Bernard J. Poiesz

Detection of Human T-Cell Lymphoma/Leukemia Virus Type I DNA and Antigen in Spinal Fluid and Blood of Patients with Chronic Progressive Myelopathy

The presence of antibodies to human T-cell lymphoma/leukemia virus Type I (HTLV-I) has been associated with chronic progressive myelopathy. We attempted to isolate the virus from the blood and spinal fluid of patients with chronic progressive myelopathy and to define the clinical, radiologic, and electrophysiologic features of this disease. Ten of 13 patients from tropical countries and 2 of 8 from the United States had serum antibodies to HTLV-I. The virus was detected in cultures of peripheral-blood lymphocytes from three of seven patients by means of Southern blot hybridization. Using a sensitive in vitro enzymatic gene-amplification technique, we detected HTLV-I sequences in fresh peripheral-blood mononuclear cells of all of 11 patients tested who were positive for the antibody, and in cell cultures of the spinal fluid from 3 of the 11 tested. Magnetic resonance imaging of the cranium revealed periventricular lesions in the white matter of 3 of the 12 antibody-positive patients. Five of these patients had mild axonal sensorimotor polyneuropathy, and one had bilateral lumbar radiculopathy. Visual evoked potentials were abnormal in three seropositive patients, and brain-stem evoked responses were abnormal in two. The detection of the DNA and proteins of HTLV-I strengthens the proposition that this virus is involved in the pathogenesis of a subset of cases of chronic progressive myelopathy.

Prognostic Significance of K-ras Codon 12 Mutations in Patients With Resected Stage I and II Non–Small-Cell Lung Cancer

PURPOSE The aim of this study was to investigate the prognostic importance of codon 12 K-ras mutations in patients with early-stage non-small-cell lung cancer (NSCLC). PATIENTS AND METHODS We identified 260 patients with surgically resected stage I (n = 193) and stage II (n = 67) NSCLC with at least a 5-year follow-up. We performed polymerase chain reaction analysis of DNA obtained from paraffin-embedded NSCLC tissue, using mutation-specific probes for codon 12 K-ras. RESULTS K-ras mutations were detected in 35 of 213 assessable specimens (16.4%). K-ras mutations were detected in 27 of 93 adenocarcinomas (29.0%), one of 61 squamous cell carcinomas (1.6%), five of 39 large-cell carcinomas (12.8%), and two of 20 adenosquamous carcinomas (10%) (P = .001). G to T transversions accounted for 71% of the mutations. There was no statistically significant difference in overall survival for all patients with K-ras mutations (median survival, 39 months) compared with patients without K-ras mutations (median survival, 53 months; P = .33). There was no statistically significant difference in overall or disease-free survival for subgroups with stage I disease, adenocarcinoma, or non-squamous cell carcinoma or for specific amino acid substitutions. The median survival time for stage II patients with K-ras mutations was 13 months, compared with 38 months for patients without K-ras mutations (P = .03). CONCLUSION Codon 12 K-ras mutations were more common in adenocarcinomas than in squamous cell carcinomas. For the subgroup with stage II NSCLC, there was a statistically significant adverse effect on survival for the presence of K-ras mutations. However, when the entire group was considered, the presence of K-ras mutations was not of prognostic significance in this cohort of patients with resected early-stage NSCLC.

Herpes zoster: a possible early clinical sign for development of acquired immunodeficiency syndrome in high-risk individuals.

Zoster is uncommon before the age of 50 years in immunologically normal individuals, but it occurs with increased frequency in people who are immunosuppressed. A retrospective review of 300 patients with acquired immunodeficiency syndrome associated with Kaposis sarcoma, revealed that 8% had prior zoster, a rate that is sevenfold greater than historic controls of the same age. We prospectively examined forty-eight patients, with no known immunodeficiency or signs of AIDS or AIDS related complex (ARC), who presented with zoster localized to the thoracic region. Forty-one patients had known risk factors for AIDS and thirty-five had antibody to the AIDS-associated virus (AAV) at the time of presentation. One seropositive subject had no known risk factors. Absolute lymphocyte counts, lymphocyte OKT4/OKT8 ratios, and lymphocyte mitogen responses were all depressed in subjects with antibody to AAV when compared with seronegative individuals. Seven of thirty-three AAV antibody-positive subjects, who could be followed longitudinally, developed AIDS from 1 to 28 months (mean = 13) after zoster. One antibody-negative subject seroconverted to become AAV seropositive 16 months after zoster and developed Kaposis sarcoma 1 month later. These eight subjects had persistently low lymphocyte OKT4/OKT8 ratios and elevated beta-2 microglobulin. In patients at risk for AIDS, the occurrence of zoster may be one sign that heralds the marked depression of cellular immunity associated with AIDS or ARC.

Rituximab as an adjunct to plasma exchange in TTP: a report of 12 cases and review of literature.

Idiopathic thrombotic thrombocytopenic purpura (TTP) is caused by the production of autoantibodies against the Von Willebrand factor cleaving enzyme. This provides a rationale for the use of rituximab in this disease. We report a retrospective review of 12 patients treated with rituximab for TTP refractory to plasma exchange. Eleven patients were treated during initial presentation, and one patient was treated for recurrent relapse. Ten patients responded to treatment. Median time to response after first dose of rituximab was 10 days (5–32). Of the 11 patients treated during initial presentation, nine remain free of relapse after a median follow‐up of 57+ months (1+–79+). Two patients died during initial treatment. One patient was lost to follow‐up 1 month after achieving complete response. The patient treated for recurrent disease during second relapse remained disease free for 2years, relapsed and was treated again with rituximab, and was in remission for 22 months. She relapsed again, was retreated, and has now been in remission for 21+ months. We conclude that rituximab is an useful addition to plasma exchange treatment in TTP, but its exact role and dosing need to be verified in prospective studies. J. Clin. Apheresis, 2008.

Epitope mapping of HTLV envelope seroreactivity in LGL leukaemia.

Sera from approximately 50% of patients with large granular lymphocyte (LGL) leukaemia react with a recombinant human T‐cell leukaemia/lymphoma virus (HTLV) transmembrane envelope protein, p21e. Two immunodominant epitopes within env p21e have been defined by reactivity against recombinant proteins GD21 and BA21. In this study sera from 41 patients with LGL leukaemia were examined for reactivity against these recombinant HTLV env proteins. Overall, 21/41 (51%) sera reacted to p21e. Only two sera reacted to GD21. The predominant immunoreactivity against p21e was directed against the BA21 epitope, with 19/41 (46%) sera being BA21 positive. Seroconversion to BA21 protein was also documented. PCR analyses confirmed the low incidence of protypical HTLV sequences (2/41, 5%). These data document an association between BA21 seroreactivity and LGL leukaemia. This finding raises the possibility that such BA21 seroreactivity could be due to cross‐reactivity to a cellular or retroviral antigen sharing some amino acid homology with the transmembrane glycoprotein of HTLV.

Multiple sclerosis, retroviruses, and PCR

Previously reported serologic and polymerase chain reaction (PCR)-based findings have suggested an association between the human retrovirus, HTLV-I, and multiple sclerosis (MS). Due to the inherent ability of PCR to produce false-positive results, we developed a set of physical and procedural safeguards to minimize the possibility of molecular carryover. These were applied as part of a blinded, large-scale, multipopulation, multiplex PCR-based study designed to examine this issue of association. Our results do not support the hypothesis that HTLV-I, which plays a role in the pathogenesis of an encephalomyeloneuropathy, HTLV-II, or closely related agents are associated with MS. A concomitant review of the current literature supports this view.

Large granular lymphocyte leukaemia occurring after renal transplantation

Post‐transplantation lymphoproliferative disorders (PTLD) are a clinicopathologically heterogenous group of lymphoid proliferations. The majority are of B‐cell origin and associated with Epstein‐Barr virus (EBV) infection. In contrast, the development of T‐cell PTLD is much less common and EBV does not appear to be involved in pathogenesis. In this report we describe three patients who developed large granular lymphocyte (LGL) leukaemia after renal transplantation. These patients had clonal expansion of CD3+, CD8+, CD57+, CD56− LGL. We were unable to detect CMV antigen or find evidence for EBV or human T‐cell leukaemia/lymphoma virus genome in the LGL from these patients. These data show that LGL leukaemia should be included as one of the types of T‐cell proliferations which can occur post transplant.

PROTOTYPICAL HTLV-I/II INFECTION IS RARE IN LGL LEUKEMIA

The etiology of LGL leukemia is not known; however, we recently detected HTLV-II in a patient with LGL leukemia. In this study, we found that sera from 6 of 28 patients with LGL leukemia were positive for HTLV-I/II using a whole virus ELISA; moreover, the ELISA-negative sera were near the positive cut-off value. Therefore, we performed additional studies on these sera using commercially available assays which can confirm and distinguish HTLV-I from HTLV-II infection. Serum from only one patient was confirmed positive using conventional criteria (HTLV-II+). Sera from 25 patients (89%) had indeterminate reactivity on Western blot assays. Of these, sera from 21 (84%) reacted to gag protein p24; 12 (48%) reacted with recombinant env protein p21e, and 10 (40%) reacted with both. We could not detect HTLV-I/II pol or pX gene sequences in these patients using polymerase chain reaction analyses, with the exception of the HTLV-II-infected patient described previously. These data show that most patients with LGL leukemia are not infected with prototypical HTLV-I or HTLV-II. The frequent reactivity of patient sera to HTLV-I/II gag protein p24 and to env protein p21e, however, suggests that a deleted or variant form of HTLV-I/II may be associated with LGL leukemia.

Association of ‘(tropical) ataxic neuropathy’ with HTLV-II

Jamaican Neuropathy of the ataxic type (tropical ataxic neuropathy [TAN] and spastic type (tropical spastic paraparesis [TSP]) have been recognized for over a century in Jamaica. The recent association of TSP with HTLV-I (TSP/HAM) is now well established. We now present evidence for a possible association between a TAN-like illness with HTLV-II in four females aged 34-49. All presented with ataxic gait and all four have prominent mental changes. Three of the four also have minor motor deficits with urinary frequency and two have nocturnal leg cramps. All have serum antibody and all had PCR evidence of HTLV-II infection. Antibody to HTLV-II is present in CSF from two subjects. The distinctive picture of prominent ataxia and altered mental status in these subjects contrasts with a predominantly myelopathic picture seen in TSP/HAM.

Low incidence of HTLV infections in random blood donors with indeterminate Western blot patterns

Peripheral blood mononuclear cells (PBMCs) were recovered from platelet units of 61 blood donors who were HTLV‐I positive and 3 blood donors who were HTLV‐I negative on enzyme‐linked immunosorbent assay (ELISA). Western blot analyses were performed on the sera and DNA was prepared from the PBMCs and analyzed by the polymerase chain reaction (PCR). Of the 61 repeatably reactive samples, 2 were positive, 26 were negative, and 33 were interpreted as indeterminate on Western blot. HTLV‐II sequences were detected by PCR in one of the Western blot‐positive samples, as well as in one Western blot‐indeterminate sample that showed reactivity to p24 only. HTLV‐I sequences were detected in the second Western blot‐positive sample. HTLV sequences were not detected in the remaining samples, which suggested that the majority of individuals with indeterminate results on Western blots that used one set of commercially available reagents are not infected with HTLV. It is demonstrated in this study that PCR can be used not only to resolve the infection status of individuals with indeterminate Western blots but also to distinguish between HTLV‐I and HTLV‐II.