A first-in-human study investigating biodistribution, safety and recommended dose of a new radiolabeled MAb targeting FZD10 in metastatic synovial sarcoma patients.

BACKGROUND: Synovial Sarcomas (SS) are rare tumors occurring predominantly in adolescent and young adults with a dismal prognosis in advanced phases. We report a first-in-human phase I of monoclonal antibody (OTSA-101) targeting FZD10, overexpressed in most SS but not present in normal tissues, labelled with radioisotopes and used as a molecular vehicle to specifically deliver radiation to FZD10 expressing SS lesions. METHODS: Patients with progressive advanced SS were included. In the first step of this trial, OTSA-101 in vivo bio-distribution and lesions uptake were evaluated by repeated whole body planar and SPECT-CT scintigraphies from H1 till H144 after IV injection of 187 MBq of 111In-OTSA-101. A 2D dosimetry study also evaluated the liver absorbed dose when using 90Y-OTSA-101. In the second step, those patients with significant tumor uptake were randomized between 370 MBq (Arm A) and 1110 MBq (Arm B) of 90Y-OTSA-101 for radionuclide therapy. RESULTS: From January 2012 to June 2015, 20 pts. (median age 43 years [21-67]) with advanced SS were enrolled. Even though 111In-OTSA-101 liver uptake appeared to be intense, estimated absorbed liver dose was less than 20 Gy for each patient. Tracer intensity was greater than mediastinum in 10 patients consistent with sufficient tumor uptake to proceed to treatment with 90Y-OTSA-101: 8 were randomized (Arm A: 3 patients and Arm B: 5 patients) and 2 were not randomized due to worsening PS. The most common Grade ≥ 3 AEs were reversible hematological disorders, which were more frequent in Arm B. No objective response was observed. Best response was stable disease in 3/8 patients lasting up to 21 weeks for 1 patient. CONCLUSIONS: Radioimmunotherapy targeting FZD10 is feasible in SS patients as all patients presented at least one lesion with 111In-OTSA-101 uptake. Tumor uptake was heterogeneous but sufficient to select 50% of pts. for 90Y-OTSA-101 treatment. The recommended activity for further clinical investigations is 1110 MBq of 90Y-OTSA-101. However, because of hematological toxicity, less energetic particle emitter radioisopotes such as Lutetium 177 may be a better option to wider the therapeutic index. TRIAL REGISTRATION: The study was registered on the NCT01469975 website with a registration code NCT01469975 on November the third, 2011.

Mutational Profile of Metastatic Breast Cancers: A Retrospective Analysis.

BACKGROUND: Major advances have been achieved in the characterization of early breast cancer (eBC) genomic profiles. Metastatic breast cancer (mBC) is associated with poor outcomes, yet limited information is available on the genomic profile of this disease. This study aims to decipher mutational profiles of mBC using next-generation sequencing. METHODS AND FINDINGS: Whole-exome sequencing was performed on 216 tumor-blood pairs from mBC patients who underwent a biopsy in the context of the SAFIR01, SAFIR02, SHIVA, or Molecular Screening for Cancer Treatment Optimization (MOSCATO) prospective trials. Mutational profiles from 772 primary breast tumors from The Cancer Genome Atlas (TCGA) were used as a reference for comparing primary and mBC mutational profiles. Twelve genes (TP53, PIK3CA, GATA3, ESR1, MAP3K1, CDH1, AKT1, MAP2K4, RB1, PTEN, CBFB, and CDKN2A) were identified as significantly mutated in mBC (false discovery rate [FDR] < 0.1). Eight genes (ESR1, FSIP2, FRAS1, OSBPL3, EDC4, PALB2, IGFN1, and AGRN) were more frequently mutated in mBC as compared to eBC (FDR < 0.01). ESR1 was identified both as a driver and as a metastatic gene (n = 22, odds ratio = 29, 95% CI [9-155], p = 1.2e-12) and also presented with focal amplification (n = 9) for a total of 31 mBCs with either ESR1 mutation or amplification, including 27 hormone receptor positive (HR+) and HER2 negative (HER2-) mBCs (19%). HR+/HER2- mBC presented a high prevalence of mutations on genes located on the mechanistic target of rapamycin (mTOR) pathway (TSC1 and TSC2) as compared to HR+/HER2- eBC (respectively 6% and 0.7%, p = 0.0004). Other actionable genes were more frequently mutated in HR+ mBC, including ERBB4 (n = 8), NOTCH3 (n = 7), and ALK (n = 7). Analysis of mutational signatures revealed a significant increase in APOBEC-mediated mutagenesis in HR+/HER2- metastatic tumors as compared to primary TCGA samples (p < 2e-16). The main limitations of this study include the absence of bone metastases and the size of the cohort, which might not have allowed the identification of rare mutations and their effect on survival. CONCLUSIONS: This work reports the results of the analysis of the first large-scale study on mutation profiles of mBC. This study revealed genomic alterations and mutational signatures involved in the resistance to therapies, including actionable mutations.

Higher percentage of CD34 + CD38- cells detected by multiparameter flow cytometry from leukapheresis products predicts unsustained complete remission in acute myeloid leukemia.

Relapse in acute myeloid leukemia (AML) after chemotherapy reflects the persistence of resistant leukemia stem cells (LSCs). These cells have been described in the CD34 + CD38- cell fraction. Leukapheresis products were harvested in 123 patients in morphological complete remission and analyzed by multiparameter flow cytometry. The CD34 + CD38- cell population showed a prognostic impact on survival. Median event-free survival (EFS) was 8.2 months (3-year EFS: 29%) for those with a higher percentage of CD34 + CD38- versus 91.9 months (3-year EFS: 62%) for those with a lower percentage for the entire cohort. These differences were confirmed in patients undergoing autologous stem cell transplant, with median EFS of 7.3 months versus 91.1 months (3-year EFS: 31% vs. 70%). Higher proportions of CD34 + CD38- cells were associated with adverse cytogenetics and with earlier relapses. Higher percentages of CD34 + CD38- cells in apheresis products reflect inadequate in vivo purging and reliably distinguish samples enriched in LSCs from those involving mainly normal cells.

Copper isotope effect in serum of cancer patients. A pilot study.

The isotope effect describes mass-dependent variations of natural isotope abundances for a particular element. In this pilot study, we measured the (65)Cu/(63)Cu ratios in the serums of 20 breast and 8 colorectal cancer patients, which correspond to, respectively, 90 and 49 samples taken at different times with molecular biomarker documentation. Copper isotope compositions were determined by multiple-collector inductively coupled plasma mass spectrometry (MC-ICP-MS). When compared with the literature data from a control group of 50 healthy blood donors, abundances of Cu isotopes predict mortality in the colorectal cancer group with a probability p = 0.018. For the breast cancer patients and the group of control women the probability goes down to p = 0.0006 and the AUC under the ROC curve is 0.75. Most patients considered in this preliminary study and with serum δ(65)Cu lower than the threshold value of -0.35‰ (per mil) did not survive. As a marker, a drop in δ(65)Cu precedes molecular biomarkers by several months. The observed decrease of δ(65)Cu in the serum of cancer patients is assigned to the extensive oxidative chelation of copper by cytosolic lactate. The potential of Cu isotope variability as a new diagnostic tool for breast and colorectal cancer seems strong. Shifts in Cu isotope compositions fingerprint cytosolic Cu chelation by lactate mono- and bidentates. This simple scheme provides a straightforward explanation for isotopically light Cu in the serum and isotopically heavy Cu in cancer cells: Cu(+) escaping chelation by lactate and excreted into the blood stream is isotopically light. Low δ(65)Cu values in serum therefore reveal the strength of lactate production by the Warburg effect.

Patients with metastatic breast cancer leading to CD4+ T cell lymphopaenia have poor outcome.

BACKGROUND: Low lymphocyte count is a prognostic factor in cancer patients including metastatic breast cancer patients (MBC) but the relative role of each lymphocyte subtype is unclear in MBC. METHODS: The impact of lymphocyte subsets was analysed in two prospective MBC patients' cohorts. Cohort A patients (n=103) were included before the first line of chemotherapy and cohort B patients (n=101) were included after at least one line of chemotherapy. Extensive phenotypic analyses were performed on fresh whole blood. Plasma cytokines levels were measured using commercially available Luminex-based multiplex kits. Prognostic value of lymphocyte subsets and circulating cytokines was analysed. RESULTS: In both cohorts, severe lymphopaenia (<0.7 Giga/L) correlated with poor overall survival (OS) (median OS: 6.6 months versus 21.7 months in cohort A and 4.5 versus 9 months in cohort B). CD8(+), CD19(+) and CD56(+) T cell counts had no significant prognostic value for OS. After stratification (≤0.2, [0.20-0.45], >0.45 Giga/L), CD4 lymphopaenia appeared to be correlated with poor OS in both cohorts. Furthermore, severe CD4(+) lymphopaenia (≤0.2 Giga/L) was strongly correlated with poor OS in both cohorts (1.2 months versus 24.9 months in cohort A and 5.7 versus 13.1 months in cohort B). In multivariate analysis, after stratification CD4(+) lymphopaenia appeared to be an independent prognostic factor for OS in both cohorts. CD4(+) lymphopaenia correlated with low plasmatic levels of CCL22 that might directly contribute to CD4(+) lymphopaenia. CONCLUSIONS: CD4(+) lymphopaenia was associated with reduced OS in MBC patients regardless of the chemotherapy line. Decreased levels of plasmatic CCL22 may contribute to CD4(+) lymphopaenia.

CD4 lymphopenia to identify end-of-life metastatic cancer patients.

BACKGROUND: Cancer patients with CD4 lymphopenia have an increased risk of severe toxicity after administration of cytotoxic chemotherapy. The impact of CD4 lymphopenia on long term overall survival (OS) of cancer patients was explored in this work. PATIENTS AND METHODS: The first prospective series (test series) included 219 patients with solid tumours, lymphomas or myelomas receiving chemotherapy within an oncology department in 1999 and 2000. A phenotypic analysis of lymphocyte subsets by flow cytometry was performed before chemotherapy on day 1. The prognostic value of total, CD4, CD8 and CD56 lymphocyte count for OS was tested in a multivariate analysis. The prognostic value of low CD4 counts was then tested in a validation series of 269 patients with metastatic solid tumours in second line treatment included in a prospective observational study in the Centre Leon Berard. RESULTS: In the test series, all patients with metastatic cancers and CD4 lymphopenia ≤ 200/µL (12% of metastatic patients) died within 18 months with a median OS of 5.9 months. CD4 count was an independent prognostic factor for OS and PFS in multivariate analysis. In the validation series, 83 (30%) of patients had CD4 count ≤ 200/µL: their median overall survival was 3.9 months with an 18-month survival rate of 6%. CD4 count was also an independent prognostic factor for overall survival in this series. CONCLUSIONS: CD4 lymphopenia <200/µL is frequent in advanced cancer patients and associated with a very short life expectancy. These patients should be proposed for specific treatment and research approaches.

Lymphopenia combined with low TCR diversity (divpenia) predicts poor overall survival in metastatic breast cancer patients.

Lymphopenia (< 1Giga/L) detected before initiation of chemotherapy is a predictive factor for death in metastatic solid tumors. Combinatorial T cell repertoire (TCR) diversity was investigated and tested either alone or in combination with lymphopenia as a prognostic factor at diagnosis for overall survival (OS) in metastatic breast cancer (MBC) patients. The combinatorial TCR diversity was measured by semi quantitative multi-N-plex PCR on blood samples before the initiation of the first line chemotherapy in a development (n = 66) and validation (n = 67) MBC patient cohorts. A prognostic score, combining lymphocyte count and TCR diversity was evaluated. Univariate and multivariate analyses of prognostic factors for OS were performed in both cohorts. Lymphopenia and severe restriction of TCR diversity called "divpenia" (diversity ≤ 33%) were independently associated with shorter OS. Lympho-divpenia combining lymphopenia and severe divpenia accurately identified patients with poor OS in both cohorts (7.6 and 10.6 vs 24.5 and 22.9 mo). In multivariate analysis including other prognostic clinical factors, lympho-divpenia was found to be an independent prognostic factor in the pooled cohort (p = 0.005) along with lack of HER2 and hormonal receptors expression (p = 0.011) and anemia (p = 0.009). Lympho-divpenia is a novel prognostic factor that will be used to improve quality of MBC patients' medical care.

Immunologic and clinical effects of injecting mature peptide-loaded dendritic cells by intralymphatic and intranodal routes in metastatic melanoma patients.

PURPOSE: A phase I/II trial was conducted to evaluate clinical and immunologic responses after intralymphatic and intranodal injections of mature dendritic cells. EXPERIMENTAL DESIGN: Fourteen patients with a metastatic melanoma received matured dendritic cells, loaded with Melan-A/MART-1 and/or NA17-A peptides and keyhole limpet hemocyanin. The cells were matured overnight with Ribomunyl, a toll-like receptor ligand, and IFN-gamma, which ensured the production of high levels of interleukin-12p70. Dendritic cells were injected at monthly intervals, first into an afferent lymphatic and then twice intranodally. Immunologic responses were monitored by tetramer staining of circulating CD8(+) lymphocytes and delayed-type hypersensitivity tests. RESULTS: Dendritic cell vaccination induced delayed-type hypersensitivity reactivity toward NA17-A-pulsed, keyhole limpet hemocyanin-pulsed, and Melan-A-pulsed dendritic cells in 6 of 10, 4 of 11, and 3 of 9 patients, respectively. Four of the 12 patients analyzed by tetramer staining showed a significantly increased frequency of Melan-A-specific T cells, including one patient vaccinated only with NA17-A-pulsed dendritic cells. Furthermore, 2 of the 12 analyzed patients had a significant increase of NA17-A-specific T cells, including one immunized after an optional additional treatment course. No objective clinical response was observed. Two patients were stabilized at 4 and 10 months and three patients are still alive at 30, 39, and 48 months. CONCLUSIONS: Injections into the lymphatic system of mature peptide-loaded dendritic cells with potential TH1 polarization capacities did not result in marked clinical results, despite immunologic responses in some patients. This highlights the need to improve our understanding of dendritic cell physiology.

Cryopreservation with hydroxyethylstarch (HES) + dimethylsulfoxide (DMSO) gives better results than DMSO alone.

This study compares the cryopreservation of PBSC by means of HES 3% + DMSO 5% ina mechanical freezer with the cryopreservation of PBSC by means of DMSO 10% in an automated rate controlled freezer. The cells harvested from the same patient were divided in two equal parts and frozen according to both methods. Cryopreservation with HES +DMSO is better than DMSO alone in terms of viability (p<0.001) and Granulocyte Macrophage Colony Forming Unit (GM-CFU) activity (p<0.05). The freezing curves differ essentially in the slope down after the surfusion peak. However, surfusion peaks are similar in both conditions. The relevance of mechanical freezing is the saving of time and the safety of such a procedure.

CD4 lymphopenia as a risk factor for febrile neutropenia and early death after cytotoxic chemotherapy in adult patients with cancer.

BACKGROUND: Lymphopenia is frequently observed in patients with cancer and correlates with the risk of febrile neutropenia and early death after chemotherapy. The phenotype of the depleted lymphocyte populations was investigated in the current study. METHODS: Peripheral blood lymphocyte subsets (CD3, CD4, CD8, CD19, CD56) were quantified on Day 1 using fluorescence-activated cell sorting in a prospective study of 213 patients with cancer treated with chemotherapy in a single oncology ward during 12 months. Correlations between lymphocyte phenotype, clinical characteristics, and the risk of febrile neutropenia and early death within 31 days after chemotherapy were investigated in univariate and multivariate analyses. RESULTS: Total lymphocyte count and CD3, CD4, and CD8 lymphocyte subsets were significantly lower in patients who experienced febrile neutropenia. Total lymphocyte count and CD3, CD4, CD8, CD19, and CD56 lymphocyte subsets were significantly lower in patients who died within 31 days after chemotherapy. Using logistic regression, CD4 lymphopenia (< 450/muL; odds ratio [OR] = 2.9, 95% confidence interval [CI] = 1.5-5.9) and the dose of chemotherapy (OR = 3,9, 95% CI = 2.0-7.8) were both identified as independent risk factors for febrile neutropenia. Fifty-four percent of patients with both risk factors experienced febrile neutropenia. CD4 lymphocyte count < 450/muL was also an independent risk factor for early death (OR = 7.7, 95% CI = 1.7-35). Thirteen percent of patients with a CD4 lymphocyte count