Brolucizumab tras fracaso de aflibercept con terapia fotodinámica en vasculopatía coroidea polipoidea: reporte de un caso / Brolucizumab after failure of aflibercept with photodynamic therapy in polypoidal choroidal vasculopathy: A case report

Describimos un caso de vasculopatía coroidea polipoidea con líquido subretiniano persistente a pesar de múltiples tratamientos intravítreos con bevacizumab, ranibizumab y aflibercept, así como aflibercept asociado a terapia fotodinámica. El paciente alcanzó la resolución completa después de la inyección intravítrea de brolucizumab, pero experimentó una recurrencia del líquido subretiniano 12 semanas después de la suspensión. Brolucizumab podría ser una opción para tratar el líquido subretiniano después del fracaso de otros agentes anti-VEGF asociados con la terapia fotodinámica. (AU)

We describe one case of polypoidal choroidal vasculopathy with persistent subretinal fluid despite multiple treatment with intravitreal bevacizumab, ranibizumab and aflibercept, as well as aflibercept associated with photodynamic therapy. The patient reached complete resolution after intravitreal brolucizumab injection, but experienced recurrence of subretinal fluid 12 weeks after discontinuation. Brolucizumab might be an option in treating subretinal fluid after failure of other anti-VEGF agents associated with photodynamic therapy. (AU)

Brolucizumab after failure of aflibercept with photodynamic therapy in polypoidal choroidal vasculopathy: A case report.

We describe one case of polypoidal choroidal vasculopathy with persistent subretinal fluid despite multiple treatment with intravitreal Bevacizumab, Ranibizumab and Aflibercept, as well as Aflibercept associated with photodynamic therapy. The patient reached complete resolution after intravitreal Brolucizumab injection, but experienced recurrence of subretinal fluid 12 weeks after discontinuation. Brolucizumab might be an option in treating subretinal fluid after failure of other anti-VEGF agents associated with photodynamic therapy.

Effect of induction chemotherapy and tandem cycles of high-dose chemotherapy on outcomes in autologous stem cell transplant for metastatic breast cancer.

We assessed the effect standard-dose induction chemotherapy and tandem cycles of high-dose chemotherapy (HDC) have on outcomes in metastatic breast cancer. One hundred and one women with metastatic breast cancer were enrolled in two non-randomized phase II studies. The first group of 64 patients (induction group) received four cycles of docetaxel 75 mg/m2 and doxorubicin 50 mg/m2. The next 37 patients did not receive induction (no induction group). Both groups received two (tandem) cycles of HDC. Blood-derived stem cells were collected after the first HDC cycle, processed using CD34+ cell selection and then reinfused after the second HDC cycle. Outcomes were compared between the two groups and also to patients participating in the Philadelphia (inter-group) randomized metastatic breast cancer transplant trial (PBT-01). Intent-to-treat analysis revealed no significant differences in complete response rates (37.5% vs 27%; P = 0.20), overall response (75% vs 71%), median progression free survival (PFS) (11.9 vs 8 months; P = 0.24) and overall survival (OS) (>36 vs 25 months; P = 0.16), in the induction vs no induction groups, respectively. Adjusting for differences in known baseline characteristics, induction group patients were found to have significantly longer PFS (P = 0.002), OS (P = 0.01) and more frequent conversion from a partial to complete response (58% vs < or = 13%, P < or = 0.0002) when compared with PBT-01 patients. Induction chemotherapy administered prior to tandem cycles of HDC does not appear to adversely affect outcomes in metastatic breast cancer patients. Outcomes in our induction group also compare favorably with those observed in PBT-01 and warrant further clinical investigation.

A phase II trial evaluating the safety and effectiveness of the AastromReplicell system for augmentation of low-dose blood stem cell transplantation.

To reduce the number of apheresis procedures and maintain the usual rate of hematopoietic recovery in patients treated with high-dose chemotherapy, we studied the effect of adding a small volume of ex vivo expanded bone marrow to low doses of CD34(+) blood stem cells. Thirty-four patients with breast cancer received G-CSF (10 microg/kg/day) priming followed by a limited volume (50-100 ml) bone marrow aspiration and standard 10-liter aphereses. Marrow was expanded ex vivo using the AastromReplicell system and infused along with low doses of blood-derived CD34(+) cells, collected in one apheresis. Thirty-one evaluable patients received a median CD34(+) blood stem cell dose of 0.7 x 10(6)/kg (range, 0.2-2.5) and 4.7 x 10(7) nucleated cells/kg (range, 1.98-8.7) of ex vivo expanded marrow. All patients recovered with normal blood counts and engrafted 500 neutrophils/microl and 20 000 platelets/microl in a median of 10 and 13 days, respectively. Multivariate analysis revealed that, in addition to CD34(+) lineage negative cell quantity, the quantity of stromal progenitors contained in the ex vivo expanded product correlated with engraftment outcome (r = 0.551, P = 0.004). Our results indicate that ex vivo expanded bone marrow is capable of facilitating engraftment when combined with low doses of mobilized blood derived CD34(+) cells.

Tumor cell depletion of peripheral blood progenitor cells using positive and positive/negative selection in metastatic breast cancer.

BACKGROUND: The clinical relevance of tumor cell purging of hematopoietic progenitor cell grafts has yet to be conclusively determined. Therefore, in addition to the demonstration that a method for graft purification is capable of removing an adequate number of tumor cells, it is critical that the procedure has as benign an impact upon the hematopoietic repopulating potential of the graft as possible. We evaluated tumor cell depletion, recovery of CD34(+) cells and post transplant engraftment kinetics as accepted measures of the effectiveness of an immunomagnetic bead (positive and positive/negative) purging methodology. METHODS: The patients received either positive selection (CD34 selection alone) or a combination of positive and negative (CD34 selection followed by breast cancer cell depletion) using the Isolex 300 (automated and semiautomated) devices. Immunocytochemistry was used to determine the degree of breast cancer cell contamination before and after the selection procedures to determine the efficacy of the procedure. CD34 enumeration was employed to evaluate the recovery and purity of the CD34-selected cellular products and engraftment indices (days to absolute neutrophil count (ANC) recovery and platelet count (Plt) recovery and transfusion requirements) were evaluated to determine the safety of the procedure. RESULTS: A total of 130 aphereses was performed on 101 patients. Ten pairs of collections were pooled before selection to increase the likelihood of achieving CD34 dose goals after selection. In all, 100 positive selections and 20 positive/negative selections were performed. Of the 10 (10.4%) ICC-positive preselection samples, 2 products showed persistent contamination after processing. The majority of patients (85.4%) required one selection procedure to achieve an adequate CD34(+) selected cell dose. Median CD34(+) cell recovery was > 50% for positive selection procedures and > 60% for the positive/negative procedures. The dose of CD34(+) cells infused ranged from 0.76 x 10(6) CD34(+) cells/kg to 27.7 x 106 CD34(+) cells/kg. There were no significant delays in neutrophil or platelet recovery or infections between any of the treatment groups. DISCUSSION: CD34 selection alone or in combination with negative selection can result in a significant reduction of contaminating tumor cells in the peripheral blood progenitor cell autograft. Although there was one engraftment failure with the CD34-positive selected cells, transplantation of the selected products after high-dose chemotherapy for metastatic breast cancer did not result in a clinically significant delay in the hematopoietic reconstitutive capacity of the autografts.

Challenges associated with the development, manufacturing, and delivery of cellular medicines.

As the potential of these exciting therapeutics approaches reality, the challenges associated with their discovery, approval, and widespread distribution are just now being realized. The current health care infrastructure is unable to accommodate the complexity of cellular therapy, which exceeds the capability of existing treatment facilities to provide ready access for patients to receive standardized collections for consistent quality of apheresis product and the most consistently acceptable and potent therapeutic agent. The goal is to develop a system for seamless international access to cellular therapies and will involve the unification of the systems for the collection of patient-specific cellular materials, the development of an international network of FDA (and other regulatory agencies) registered facilities, the preservation of patient-specific product control from acquisition to re-infusion, the integration of a sophisticated transportation network to meet the time and environment-critical nature of the raw materials and the finished product, and the development of a comprehensive informatics system to coordinate all of the elements.

Treatment of leukemic relapse following unrelated umbilical cord blood transplantation with interleukin-2: potential for augmenting graft-versus-leukemia and graft-versus-host effects with cytokines.

In comparison to bone marrow, umbilical cord blood has decreased intrinsic immune responsiveness allowing transplantation across HLA barriers with lower rates of graft-versus-host disease. However, laboratory models have also suggested that cord blood may be extremely sensitive to stimulation by cytokines. We report an adult recipient of an ex vivo expanded, HLA-mismatched, unrelated cord blood transplant who experienced a late extramedullary relapse while still in hematologic remission. Despite demonstrating immune tolerance on minimal immunosuppressive agents, a brief course of intravenous interleukin-2 resulted in rapid, aggressive graft-versus-host and graft-versus-leukemia reactions. This case highlights the potential of cytokine immunomodulation following cord blood transplantation, but also suggests caution in stimulating these cells.

High-dose chemotherapy and stem cell transplantation for patients with stage IV breast cancer without clinically evident disease: correlation of CD34+ selection to clinical outcome.

Forty-five patients with metastatic breast cancer without clinically evident disease were treated with thiotepa 750 mg/m2, mitoxantrone 40 mg/m2 and carboplatin 1000 mg/m2 followed by stem cell transplantation to determine the safety and efficacy of CD34+ selection of peripheral blood stem cells. Of these, 15 patients' (group I) stem cells were processed through Baxter Isolex 300 device for CD34+ selection, whereas 30 patients (group II) received unmanipulated stem cells. Toxicity, progression-free survival and survival were compared between these two groups. There was no difference in transfusion requirements, white cell count and platelet recovery and non-hematologic toxicity between the two groups. The survival of patients in group I was 27 months compared to 38 months in group II (P = 0.8). The progression-free survival was 12 months and 13.5 months for group I and group II patients, respectively (P = 0.6). Our results indicate that while there is no adverse effect, there is also no significant advantage of CD34+ selection in terms of progression-free survival and survival in patients with metastatic breast cancer without clinically evident disease. Bone Marrow Transplantation (2000).

Transplantation of CD34+ peripheral blood cells selected using a fully automated immunomagnetic system in patients with high-risk breast cancer: results of a prospective randomized multicenter clinical trial.

Tumor contamination of autologous peripheral blood stem/progenitor cell grafts occurs in a substantial proportion of high-risk breast cancer patients, and the possibility that such contamination may contribute to relapse has focused attention on methods for removal of the contaminating cells prior to transplantation. One such approach is positive selection of CD34+ cells. A fully automated immunomagnetic cell selection system has recently been introduced to facilitate the positive selection process. A multicenter randomized clinical trial was designed to evaluate the capacity of CD34+ cells isolated using the fully automated system to support prompt hematopoietic reconstitution following high-dose chemotherapy in high-risk breast cancer patients, as well as to assess the safety and tolerability of the CD34+ cell transplants. In recipients of isolated CD34+ cells, the median time to an absolute neutrophil count > or =500/microl was 10 days, a value identical to that observed in patients receiving unfractionated apheresis collections. In the isolated CD34+ cell recipients median time to a platelet count > or =20 000/microl was 12 days, compared with 10 days in the unfractionated cell group. There were no statistically significant differences between the groups in median time to neutrophil or platelet engraftment. Infusion of autologous cells was well tolerated by the study groups. There were no inter-group differences in the incidence of infections, need for platelet transfusions, or duration of hospitalization. Isolated CD34+ cells were high in purity and sufficient in number for use in autologous transplantation. The fully automated immunomagnetic cell selection system affords an efficient and time-saving option for isolation of CD34+cells to be used as autologous grafts in high-risk breast cancer patients, and the isolated CD34+ cells support undelayed hematopoietic reconstitution.

Prompt and durable engraftment in two older adult patients with high risk chronic myelogenous leukemia (CML) using ex vivo expanded and unmanipulated unrelated umbilical cord blood.

Delayed engraftment, graft failure, and adverse transplant-related events have been observed in unrelated umbilical cord blood (UCB) recipients, particularly in those receiving a low leukocyte cell dose and in CML patients. We report the outcomes of two older adult patients with high risk CML who received a low leukocyte cell dose of unmanipulated UCB cells supplemented with ex vivo expanded (AastromReplicell System) UCB cells. Each engrafted promptly and neither patient experienced GVHD or life-threatening infection. Both remain engrafted with cells exclusively of donor origin and are in cytogenetic remission at 19 and 8 months follow-up. Ex vivo expanded UCB cells appear to facilitate hematopoietic recovery and therefore may increase the number of CML patients eligible for unrelated UCB transplant.

Process validation.

Process validation is a required element in the clinical good manufacturing practice equation that helps to ensure that systems are performing in the intended manner and that the product we manufacture has the required potency, purity and safety. The parameters defined during the validation process help to define the elements of process control and provide a mechanism for on-going quality assurance, quality control, training, competency review and continuous improvement.

Effects of long-term storage at -90 degrees C of bone marrow and PBPC on cell recovery, viability, and clonogenic potential.

Autologous BM and PB HPC are usually stored from weeks to months until reinfusion after myeloablative chemotherapy. HPC have been stored for up to 16 months at -90 degrees C, using a mixture of 5% DMSO, 6% hydroxyethyl starch (HES), and 4% HSA as a cryoprotectant. Long-term storage (LTS) has usually entailed rate-controlled freezing using 10% DMSO and preservation in liquid nitrogen. The effects of LTS at -90 degrees C on the in vitro cell recovery, viability, and colony-forming unit-granulocyte macrophage (CFU-GM) clonogenic potential of autologous HPC that were not transplanted was studied. Sixteen BM and sixteen PB HPC had been cryopreserved for a median of 53 months (range 27-71) and 35 months (range 26-78), respectively. Samples of frozen HPC were thawed after 48 h, and the nucleated cell count, viability by trypan blue exclusion, and culture for CFU-GM were obtained. Following LTS, the cells were thawed and examined using the same assays. No difference in the median percentage recovery of nucleated cells was found in either the BM or PB HPC between the samples stored for 48 h and after LTS (5.73 x 10(9) versus 5.61 x 10(9) and 6.20 x 10(9) versus 5.78 x 10(9), respectively). In addition, no difference in median percentage viability was found in either the BM or PB HPC sampled at 48 h and at the end of LTS (75% versus 74% and 75% versus 76%, respectively). Finally, the median number of CFU-GM cultured from BM HPC at 48 h was 2.41 x 10(5) (range 0.33-11.01 x 10(5)) and at the end of LTS was 1.93 x 10(5) (range 0.32-10.55), representing a median recovery of 93% (range 19%-308%). Similarly, the median number of CFU-GM cultured from PB HPC was 1.66 x 10(5) (range 0-50.57) and at the end of LTS was 0.93 x 10(5) (range 0-44.9), representing a median recovery of 80% (range 36%-165%). This difference in percentage recovery was not significant (p = 0.514). There was poor correlation between the number of nucleated cells harvested and the percentage recovery of nucleated cells, cell viability, or CFU-GM for either the BM or PB HPC. Similarly, there was poor correlation between the number of CFU-GM in the harvest and their percentage recovery following LTS for both BM and PB HPC. Finally, there was poor correlation between the storage time of the BM or PB HPC and the percentage recovery of nucleated cells, cell viability, and CFU-GM. These data suggest that LTS of HPC at -90 degrees C is not associated with decreased recovery of nucleated cells or in vitro viability and is associated with only a modest decrease in clonogenic potential. This indicates that storage of HPC at -90 degrees C for periods in excess of 3 years is possible.

CD34+CD33- cells influence days to engraftment and transfusion requirements in autologous blood stem-cell recipients.

PURPOSE: To evaluate the reliability of CD34/CD33 subset enumeration as a predictor of hematopoietic repopulating potential in autologous blood stem-cell transplantation and to determine which patient and treatment-related factors affect the timing, quantity, and type of blood stem cells mobilized. PATIENTS AND METHODS: We analyzed blood stem-cell collections from 410 consecutive cancer patients who received mobilization therapy and evaluated factors, including CD34+ subset quantities, that might influence engraftment kinetics and transfusion requirements in autologous blood stem-cell recipients. RESULTS: The majority of patients (97%) mobilized CD34+33- cells, which were usually collected in the greatest quantity on the first day of apheresis. Patients who received only growth factor mobilized the highest percentage of CD34+33- cells. Extensive prior chemotherapy limited the collection of CD34+33- cells. In addition to patient diagnosis (P < .006) and total CD34+ cell dose (P = .0001), CD34+33- cell dose (P < .005) and percentage of CD34+33- cells (P < .005) were identified as independent factors significantly predictive of engraftment kinetics. CD34+33- cell dose (R2 < or = .177; P < .0001) was a strong and the only significant predictor of RBC and platelet transfusion requirements. Furthermore, independent of the total CD34+ cell dose, as the CD34+33- cell dose increased, days to neutrophil recovery, days to platelet recovery, and transfusion requirements decreased. CONCLUSION: These findings show that CD34+33- cells are readily collected in most cancer patients and significantly influence engraftment kinetics and transfusion requirements in autologous blood stem-cell recipients. CD34+33- cell quantity of the blood stem-cell graft appears to be a more reliable predictor of hematopoietic recovery rates than total CD34+ cell quantity in this setting.

Effect of interface/offset (I/O) adjustment on collection efficiency using the Fenwal CS3000 Plus Blood Cell Separator for peripheral blood progenitor cell collection.

Peripheral blood progenitor cells (PBPC) reside within the mononuclear cell (MNC) component of the blood and can be collected using a number of apheresis devices, including the Fenwal CS3000 Plus Blood Cell Separator. Increased MNC collection efficiency, therefore, may reduce the number of apheresis required to achieve collection goals. In this study, patients were divided into groups by absolute MNC count to determine the effect of interface detector offset (I/O) adjustment on MNC collection efficiency. Apheresis products from 104 procedures collected using a standard I/O setting of 100 were compared with 121 collections for which the I/O setting was adjusted according to the preapheresis MNC count. Adjustment of the I/O setting in this manner had no statistically significant impact on the per kilogram dose of MNC collected. The data did show that MNC collection efficiency was reduced as both the MNC count and I/O setting increased, as the collection efficiency was greatest for patients with the lowest peripheral MNC counts and was inversely correlated with the preapheresis MNC count. Although contamination of the product with platelets was drastically reduced at higher I/O settings, there was a concomitant rise in RBC contamination. We conclude that a standard setting of 100 is preferable to adjustment of the I/O setting as a function of the preapheresis MNC count.

An intralaboratory quality control program for quantitation of CD34+ cells by flow cytometry.

This study was undertaken to develop a quality control protocol to monitor instrument, operator, and CD34 assay performance. A dual level control system was established by cryopreserving aliquots of cells from peripheral blood progenitor cell (PBPC) collections exhibiting different percentages of CD34+ cells. Twenty-five samples from each control specimen were assayed to establish a control range (mean +/- 2 SD). Levey-Jennings graphs were prepared for each control specimen to plot multiple measurements of CD34%. No significant differences were observed between fresh or cryopreserved PBPC aliquots in terms of light scatter properties or CD34 antigen density within the gated cell population. Cryopreserved PBPC samples are ideal for serving as a positive methodology control for daily CD34 analysis. Furthermore, such a system can help identify problems with assay reagents, sample preparation technique, or incorrect data analysis.

Single and double autotransplants for relapsing/refractory Hodgkin's disease: results of two consecutive trials.

To evaluate a strategy of one cycle of dose-intensive chemotherapy for patients with Hodgkin's disease in sensitive relapse and two cycles for those with refractory disease, 122 patients received dose-intensive chemotherapy followed by autotransplant in two consecutive studies. Patients with refractory disease were offered a second transplant with different conditioning in the absence of progression or excessive toxicity. CR was present after treatment in 46% while 16% died in the peritransplant period. Of 41 patients with primary refractory disease and 42 with refractory relapse, 24 and 21 respectively received a second cycle. Of these 45 refractory patients, 12 were in CR and 11 in PR after the first cycle and 10 of these 11 in PR achieved a durable CR with the second transplant. The CR rate is 37% in patients with refractory relapse and 19% in those with primary refractory disease. At a median follow-up of 4 years, median survival is 45 months. Progression-free survival of the refractory patients who could receive a second cycle was similar to that of patients with sensitive disease. A sequential transplant strategy is feasible. A subgroup of patients with refractory disease can achieve long-term survival after sequential BMT.

Optimization of CD34+ cell selection using immunomagnetic beads: implications for use in cryopreserved peripheral blood stem cell collections.

Isolation of CD34+ cells from bone marrow, umbilical cord blood, and mobilized peripheral blood stem cell (PBSC) collections has many potential clinical benefits. The aim of this study was to evaluate the use of the ISOLEX 300 system to select hematopoietic precursors and determine the effectiveness at depleting contaminating tumor cells from cryopreserved/thawed PBSC. Median recovery of CD34+ cells and CFU-GM colonies was 71% and 51.5%, respectively, using a protocol optimized for our laboratory. A mean 2.9 log10 decrease in contaminating breast carcinoma cells was seen after the selection process. Selected CD34+ cells underwent a second round of cryopreservation/thawing while retaining 85.6% viability and 72.3% recovery of CFU-GM colonies.

New York State guidelines for cord blood banking. Department of Health.

In response to the profound interest in the transplantation of umbilical cord blood, the New York State (NYS) Department of Health embarked on a regulatory strategy designed to provide guidance to the practitioner while allowing for the flexibility required by the inherent developmental nature of the technology. The result was the development of the Guidelines for Collection, Processing, and Storage of Cord Blood Stem Cells, adherence to which is optional. It is anticipated that through the use of such guidelines as a supplement to the existing hematopoietic progenitor cell (HPC) regulations for tissue banks, practitioners may provide to potential patients the safest and most efficacious cord blood products currently available, and information gained through the use of these products will be of optimal value toward the goal of continuous improvement in the delivery of this new technology.

Va-es-bj - an epithelioid sarcoma cell-line.

A tumor cell line was derived and established from a bone marrow aspirate of a 41-year-old male who presented with motor symptoms of cord compression and an epidural tumor with osseous, pulmonary and hepatic metastases. The epidural tumor was an epithelioid sarcoma staining positive for cytokeratin markers of epithelial differentiation AE-1 and AE-3. This cell line was composed of lowly refractile, multinucleated giant or mononuclear round or elongated cells with monopolar or bipolar short processes. When permitted to grow beyond confluence these cells clumped up and formed mounds progressing to free floating spheroids. The cells were characterized by chromosomal triploidy with marker chromosomes, rapid growth in nude mice and secretion of immunoreactive and biologically functional granulocyte macrophage colony stimulating factor. This epithelioid sarcoma cell line stained positive for AE-1, AE-3, vimentin, epithelial membrane antigen and was designated VA-ES-BJ.