A phase III placebo-controlled study in advanced head and neck cancer using intratumoural cisplatin/epinephrine gel.

Patients with recurrent or refractory head and neck squamous cell carcinoma received cisplatin/epinephrine injectable gel or placebo gel injected directly into the clinically dominant tumour. The double-blind phase III trial comprised of up to 6 weekly treatments over 8 weeks, 4 weekly evaluation visits, and then monthly follow-up; open-label dosing began as needed after three blinded treatments. Tumour response was defined as complete (100% regression) or partial (50-99% regression) sustained for > or =28 day, and patient benefit as attainment of palliative or preventive goals prospectively selected by investigators and patients. With cisplatin/epinephrine gel, 25% (14 out of 57) of tumours responded (16% complete regression, 9% partial regression), vs 3% (one out of 35, complete regression) with placebo (P=0.007). Patient benefit was positively associated with target tumour response in the blinded period among cisplatin/epinephrine gel recipients (P=0.024): 43% (six out of 14) of responders benefited, vs 12% (five out of 43) of non-responders. The most frequent adverse event was pain during injection and the next most frequent was local cytotoxic effects consistent with the gel's mode of action. Systemic adverse events typical of intravenous cisplatin were uncommon. Intratumoural therapy with cisplatin/epinephrine gel provided safe, well-tolerated, effective palliative treatment for patients with locally advanced head and neck squamous cell carcinoma, who lack other satisfactory treatment options.

The significance of CD34 and TdT determinations in patients with untreated de novo acute myeloid leukemia.

The results of intensive chemotherapy given to 247 adults at the University of Maryland Cancer Center with previously untreated de novo acute myeloid leukemia (AML) were reviewed with respect to expression of terminal deoxynucleotidyl transferase (TdT) and CD34. Of the 228 patients with data for TdT, 32 (14%) had > 5% of the leukemia cells positive by an immunofluorescence assay. The median age of the TdT-positive patients was approximately 10 years less than the TdT-negative patients (50 versus 60 years). Patients with TdT-positive AML had similar median survival (12 versus 10.5 months) and complete remission (CR) rates (53 versus 59%), but a greater frequency of long-term complete responders (60 of complete remitters versus 20%, p = 0.08) than TdT-negative patients. Of 126 patients tested, 59% were CD34-negative (< 20% reactivity with leukemia cells). These 74 patients (median age 60 years) had a greater CR rate (71 versus 48%, p = 0.008) than the 52 CD34-positive patients (median age 60 years), and improved survival (p = 0.013 by Wilcoxon) although there was no difference in the duration of CR between the CD34-positive and negative groups. Of CD34-positive patients 12/52 remain in continuous CR, and 16/74 CD34-negative patients remain in continuous CR. None of eight patients strongly positive for CD34 (> 70% reactivity) remain disease-free. Positivity for TdT or CD34 was associated with less differentiated AML. Of CD34-positive patients, 44% had FAB M0/M1 morphology versus 13% of CD34-negative patients (p = 0.0001); similarly, 47% of TdT-positive patients were FAB M0/ML1 versus 25% of TdT-negative patients (p = 0.01). Of seven patients with FAB M4E0, five were CD34-positive. Of the 12 CD34-positive survivors, four had FAB M4E0. Thus CD34 expression predicts for CR rate and overall survival in adults with AML. TdT expression does not significantly affect overall outcome but may be associated with longer CR durations.

Minimally differentiated acute nonlymphocytic leukemia: a distinct entity.

Ten of 136 consecutive adult patients with previously untreated acute leukemia had morphologically undifferentiated leukemia by light microscopy. Leukemic cells from these patients were characterized by agranular cytoplasm, negative histochemical staining with sudan black (SB) and nonspecific esterase, and absent lymphoid cell surface markers and therefore were not classifiable according to the French-American-British (FAB) system. Electron microscopy with myeloperoxidase (MPO) staining revealed the presence of peroxidase positive cytoplasmic granules and endoplasmic reticulum in eight of the nine patients studied. Cells from the patient who was negative for MPO were also negative for platelet peroxidase. A series of monoclonal antibodies to myeloid antigens also revealed myeloid features with all patients having at least one myeloid differentiation antigen present on the surface of their cells. Common acute lymphoblastic leukemia (ALL) antigen was absent in the nine patients tested. Cytogenetic analysis of blast cells was abnormal in seven patients on whom adequately banded chromosomes were obtained although there were no consistent abnormalities. No patient had a Ph1 chromosome. Only two of the ten patients achieved a complete remission. Morphologically undifferentiated leukemia may have myeloid features when studied by transmission electron microscopy or with monoclonal antibodies for cell surface markers. Such studies should be performed when the leukemia cannot be classified using either light microscopy or lymphoid cell surface markers. Such patients infrequently achieve remission with standard therapy and constitute a distinct entity.

Effects of interferon and retinoic acid on the growth and differentiation of clonogenic leukemic cells from acute myelogenous leukemia patients treated with recombinant leukocyte-alpha A interferon.

Studies with human myeloid leukemia cell lines indicate that combined interferon (INF) and retinoic acid (RA) have greater effects in inhibiting cell growth and in inducing terminal differentiation than either agent alone. Consequently, we studied the effects of these agents, singly and in combination, on fresh leukemic blast cells obtained from 13 acute myelogenous leukemia (AML) patients, most of whom were subsequently treated with recombinant leukocyte-alpha A interferon (rINF-alpha A). The in-vitro response to rINF-alpha A and RA was assessed in an established myeloid leukemic blast cell clonogenic assay containing conditioned medium from phytohemagglutinin-stimulated lymphocytes. Strong inhibition of colony cell growth (greater than or equal to 50%) was observed in 4/10 cases treated with rINF-alpha A alone, but only at high concentration (greater than or equal to 2500 U/ml) and in 4/10 cases treated with RA alone (5 X 10(-8) M or 5 X 10(-7) M). Combined rINF-alpha A and RA augmented the inhibition of primary or secondary colony cell growth in 5/8 evaluable cases. Stimulation of leukemic cell differentiation was observed in 1/8 cases by rINF-alpha A alone and in 4/7 cases by RA alone. Combined rINF-alpha A and RA enhanced cell differentiation in 4/7 cases. In addition, increased inhibition of clonal cell growth and/or differentiation by RA alone was observed in 2/5 cases following in-vivo rINF-alpha A treatment. These results suggest that treatment with combined rINF-alpha A and RA may be rewarding in some cases of AML.

Acquired chromosome rearrangements, including fine interstitial deletions, in a patient with Down syndrome and monoblastic leukemia.

A female child with Down syndrome who developed acute monoblastic leukemia is reported. Anemia associated with milk leukopenia was first recognized when the patient was 14 months old. Acute monoblastic leukemia was diagnosed 1 year later; cytogenetic studies were performed on circulating leukemic cells at this time. Analysis of elongated, finely banded chromosomes revealed three structural rearrangements, including two rather subtle interstitial deletions, in addition to trisomy 21 which was representative of the patient's constitutional karyotype. The karyotype of the leukemic cells was 47,XX,+21,t(3;18)(p23;q11.2), del(7)(q31.1q31.3), del(9)(p22p24 or p21p23). The patient received no cytostatic chemotherapy and died 4 months after the diagnosis of acute leukemia was made.

Unusual karyotypic changes and B cell involvement in a case of lymph node blast crisis of chronic myelogenous leukemia.

A patient with Philadelphia chromosome (Ph1) positive chronic myelogenous leukemia (CML) entered a blast crisis localized to lymph nodes. On light microscopy, by morphology and histochemical staining, the blasts were undifferentiated. In spite of terminal deoxynucleotidyl transferase positivity, some of the lymph node cells expressed a myeloid differentiation antigen, OKM1, and were peroxidase positive by transmission electron microscopy (TEM). However, the majority of cells were peroxidase negative on TEM and expressed OKT-10, a marker found on both primitive myeloid and lymphoid cells. Cultures of lymph node cells stimulated with Epstein-Barr virus or lipopolysaccharide (LPS) revealed the Ph1, indicating B cell involvement in the CML. T cells from cultures stimulated with L4-phytohemagglutinin and T cell growth factor were negative for the Ph1. In unstimulated lymph node cells, the uncomplicated Ph1 could not be demonstrated; instead, a unique complex karyotype involving a masked Ph1 was identified in these and the LPS cultures. This karyotype was not found in bone marrow (BM) metaphase cells. Instead, BM cells showed either the simple Ph1 or the Ph1 with a rearrangement involving chromosomes 13 and 20. The patient had transient responses to three chemotherapy regimens, two of which were designed to treat acute lymphocytic leukemia, but he died 8 months after disease acceleration without BM blast crisis. These findings are compatible with an extramedullary blast crisis originating in a primitive cell with both myeloid and lymphoid characteristics.

Mitogenic and mitogenically defective phytohemagglutinin isolectins stimulate T-cell growth factor (interleukin 2) production and response in fresh and cultured human T lymphocytes.

L4-PHA (L4) and E4-PHA (E4) lectins isolated from Phaseolus vulgaris have different mitogenic properties. The mechanisms of the differences in mitogenic behavior were sought in the interaction of lectin, lymphocyte subsets, and T-cell growt factor (TCGF) also known as interleukin 2 (IL-2). TCGF activity in culture supernatants ( L4S ; E4S ) from L4- and E4-stimulated, freshly isolated lymphocytes was assayed as stimulation of DNA synthesis in TCGF-dependent continuous T-cell cultures (CTC). E4S contained less TCGF than did L4S . Addition of partially purified TCGF does not increase the stimulation of fresh lymphocytes by L4 or E4. L4 and E4 equally stimulate both helper (OKT4+) and suppressor (OKT8+) cells. The ability of L4 to further stimulate CTC is slowly lost (15 greater than 30 greater than 45 days). It is concluded that production of TCGF is not rate limiting in E4 and L4 stimulation of lymphocytes. The growth of CTC, which requires the presence of TCGF, remains sensitive to, but not dependent on, L4 for at least 30 days.

Searching for the cause of the acquired immune deficiency syndrome.

An outbreak of unexplained immune deficiency associated with opportunistic infection and Kaposi's sarcoma is occurring in the USA and other parts of the world. Affected individuals with what had come to be known as the acquired immune deficiency syndrome (AIDS) have a high mortality. Epidemiological features suggest the presence of a transmissable agent, but no responsible agent has yet been identified. Homosexual and bisexual men make up 75% of these affected individuals. Cytomegalovirus, Epstein Barr and herpes simplex viruses, organisms that commonly affect male homosexuals, may produce some features of AIDS. Individually or collectively, however, they can not account for the emergence of a previously unrecognized clinical syndrome. Hepatitis B is prevalent in patients with AIDS and may play a role as a co-factor in the disease. The properties of a number of other known viruses may provide a model for the pathogenesis of some features of the AIDS immunodeficiency. Newly described simian acquired immune deficiency syndrome (SAIDS) is the best available animal model. In man, the retrovirus, human T-cell leukemia virus (HTLV) may play a role in AIDS. However, HTLV or any other known virus cannot yet be assumed to cause AIDS. It is likely that an as yet unrecognized agent is the key causative agent of AIDS.

Acquired immune deficiency syndrome presenting as oral pharyngeal and cutaneous Kaposi's sarcoma.

A 25-year-old black male homosexual with AIDS presented with Kaposi's sarcoma of the tongue, palate and skin. The definition, epidemiology, diagnosis, treatment and prognosis of AIDS are discussed. The role of the otolaryngologist-head and neck surgeon in diagnosing this disease is outlined.

Phytohemagglutinin isolectin subunit composition.

The subunit compositions of individual phytohemagglutinin isolectins from red kidney bean Phaseolus vulgaris were examined by isoelectric focusing and sodium dodecyl sulfate electrophoresis on polyacrylamide gels. Isoelectric focusing reveals heterogeneous but unique and non-overlapping protein band patterns for each of the homotetrameric isolectins, E4 and L4. Isoelectric focusing of the intermediate isolectins which contain both subunits (E3L1, E2L2, and E1L3) show all the protein bands common to isolectins E4 or L4 in proportions relative to their suggested subunit compositions. Polyacrylamide gel electrophoresis in a continuous sodium dodecyl sulfate buffer system gives a single protein band for all of the isolectins. In contrast, a discontinuous sodium dodecyl sulfate buffer procedure resolves isolectins E4 and L4 into single major protein bands of apparent molecular weights 31 700 (+/-600) and 29 900 (+/-200), respectively. Each of the intermediate isolectins contained both protein bands and their relative proportion, as determined by absorbance scanning, confirms the phytohemagglutinin isolectin subunit compositions as E4, E3L1, E2L2, E1L3, and L4.

Phaseolus vulgaris isolectin binding to human erythrocytes.

The Phaseolus vulgaris isolectins L4,L3E1, L2E2, L1E3, and E4 were isolated by affinity and ion exchange chromatography. Pure isolectins were radiolabeled by the chloramine-T method with Na125IO4 and their binding to human erythrocytes was studied. A normal erythrocyte has approximately 8 times 10(5) receptor sites for each isolectin; however, the association constants (Ka) of binding increased from 1.1 times 10(7) M-1 to 3.8 times 10(8) M-1, with increasing number of E subunits per tetrameric isolectin molecule. Isolectin to erythrocyte binding reached equilibrium rapidly and was reversed by fetuin. All isolectins competed with 125I-E4 for erythrocyte binding sites, with a constant (KI) similar to the Ka calculated for each respective radiolabeled isolectin. When isolectin binding at 0 degrees C, 4 degrees C, or 8 degrees C was compared to that at 25 degrees C, there was no reduction in the number of binding sites per cell, but the Ka of E4 was reduced to 3 times 10(7) M-1. Fixed erythrocytes displayed similar isolectin binding characteristics.

Biological and biochemical properties of Phaseolus vulgaris isolectins.

Affinity-purified phytohemagglutinin from red kidney bean resolves into five isolectins by SP-Sephadex ion exchange chromatography. Recoveries ranging from 30 to 130 mg of protein for each isolectin are easily achieved. The isolectins have similar amino acid compositions which differ only in threonine, lysine, and arginine. A distinguishing feature of the amino acid composition is the total lack of sulfur-containing amino acids. Each isolectin contains about 4% mannose and 2.2% N-acetyl-D-glucosamine. All isolectins on electrophoresis form single protein bands under denaturing and nondenaturing conditions in polyacrylamide gels, and all have apparent subunit molecular weights of 33,000 by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The isolectins are also homogeneous by ultracentrifugation and have apparent native molecular weights of 115,000 +/- 4,130, suggesting tetrameric quaternary structures. Whereas 80% of the starting erythroagglutinin activity is recovered, one of the five isolectins possesses 50% of that original activity. As sequentially eluted from the ion exchange column, each isolectin displays progressively higher erythroagglutinin and lower lymphocyte mitogenic activities. Based on their relative biological activities, the isolectins are assigned the structures L4, L3E1, L2E2, L1E3, and E4, where L and E represent lymphocyte- and erythrocyte-reactive subunits, respectively, and the subscripts represent the proposed subunit composition.

Recombinations of subunits of Phaseolus vulgaris isolectins.

Phaseolus vulgaris phytohemagglutinin is formed in vivo by the combination of erythrocyte (E)-reactive and lymphocyte (L)-reactive subunits into five tetrameric isolectins:L4,L3E1, L2E2, L1E3, and E4. Evidence for phytohemagglutinin subunit structure is obtained by in vitro dissociation of native isolectins in 6 M guanidine HCl followed by removal of dissociating agents to allow subunit recombination. Dissociation and recombination of L4 yielded a single protein, electrophoretically indistinguishable from the native L4. Similar treatment of E4 also yielded a single protein indistinguishable from native E4. Treatment of L3E1, L2E2, L1E3, or a mixture of L4 and E4, yielded five distinct proteins electrophoretically similar to all five native phytohemagglutinin isolectins. Milligram quantities of all five recombinant isolectins were prepared either from L2E2 or a mixture of L4 and L1E3 proportioned to yield equimolar quantitives of the two subunits on dissociation. The recombinant isolectins were purified by affinity and SP-Sephadex ion exchange chromatography. Electrophoretic and chromatographic properties and the erythroagglutinating and mitogenic activities of recombinant isolectins were essentially identical with the native isolectins. The inclusion of 125I-labeled L4 in the dissociation results in a distribution of 125I-labeled L subunit among the purified recombinant isolectins proportional to their proposed subunit structures.

Purification of the phytohemagglutinin family of proteins from red kidney beans (Phaseolus vulgaris) by affinity chromatography.

Half-gram quantities of phytohemagglutinin lectins are purified from saline extracts of red kidney beans (Phaseolus vulgaris) by affinity absorption on porcine thyroglobulin-Sepharose. All of the mitogenic and erythroagglutinin activity of the saline extract is removed by this absorbent, and 74% of the original erythroagglutinating activity elutes from the affinity absorbent representing a 25-fold purification. Five distinct proteins appear in the polyacrylamide gel electrophoresis of the affinity absorbent eluate. Although all five proteins specifically bind to porcine thyroglobulin, the cathodal migrating proteins bind more strongly than the anodal migrating proteins. The most cathodal proteins are potent erythroagglutinins. This simple, efficient method is used to prepare all the active components of the phytohemagglutinin family in large yield and high purity.