Expression Level of Membrane Markers CD5, CD19, and CD20 in B Cells after UV-Irradiation and Incubation in the Presence of Autologous Plasma.

The effect of UV-light (240-390 nm) in doses of 151 and 755 J/m2 on the expression of membrane markers CD5, CD19, CD20 in human peripheral blood B cells was studied by flow cytometry. In 24 h after exposure to UV light, we observed activation of processes accompanied by structural rearrangements of B-cell membranes leading to changes in the expression of receptor molecules: the content of of CD19 and CD20 increased due to activation of the synthesis of these proteins, while the content of CD5 decreased. The percentage of CD5+ cells decreased over 24 h after UV-irradiation of lymphocytes, while addition of autologous plasma to the incubation medium produced a photoprotective effect on CD5+ cells.

Indications for Rituximab Use in an Integrated Health Care Delivery System.

BACKGROUND: Rituximab is a top-selling biologic that was first approved by the FDA in 1997 for a non-Hodgkin lymphoma orphan indication. It has since been approved for additional orphan indications, with rheumatoid arthritis as the only FDA-approved, nonorphan indication. Evidence suggests that rituximab is frequently used off-label, but information on its use over time and indications for use in the United States is limited. OBJECTIVE: To assess incident rituximab use over time in an integrated health care delivery system. METHODS: This was a cross-sectional, retrospective study. Data were collected from administrative databases and manual chart reviews. Patients who received their first rituximab infusion between October 1, 2009, and December 31, 2017, and who were not a part of a clinical trial were included. Indication for use (FDA-approved orphan/nonorphan, off-label) was determined. Proportions of use were assessed over time. Multivariable logistic regression modeling was performed to assess factors associated with receiving rituximab for an FDA-approved indication. RESULTS: A total of 1,674 patients were included. The majority (66.4%) of patients had an FDA-approved indication, with lymphoma being the most common approved indication (66.4%). The most common indication for off-label use was neurologic conditions (72.7%), predominantly demyelinating diseases. Off-label indication use increased from 1.2% in 2009 to 55.6% in 2017. Factors associated with rituximab use for an FDA-approved indication included increased age (adjusted odds ratio [AOR] = 1.05, 95% CI = 1.04-1.07) and increased burden of chronic disease (chronic disease score: AOR = 1.07, 95% CI = 1.02-1.12; Charlson Comorbidity Index score: AOR = 3.52, 95% CI = 3.03-4.10). CONCLUSIONS: Off-label use of rituximab grew dramatically over the course of the study. With the recent FDA approval of the rituximab biosimilar and its expected lower price, off-label use will likely continue to rise. Opportunities for cost savings and to ensure appropriate use of these medications should be evaluated. DISCLOSURES: This study was funded by Kaiser Permanente. All authors except Hansen are employed by Kaiser Permanente. Hansen has nothing to disclose. Preliminary results were presented at the Mountain States Conference for Residents and Preceptors in May 2019 in Salt Lake City, UT, and at an encore presentation October 2019 at the American College of Clinical Pharmacy Annual Meeting in New York, NY.

The role of obinutuzumab in the management of follicular lymphoma.

The outcomes for follicular lymphoma (FL) have improved significantly in recent years. This has been driven by an improved understanding of the pathobiology of FL and the development of therapeutic anti-CD20 antibodies. Combining rituximab with chemotherapy, coupled with its use as maintenance therapy, has contributed to significant improvements in disease control and progression-free survival. However, FL remains incurable and almost all patients invariably relapse. Therefore, there remains a need to develop novel therapeutic options and optimize existing regimens. Obinutuzumab (a first-in-class, glycoengineered, humanized type 2 anti-CD20 antibody) has been evaluated in a number of clinical trials. In this review, we will summarize the evaluable results of clinical trials investigating the efficacy of obinutuzumab in the treatment of FL.

The effect of hexanic extract of Serenoa repens on prostatic inflammation: results from a randomized biopsy study.

PURPOSE: To evaluate the effect of hexanic extract of Serenoa repens (HESr) on prostatic inflammation in patients with diagnosed prostatic inflammation. METHODS: Patients with prostatic inflammation histologically confirmed by TRUS prostatic biopsy were randomized either to receive HESr (320 mg/day) or no treatment. A second biopsy was performed 6 months later according to standard clinical practice. Inflammation was assessed by the Irani's score and immunohistochemical staining using the CD3, CD4 and CD8 (for T-leucocytes), CD20 (for B-leucocytes) and CD163 (for macrophages) antibodies. RESULTS: Overall 97 patients were eligible for analysis. In the HESr group the mean inflammation grading and aggressiveness grading score significantly decreased from 1.55 and 1.55 at baseline to 0.79 (p = 0.001) and 0.87 (p = 0.001) at the second biopsy, respectively. In the control group the mean inflammation grading score was 1.44 at first biopsy and 1.23 at the second biopsy. The mean aggressiveness gradings core was 1.09 and 0.89, respectively. No statistical significance was found (p = 0.09 and p = 0.74).The mean decrease in all inflammation scores was statistically higher in the HESr patients compared to controls. The immunohistochemical staining showed a significant change in the expression of the analyzed antibodies for the HESr patients compared to the first biopsy. In the nontreatment group, no significant difference was found at the second biopsy. The change in expression of each antibody in the HESr group was statistical significant compared to control. CONCLUSIONS: HESr seems to reduce prostatic inflammation in terms of histological and immunohistochemical parameters in this specific patients population.

Combination of 177 Lu-lilotomab with rituximab significantly improves the therapeutic outcome in preclinical models of non-Hodgkin's lymphoma.

OBJECTIVES: To investigate the therapeutic potential of the next-generation anti-CD37 radioimmunoconjugate 177 Lu-lilotomab satetraxetan (177 Lu-lilotomab) in combination with the anti-CD20 antibody rituximab for treatment of mice with non-Hodgkin's lymphoma (NHL) xenografts. METHODS: Nude mice with subcutaneous (s.c.) Burkitt's lymphoma Daudi xenografts and SCID mice intravenously (i.v.) injected with Mantle cell lymphoma Rec-1 cells were treated with either 177 Lu-lilotomab or rituximab alone or with the combination of both treatments. Tumour volume, body weight, blood counts and clinical status were monitored. CD20 expression was measured using flow cytometry with fluorescence-labelled rituximab. RESULTS: The combination of 177 Lu-lilotomab and rituximab was synergistic for treatment of nude mice with s.c. Daudi xenografts while it was additive for treatment of SCID mice with i.v. injected Rec-1 cells. Binding of rituximab to NHL cells in-vitro was increased by pretreatment with 177 Lu-lilotomab. CONCLUSIONS: Treatment of mice with NHL xenografts with 177 Lu-lilotomab synergistically increased tumour suppression of subsequent anti-CD20 immunotherapy and improved survival. If the same effect is confirmed in a recently started clinical study, it could change the way radioimmunotherapy and CD20 immunotherapy would be used in the future.

Hypersensitivity Reactions to Obinutuzumab in Cynomolgus Monkeys and Relevance to Humans.

Obinutuzumab (GA101, Gazyva™, Gazyvaro®, F. Hoffmann-La Roche AG, Basel, Switzerland) is a humanized, glycoengineered type II antibody targeted against CD20. The preclinical safety evaluation required to support clinical development and marketing authorization of obinutuzumab included repeat-dose toxicity studies in cynomolgus monkeys for up to 6-month dosing with a 9-month recovery period. Results from those studies showed decreases in circulating B cells and corresponding B-cell depletion in lymphoid tissues, consistent with the desired pharmacology of obinutuzumab. Hypersensitivity reactions were noted at all doses in the 6-month study and were attributed to the foreign recognition of the drug construct in cynomolgus monkeys. Findings in monkeys were classified as acute hypersensitivity reactions that were evident immediately after dosing, such as excessive salivation, erythema, pruritus, irregular respiration, or ataxia, or chronic hypersensitivity reactions characterized by glomerulonephritis, arteritis/periarteritis, and inflammation in several tissues including serosal/adventitial inflammation. Immune complex deposits were demonstrated in tissues by immunohistochemistry, immunofluorescence, and electron microscopy. Some of, but not all, the animals that developed these reactions had detectable antidrug antibodies or circulating immune complexes accompanied by loss of drug exposure and pharmacodynamic effect. On the basis of clinical evidence to date, hypersensitivity reactions following obinutuzumab are rare, further supporting the general view that incidence and manifestation of immunogenicity in nonclinical species are generally not predictive for humans.

Near infrared photoimmunotherapy of B-cell lymphoma.

Near infrared photoimmunotherapy (NIR-PIT) is a new, highly-selective cancer theranostics that employs an antibody-photo absorber conjugate (APC). NIR-PIT has successfully treated preclinical tumor models with APCs and is now in the first-in-human phase 1 clinical trial for head and neck cancer patients against EGFR. CD20 is highly expressed in many B-cell lymphomas and is emerging as a molecular target for this disease. Here, we describe the use of the anti-CD20 monoclonal antibody (mAb), rituximab-IR700 APC for NIR-PIT of B-cell lymphoma in two CD20-expressing lymphoma mouse models. CD20 expressing B-cell lymphoma cell lines (Daudi and Ramos) were used in this study. Rituximab-IR700, rituximab conjugated with IRDye700DX, showed specific binding, and cell-specific killing only after exposure of NIR light to both cells in vitro. To evaluate effects of NIR-PIT in vivo, tumor-bearing mice were separated into 4 groups: (1) control; (2) APC i.v. only; (3) NIR light exposure only; (4) APC and NIR light (NIR-PIT). These were performed every week for up to 3 weeks. Rituximab-IR700 showed high tumor accumulation and high target-to-background ratio in vivo. Tumor growth was significantly inhibited by NIR-PIT in comparison with the other groups (p < 0.001 for both tumors), and survival was significantly prolonged in both tumors (p < 0.001 for Daudi tumors and p < 0.0001 for Ramos tumors vs other groups). More than half of tumors were cured with this single regimen of NIR-PIT. In conclusion, anti-CD20 rituximab-IR700 works as a highly effective APC for NIR-PIT against B-cell lymphoma.

B cell-targeted immunotherapy for type 1 diabetes: What can make it work?

Immunotherapy has revolutionized treatment of cancers and autoimmune diseases. Bucking the trend, however, is type 1 diabetes (T1D), although it is one of best understood autoimmune diseases and individuals at genetic risk are identifiable with high certainty. Here we review the major obstacles associated with pan-B-cell-depletion using rituximab (RTX) and discuss the notion that B cell-directed therapy may be most effective as a preventive measure. We suggest that it will be more productive to aim at identifying and targeting autoreactive B cells rather than making adjustments to pan-B cell depletion and that non-conventional alternative therapies such as antibody blockade of FasL to bolster IL-10-producing Breg cells, which work successfully in mice, should be considered.

Prognostic impact of immune response in resectable colorectal liver metastases treated by surgery alone or surgery with perioperative FOLFOX in the randomised EORTC study 40983.

AIM: To investigate whether the immune response in colorectal liver metastases is related to progression free survival (PFS) and if this may be influenced by systemic therapy. METHODS: A retrospective central collection of tumour tissue was organised for the European Organisation for Research and Treatment of Cancer (EORTC) study 40983, where patients with colorectal liver metastases were treated by either resection alone or resection with perioperative FOLFOX. Immunostaining on whole slides was performed to recognise T-lymphocytes (CD3+, CD4+, CD8+), B-lymphocytes (CD20+), macrophages (CD68+) and mast cells (CD117+) inside the tumour, at the tumour border (TNI) and in normal liver tissue surrounding the tumour (0.5-2mm from the TNI). Immunological response was compared between treatment arms and correlated to PFS. RESULTS: Tumour tissue and immune response profiles were available for 82 resected patients, 38 in the perioperative chemotherapy arm and 44 in the surgery alone arm. Baseline patient and disease characteristics were similar between the treatment arms. In response to chemotherapy, we observed increased CD3+ lymphocyte and mast cell counts inside the tumour (p<0.01), lower CD4+ lymphocytes in the normal liver tissue (p=0.02) and lower macrophage counts in normal tissue (p<0.01) and at the TNI (p=0.02). High number of CD3+ lymphocyte and mast cells, and high T-cell score were correlated with tumour regression grade (TRG). Prolonged PFS correlated with the presence of mast cells in the tumour (9.8 versus 16.5 months, Hazard ratio (HR) 0.54 p=0.03), higher CD3+ lymphocyte count at the TNI (10.8 versus 22.8 months, HR 0.57, p=0.03) and T-cell score >2 (10.8 versus 38.6 months, HR 0.51, p=0.04). CONCLUSION: Our analyses in the context of a randomised study suggest that chemotherapy influences immune cell profiles, independent of patient characteristics. Immune responses of lymphocytes and mast cells were associated with pathological response to chemotherapy and to increased PFS. High CD3+ lymphocytes at the tumour front and intratumoural mast cells appear to be prognostic for patients with colorectal liver metastases.

Ofatumumab Exhibits Enhanced In Vitro and In Vivo Activity Compared to Rituximab in Preclinical Models of Mantle Cell Lymphoma.

PURPOSE: Mantle cell lymphoma (MCL) is a mature B-cell lymphoma considered to be incurable with current treatments, including first-line rituximab in combination with multiagent chemotherapy and for those eligible, high-dose chemotherapy and stem cell support or rituximab maintenance. On the other hand, achieving a complete remission by high-sensitive flow cytometry is associated with prolonged duration of remission, stressing the need to develop and/or incorporate novel agents into the management of MCL. To this end, we examined the activity of ofatumumab, an anti-CD20 monoclonal antibody with distinct binding and immunologic properties compared to rituximab, in MCL preclinical models. EXPERIMENTAL DESIGN: MCL cells were labeled with (51)Cr before incubation with rituximab or ofatumumab (10 µg/mL) plus human serum or effector cells. (51)Cr-release was measured and the percentage of lysis was calculated. Surface CD20, CD55, and CD59 were measured by Imagestream analysis. SCID mice inoculated subcutaneously with Z138 cells were assigned to control versus four doses of ofatumumab or rituximab (10 mg/kg/dose). RESULTS: Ofatumumab exhibited enhanced in vitro complement-dependent cytotoxicity activity compared with rituximab in MCL cell lines, despite a high degree of in vitro resistance to rituximab associated with low CD20 levels and/or high expression of complement inhibitory proteins. Ofatumumab also delayed tumor progression and prolonged survival in a murine model of MCL. CONCLUSIONS: Our results demonstrate that ofatumumab is more effective than rituximab in MCL preclinical models, including in the presence of rituximab resistance, and support the clinical investigation of ofatumumab in combination with standard systemic chemotherapy in MCL (NCT01527149).

Pathological study of chronic pulmonary toxicity induced by intratracheally instilled Asian sand dust (kosa).

Asian sand dust (ASD) events are associated with an increase in pulmonary morbidity and mortality. The number of ASD events has increased rapidly in the east Asian region since 2000. To study the chronic lung toxicity of ASD, saline suspensions of low doses (200 and 400 µg) and high doses (800 and 3,000 µg) of ASD were intratracheally instilled into ICR mice. Animals were sacrificed at 24 hr, 1 week, or 1, 2, or 3 months after instillation. Histopathological examination revealed that ASD induced acute inflammation at 24 hr after instillation. The acute inflammation was transient and subsided at 1 week and 1 month after instillation. At 2 and 3 months after instillation, focal infiltration of lymphocytes with accumulation of epithelioid macrophages, which is a suggestive finding of transformation to granuloma, and granuloma formation were occasionally observed. Aggregation of macrophages containing particles was observed in the pulmonary lymph nodes at 3 months after instillation in high-dose groups. Prolonged inflammatory foci (granuloma) and presence of ASD particles in pulmonary lymph nodes would have a chance to induce immunological modulation leading to adverse health effects in the exposed animals.

[Clinical characteristics and diagnosis of pulmonary mucosa-associated lymphoid tissue-derived (MALT) lymphoma: a retrospective analysis of 29 cases].

OBJECTIVE: To study the clinical manifestations and radiological characteristics, diagnostic methods and outcomes of pulmonary mucosa-associated lymphoid tissue-derived(MALT) lymphoma. METHODS: A retrospective review of clinical, radiological and follow-up data of 29 pulmonary MALT lymphoma cases at Shanghai Pulmonary Hospital affiliated to Tong Ji University from January 2002 to June 2010 was performed. RESULTS: Among these patients, there were 19(65.5%) males and 10 (34.5%) females aged from 27 to 73 (median 53) years old. Common clinical manifestations were cough (51.7%), fever (20.7%), apnea (17.2%), chest pain (17.2%), fatigue (13.8%) and weight loss (13.8%), while 9(31.0%) cases had no symptoms at diagnosis. The characteristics of the chest CT showed that 22 (75.9%) of the cases had patch infiltration or consolidation of the lung, 7(24.1%) of the cases had mass, and 15 (51.7%) unilateral and 14(48.3%) bilateral lesions. Their diagnosis duration varied between 0.5 and 96 months. 18(62.1%) cases were confirmed by surgery (15 open lung and 7 video-assisted thoracic surgery, VAST), 4 (13.8%) by percutaneous lung biopsy, 5 (17.2%) by bronchoscopic biopsy, and 2 (6.9%) by peripheral lymph node biopsy. The treatment methods included surgery, combined chemotherapy, radiotherapy and Chinese herbal medicine. The 1- and 3-year-survival rates were 92.3% and 87.4%, respectively. CONCLUSIONS: Pulmonary MALT lymphoma is atypical in clinical manifestations and radiological characteristics, and easy to be misdiagnosed. Local diseases are mainly treated by operation while extensive diseases receive combined chemotherapy. A proper diagnosis is mainly based on pathological biopsy. Patients with MALT lymphoma have a favorable outcome. Poor prognosis may be connected with poor performance status and long diagnosis duration.

Galiximab signals B-NHL cells and inhibits the activities of NF-κB-induced YY1- and snail-resistant factors: mechanism of sensitization to apoptosis by chemoimmunotherapeutic drugs.

Galiximab (anti-CD80 monoclonal antibody) is a primatized (human IgG1 constant regions and cynomologus macaque variable regions) monoclonal antibody that is currently in clinical trials. Galiximab inhibits tumor cell proliferation through possibly cell signaling-mediated effects. Thus, we hypothesized that galiximab may signal the tumor cells and modify intracellular survival/antiapoptotic pathways such as the NF-κB pathway. This hypothesis was tested using various CD80(+) Burkitt B-NHL (non-Hodgkin lymphomas) cell lines as models. Treatment of B-NHL cells with galiximab (25-100 µg/mL) resulted in significant inhibition of NF-κB activity and its target resistant factors such as YY1, Snail, and Bcl-2/Bcl-XL. Treatment of B-NHL cells with galiximab sensitized the tumor cells to both cis-diamminedichloroplatinum(II) (CDDP)- and TRAIL-induced apoptosis. The important roles of YY1- and Snail-induced inhibition by galiximab in the sensitization to CCDP and TRAIL were corroborated following transfection of Raji cells with YY1 or Snail short interfering RNA. The transfected cells were shown to become sensitive to both CCDP- and TRAIL-induced apoptosis in the absence of galiximab. Furthermore, knockdown of YY1 or Snail inhibited Bcl-XL. The involvement of Bcl-XL inhibition in sensitization was corroborated by the use of the pan-Bcl-2 inhibitor 2MAM-3 whereby the treated cells were sensitive to both CDDP- and TRAIL-induced apoptosis. These findings show that galiximab inhibits the NF-κB/Snail/YY1/Bcl-XL circuit that regulates drug resistance in B-NHL and in combination with cytotoxic drugs results in apoptosis. The findings also support the therapeutic application of the combination of galiximab and cytotoxic drugs in the treatment of drug-resistant CD80-positive B-cell malignancies.

B-cell depletion attenuates white and gray matter pathology in marmoset experimental autoimmune encephalomyelitis.

This study investigated the effect of CD20-positive B-cell depletion on central nervous system (CNS) white and gray matter pathology in experimental autoimmune encephalomyelitis in common marmosets, a relevant preclinical model of multiple sclerosis. Experimental autoimmune encephalomyelitis was induced in 14 marmosets by immunization with recombinant human myelin oligodendrocyte glycoprotein in complete Freund adjuvant. At 21 days after immunization, B-cell depletion was achieved by weekly intravenous injections of HuMab 7D8, a human-anti-human CD20 antibody that cross-reacts with marmoset CD20. In vivo magnetic resonance imaging showed widespread brain white matter demyelination in control marmosets that was absent in CD20 antibody-treated marmosets. High-contrast postmortem magnetic resonance imaging showed white matter lesions in 4of the 7 antibody-treated marmosets, but these were significantly smaller than those in controls. The same technique revealed gray matter lesions in 5 control marmosets, but none in antibody-treated marmosets. Histologic analysis confirmed that inflammation, demyelination, and axonal damage were substantially reduced in brain, spinal cord, and optic nerves of CD20 antibody-treated marmosets. In conclusion, CD20-postive B-cell depletion by HuMab 7D8 profoundly reduced the development of both white and gray matter lesions in the marmoset CNS. These data underline the central role of B cells in CNS inflammatory-demyelinating disease.

Transforming growth factor ß-receptor II protein expression in benign prostatic hyperplasia is associated with prostate volume and inflammation.

OBJECTIVE: To assess transforming growth factor ß-receptor II (TGFBRII) protein expression in benign prostatic hyperplasia (BPH) using immunohistochemistry analysis, and to compare the analysis with phenotypic properties. METHODS: TGFBRII protein expression was profiled using three clinical outcome tissue microarrays (TMAs), sampled from 231 patients who underwent surgery for BPH. Using these TMAs, five inflammatory cell markers were also assessed, including CD3, CD4, CD8, CD20, and CD163. The surgical procedure was open prostatectomy in 95 patients and transurethral resection of the prostate in 136 patients. RESULTS: TGFBRII protein expression was found in BPH epithelium cells for both basal and secretory cells, as well as in fibromuscular stromal cells. TGFBRII staining was also strong in most of the lymphocytes infiltrating the prostate. TGFBRII stromal staining was found to be significantly associated with prostate volume (P = 0.04), whereas TGFBRII epithelial staining was found to be significantly associated with 5-α-reductase-inhibitor medical therapy received by patients before surgery (P = 0.004). Both stromal and epithelial TGFBRII staining were found to be associated with CD4 T-lymphocyte infiltrate, independently of prostate volume (P < 0.001 and P = 0.002). CONCLUSIONS: TGFBRII protein expression in BPH is associated with prostate gland volume and with CD4 T-lymphocyte prostatitis. TGFBRII might be a promising therapeutic target to prevent prostate enlargement or even to decrease prostate volume.

A mechanistic rationale for combining alemtuzumab and rituximab in the treatment of ALL.

B-lineage acute lymphoblastic leukemia (ALL) may express CD52 and CD20. Alemtuzumab (ALM) and rituximab (RTX) are therapeutic antibodies directed against CD52 and CD20, respectively, but showed limited activity against ALL in clinical trials. The mechanisms for the impaired responses remained unclear. We studied expression of CD52 and CD20 on ALL cells and found that most cases coexpressed CD52 and CD20. However, distinct CD52-negative (CD52(-)) subpopulations were detected in most cases as the result of defective glycophosphatidyl-inositol anchoring. Although ALM efficiently eradicated CD52-positive (CD52(+)) cells in NOD/scid mice engrafted with primary human ALL, CD52(-) subclones escaped therapy. In the same model, RTX showed limited activity resulting from occurrence of CD20 down-modulation. However, CD52(-) cells concurrently lacked the glycophosphatidyl-inositol-anchored complement regulators CD55 and CD59 and showed increased susceptibility to RTX-mediated complement-dependent cytotoxicity in vitro. At the same time, ALM was shown to inhibit down-modulation of CD20 in response to RTX by depleting the trogocytic capacity of phagocytic cells. Probably because of these complementary mechanisms, combined administration of ALM and RTX induced complete responses in vivo. Based on these data, we propose a mechanistic rationale for combined application of RTX and ALM in ALL.

Type II (tositumomab) anti-CD20 monoclonal antibody out performs type I (rituximab-like) reagents in B-cell depletion regardless of complement activation.

Anti-CD20 monoclonal antibodies (mAbs) are classified into type I (rituximab-like) or type II (tositumomab-like) based on their ability to redistribute CD20 molecules in the plasma membrane and activate various effector functions. To compare type I and II mAbs directly in vivo and maximize Fc effector function, we selected and engineered mAbs with the same mouse IgG(2)a isotype and assessed their B-cell depleting activity in human CD20 transgenic mice. Despite being the same isotype, having similar affinity, opsonizing activity for phagocytosis, and in vivo half-life, the type II mAb tositumomab (B1) provided substantially longer depletion of B cells from the peripheral blood compared with the type I mAb rituximab (Rit m2a), and 1F5. This difference was also evident within the secondary lymphoid organs, in particular, the spleen. Failure to engage complement did not explain the efficacy of the type II reagents because type I mAbs mutated in the Fc domain (K322A) to prevent C1q binding still did not display equivalent efficacy. These results give support for the use of type II CD20 mAbs in human B-cell diseases.

[Adriamycin combined with hyperthermia inhibits transplanted B cell lymphoma in nude mice].

OBJECTIVE: To study the effect of adriamycin combined with hyperthermia on tumor formation and growth of human B lymphoma cell line (Raji) in nude mice. METHODS: Twenty-four BALB/C nude mice were divided into control group (37 degrees celsius;), chemotherapy group (37 degrees celsius;+ADM), hyperthermia group (42 degrees celsius;) and thermochemotherapy group (42 degrees celsius;+ADM), and in each mouse, 5 x 10(6) Raji cells were injected subcutaneously. The time and rate of tumor formation were observed, the tumor diameter was measured every 3 days and the tumor volume calculated to obtain the growth curves of the tumors. Three weeks after tumor formation, all the mice were executed to observe the histopathological changes of the tumor with HE staining. RESULTS: The time of tumor formation in the control, chemotherapy, hyperthermia and thermochemotherapy groups were 11.2-/+1.7, 20.2-/+2.3, 15.3-/+1.6 and 23.8-/+1.7 days, respectively. Three weeks after tumor formation, the average weight of the tumors were 3.33-/+0.57, 2.26-/+0.28, 2.76-/+0. 26 and 1.73-/+0. 33 g, respectively, and the tumor growth inhibition rate was 48.0% in the thermochemotherapy group. CONCLUSION: Thermochemotherapy can be effective in inhibiting tumor growth and provides an alternative of adjuvant therapy for malignant B cell lymphoma.

Induction of apoptosis and effect on CD20+ using rituximab on autologous peripheral blood stem cell harvests from patients with B cell lymphomas.

Purging of neoplastic cells for autologous stem cell transplantation is usually done in vivo by administering chemotherapy and/or other agents before harvesting. It is also possible to decrease malignant cells counts directly in the cell harvest. In this study, we ascertained the effect of anti-CD20 monoclonal antibody and rituximab administration on peripheral blood hematopoietic stem cells. Five samples of stem cell harvests from different patients with B cell lymphoma were obtained. Each sample was divided in two tubes with calcium gluconate (20 mEq/50 microl). Rituximab (1 mg/600,000 mononuclear cells) was added to one of the tubes. Using flow cytometry, CD19, CD20 (B cell markers), and CD95 (apoptosis marker), expression was measured at baseline and 24 h after the addition of rituximab. A one-sided t-test with equal variances was used to analyze the results. Immediately after rituximab addition, CD20 expression became null. No significant difference in variation of CD19 expression was detected after the addition of rituximab (-3.64% control vs. 0.63% rituximab, p = 0.69). Mean variations of percentage of CD95 expression were 2.9% (controls) and 10.52% (rituximab tubes) (p = 0.06). We conclude that rituximab is capable of initiating apoptosis in vitro. We found no decrease in the CD19+ cell count, used as a surrogate marker for CD20+ cells, meaning that, at least in 24 h, apoptosis activation is not capable of decreasing CD20+ cell numbers. In vitro purging of peripheral blood stem cells harvests with rituximab could be part of a broader therapeutic strategy to be offered to lymphoproliferative disorder patients.