to single agent 5’-Aza (OS 767 and 579 days) despite having adverse risk AML and poor performance status. Our data argue in favor of using hypomethylating agents for chemoresistant disease or as first line therapy in patients with biallelic TET2-mutated AML and demonstrate the importance of considering mutant allele dosage in the implementation of precision medicine for cancer patients.Key PointsMutant TET2 allele dosage affects response to 5’-azacitidine in acute myeloid leukemia in vitro and in a xenograft model.Our data highlight the importance for screening of biallelic mutations to predict response to therapy in acute myeloid leukemia.

Background: Clonal evolution in AML originates long before diagnosis and is a highly dynamic process. Having a greater understanding of leukemogenesis may contribute to develop treatment strategies that target the tumor evolutionary process. However, dissecting leukemic transformation at the onset of AML is challenging without single-cell sequencing, and most clinical laboratories do not have infrastructure to perform these studies routinely. Patients with newly diagnosed AML may present dysplasia. If these residual, mature, dysplastic cells were generated before the differentiation blockage of blasts preceding leukemic transformation, it could be hypothesized that studying the genetic landscape of dysplastic cells and blasts could uncover the evolutionary process from dysplasia to AML. This hypothesis has never been investigated. Aim: Reconstruct clonal evolution from dysplasia to AML based on the genetic signature of dysplastic cells and leukemic blasts, analyzed using integrated MFC immunophenotyping and sorting with NGS. Methods: Presence of dysplasia according to aberrant phenotypic differentiation of the neutrophil, monocytic and erythroid lineages was investigated using MFC and EuroFlow MDS/AML panels in 283 newly diagnosed AML patients (median age 74; range 29-90). Patient-specific phenotypes were leveraged to isolate a total of 99 cell types from 22 AML cases for targeted (48 MDS/AML related genes) and whole-exome sequencing (WES), with a mean depth of 3246x and 141x, respectively. In patients with measurable residual disease (MRD) by MFC at the time of complete remission, tumor resistant cells were FACSorted for WES using patient-specific aberrant phenotypes. T cells were used as germline control in both approaches. Mutations were considered if ≥0.05 allele frequency in leukemic blasts or dysplastic cells and ≤0.2 in T cells. Results: We first assessed the applicability of our hypothesis by investigating how many patients show dysplasia at the onset of AML. Dysplastic cells were observed in 252 of 283 (89%) cases. Phenotypic abnormalities were more frequently noted in the neutrophil lineage (47%), followed by the monocytic (40%) and erythroid cells (13%). Up to 169/283 (60%) patients showed multi-lineage dysplasia. Only nine cases showed no signs of dysplasia, whereas the remaining 22 had undetectable hematopoiesis. Targeted sequencing of dysplastic cells and blasts in 16 patients uncovered three evolutionary patterns of leukemogenesis. Stable transition in those displaying identical mutational landscapes in blasts and residual mature dysplastic cells (9/16); clonal selection in cases where blasts originated from leukemic stem cells other than the ones driving dysplasia, due to mutations absent in blasts and present in dysplastic cells (4/16); and clonal evolution in cases showing new mutations in blasts onto mutations shared between these and dysplastic cells (3/16). Interestingly, most patients displaying stable transition from dysplasia to AML had mutated ASXL1, RUNX1 and/or TP53 (8/9). Mutations present in dysplastic cells while absent in blasts from patients showing a clonal selection evolutionary pattern, were more frequently detected in genes related to signaling pathways (eg JAK2, KRAS and NRAS). By contrast, clonal evolution was characterized by new mutations affecting FLT3ITD and STAG2. The higher throughput of WES of dysplastic cells and blasts from six patients unveiled a more complex dynamic process of leukemogenesis, with all three evolutionary patterns being detectable in nearly all cases. Most interestingly, we found patients with mutations in dysplastic cells and blasts at diagnosis, but not in MRD cells (eg NBPF1 and ZNF717); and patients showing mutations in dysplastic and MRD cells, but not in blasts at diagnosis (eg MUC2 and KIR2DL3). These findings uncover that genetic alterations that are critical in leukemic transformation and chemoresistance, may not overlap (Figure). Conclusions: We showed for the first time that it is possible to reconstruct leukemogenesis in nearly 90% of newly-diagnosed AML patients, using techniques that are commonly available in clinical laboratories. The possibility to identify the genetic drivers of leukemic transformation and chemoresistance, could be clinically meaningful to develop tailored treatment strategies aiming at the eradication of genetically diverse leukemic clones. Figure 1 Figure 1. Disclosures Prósper: Oryzon: Honoraria; Janssen: Honoraria; BMS-Celgene: Honoraria, Research Funding. Ayala: Incyte Corporation: Membership on an entity's Board of Directors or advisory committees; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees; Astellas: Honoraria; Celgene: Honoraria. Perez-Simon: JANSSEN, TAKEDA, PFIZER, JAZZ, BMS, AMGEN, GILEAD: Other: honorarium or budget for research projects and/or participation in advisory boards and / or learning activities and / or conferences. San-Miguel: AbbVie, Amgen, Bristol-Myers Squibb, Celgene, GlaxoSmithKline, Janssen, Karyopharm, Merck Sharpe & Dohme, Novartis, Regeneron, Roche, Sanofi, SecuraBio, and Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees. Montesinos: Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Pfizer: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Daiichi Sankyo: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Sanofi: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Incyte: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Karyopharm: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Stemline/Menarini: Consultancy; Teva: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Janssen: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Agios: Consultancy; Tolero Pharmaceutical: Consultancy; Forma Therapeutics: Consultancy; Glycomimetics: Consultancy; AbbVie: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Astellas Pharma, Inc.: Consultancy, Honoraria, Other: Advisory board, Research Funding, Speakers Bureau. Paiva: Celgene, EngMab, Roche, Sanofi, Takeda: Research Funding; Adaptive, Amgen, Bristol-Myers Squibb-Celgene, Janssen, Kite Pharma, Sanofi and Takeda: Honoraria; Bristol-Myers Squibb-Celgene, Janssen, and Sanofi: Consultancy.

Background: Allogeneic hematopoietic stem cell transplantation (HSCT) remains the only curative therapy in patients with primary or secondary myelofibrosis. Ruxolitinib, a JAK-inhibitor, has been approved in myelofibrosis patients and is effective to alleviate symptoms of the disease in a substantial proportion of patients. In a French phase 2 study, the use of ruxolitinib as a bridge to HSCT was followed by a high transplantation rate, with one third of patients presenting clinical improvement before HSCT (partial response or spleen size reduction) (BMT 2021, Robin et al). The aim of the current study is to compare post-transplantation outcomes between patients who received ruxolitinib before HSCT and those who did not. Methods: This retrospective study includes patients with primary or secondary myelofibrosis registered in the SFGM-TC registry including patients from France, Belgium and Switzerland. Patients who received HSCT between July 2010 and December 2019 with available information regarding ruxolitinib treatment were included. The impact of ruxolitinib was analyzed by multivariable Cox regression and a propensity score. Results: 305 patients could be included, of whom 143 received ruxolitinib prior to HSCT (RuxoG). All patient characteristics are given in RuxoG followed by no ruxolitinib group (noRuxo). The diagnosis before HSCT was primary myelofibrosis (83% vs 75%), secondary myelofibrosis (13% vs 19%) or transformed into AML (4% vs 6%). Karnofsky score at transplant was < 70% in 9% and 11%. In RuxoG. median age was older (60.29 vs. 58.75, p=0.069) and delay from diagnosis longer (20.42 vs 13.26 months, p=0.040). RuxoG patients had less frequently anemia < 10 g/dl (84% vs 98%, p=0.027) or thrombocytopenia < 50 G/L (22% vs 32%, p=0.066) and presented more frequently splenomegaly or pre-graft splenectomy (83% vs 63%, p=0.0007). DIPSS score was well balanced in both groups, being high or intermediate-2 in 100 patients (79% vs 86%, p=0.23), from the 235 patients with available data. Myeloablative conditioning regimen was used in 16% vs 26% (p=0.049), and conditioning regimen consisted of on fludarabine-busulfan in 33% vs 61%, fludarabine-melphalan in 47% vs 22% or other in 20% vs 16% (p Conclusion: Patients who received ruxolitinib before HSCT were at higher risk of acute GVHD and post-transplant mortality, but this was not observed in patients responding to ruxolitinib and it does not translate into higher risk of chronic GVHD. Main hypotheses to explain these finding could be: 1) specific disease, patient or transplant characteristics in RuxoG; 2) the longer delay to transplantation or 3) a specific effect of ruxolitinib. This study suggests that myelofibrosis patients responding to ruxolitinib should have the transplant without waiting disease progression. Figure 1 Figure 1. Disclosures Forcade: Novartis: Other: travel grant. Loschi: AbbVie: Ended employment in the past 24 months, Honoraria; CELGENE/BMS: Honoraria; Gilead: Ended employment in the past 24 months, Honoraria; Novartis: Ended employment in the past 24 months, Honoraria; Servier: Ended employment in the past 24 months, Honoraria; MSD: Honoraria. Duléry: Takeda Gilead Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Robin: NEOVII MEDAC NOVARTIS: Research Funding.

Acute myeloid leukemia (AML) is a hematologic neoplasm characterized by a remarkable phenotypic and genomic heterogeneity. The recent characterization of genomic subtypes of AML based on large sequencing studies has provided the rationale for the development of targeted therapies based on the presence of specific genomic abnormalities. However, long term survival particularly in older patients remains a unmet medicalneed. Additionally, recent studies using RNA interference (RNAi) libraries have determined the existence of genes that are essential for the survival of multiple cancer cells. Understanding the effect of genomic alterations (mutations, deletions, translocations) on gene essentiality could favor the development of targeted therapies for specific subgroups of AML patients. However, current statistical methods such as the Benjamini-Hochberg (BH) procedure have shown limitations for controlling the false discovery rate (FDR) and have suboptimal sensitivity (recall of true positives) because the P-value correction does not include any prior information of individual tests. For this reason, in this study we developed a new large-scale statistical algorithm, which combine the RNAi libraries (more than 17.000 genes) data with mutational profiles, to identify gene essentialities associated with specific genomic mutations in order to explore this approach in AML. We adapted the Independent Hypothesis Weighting (IHW) procedure to the problem of identifying mutations as surrogate markers of gene essentiality, by using the gene mutation state in each cell line as prior information of a IHW problem. This approach was tested in 19 tumor subtypes, of the Cancer Cell Line Encyclopedia (CCLE) showing that it recalls new discoveries that cannot be identified with standard procedures in 17 out of 19 tumors, including the identification of up to 1,000 discoveries in tumor types in which BH recalls no discovery. These results demonstrated the accuracy of the IHW-based approach to identify gene mutations as surrogate markers of gene essentiality in the future. Once validated, we applied this computational model to the15 AMLcell lines of CCLE. The number of discoveries with an FDR of 20% increases from 2 (using the traditional BH correction), to 38 using our procedure, showing NRAS as the top mutation biomarker in the ranking. Interestingly, the algorithm identified one essential gene (NRAS) for NRAS mutated (NRAS-mut) and another essential gene (PTPN11) for NRAS wild type (NRAS-wt) AML cells, covering all samples of AMLs. To validate this hypothesis, we examined the effect of two different specific siRNAs for each gene (siPTPN11 and siNRAS) on cell proliferation of four AML cell lines: two lines with NRAS-mut (HL-60 and OCIAML3) and two with NRAS-wt (MV4-11 and HEL). Downregulation of NRAS expression significantly decreases the cell proliferation only in the 2 NRAS-mutated AML cell lines. Whereas the inhibition of PTPN11expression produced an equivalent effect, but specifically in the 2 NRAS-wt AML cell lines (Figure 1). These results confirmed our predictions and showed the essential role of NRAS or PTNPN11 in AML cell lines either with NRAS mutated or wild type, respectively. These results demonstrate that the application of our algorithm in the context of specific gene mutation not only may allow identification of directed therapies based on the mutation but can also define new gene essentialities amenable for targeted therapies providing new therapeutic strategies in patients with AML and potentially in other tumors. Disclosures Paiva: Amgen, Bristol-Myers Squibb, Celgene, Janssen, Merck, Novartis, Roche and Sanofi: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene, Janssen, Sanofi and Takeda: Consultancy. San-Miguel:Amgen, Bristol-Myers Squibb, Celgene, Janssen, MSD, Novartis, Roche, Sanofi, and Takeda: Consultancy, Honoraria.

Acute myeloid leukemia (AML) is a malignant disease characterized by uncontrolled proliferation, differentiation arrest and accumulation of immature myeloid progenitors. Despite recent developments and the approval of new therapeutic agents in the last few years, long term survival of AML, particularly in elderly patients remains an unmet medical need.The use of all-trans retinoic acid (ATRA) in Acute Promyelocytic Leukemia has proven that differentiation therapy may significantly change the survival of AML patients, however the success in APL has not been translated to other groups of AML. Therefore, the identification of new therapeutic agents that may induce the differentiation of AML blasts represents an attractive new target. Furthermore, it is well known that epigenetic alterations have an important role in the development and maintenance of cancer and AML in particular. Thus, our aim was to develop new small molecules targeting epigenetic modifying enzymes like DNA methyltransferases (DNMT), histone methyltransferases or histone deacetylase (HDAC) with the aim of inducing differentiation in AML. We performed a screening of over 50 small molecules synthesized by our group. The design was performed in-house using a knowledge and structure based strategy and the read out of the screening was based on changes in expression of CD11b (a well described marker of myeloid differentiation) after in vitro treatment of AML cells lines. Interestingly, we found several compounds with high capacity to promote the differentiation of leukemic cells in AML cells lines at low non-cytotoxic doses, selecting CM-444 and CM-1758 as our lead compounds (Figure 1a).A complete biochemical characterization showed that both compounds are specific pan-HDACs inhibitors (HDACi). CM-444 and CM-1758 induced in vitro cell differentiation in all subtypes of AML, independently of the AML genetic subgroups or the presence of mutations, which was significantly more pronounced that differentiation induced by reference compounds such as Panobinostat, Vorinostat, Entinostat, Tubastatin or Quisinostat, previously described HDACi. CM-444 and CM-1758 also induced in vivo differentiation in xenogeneic models of AML. AML differentiation was associated with induction of CD11b, downregulation of c-MYC, overexpression of transcription factors that govern the myeloid differentiation and morphologic changes. In addition, these compounds promoted in vitro differentiation of patient-derived AML blasts. The complete transcriptome analysis by RNA-Seq carried out in AML cell lines after CM-444, CM-1758, Panobinostat or Vorinostat treatment showed changes in genes implicated in differentiation, but without explaining the differences among the different HDACi. Analysis of the complete acetylome and proteome before and after treatment with CM-444 and CM-1758 in comparison with other HDACi showed differential acetylation of non-histone proteins included in the GO categories of Zn binding proteins and nucleic acid binding proteins (Figure 1b). Most of these proteins are epigenetic enzymes and have been related to AML and myeloid differentiation, such as MLL2, EP300 or BRD4. In summary, we have developed and characterized novel epigenetic small molecules with a high in vitro and in vivo capacity of differentiating AML cells. These compounds might be an effective differentiation-based therapy to be tested in AML. Besides, the mechanism of differentiation of these compounds is due, at least in part, to the acetylation of non-histone epigenetic proteins, which are key in the myeloid differentiation. Disclosures Paiva: Celgene, Janssen, Sanofi and Takeda: Consultancy; Amgen, Bristol-Myers Squibb, Celgene, Janssen, Merck, Novartis, Roche and Sanofi: Honoraria, Membership on an entity's Board of Directors or advisory committees. San-Miguel:Amgen, Bristol-Myers Squibb, Celgene, Janssen, MSD, Novartis, Roche, Sanofi, and Takeda: Consultancy, Honoraria.

To assess the effectiveness of simplification to a dual antiretroviral regimen containing a ritonavir-boosted protease inhibitor (PI/r) in treatment-experienced HIV-1-infected patients.Retrospective analysis of 131 HIV-1-infected patients on suppressive antiretroviral treatment (HIV-RNA50 copies/mL) who switched to a maintenance dual antiretroviral regimen, containing a PI/r, in three hospitals in Spain. Virological failure was defined as confirmed HIV-RNA50 copies/mL. The percentage of patients remaining free of therapeutic failure was estimated using the time-to-loss-of-therapeutic-response algorithm, by intent-to-treat analysis.Median baseline characteristics of the patients were 14 years on antiretroviral therapy, five prior HAART regimens and 10 different drugs, 24 months on a suppressive regimen and 522 CD4+ cells/mL. Reasons for simplification to dual therapy were nucleoside reverse transcriptase inhibitor-related toxicity (46.6%), removal of lamivudine/emtricitabine due to resistance (16.8%), simplification from regimens containing a dual PI, enfuvirtide or tipranavir (20.6%) and simplification from other complex regimens (16.0%). Darunavir (58.0%), lopinavir (16.8%) or atazanavir (13.0%) were the preferred PIs, used in combination with tenofovir (50.4%), raltegravir (22.1%) or etravirine (12.2%). At the end of follow-up (median 14 months), 90.1% of patients remained free of therapeutic failure; corresponding data at treatment weeks 24, 48 and 96 were 93.6% (95% CI, 89.3-97.9), 90.9% (95% CI, 84.9-95.9) and 87.4% (95% CI, 80.7-94.1), respectively. Two (1.5%) patients had virological failure and 11 (8.4%) discontinued treatment due to side effects or were lost to follow-up.Simplification to a dual-therapy regimen including a PI/r might be useful to enhance convenience and/or diminish toxicity in selected treatment-experienced patients.

Background. To study the feasibility of a response-guided therapy for chronic hepatitis C virus (HCV) infection in patients coinfected with human immunodeficiency virus (HIV) in a tertiary care hospital. Methods. Treatment duration was individualized on the basis of week 4 and week 12 virologic response. Sixty patients were enrolled and received pegylated interferon alfa-2b (1.5 μg/kg per week) plus weight-based ribavirin (800-1400 mg/day). Patients who achieved a rapid virologic response, defined as viral load

Objectives: The aim of this study was to assess the influence of hepatitis B virus or hepatitis C virus co-infection and the extent of liver fibrosis on saquinavir and ritonavir pharmacokinetics in HIV-infected subjects without liver function impairment. Methods: A cross-sectional, comparative study enrolling HIV-infected adults receiving saquinavir/ ritonavir 1000/100 mg twice daily or 1500/100 mg once daily was conducted. Patients with chronic viral hepatitis (HEP+) were grouped as having advanced liver fibrosis (HEP+/FIB+) or not (HEP+/FIB-) based on the FIB-4 index. Saquinavir and ritonavir trough concentrations (C trough ) in plasma were determined by HPLC. The geometric mean ratio (GMR) was used to compare saquinavir and ritonavir C trough between HEP― and HEP+ patients, and the influence of the extent of liver fibrosis on saquinavir and ritonavir pharmacokinetics was explored using analysis of variance. Results: One hundred and thirty-eight patients on twice-daily saquinavir/ritonavir (67 HEP―, 71 HEP+) and 36 patients on once-daily saquinavir/ritonavir (12 HEP―, 24 HEP+) were included. Saquinavir C trough was comparable between HEP― and HEP+ patients receiving either saquinavir/ritonavir 1000/ 100 mg twice daily [GMR 0.91, 95% confidence interval (CI) 0.60―1.37; P=0.655] or 1500/100 mg once daily (GMR 0.88, 95% CI 0.39―1.97; P=0.752). Similarly, ritonavir C trough was also comparable between HEP― and HEP+ patients. The extent of liver fibrosis was not significantly related to saquinavir or ritonavir C trough in patients receiving either of the two studied doses. Conclusions: Saquinavir C trough was not increased in HIV-infected patients with chronic viral hepatitis in the absence of liver function impairment. These results confirm that no specific dose modification of saquinavir/ritonavir should be recommended in this setting.

Background The aim of this study was to analyze the incidence of new cases, survival of HIV-1-infected patients with progressive multifocal leukoencephalopathy (PML), and the characteristics of PML-associated immune reconstitution inflammatory syndrome (IRIS). Methods Multicenter observational cohort study of all HIV-1-infected patients newly diagnosed of PML in 7 hospitals in Barcelona (Spain) from 2002 to 2006. The annual incidence of PML was calculated. Survival was estimated using the Kaplan-Meier method. IRIS was defined as new onset or rapid worsening of PML shortly after initiation of highly active antiretroviral therapy together with a decline in HIV-1 viral load and rising of CD4 lymphocytes. Results Sixty-one new cases of PML were diagnosed. The mean survival time was 15 months [95% confidence interval (CI), 11 to 19]. The Kaplan-Meier estimates of the probability of survival were 47.7% (95% CI, 35 to 59) at 6 months, 38.6% (95% CI, 25 to 51) at 12 months, 35.1% (95% CI, 22 to 48) at 24 months, and 25.1% (95% CI, 10 to 40) at 36 months. IRIS was diagnosed in 14 (23%) cases. Mortality was similar in patients with and without IRIS. Conclusions PML continues to be one of the deadliest opportunistic infections in acquired immunodeficiency syndrome patients. The development of PML-associated IRIS has no influence on prognosis.

Latent parasitic infections can reactivate because of immunosuppression. We conducted a prospective observational study of all human immunodeficiency virus (HIV)-infected immigrants who visited the Infectious Diseases Department of the Hospital Universitari Vall d'Hebron, Barcelona, Spain, during June 2010-May 2011. Screening of the most prevalent tropical diseases (intestinal parasitosis, Chagas disease, leishmaniasis, malaria, schistosomiasis, and strongyloidiasis) was performed according to geographic origin. A total of 190 patients were included: 141 (74.2%) from Latin America, 41 (21.6%) from sub-Saharan Africa, and 8 (4.2%) from northern Africa. Overall, 36.8% (70 of 190) of the patients had at least one positive result for any parasitic disease: 5 patients with positive Trypanosoma cruzi serology, 11 patients with positive Schistosoma mansoni serology, 35 patients with positive Strongyloides stercoralis serology, 7 patients with positive Leishmania infantum serology, intestinal parasitosis were detected in 37 patients, malaria was diagnosed in one symptomatic patient. We propose a screening and management strategy of latent parasitic infections in immigrant patients infected with HIV.

Background: Eligibility criteria might explain differences in viral response to combined antiretroviral treatment (cART) between clinical trials and routine care setting. Our aim was to assess the effectiveness of cART and factors associated to therapeutic failure (TF) in real clinical conditions. Methods: A prospective cohort analysis including HIV-1 infected patients who started a cART between January 2004 and December 2009, at Vall d’Hebron Hospital. Effectiveness was evaluated as time to TF defined as the first of either virologic failure, treatment discontinuation whatever the reason other than switching, loss to follow-up, or death. The Kaplan-Meier method was used to estimate time-to-event distributions and Cox regression modelling to identify factors associated with TF. Results: We analyzed 232 patients; median CD4+ cell count was 229 cells/mm3 and median viral load 4.89 log10. Almost a third of patients was co-infected with HCV and/or HBV. Tenofovir plus lamivudine/emtricitabine (67%) was the commonest backbone, and efavirenz (77%) the preferred third drug. The proportion of patients with no TF at month 12, 24 and 36 was 82.9%, 78.5% and 76% respectively. TF occurred in 57 (24.6%) patients, mainly due to intolerance or toxicity. The risk of TF was higher in patients starting cART before 2006 and in those with a protease inhibitor based regimen. Conclusions: After a median follow-up of 36.5 months, three-fourth of patients who started first-line cART remained free of TF. Treatment discontinuation stands as the leading cause of TF.

Late diagnosis of HIV-1 infection is quite frequent in Western countries. Very few randomized clinical trials to determine the best antiretroviral treatment in patients with advanced HIV-1 infection have been performed. To compare immune reconstitution in two groups of very immunosuppressed (less than 100 CD4(+) cells/microl), antiretroviral-naive HIV-1-infected adults, 65 patients were randomly assigned in a 1:1 ratio to receive zidovudine + lamivudine + efavirenz (group A, 34 patients) or zidovudine + lamivudine + ritonavir-boosted indinavir (group B, 31 patients). The median (interquartile range) CD4(+) cell increase after 12 and 36 months was +199 (101, 258) and +299 (170, 464) cells/microl in the efavirenz arm and +136 (57, 235) and +228 (119, 465) cells/microl in the ritonavir-boosted indinavir arm (p0.05 for all time points). The proportion (95% confidence interval) of patients achieving HIV-1 RNA levels under 50 copies/ml was significantly greater in the efavirenz arm at 3 years by the intention-to-treat analysis [59% (41%, 75%) vs. 23% (10%, 41%)], whereas no differences were found in the on-treatment analysis. Immune activation (CD8(+)CD38(+) and CD8(+)CD38DR(+) T cells) was significantly lower for the efavirenz arm from month 6 to month 24. Adverse events were more frequent in the ritonavir-boosted indinavir arm. Almost all cases of disease progression and death were observed in the first year of treatment, with no significant differences between the two arms (p = 0.79 by the log-rank test). At 1 and 3 years, the immune reconstitution induced by an efavirenz-based regimen in very immunosuppressed patients was at least as potent as that induced by a ritonavir-boosted protease inhibitor-based antiretroviral regimen.

Background: Boosted darunavir (DRV/r) plus etravirine (ETR), in DUET trials, and raltegravir, in BENCHMRK trials, showed high rates of virologic response in patients with multidrug-resistant HIV-1 infection, particularly when associated with two more fully active antiretroviral drugs. No data from clinical trials, about this combination, are available. Patients and Methods: Thirty-two consecutive heavily pretreated patients with multidrug-resistant HIV-1 infection who started a new salvage regimen with RAL (400 mg twice daily), ETR (200 mg twice daily), and DRV/r (600/100 mg twice daily) were studied. Clinical evaluation and immunologic, virologic, and biochemical parameters were collected at baseline and at Weeks 4, 12, and 24. Results: Median baseline characteristics were age 44 years, 13 years on antiretroviral therapy, nine prior highly active antiretroviral therapy regimens, 261 CD4 cells/mL, and HIV-1 RNA 4.2 log 10 copies/mL. Sixteen (50%) and 14 (44%) patients were enfuvirtide-and tipranavir-experienced, respectively. Three-class resistance mutations were present in all patients. Three patients (9%) had isolates with three ETR resistance mutations. All patients were DRV-naive with a median of one DRV resistance mutation. At Weeks 4, 12, and 24, respectively, 63%, 81%, and 94% of patients achieved HIV1-RNA less than 50 copies/mL. Median CD4 cell count increased 30, 73, and 103 cells/mL at Weeks 4, 12, and 24, respectively. No patient had adverse events leading to discontinuation of the regimen. Conclusion: The combination of raltegravir, ETR, and DRV/r was a highly effective and well-tolerated antiretroviral salvage regimen in patients infected with multidrug-resistant HIV-1.

Lipoatrophy is the most stigmatizing side effect of stavudine therapy. We assessed the long-term effects of replacing stavudine with tenofovir in HIV-infected patients with lipoatrophy.Prospective switch study. Sixty-two clinically stable patients with antiretroviral therapy (ART) containing stavudine, HIV-1 RNA50 copies/mL, and lipoatrophy at least in the face on physical examination were included. All patients switched from stavudine to tenofovir without changing any other drug. Objective (malar ultasonography, bioelectrical impedance analysis) and subjective measures of lipoatrophy were assessed.Median age at baseline was 40 years, 44 patients (71%) were male, and median time on stavudine was 4.8 years. Median malar fat thickness increased 0.8 mm (25%) 24 months after switching. Total fat mass increased 3.9 kg (21%). Plasma lactate levels decreased significantly, mainly in patients with baseline hyperlactatemia (from 3.05 to 1.19 mmol/L). Significant improvement in total cholesterol (-12%), triglycerides (-31%), and total cholesterol/HDL cholesterol ratio (-11%) was observed at Month 24.In this study, switching from stavudine to tenofovir maintained durable virologic suppression when the HAART regimen included a protease inhibitor or a non-nucleoside reverse transcriptase inhibitor, led to a slow improvement of lipoatrophy, and improved the lipid profile and lactate levels with excellent tolerability. These results support the proactive change of stavudine to tenofovir.

Background Emerging data suggest that higher ribavirin (RBV) exposure could improve early hepatitis C virus (HCV) response. Furthermore, interindividual RBV bioavailability shows high variation, and dose-limiting haemolytic anaemia is a common adverse event. Therefore, it has been suggested that monitoring RBV serum levels could be used to drive dose modification and to optimize management of HCV-infected patients receiving combination treatment. Methods To assess the effect of RBV serum levels on HCV RNA clearance at week 4 and 12 of treatment, and to determine the correlation between RBV serum concentration and haemoglobin decrease, RBV trough levels were measured by HPLC in stored serum samples obtained from 94 HCV-HIV-coinfected patients at week 4 and 12 of treatment with peginterferon-α2b (1.5 μg/kg/weekly) plus ribavirin (800–1,200 mg/day). Results The median RBV levels increased from 1.70 μg/ml at week 4 to 1.97 μg/ml at week 12 of treatment ( P=0.001) and were independently predicted by weight-adjusted dose of RBV and co-administration of tenofovir. Haemoglobin drop was higher among patients who received zidovudine and weakly correlated with RBV level. Although RBV concentration was lower in genotype 1 or 4 HCV-infected patients who cleared the virus at treatment week 4, the ability of this parameter to discriminate between responders and non-responders at treatment week 4 and 12 was poor. Conclusion Intracellular RBV accumulation early in treatment might improve the kinetics of HCV response in difficult to treat patients. Although this hypothesis and the potential interaction between RBV and tenofovir warrant further research, our data do not support RBV serum monitoring as a tool to optimize treatment in HCV-HIV-coinfected patients.