Biomedical subjects
Publications and source records attributed to N E Kay.
This article reviews pertinent immunobiologic features of the malignant B cell and the circulating immunoregulatory cells (T and NK) that characterize B-CLL.
Previous work has demonstrated that large granular lymphocytes (LGL) may be suppressive for B cell function. We have investigated the impact of blood LGL from B-CLL patients with and without hypogammaglobulinemia on normal B cell Ig synthesis and proliferation. Purified blood LGL from controls (age and sex matched) and B-CLL patients were added in various concentrations to isolated normal B cells. The cell co-cultures plus mitogens were incubated at 37 degrees C for 5-7 days and both Ig levels in culture supernatants or cell proliferation determined by ELISA or thymidine incorporation respectively. Percoll purified LGL from controls and B-CLL patients were evaluated for their effect on both PWM and anti-Ig/staph protein A (SPA) induced B-cell proliferation. CLL LGL from certain patients were significantly down regulatory only for anti-Ig/SPA induced B cell proliferation. These B-CLL patients were patients with obvious hypogammaglobulinemia. Subsequently, we purified blood LGL subsets from 5-CLL patients with and without hypogammaglobulinemia. Two LGL subsets; CD16+, CD3- and CD16+, CD3+ were purified by flow cytometry. These LGL were then added to control B cells in presence of PWM and Ig levels determined at day 5 of culture. CLL LGL (CD16+, CD3-) from 3 B-CLL patients with hypogammaglobulinemia were clearly down regulatory for Ig levels. The LGL from 3 B-CLL patients with normal serum Ig levels were not suppressive of mitogen induced B cell Ig synthesis/secretion, nor were the LGL (CD16+, CD3- or CD16+, CD3+) from age and sex matched controls (n = 2).(ABSTRACT TRUNCATED AT 250 WORDS)
Both membrane (p55) and soluble (p45) forms of TAC-reactive interleukin-2 receptor (IL-2R) are expressed and/or released by activated lymphocytes or monocytes. Previous work has detected increased levels of circulating, TAC-soluble IL-2R (soluble TAC antigen) in the serum of most B-cell chronic lymphocytic leukemia (B-CLL) patients. We detected soluble TAC antigen in B-CLL patients (mean of 3,332 U/mL v 410 for controls). Serum soluble TAC antigen levels increased with stage (mean value of 1,187 U/mL for stage 0 v 2,527 for stage 2 and 5,410 for stages 3 and 4). We next attempted to determine whether the elevated serum levels of soluble TAC antigen in B-CLL patients might result from shedding or secretion of the receptor from the circulating, malignant B cells. Purified, malignant B cells from B-CLL patients were capable of producing easily detectable soluble TAC antigen after 48 hours of in vitro culture (range of 60 to 1,563 U/mL). IL-2R production by CLL B cells was dose dependent in most patients over a concentration of 10 x 10(6) to 60 x 10(6)/mL. In contrast, there was little or no detectable soluble TAC antigen when highly purified T cells from the same patients were cultured. Finally, despite elaboration of soluble IL-2R by CLL B cells, membrane expression of B-cell IL-2R was detected in only six of 11 patients. Thus, the cellular source of the elevated serum IL-2R levels is the malignant CLL B cell. Taken together these data suggest that (a) the malignant CLL B cell is "activated" in terms of release of soluble IL-2R and may serve as a tumor marker in this disease and (b) the elevated levels of circulating IL-2R may be an associated factor in the cellular immunodeficiency noted in B-CLL patients.
Elevation of intracellular free calcium (Ca++) is an early activation event that occurs as a result of ligand binding in several human cell systems. In this report, erythropoietin, the major hormone governing erythroid differentiation, was found to elicit an increase in Ca++ in human bone marrow mononuclear cells. Two chelators of intracellular calcium, quin 2 and the more specific and sensitive analogue, fura-2, were used to characterize the response evoked by both recombinant and native hormone. Erythropoietin caused a rapid, dose-dependent rise (within seconds) in Ca++ in bone marrow mononuclear cells, which could be prevented by preincubation of hormone with a rabbit erythropoietin antiserum. The erythropoietin response did not occur in purified populations of T- or B-lymphocytes. These studies suggested that increased Ca++ on erythropoietin binding may be an early transmembrane signal in hormone action.
Recombinant interleukin-2 (rIL-2) is a biologic response modifier that is capable of enhancing or restoring the cytolytic capacity of large granular lymphocytes (LGL). We utilized this biologic response modifier in the treatment of B-chronic lymphocytic leukemia (B-CLL), a disease frequently characterized by deficient or absent natural killer activity. B-CLL (n = 12) patients previously refractory to chemotherapy or with progressive disease post cessation of chemotherapy were eligible. rIL-2 was given as i.v. infusion (2 x 10(6) units/m2) over 2 h 5 times per week for 3 weeks as induction. Responding patients were placed on maintenance therapy. Although there were no complete or partial responses (by ECOG criteria) there was clear evidence of tumor reduction. Seven of 10 evaluable patients had a reduction of the peripheral blood B cell clone, 3 had node reduction and 2 had reduction in their splenomegaly. All patients experienced mild to moderate toxicity and 1 patient died while on induction therapy. Three B-CLL patients following induction rIL-2 treatment were placed back on chemotherapy because of progressive disease. Interestingly, these 2 B-CLL patients achieved extremely rapid and complete responses to chemotherapy which had previously been ineffective. These data suggest a possible role for rIL-2 in treatment of B-CLL.
There is increasing evidence for a bidirectional communications system between the immune system and the brain. Many of the substances involved in this communication appear to be neuropeptides. These findings have given biochemical validity to the clinical and epidemiological studies that have suggested that psychosocial factors can modulate the response to infections and neoplasms.
Aberrations in chromosome number, ploidy abnormalities, have been associated with malignancy and are predictive of outcome. Automated flow cytometry has made DNA analysis applicable to many solid tumors. Analysis can be performed on fixed specimens, allowing archival retrieval. The techniques, however, are unique and must be individually tested for each tumor type. Presently, few studies have been applied to head and neck cancers. This series of flow cytometric DNA analyses compares the results of 17 fresh and fixed head and neck squamous cell carcinoma specimens. Aneuploidy was present to a significant degree (47%). The method produced interpretable results in 100% of cases, with 100% reproducibility. Fresh and fixed tumor specimens yielded comparable results 76% of the time and, in fact, interpretability of fixed specimens was superior. This series demonstrates a practical and accurate flow cytometric DNA assay for fixed squamous cell carcinoma specimens, facilitating rapid retrospective ploidy analysis.
Natural killer (NK) function in the majority of B-cell chronic lymphocytic leukemia patients is markedly deficient. This study was undertaken to determine if the biological response modifier interleukin-2 (IL-2), which is a potent augmenter of normal individuals' NK activity, could augment the low NK activity in these patients. Peripheral blood lymphocytes (PBL), depleted of B-cells, from most B-CLL patients exhibited low natural killer activity against NK-sensitive K562 cells and against herpes simplex virus (HSV)-infected lymphoblastoid cell lines (LCL). Incubation of patients' B-cell-depleted PBL with recombinant IL-2 resulted in augmentation of their NK activity against both K562 cells and HSV-infected cells. Furthermore, whereas large granular lymphocytes (LGL) isolated from CLL patients are deficient in cytoplasmic granules, which are thought to play a role in NK-cell-mediated lysis, treatment of patients' LGL resulted in increased granulation by 4 hr after treatment with IL-2 and showed a concomitant increase in lytic activity comparable to that of normal individuals.
Lymphocytes exposed to cuprammonium cellulose membranes have been shown to exhibit depressed natural killer (NK) function. In the present study we investigated the extent to which three dialyzer membranes of different compositions suppressed human lymphocyte NK activity. Peripheral blood lymphocytes or T cells from normal donors were exposed in vitro to cuprammonium cellulose, cellulose acetate, or polycarbonate dialyzer membranes. After exposure to the membranes, NK activity of the cells was studied by using the NK-sensitive cell line K562 as targets. All three membranes adversely affected human lymphocyte NK function, with cuprammonium cellulose producing the most (70-80%) and polycarbonate producing the least (10-15%) suppression. Our results suggest that the composition of dialyzer membranes affects the extent to which the membranes impair human lymphocyte function. The use of more biocompatible membranes might lessen the potential clinical impact of abnormal NK function in hemodialysis patients.
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The functional importance of interleukin 2 (IL-2) receptors in the regulation of malignant B cell proliferation still remains to be clarified. We studied malignant CLL B cells for the presence of IL-2 receptors and responsiveness to IL-2 with respect to proliferation and B cell colony formation. Seven of 25 B-cLL patients studied had reactivity for anti-Tac (mean, 11.8%; range 4-31%). Purified control T cells expressed less than 2% reactivity to anti-Tac. CLL B cell colony forming cells were reactive with anti-Tac in all five patients studied (mean, 24.8%; range, 17-31%). The proliferative response of control and CLL B cells to both a partially purified preparation of IL-2 and recombinant IL-2 (rIL-2) was also examined. Control B cells demonstrated a dose-dependent, enhanced proliferative response to IL-2. At lower IL-2 concentrations (2-200 units/ml), rIL-2 appeared to have a more significant proliferative enhancing effect on control B cells than did the partially purified IL-2 preparation. In contrast, no concentration of the IL-2 preparations enhanced CLL B cells' proliferative response. The monoclonal antibody anti-Tac was capable of inhibiting control B cell responsiveness to IL-2, rIL-2 did not support CLL B cell colony formation. Thus, malignant B cell populations may spontaneously express the Tac antigen in the absence of a functional response to IL-2. The significance of this in the treatment of lymphomas is underscored by the development of new therapeutic strategies which would seek to incorporate the use of immunoregulatory lymphokines such as IL-2.
Peripheral blood natural killer (NK) activity in patients with B-cell chronic lymphocytic leukemia (B-CLL) is frequently low or absent. Because cimetidine (a histamine-2 antagonist) has been shown to alter human lymphocyte function in vitro, we decided to study cimetidine's effect on peripheral blood NK activity of patients with B-CLL and controls. We administered cimetidine orally (1.2 gm per day) to seven patients with B-CLL and 12 controls for up to 28 days. Peripheral blood NK activity of patients with B-CLL rose from a pretreatment level of 0.7 +/- 0.5 (mean +/- SEM) lytic units/10(6) cells (LU) to 8.7 +/- 2.4 LU (P less than 0.05) at day 28. Peripheral blood NK activity of controls decreased after 14 days of cimetidine treatment but returned to pretreatment levels by day 28. When peripheral blood cells from controls were exposed to cimetidine during in vitro incubation (10 micrograms/ml), mean NK activity was increased at 48 hours (54% +/- 22% increase over controls, n = 5, P less than 0.05). Single cell cytotoxicity assays revealed increased killing of target cells (but not effector-target conjugation) with cimetidine-exposed effector cells. These data suggest that cimetidine may be useful to augment peripheral blood NK activity for patients with B-CLL.
While developing monoclonal antibodies (MoAb) to colorectal carcinoma (CRC) cells, we noted that one MoAb, termed CJA3, down-regulated natural cell-mediated cytotoxicity (NCMC) against CRC cell lines SW480 and SW620. The MoAb CJA3 was developed by immunizing a BALB/c mouse with fresh human colorectal adenocarcinoma cells. The antigen recognized by the MoAb CJA3 was expressed on several solid tumor cell lines and on one of the six lymphoreticular cell lines tested, but was not detected on normal peripheral blood lymphocytes (PBL). SDS-PAGE analysis of the antigen immunoprecipitated by the MoAb CJA3 from the CRC cell lines SW480 and SW620 and from the melanoma cell line MALME-3M revealed a component with a m.w. of 150,000. Preincubation of CRC cell lines SW480 and SW620 with the MoAb CJA3 for 16 hr reduced their susceptibility to NCMC by about 50%. Kinetic experiments showed that prolongation of the incubation of target cells with the MoAb CJA3 resulted in a time-dependent increase in the amount of MoAb bound. Maximum binding of the MoAb CJA3 was reached after 4 hr of incubation. The increase in antigen expression chronologically paralleled the decrease in NCMC target cell sensitivity, suggesting that the membrane alterations induced by the MoAb CJA3 were important for NCMC against these two cell lines.
Significant alterations in T cell subpopulations and function occur in chronic lymphocytic leukemia (CLL) patients. We studied whether abnormalities in peripheral blood T cell parameters were present in 15 untreated early stage CLL patients (ie, Rai stage 0, 1, 2). Seven of the nine patients showed decreased T helper support as compared to control T cells for pokeweed mitogen (PWM)-induced control B cell proliferation (ie, patient 6,063 +/- 1,434 cpm vs control 14,894 +/- 121 cpm). All stage 0 and 1 patients showed a marked impairment of T helper activity for control B cell proliferation (patient T = 7,752 +/- 1,137 cpm vs control T = 14,894 +/- 121 cpm). In a separate assay system, six of nine CLL patients showed T suppressor activity for control B cell proliferation greater than control T cell suppressor activity. Four patients were stage 0 and 1. CLL patients demonstrated markedly impaired T cell support for control B cell immunoglobulin synthesis compared to control T cells (188 +/- 28 vs 869 +/- 56 hemolytic plaque-forming cells (HePFC)/culture, respectively). Control T cells showed increasing support for control B cell immunoglobulin synthesis with increasing T:B cell ratios (869 +/- 56 vs 1,265 +/- 48 HePFC/culture, at 1:1 and 2:1 T:B cell ratios, respectively). In contrast, five of eight CLL patients' T cells showed no improvement in control B cell immunoglobulin synthesis with increasing T:B cell ratios (795 +/- 56 vs 569 +/- 48 HePFC/culture, at 1:1 and 2:1 T:B cell ratios, respectively). There was no direct correlation with CLL T cell-mediated suppression of B cell proliferation and suppression of B cell immunoglobulin synthesis. These studies suggest there is a complex array of abnormal immunoregulatory T cell function in early stage CLL. These include a prominent T helper dysfunction and more variable excessive suppressor activity. The relationship of these findings to the basic disease process remains to be elucidated.
Acute myeloblastic leukemia occurs as a complication of myelodysplastic syndromes, but the appearance of an acute lymphoblastic leukemia (ALL) has only been reported once. We describe a case in which lymphoblastic transformation occurred in the setting of a dysmyelopoietic syndrome. This leukemia was characterized by lymphoid morphology, terminal deoxynucleotidyl transferase (TdT) positivity, cytogenetic abnormalities, and immunoglobulin gene rearrangements. The patient responded to conventional therapy for this leukemia (vincristine and prednisone). Our case supports the hypothesis of a common lymphohemopoietic progenitor and suggests that in vitro tests may help identify a subset of these patients and be important in selecting appropriate therapy.
Normal human B cell proliferation is controlled by various immunoregulatory signals including the T cell-derived lymphokine B cell growth factor (BCGF). Human BCGF provides the final proliferative signal to normal, activated B cells. We herein show that anti-CR2 monoclonal antibodies inhibit human B cell responsiveness to purified BCGF. Addition of anti-CR2 antibody, AB5, was capable of completely inhibiting BCGF-mediated enhancement of either anti-mu or staphylococcal protein A-activated human B cells (191 +/- 21 cpm vs. 3942 +/- 622 cpm, mean +/- SEM). Inhibition of B cell response to BCGF by AB5 occurred in a dose-dependent manner. Monoclonal antibody anti-B2, which recognizes the same 140-kDa glycoprotein as AB5, in comparable concentrations also inhibited B cell responsiveness to BCGF. Monoclonal antibodies of the same subclass (IgG1) showed no inhibitory effect on BCGF enhancement of B cell proliferation. The F(ab')2 fragment of AB5 generated by pepsin digestion was similarly inhibitory as was the intact Ig. AB5-mediated inhibition was independent of the target B cell state of activation. Both resting and activated B cells (anti-mu or staphylococcal protein A activated) incubated with similar concentrations of AB5 were unresponsive to BCGF. The ability of anti-CR2 antibodies to block BCGF-dependent B cell proliferation suggests that occupancy of C3d membrane receptors may result in modulation of B cell proliferation in physiologic or clinical disease states.
Immune cell populations in 8 diffuse histology B-cell lymphomas were analysed in frozen tissue sections by indirect immunofluorescence to gain insight into their possible modulating influence in these tumors. Use of monoclonal antibodies to identify cellular and extracellular antigens combined with nuclear counterstaining allowed precise quantitation, localization and comparison of T- and B-lymphocyte populations. T lymphocytes clustered in non-random fashion. Areas of high T-lymphocyte density manifested higher T4:T8 ratios than locales with fewer T lymphocytes (p less than 0.05). Few cells had surface antigens (Leu 7, 73.1, OKM1) associated with natural killing. Cells strongly reactive with anti-TAC (Interleukin-2 receptor, associated with T-lymphocyte activation) were also T11 reactive and were usually helper (Leu 3) phenotype. In addition, B-lineage lymphoma cells in some tissues reacted with anti-TAC. The pattern of tumor cell reactivity with anti-TAC correlated with Rappaport histologic classification. These findings suggest that non-malignant T lymphocytes modulate B-lymphoma cell growth in situ, and that in some lymphomas the T-cell product IL-2 may be an important local growth factor.
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