Industry's Giant Leap Into Cellular Therapy: Catalyzing Chimeric Antigen Receptor T Cell (CAR-T) Immunotherapy.

PURPOSE OF REVIEW: We describe the significant technological leap from bench to bedside that was achieved through a strong academic-industry collaboration between dedicated clinicians and researchers at the University of Pennsylvania, the Children's Hospital of Philadelphia, and Novartis to commercialize the chimeric antigen receptor T cell (CAR-T) therapy tisagenlecleucel (CTL019; Kymriah®; Novartis Pharma AG, Basel, Switzerland). RECENT FINDINGS: Tisagenlecleucel was the first CAR-T therapy and the first gene therapy to receive US Food and Drug Administration approval in 2017, with an initial indication for pediatric and young adult patients with relapsed or refractory (r/r) acute lymphoblastic leukemia, followed by approval in May 2018 for a second indication in adult patients with r/r diffuse large B cell lymphoma. Subsequent approvals in the European Union, Switzerland, and Canada soon followed. The tisagenlecleucel success story represents the development and commercialization of a first-of-its-kind personalized cellular therapy with a manufacturing process that supports commercial production and ongoing global clinical trials in a growing number of countries.

Building blocks for institutional preparation of CTL019 delivery.

Chimeric antigen receptor (CAR) T-cell therapy is an investigational immunocellular therapy that reprograms a patient's cytotoxic T cells to engage and eliminate malignant cells. CAR T-cell therapies targeting the CD19 antigen have demonstrated high efficacy in clinical trials for patients with B-cell malignancies and may potentially be available on a broader scale in the future. CAR T-cell therapy begins with the collection of a sufficient number of T cells from a patient's peripheral blood through leukapheresis. Several factors must be considered when patients undergo leukapheresis for CAR T-cell therapy, including age and prior therapies. The leukapheresis material is shipped to a manufacturing facility, followed by return of the CAR T cells to the treatment center. Careful coordination of a multidisciplinary team composed of physicians, nurses, pharmacists and other hospital personnel is critical for the proper care of the patient before, during and after CAR T-cell therapy. CAR T-cell therapy has been associated with adverse events (AEs) such as cytokine release syndrome, which requires rapid attention by the emergency department, intensive care unit and hospital pharmacy. In this review, we discuss several aspects of institutional preparation for leukapheresis, CAR T-cell infusion and AE management based on our experience with clinical trials of the CD19 CAR T-cell therapy CTL019.

Exploratory study on the impact of switching to nilotinib in 18 patients with chronic myeloid leukemia in chronic phase with suboptimal response to imatinib.

BACKGROUND: The phase II, exploratory, open-label Exploring Nilotinib BCR-ABL Effects (ENABL) study [ClinicalTrials.gov identifier: NCT00644878] assessed the impact of switching to nilotinib therapy in patients with chronic myeloid leukemia in chronic phase (CML-CP) who had a suboptimal molecular response with imatinib. METHODS: Patients with CML-CP who had previously achieved a complete cytogenetic response (CCyR), but had a suboptimal molecular response, with frontline imatinib therapy (N = 18) were assigned to receive nilotinib 300 mg twice daily. The primary endpoint was the change in BCR-ABL1 transcript levels from baseline after 12 months; rates of major molecular response (MMR) and safety were also assessed. RESULTS: At 3 months after switching to nilotinib, 10 of 17 (59%) evaluable patients had achieved MMR. At 12 months, 9 of 12 (75%) evaluable patients had achieved MMR, and the median BCR-ABL1 level among all patients remaining in the study was 0.020% on the International Scale (IS), equivalent to a 3.7-log reduction from the standardized IS baseline (primary endpoint). Adverse events (AEs) were typically grade 1/2 and manageable with dose interruptions. A total of three patients experienced serious study drug-related AEs, including pancreatitis, bradycardia, and vertigo. No deaths were reported. CONCLUSIONS: Overall, results from this exploratory study suggest that switching to nilotinib due to suboptimal molecular response with imatinib can result in improved molecular response for patients with CML-CP.

Bone Marker-Directed Dosing of Zoledronic Acid for the Prevention of Skeletal Complications in Patients with Multiple Myeloma: Results of the Z-MARK Study.

PURPOSE: Zoledronic acid (ZOL) given every 3 to 4 weeks can reduce skeletal-related events (SRE) in patients with bone lesions from multiple myeloma. This study evaluated efficacy and safety of less-frequent ZOL dosing based on bone turnover markers in patients with 1 to 2 years of prior bisphosphonate therapy. EXPERIMENTAL DESIGN: Patients received ZOL (4 mg) every 4 or 12 weeks based on urinary N-telopeptide of type 1 collagen (uNTX) levels (every 4 weeks if uNTX ≥50 nmol/mmol creatinine, every 12 weeks if uNTX < 50). RESULTS: Of 121 patients enrolled (mean age, 63.8 years; median follow-up, 21 months), 4 patients started ZOL every 4 weeks and 117 received ZOL every 12 weeks based on uNTX at study entry. All 4 patients who initiated ZOL every 4 weeks switched to every 12 weeks due to decreased uNTX. Thirty-eight of 117 patients who initiated ZOL every 12 weeks switched to ZOL every 4 weeks due to disease progression (n = 20), increased uNTX (n = 14), and SREs (n = 4). Overall SRE incidence was low; 7 (5.8%) and 5 (4.9%) patients experienced an SRE during years 1 and 2, respectively. Mean (SD) SRE rate at year 2 was 0.01 (0.03) per person-year. The 2-year incidence rate for osteonecrosis of jaw was 3.3%. Four deaths were reported, none related to ZOL. CONCLUSIONS: Less frequent ZOL dosing (every 12 weeks over 2 years) maintains a low SRE rate and can be safely administered for up to 4 years.

Correlations between cytogenetic and molecular monitoring among patients with newly diagnosed chronic myeloid leukemia in chronic phase: post hoc analyses of the Rationale and Insight for Gleevec High-Dose Therapy study.

CONTEXT: Although bone marrow (BM) karyotyping has been the standard in monitoring patients with chronic myeloid leukemia, peripheral blood (PB) monitoring methods may be more convenient. OBJECTIVE: To conduct post hoc analyses of the Rationale and Insight for Gleevec High-Dose Therapy study to evaluate correlations between results of cytogenetic testing and molecular monitoring from BM and PB during the first 18 months of high-dose imatinib therapy, and between early and late molecular responses. DESIGN: Newly diagnosed patients with chronic-phase chronic myeloid leukemia received imatinib 400 mg twice daily and were monitored quarterly for up to 18 months. Cytogenetic testing was performed by karyotyping using BM or by fluorescence in situ hybridization using PB. Molecular testing was performed by quantitative reverse transcriptase polymerase chain reaction using BM and PB. RESULTS: Significant pairwise correlations were found between results obtained by karyotyping, fluorescence in situ hybridization, and quantitative reverse transcriptase polymerase chain reaction using PB or BM (all pairwise correlations >0.8; P < .001). At 12 months, cytogenetic response by karyotyping correlated well with response by fluorescence in situ hybridization. A median 2.579-log reduction in BCR-ABL1 level from a standardized baseline correlated with fluorescence in situ hybridization-negative status. Patients with greater than 2-log reduction in BCR-ABL1 level at 3, 6, and 9 months were more likely to achieve major molecular response at 18 months than those with 2-log reduction or less. CONCLUSIONS: Our findings support the feasibility of molecular monitoring using PB and suggest that molecular monitoring conducted at a single reliable reference laboratory can adequately track response without invasive BM testing. Our findings are consistent with other work indicating that early response to imatinib predicts favorable long-term outcome.

Assessment of treatment and monitoring patterns and subsequent outcomes among patients with chronic myeloid leukemia treated with imatinib in a community setting.

INTRODUCTION: Real-world treatment and monitoring patterns have not been well documented among imatinib-treated chronic phase chronic myeloid leukemia (CP-CML) patients. Thus, we evaluated these patterns and responses to imatinib in CP-CML patients. METHODS: This retrospective study, based on the Georgia Cancer Specialists' electronic medical record (EMR) system, identified CP-CML patients initiating treatment with imatinib from 01/01/2002 to 11/01/2011 who were subsequently followed for ≥6 months. RESULTS: A total of 177 patients met the study criteria. Imatinib dose modification occurred in 59 patients (33%). Rates of treatment interruption, discontinuation, and switching to another therapy were 16%, 24%, and 23%, respectively. Of 27 patients discontinuing imatinib for lack of efficacy, 9 (33%) had initial dose escalation; 26 patients (96%) eventually switched to a second-generation tyrosine kinase inhibitor. By 3 months, 168 patients remained on imatinib, of whom 96 (57%) had undergone cytogenetic and/or molecular testing. The frequency of response monitoring fluctuated over time, with rates as high as 28% for cytogenetic and 69% for molecular testing. Cumulative response rates steadily increased; 18 month rates were 47% for complete cytogenetic response and 26% for major or complete molecular response. There were no cases of progression and/or death among 38 patients who were regularly monitored for molecular response within the first 12 months of imatinib. Ten of 98 patients (10%) not regularly monitored had progressed or died. CONCLUSIONS: Almost one-third of patients initiating imatinib for CP-CML required dose modification, treatment interruption, or discontinuation. Opportunities for improved monitoring in this setting were identified. Limitations include those inherent to retrospective analyses based on EMR and the uncertain extrapolability of the results.

Association between molecular monitoring and long-term outcomes in chronic myelogenous leukemia patients treated with first line imatinib.

OBJECTIVE: Molecular monitoring using quantitative polymerase chain reaction (qPCR) of BCR-ABL mRNA transcripts using the international scale (IS) is recommended by the National Comprehensive Cancer Network and the European LeukemiaNet for patients with chronic myelogenous leukemia in chronic phase (CML-CP). This study assessed the impact of the frequency of qPCR testing on progression-free survival (PFS). RESEARCH DESIGN AND METHODS: This retrospective chart review of 402 CML-CP patients on first line imatinib therapy, performed by 38 community-based US physicians, analyzed the impact of the frequency of molecular monitoring on the risk of progression and PFS. MAIN OUTCOME MEASURES: Time to progression and progression-free survival. RESULTS: Over the 3 year study, 13.2% of patients did not have any qPCR monitoring and 46.3% had 3-4 qPCR tests per year; 5.7% of CML-CP patients progressed to accelerated/blast phase or died. Compared to patients with no qPCR monitoring, those with 3-4 qPCR tests per year had a lower risk of progression (HR = 0.085; p = 0.001) and longer PFS (HR = 0.088; p = 0.001) after adjusting for potential confounders, as did those patients with 1-2 qPCR tests per year (both p < 0.02). Results were consistent after adjusting for Sokal score when available. CONCLUSION: This is the first study to document the clinical impact of frequent molecular monitoring, and the findings underscore the importance of regular molecular monitoring in delivering quality care for CML. These findings could be subject to unobserved confounders.

Monitoring and switching patterns of patients with chronic myeloid leukemia treated with imatinib in community settings: a chart review analysis.

OBJECTIVES: Monitoring treatment response is an integral part of chronic myeloid leukemia (CML) treatment. The guidelines recommend regular monitoring using standard methods (e.g., real-time quantitative polymerase chain reaction based on the international scale for molecular response) and treatment adjustment when failure is detected among patients treated with imatinib. The objective of this study was to assess the real-world monitoring and therapy adjustment in this patient population in the US. METHODS: Twenty-nine physicians from community practices across the US participated in an online chart review. Adult patients with chronic phase CML who initiated imatinib as first-line therapy during 2006-2010 were selected. Information was collected up to 36 months after imatinib initiation, including response monitoring, response status, and therapy adjustment upon treatment failure. RESULTS: The study included 297 eligible patients. By 18 months, 47% of patients had received cytogenetic response assessment continuously as recommended by the guidelines. The corresponding proportion was 39% for continuous molecular response assessment. Among patients who experienced treatment failure by 18 months, only 14%-38% of patients switched to a second-generation tyrosine kinase inhibitor as recommended by the National Comprehensive Cancer Network and the European Leukemia Net guidelines. LIMITATIONS: Major limitations included limited generalizability and the inability to accurately assess molecular response due to the variations in testing methods during the study period. CONCLUSIONS: Based on the guidelines, the rates of treatment monitoring and switching upon failure were low, demonstrating the need for improvement in CML care in community settings in the US.

Final 5-year results of Z-FAST trial: adjuvant zoledronic acid maintains bone mass in postmenopausal breast cancer patients receiving letrozole.

BACKGROUND: Postmenopausal breast cancer (BC) patients receiving adjuvant aromatase inhibitor therapy are at risk of progressive bone loss and fractures. Zoledronic acid inhibits osteoclastic bone resorption, is effective in maintaining bone health, and may therefore be beneficial in this setting. METHODS: Overall, 602 postmenopausal women with early, hormone receptor-positive BC receiving adjuvant letrozole were randomized (301 each group) to receive upfront or delayed-start zoledronic acid (4 mg intravenously every 6 months) for 5 years. The primary endpoint was the change in lumbar spine (LS) bone mineral density (BMD) at month 12. Secondary endpoints included changes in LS BMD, total hip BMD, and bone turnover markers at 2, 3, and 5 years; fracture incidence at 3 years; and time to disease recurrence. RESULTS: At month 61, the adjusted mean difference in LS and total hip BMDs between the upfront and delayed groups was 8.9% and 6.7%, respectively (P < .0001, for both). Approximately 25% of delayed patients received zoledronic acid by month 61. Only 1 patient experienced grade 4 renal dysfunction; no confirmed cases of osteonecrosis of the jaw were reported. Fracture rates (upfront, 28 [9.3%]; delayed, 33 [11%]; P = .3803) and Kaplan-Meier disease recurrence rates (upfront, 9.8 [95% confidence interval (CI), 6.0-10.3]; delayed, 10.5 [95% CI, 6.6-14.4]; P = .6283) were similar at month 61. CONCLUSIONS: Upfront zoledronic acid seems to be the preferred treatment strategy versus delayed administration, as it significantly and progressively increases BMD in postmenopausal women with early BC receiving letrozole for 5 years, and long-term coadministration of letrozole and zoledronic acid is well tolerated.

Comparison of skeletal complications and treatment patterns associated with early vs. delayed zoledronic acid therapy in multiple myeloma.

BACKGROUND: This study retrospectively compared the risks of skeletal-related events (SREs) and zoledronic acid (ZOL) treatment discontinuation associated with early vs. delayed ZOL therapy for patients with symptomatic multiple myeloma (MM). PATIENTS AND METHODS: Data were collected from a physician-administered medical chart review among US patients with a confirmed diagnosis of symptomatic MM treated after 01/01/2002. Early and delayed ZOL therapy were defined, respectively, as initiating ZOL ≤ 60 days (N = 126) vs. > 60 days (N = 186) after the first symptomatic MM diagnosis. Kaplan-Meier analysis with a log-rank test was performed to compare the risk of SREs between the cohorts. Cox proportional hazard modeling compared the risk of SREs associated with early vs. delayed ZOL treatment, controlling for demographic factors, stage of MM, bone health status, and presence of major comorbidities at diagnosis. Time to ZOL discontinuation was evaluated using the Kaplan-Meier method, following patients from the date of ZOL initiation. RESULTS: Time to the first SRE was significantly longer for patients who received early treatment with ZOL (P = .005). At 2 years after diagnosis, the SRE-free rate was 74.6% vs. 56.5% in the early vs. delayed treatment group, respectively. Early ZOL therapy was associated with a significantly lower risk of any SRE (hazard rate [HR] = .625 vs. delayed ZOL therapy; P = .029). At 2 years from ZOL therapy initiation, rates of ZOL discontinuation were 9.6% vs. 16.4% among patients with early vs. delayed therapy, respectively (P < .05). CONCLUSION: Early treatment with ZOL was associated with significantly reduced risks of SREs and with better treatment persistence compared with delayed treatment.

Results of a multicenter open-label randomized trial evaluating infusion duration of zoledronic acid in multiple myeloma patients (the ZMAX trial).

Zoledronic acid, an intravenous (IV) bisphosphonate, is a standard treatment for multiple myeloma (MM) but may exacerbate preexisting renal dysfunction. The incidence of zoledronic acid-induced renal dysfunction may correlate with infusion duration. In this randomized, multicenter, open-label study, 176 patients with MM, at least one bone lesion, and stable renal function with a serum creatinine (SCr) level < 3 mg/dL received zoledronic acid 4 mg (in 250 mL) as a 15- or 30-minute IV infusion every 3-4 weeks. At month 12, 20% (17 patients) in the 15-minute and 16% (13 patients) in the 30-minute arm experienced a clinically relevant but nonsignificant SCr-level increase (P = 0.44). By 24 months, the proportion of patients with a clinically relevant SCr-level increase was similar between arms (15-minute 28% [24 patients] vs 30-minute 27% [23 patients], P = 0.9014). Median zoledronic acid end-of-infusion concentrations were higher with the shorter infusion (15-minute 249 ng/mL vs 30-minute 172 ng/mL), and prolonging the infusion beyond 15 minutes did not influence adverse events related to zoledronic acid. For patients with MM, the safety profile of IV zoledronic acid is similar between those receiving a 15- or 30-minute infusion; therefore, determining the appropriate infusion duration of zoledronic acid should be based on individual patient considerations.

Multicenter, randomized, phase 2 study of zoledronic acid in combination with docetaxel and carboplatin in patients with unresectable stage IIIB or stage IV non-small cell lung cancer.

This study was designed to evaluate the efficacy and safety of combined zoledronic acid and docetaxel/carboplatin in patients with non-small cell lung cancer (NSCLC) as preclinical studies showed synergistic antitumoral activity with bisphosphonates and docetaxel. Patients with inoperable stage IIIB or stage IV NSCLC were randomized 2:1 to receive docetaxel 75mg/m(2) and carboplatin area under the concentration time curve 6 with (Arm A) or without (Arm B) zoledronic acid 4mg every 3 weeks for 6 cycles. Patients responding in Arm A were rerandomized to receive monthly zoledronic acid (maximum: 12 months [Arm A1] or no zoledronic acid [Arm A2]). Patients responding in Arm B entered Arm B1 for follow-up evaluation only. The primary endpoint was the proportion of patients without disease progression; secondary endpoints were time to disease progression (TTP), TTP in bone, best overall response rate, 1-year overall survival (OS) time, and safety; study not powered to detect endpoint differences. Of 150 patients, 98 were randomized to Arm A and 52 to Arm B. In the treatment phase, results were similar between groups in the proportion of patients without disease progression (40.9% vs 38.8%; P=.8096) and median TTP (132d vs 132d; P=.9622). One-year OS times and best overall response rates were 266d vs 206d (P=.4855) and 64.1% vs 72% (P=.3423), respectively; the study was not powered to detect differences. In the follow-up phase, TTP and OS time were similar. Adding zoledronic acid to docetaxel/carboplatin in advanced stage NSCLC patients was well tolerated, but provided little to no effect on disease progression endpoints.

Zoledronic acid effectively prevents aromatase inhibitor-associated bone loss in postmenopausal women with early breast cancer receiving adjuvant letrozole: Z-FAST study 36-month follow-up results.

BACKGROUND: Postmenopausal women with breast cancer receiving adjuvant aromatase inhibitors (AIs) are at risk for accelerated bone loss and subsequent fractures. The ongoing Zometa-Femara Adjuvant Synergy Trial (Z-FAST) is evaluating the efficacy and safety of zoledronic acid in preventing such bone loss. PATIENTS AND METHODS: In this multicenter study, postmenopausal women with early hormone receptor-positive breast cancer receiving adjuvant letrozole were randomized to receive up-front or delayed-start zoledronic acid (ZA; 4 mg intravenously every 6 months) for 5 years. Delayed-start ZA was administered if the lumbar spine (LS) or total hip (TH) T score fell below -2.0 or a nontraumatic fracture occurred. The primary endpoint was to compare the change from baseline in LS bone mineral density (BMD) between groups at month 12; secondary endpoints, measured at other predetermined timepoints, included comparing changes in TH BMD, LS BMD, and markers of bone turnover, fracture incidence, and time to disease recurrence. Herein, we report the results of the 36-month interim analysis. RESULTS: Overall, 301 patients were randomized to each group. At month 36, the absolute difference in mean LS and TH BMDs between the up-front and delayed groups was 6.7% and 5.2%, respectively (P < .0001 for both). Although this study was not designed to show antifracture efficacy, the incidence of fractures was slightly higher in the delayed group (up-front, 17 [5.7%] vs. delayed, 19 [6.3%]) but not statistically significant (P = .8638). Pyrexia (27 [9%] vs. 6 [2%]; P = .0002) and bone pain (39 [13%] vs. 20 [6.7%]; P = .01) were more common in up-front patients; cough (13 [4.3%] vs. 27 [9%]; P = .03) was more common in delayed patients. No severe renal dysfunction or confirmed cases of osteonecrosis of the jaw were reported. Disease recurrence was reported in 9 up-front (3.0%) and 16 delayed (5.3%) patients (Kaplan-Meier analysis, P = .127), with an absolute decrease of 2.3%. CONCLUSION: Up-front ZA more effectively prevents AI-associated bone loss in postmenopausal women with early breast cancer than delaying therapy until substantial bone loss or fracture occurs.

Integrated analysis of zoledronic acid for prevention of aromatase inhibitor-associated bone loss in postmenopausal women with early breast cancer receiving adjuvant letrozole.

BACKGROUND: The interim (12-month) results of two similarly designed, ongoing studies (the Zometa-Femara Adjuvant Synergy Trials [Z-FAST and ZO-FAST]) suggest that zoledronic acid (4 mg intravenously every 6 months) when initiated with adjuvant letrozole increases bone mineral density (BMD) of the lumbar spine (LS) in postmenopausal women with early-stage breast cancer compared with patients who receive zoledronic acid only when bone loss became clinically significant or a fragility fracture occurred. METHODS: An integrated analysis was performed to maximize the value of the large pool of data from the two studies in answering clinically relevant questions. The primary objective was to compare the change in LS BMD at month 12. Secondary objectives included comparing (a) the change in total hip (TH) BMD, (b) changes in bone turnover marker concentrations, (c) time to disease recurrence, and (d) safety at month 12. FINDINGS: The integrated analysis included 1,667 patients. At month 12, LS BMD was 5.2% higher in the upfront group than in the delayed group; TH BMD was 3.5% higher. N-telopeptide and bone-specific alkaline phosphatase concentrations decreased by 21.3% and 12.8% in the upfront group and increased by 21.7% and 24.9% in the delayed group, respectively (p < .0001 for intergroup comparisons). Fewer patients receiving upfront zoledronic acid experienced disease recurrence than patients in the delayed group-seven patients (0.84%) versus 17 patients (1.9%) (p = .0401). Fracture rates were similar. No confirmed osteonecrosis of the jaw was reported. CONCLUSIONS: The results of this analysis strengthen the statistical validity of the preliminary results of the Z-FAST and ZO-FAST studies, showing that upfront zoledronic acid prevents aromatase inhibitor-associated bone loss more effectively than delayed-start zoledronic acid in postmenopausal women with early-stage breast cancer receiving letrozole. Additionally, disease recurrence appears to be lower with upfront zoledronic acid, but further follow-up is needed to confirm these interim results.

Cyclosporin a and tacrolimus (FK506) differentially alter T-cell receptor expression in vivo.

Cyclosporin A (CSA) and tacrolimus (FK506) are two common immunosuppressive agents used post blood and marrow transplantation. Despite similarity in their accepted modes of action, we observed polarized effects of CSA and FK506 on the in vivo human T cell repertoire. To determine the possible mechanism for this difference, the effects of CSA and FK506 on cell viability, cell proliferation, interleukin-2 production, and calcineurin inhibition were determined in vitro. Our data suggest that a secondary mechanism of action exists for the different T-cell repertoire induced by exposure to CSA and FK506.

Complete response after high-dose chemotherapy and autologous hemopoietic stem cell transplatation in metastatic breast cancer results in survival benefit.

Metastatic breast cancer is an incurable disease even with high-dose chemotherapy (HDC) and autologous hematopoietic stem cell transplantation (ASCT). Even though phase III studies have not shown a survival advantage for this group as a whole, it is possible that a small subset of patients may benefit from HDC/ASCT with careful patient selection. A total of 198 patients from three different institutions were treated with HDC/ASCT. After complete staging, patients with central nervous system or bone marrow involvement were excluded. The HDC regimen consisted of: Carboplatin 600 mg/m(2) IV infusion over 48 hours, Thiotepa 300 mg/m(2) IV infusion over 2 hours, and Cytoxan 60 mg/kg IV infusion given over 2 hours x3 days. The median age at the time of transplant was 46 (24-62) years and median follow-up was 20 months. Hormone receptor status was known in 148 patients, of whom 84 had estrogen receptor (ER) and/or progestrone receptor (PgR)-positive tumors. Eighty patients had no evidence of disease at the time of HDC/ASCT (CR1). At the completion of HDC and ASCT, complete responses (CR) were seen in an additional 57 patients (CR2). Using Kaplan-Meier analysis, the median relapse-free survival (RFS) for the entire group was 15 months and overall survival (OS) was 27 months. The patients in CR1 had a median RFS and OS of 20.7 and 50.6 months, respectively. This was very similar to the RFS and OS in patients achieving CR2 after HDC/ASCT (p < 0.001; median: 19 and 40 months, respectively). In contrast, the patients with persistent residual disease had an RFS and OS of 7 and 12 months (p < 0.001). These data show that patients achieving a CR after HDC/ASCT have a better relapse-free and OS, when compared to patients with persistent residual disease after HDC/ASCT. This study suggests that a subset of patients with residual metastatic breast cancer after standard chemotherapy can achieve CR with HDC and ASCT which may result in better long-term outcome.

An improved methodology to detect human T cell receptor beta variable family gene expression patterns.

Comprehensive gene expression analysis of the T cell receptor repertoire of an individual can be very useful in evaluating the immune response in a variety of conditions. Antibody-based analysis methods can detect approximately 60% of the human T cell receptor beta variable (TCRBV) proteins, while gene expression analysis, primarily through employment of the polymerase chain reaction (PCR), has had somewhat greater success in the detection of additional TCRBV families. Many of these previous PCR methods, however, have been unable to detect all 91 alleles of the human TCRBV genes. This is primarily due to either deficiencies in the amplification of all of the variable beta families, subfamilies, and alleles, or the prior lack of a systematic classification of the TCR variable family gene segment sequences. We describe here a real-time reverse transcription polymerase chain reaction-based method, which allows efficient automation and integration of amplification, detection, and analysis with sequence-specific detection of all T cell receptor beta variable gene families, subfamilies, and alleles. This method, which in itself contributes significant improvements over existing technologies through its comprehensiveness and efficiency, also functions independently of variables such as sample source and sample processing and has the ability to run on multiple real-time PCR platforms, affording one the implementation of personal preferences.

Vanadate stimulates monocytic differentiation activity of IL-6 by enhancing actin filament polymerization in HL-60 cells.

IL-6 is involved in the control of differentiation of the acute promyelocytic leukemia cell line, HL-60. However, the participation of protein tyrosine phosphatase (PTP) in the monocytic differentiation activity of IL-6 at low concentrations has not been well clarified. In the present study, we demonstrate that IL-6 (10 ng/ml) alone increased cell growth without differentiation. In the presence of vanadate (10 microM), a PTP inhibitor, IL-6 induced pronounced G0/G1 cell cycle arrest; this effect was associated with CD14+ monocytic differentiation as well as F-actin filament polymerization. Furthermore, vanadate potentiated IL-6-signaling pathway by increasing the tyrosine phosphorylated levels of STAT3 (Tyr705), and Lyn. Such induction of Lyn kinase activity resulted from hypophosphorylated tyrosine (Tyr507) at its negative regulatory site. Vanadate also cooperated with IL-6 to form a protein complex containing Lyn and an actin-associated protein, AFAP110. A complex between Lyn and AFAP110 may serve to regulate F-actin filament polymerization. In conclusion, inhibition of PTP by vanadate promotes hematopoietic differentiation activity of IL-6 through modulating multiple signalings, particularly actin filament polymerization.