Ronald T. Mitsuyasu

Effect of Recombinant Human Granulocyte–Macrophage Colony-Stimulating Factor on Myelopoiesis in the Acquired Immunodeficiency Syndrome

We administered recombinant (biosynthetic) human granulocyte-macrophage colony-stimulating factor (GM-CSF) to 16 patients with the acquired immunodeficiency syndrome (AIDS) and leukopenia (2225 +/- 614 cells per microliter [mean +/- SD]). Each patient first received a single intravenous dose; 48 hours later a 14-day continuous intravenous infusion of the agent was begun. The doses used were 1.3 X 10(3) (n = 4), 2.6 X 10(3) (n = 4), 5.2 X 10(3) (n = 4), 1.0 X 10(4) (n = 3), or 2.0 X 10(4) (n = 1) U per kilogram of body weight per day. Administration of recombinant GM-CSF resulted in dose-dependent increases in circulating leukocytes and in increases in circulating neutrophils, eosinophils, and monocytes. The peak leukocyte count ranged from 4575 +/- 2397 cells per microliter at the lowest dose, to 48,700 in the patient receiving the highest dose. Mild side effects--low-grade fever, myalgia, phlebitis, and flushing--were observed in some patients; there were no life-threatening toxic reactions. Our data demonstrate that recombinant human GM-CSF is well tolerated and biologically active in leukopenic patients with AIDS. Strategies to increase the number and function of circulating leukocytes may reduce the morbidity and mortality of infections in these and other patients with leukopenia.

Initial increase in blood CD4+ lymphocytes after HIV antiretroviral therapy reflects redistribution from lymphoid tissues

Previous studies proposed a dynamic, steady-state relationship between HIV-mediated cell killing and T-cell proliferation, whereby highly active antiretroviral therapy (HAART) blocks viral replication and tips the balance toward CD4(+) cell repopulation. In this report, we have analyzed blood and lymph node tissues obtained concurrently from HIV-infected patients before and after initiation of HAART. Activated T cells were significantly more frequent in lymph node tissue compared with blood at both time points. Ten weeks after HAART, the absolute number of lymphocytes per excised lymph node decreased, whereas the number of lymphocytes in the blood tended to increase. The relative proportions of lymphoid subsets were not significantly changed in tissue or blood by HAART. The expression levels of mRNA for several proinflammatory cytokines (IFN-gamma, IL-1beta, IL-6, and macrophage inflammatory protein-1alpha) were lower after HAART. After therapy, the expression of VCAM-1 and ICAM-1 -- adhesion molecules known to mediate lymphocyte sequestration in lymphoid tissue -- was also dramatically reduced. These data provide evidence suggesting that initial increases in blood CD4(+) cell counts on HAART are due to redistribution and that this redistribution is mediated by resolution of the immune activation that had sequestered T cells within lymphoid tissues.

Decade-Long Safety and Function of Retroviral-Modified Chimeric Antigen Receptor T Cells

Adoptively transferred chimeric antigen receptor T cells have stable stem cell–like persistence for at least a decade and more than 500 years of patient safety. Standing the Test of Time Retroviral vectors were once the mainstay of gene transfer because they could stably integrate into the host genome. However, some patients in early trials developed leukemia because of insertional mutagenesis. Now, Scholler et al. report that retroviral vector–mediated gene transfer in T cells may not have the same safety concerns, and that these cells may persist over a decade in patients. The authors followed patients from three clinical trials who received T cells transduced with gammaretroviruses carrying a chimeric antigen receptor. They found that these cells were present in recipients over a decade after infusion at levels higher than those induced by standard vaccines. These cells were still functional, had stable levels of engraftment, and did not require host immunosuppression before transplant. Moreover, the authors found no evidence of integration-induced immortalization, with no observable enrichment of integration sites near genes involved in growth control or transformation. Thus, the safety of retroviral vectors may be cell type–specific, opening up engineered T cells as a delivery platform for therapeutics. The success of adoptive T cell gene transfer for treatment of cancer and HIV is predicated on generating a response that is both durable and safe. We report long-term results from three clinical trials to evaluate gammaretroviral vector–engineered T cells for HIV. The vector encoded a chimeric antigen receptor (CAR) composed of CD4 linked to the CD3ζ signaling chain (CD4ζ). CAR T cells were detected in 98% of samples tested for at least 11 years after infusion at frequencies that exceeded average T cell levels after most vaccine approaches. The CD4ζ transgene retained expression and function. There was no evidence of vector-induced immortalization of cells; integration site distributions showed no evidence of persistent clonal expansion or enrichment for integration sites near genes implicated in growth control or transformation. The CD4ζ T cells had stable levels of engraftment, with decay half-lives that exceeded 16 years, in marked contrast to previous trials testing engineered T cells. These findings indicate that host immunosuppression before T cell transfer is not required to achieve long-term persistence of gene-modified T cells. Further, our results emphasize the safety of T cells modified by retroviral gene transfer in clinical application, as measured in >500 patient-years of follow-up. Thus, previous safety issues with integrating viral vectors are hematopoietic stem cell or transgene intrinsic, and not a general feature of retroviral vectors. Engineered T cells are a promising form of synthetic biology for long-term delivery of protein-based therapeutics. These results provide a framework to guide the therapy of a wide spectrum of human diseases.

Interleukin-2 therapy in patients with HIV infection.

BACKGROUND Used in combination with antiretroviral therapy, subcutaneous recombinant interleukin-2 raises CD4+ cell counts more than does antiretroviral therapy alone. The clinical implication of these increases is not known. METHODS We conducted two trials: the Subcutaneous Recombinant, Human Interleukin-2 in HIV-Infected Patients with Low CD4+ Counts under Active Antiretroviral Therapy (SILCAAT) study and the Evaluation of Subcutaneous Proleukin in a Randomized International Trial (ESPRIT). In each, patients infected with the human immunodeficiency virus (HIV) who had CD4+ cell counts of either 50 to 299 per cubic millimeter (SILCAAT) or 300 or more per cubic millimeter (ESPRIT) were randomly assigned to receive interleukin-2 plus antiretroviral therapy or antiretroviral therapy alone. The interleukin-2 regimen consisted of cycles of 5 consecutive days each, administered at 8-week intervals. The SILCAAT study involved six cycles and a dose of 4.5 million IU of interleukin-2 twice daily; ESPRIT involved three cycles and a dose of 7.5 million IU twice daily. Additional cycles were recommended to maintain the CD4+ cell count above predefined target levels. The primary end point of both studies was opportunistic disease or death from any cause. RESULTS In the SILCAAT study, 1695 patients (849 receiving interleukin-2 plus antiretroviral therapy and 846 receiving antiretroviral therapy alone) who had a median CD4+ cell count of 202 cells per cubic millimeter were enrolled; in ESPRIT, 4111 patients (2071 receiving interleukin-2 plus antiretroviral therapy and 2040 receiving antiretroviral therapy alone) who had a median CD4+ cell count of 457 cells per cubic millimeter were enrolled. Over a median follow-up period of 7 to 8 years, the CD4+ cell count was higher in the interleukin-2 group than in the group receiving antiretroviral therapy alone--by 53 and 159 cells per cubic millimeter, on average, in the SILCAAT study and ESPRIT, respectively. Hazard ratios for opportunistic disease or death from any cause with interleukin-2 plus antiretroviral therapy (vs. antiretroviral therapy alone) were 0.91 (95% confidence interval [CI], 0.70 to 1.18; P=0.47) in the SILCAAT study and 0.94 (95% CI, 0.75 to 1.16; P=0.55) in ESPRIT. The hazard ratios for death from any cause and for grade 4 clinical events were 1.06 (P=0.73) and 1.10 (P=0.35), respectively, in the SILCAAT study and 0.90 (P=0.42) and 1.23 (P=0.003), respectively, in ESPRIT. CONCLUSIONS Despite a substantial and sustained increase in the CD4+ cell count, as compared with antiretroviral therapy alone, interleukin-2 plus antiretroviral therapy yielded no clinical benefit in either study. (ClinicalTrials.gov numbers, NCT00004978 [ESPRIT] and NCT00013611 [SILCAAT study].)

Recombinant Alpha-2 Interferon Therapy for Kaposi's Sarcoma Associated with the Acquired Immunodeficiency Syndrome

In a randomized prospective study we tested the toxicity and efficacy of recombinant alpha-2 interferon in the treatment of Kaposis sarcoma associated with the acquired immunodeficiency syndrome. High doses (50 X 10(6) U/m2 body surface area, intravenously) or low doses (1 X 10(6) U/m2, subcutaneously) of recombinant alpha-2 interferon were administered to 20 patients for 5 days/wk, every other week, for four treatment cycles. Therapy was well tolerated subjectively and caused only mild hematologic and hepatic toxicity at both dose levels. No consistent or sustained changes were seen in immunologic variables during or after treatment. Six patients with Kaposis sarcoma, four at high dose and two at low dose, had objective responses (complete or partial) to treatment. However, therapy did not appear to eradicate cytomegalovirus carriage or prevent opportunistic infections related to cytomegalovirus.

Treatment of Donor Bone Marrow with Monoclonal Anti-T-Cell Antibody and Complement for the Prevention of Graft-Versus-Host Disease: A Prospective, Randomized, Double-Blind Trial

The effects of ex-vivo depletion of T lymphocytes from donor bone marrow using a monoclonal anti-T-cell antibody (CT-2) and complement on the outcome of allogeneic bone marrow transplantation was evaluated in a prospective, randomized, double-blind study of 40 patients with leukemia. Patients receiving T-cell-depleted bone marrow had a lower incidence of acute graft-versus-host disease than control patients (3 of 20 compared with 13 of 20; p = 0.004), and mortality due to acute graft-versus-host disease was reduced. Five patients in the T-cell-depletion group developed graft failure; all control patients had sustained engraftment (p less than 0.05). Clinically apparent relapse of leukemia occurred in 7 patients from the T-cell-depletion group and in 2 controls (p, not significant). Cytogenetic evidence of residual leukemia was also detected in the 5 patients with graft failure without overt relapse. Infections and overall survival were similar in the two groups. The effects of T-cell depletion on engraftment and recurrence of leukemia require further evaluation.

Phase 2 gene therapy trial of an anti-HIV ribozyme in autologous CD34+ cells

Gene transfer has potential as a once-only treatment that reduces viral load, preserves the immune system and avoids lifetime highly active antiretroviral therapy. This study, which is to our knowledge the first randomized, double-blind, placebo-controlled, phase 2 cell-delivered gene transfer clinical trial, was conducted in 74 HIV-1-infected adults who received a tat-vpr-specific anti-HIV ribozyme (OZ1) or placebo delivered in autologous CD34+ hematopoietic progenitor cells. There were no OZ1-related adverse events. There was no statistically significant difference in viral load between the OZ1 and placebo group at the primary end point (average at weeks 47 and 48), but time-weighted areas under the curve from weeks 40-48 and 40-100 were significantly lower in the OZ1 group. Throughout the 100 weeks, CD4+ lymphocyte counts were higher in the OZ1 group. This study indicates that cell-delivered gene transfer is safe and biologically active in individuals with HIV and can be developed as a conventional therapeutic product.

Safety and Immunogenicity of the Quadrivalent Human Papillomavirus Vaccine in HIV-1-Infected Men

BACKGROUND Human immunodeficiency virus type 1 (HIV-1)-infected men are at increased risk for anal cancer. Human papillomavirus (HPV) vaccination may prevent anal cancer caused by vaccine types. METHODS AIDS Malignancy Consortium Protocol 052 is a single-arm, open-label, multicenter clinical trial to assess the safety and immunogenicity of the quadrivalent HPV (types 6, 11, 16, and 18) vaccine in HIV-1-infected men. Men with high-grade anal intraepithelial neoplasia or anal cancer by history or by screening cytology or histology were excluded. Men received 0.5 mL intramuscularly at entry, week 8, and week 24. The primary end points were seroconversion to vaccine types at week 28, in men who were seronegative and without anal infection with the relevant HPV type at entry, and grade 3 or higher adverse events related to vaccination. RESULTS There were no grade 3 or greater adverse events attributable to vaccination among the 109 men who received at least 1 vaccine dose. Seroconversion was observed for all 4 types: type 6 (59 [98%] of 60), type 11 (67 [99%] of 68), type 16 (62 [100%] of 62), and type 18 (74 [95%] of 78). No adverse effects on CD4 counts and plasma HIV-1 RNA levels were observed. CONCLUSIONS The quadrivalent HPV vaccine appears safe and highly immunogenic in HIV-1-infected men. Efficacy studies in HIV-1-infected men are warranted. Clinical trials registration. NCT 00513526.

Rituximab plus concurrent infusional EPOCH chemotherapy is highly effective in HIV-associated B-cell non-Hodgkin lymphoma

Rituximab plus intravenous bolus chemotherapy is a standard treatment for immunocompetent patients with B-cell non-Hodgkin lymphoma (NHL). Some studies have suggested that rituximab is associated with excessive toxicity in HIV-associated NHL, and that infusional chemotherapy may be more effective. We performed a randomized phase 2 trial of rituximab (375 mg/m(2)) given either concurrently before each infusional etoposide, vincristine, doxorubicin, cyclophosphamide, and prednisone (EPOCH) chemotherapy cycle or sequentially (weekly for 6 weeks) after completion of all chemotherapy in HIV-associated NHL. EPOCH consisted of a 96-hour intravenous infusion of etoposide, doxorubicin, and vincristine plus oral prednisone followed by intravenous bolus cyclophosphamide given every 21 days for 4 to 6 cycles. In the concurrent arm, 35 of 48 evaluable patients (73%; 95% confidence interval, 58%-85%) had a complete response. In the sequential arm, 29 of 53 evaluable patients (55%; 95% confidence interval, 41%-68%) had a complete response. The primary efficacy endpoint was met for the concurrent arm only. Toxicity was comparable in the 2 arms, although patients with a baseline CD4 count less than 50/microL had a high infectious death rate in the concurrent arm. We conclude that concurrent rituximab plus infusional EPOCH is an effective regimen for HIV-associated lymphoma.

Treatment of acute myelogenous leukemia. A prospective controlled trial of bone marrow transplantation versus consolidation chemotherapy.

In a prospective controlled trial, the relative effectiveness of allogeneic bone marrow transplantation and postremission chemotherapy was assessed for adult patients with acute myelogenous leukemia in first complete remission. Twenty-three patients, 15 to 45 years of age, who had an HLA-identical sibling donor were designated to receive bone marrow transplantation. Forty-four patients who either lacked an HLA-identical sibling or were over 45 years of age were designated to receive intensive consolidation chemotherapy. The actuarial rate of leukemia relapse was significantly lower in the transplantation group than in the chemotherapy group (40 +/- 25% [95% confidence interval] compared with 71 +/- 14%, p = 0.01). Actuarial survival at greater than 4 years was not significantly different (40 +/- 21% compared with 27 +/- 14%, p greater than 0.4). These data show that bone marrow transplantation is more effective than consolidation chemotherapy in preventing leukemia relapse, but overall survival was not improved in this study.