Analysis of a family of cyclin-dependent kinase inhibitors: p15/MTS2/INK4B, p16/MTS1/INK4A, and p18 genes in acute lymphoblastic leukemia of childhood

A newly recognized family of proteins that inhibit cyclin-dependent kinases (CDKs) termed cyclin-dependent kinase inhibitors (CDKI) have an important role in regulation of cell-cycle progression. A subfamily of these CDKIs (p15INK4B/MTS2, p16INK4/MTS1, and p18) have a high degree of structural and functional homology and are candidate tumor- suppressor genes. We evaluated the mutational status of the p15, p16, and p18 genes in 103 childhood acute lymphoblastic leukemia (ALL) samples and correlated these results with both their clinical data and additional results concerning their loss of heterozygosity in the region of the p15/p16 genes. Homozygous deletions of the p16 gene occurred extremely frequently in T-ALLs (17/22; 77%), and it was also frequent in precursor-B ALLs (12/81; 15%). Homozygous deletions of the p15 gene were also very frequent in T-ALLs (9/22; 41%), and it occurred in 5 of 81 (6%) precursor-B ALL samples. No deletions of p18 was found in any of the 103 ALL samples. Also, no point mutations of the p15, p16, and p18 genes were detected. We correlated p15/p16 alterations at diagnosis with their clinical characteristics as compared with 2,927 other patients treated similarly. Those with p15/p16 alterations were older; had higher white blood cell counts, often with T-cell ALL phenotype; and more frequently had a mediastinal mass at presentation; but they had the same nonremission, relapse, and survival rates at 5 years as did those patients whose blast cells did not have a p15/p16 deletion. To better understand the extent of alterations affecting chromosome 9p21 (location of the p15/p16 genes), loss of heterozygosity (LOH) was examined at D9S171, which is about 1 megabase proximal to the p15/p16 genes. LOH was detected in 15 of 37 (41%) informative samples. Interestingly, of the 24 informative samples that had no detectable alteration of the p15/p16 genes, 7 samples (29%) had LOH at D9S171. In summary, we show in a very large study that p15 and p16, but not p18, CDKI genes are very frequently altered in ALL; those with p15/p16 alterations are more frequently older children, have higher white blood cells at presentation, and often have a T-cell ALL phenotype. The LOH analysis suggests that another tumor-suppressor gene important in ALL also is present on chromosome 9p21.     

Acquired erythropoietin responsiveness of interleukin-2-dependent T lymphocytes retrovirally transduced with genes encoding chimeric erythropoietin/interleukin-2 receptors

Adoptive immunotherapy with tumor-infiltrating lymphocytes (TILs) causes regression of some human tumors. However, the sustained proliferation and antitumor activity of TILs requires the coadministration of potentially toxic amounts of interleukin-2 (IL-2). In an effort to overcome the requirement by T cells for IL-2, we have introduced alternative growth factor receptors that use the relatively nontoxic cytokine erythropoietin (Epo) as a ligand. In our model system, the coexpression of chimeric receptors consisting of the extracellular portion of the Epo receptor (EpoR) and the intracellular portions of the IL-2 receptor subunits, beta and gamma, conferred Epo responsiveness on a T-cell line. By contrast, cells expressing the wild- type EpoR did not proliferate in response to Epo. This suggested that Epo binding caused the activation of an IL-2 signal pathway mediated by the chimeric receptors. This approach can be used to minimize toxicity and potentially improve cancer immunotherapy with TILs.     

Bone mineral densities in individuals with Gilbert’s syndrome: A cross-sectional,case-control pilot study

BACKGROUND

Unconjugated bilirubin inhibits osteoblastic proliferative activity in vitro, raising the possibility that Gilbert’s syndrome (GS) patients are at increased risk of osteoporosis.

OBJECTIVES

To compare bone mineral density (BMD), serum parathyroid hormone (PTH), C-telopeptide (CTX) and osteocalcin levels in GS subjects versus matched controls in a cross-sectional, case-control study.

METHODS

BMD determinations were obtained with central dual-energy x-ray absorptiometry. Serum PTH, CTX and osteocalcin levels were measured by enzyme immunoassay.

RESULTS

A total of 17 GS and 30 control subjects were studied. Overall, there were no significant differences in BMD, PTH, CTX or osteocalcin levels between the two groups. However, when older (older than 40 years of age) and younger (40 years of age and younger) cohorts were considered separately, the older GS cohort had significantly decreased total hip BMD, T scores and Z scores, and femoral neck BMD, T scores and Z scores (P<0.005 for each parameter, respectively) compared with older control subjects. Serum osteocalcin levels were lower in the older versus younger GS cohort (P=0.006). An inverse correlation existed between all subjects’ serum unconjugated bilirubin levels and total body BMD determinations (r=−0.42; P=0.04). On univariate analysis, the association between serum unconjugated bilirubin and total body BMD was not significant (P=0.066), nor was serum unconjugated bilirubin identified as a risk factor for low BMD when entered into multivariate analyses.

CONCLUSIONS

The results of the present pilot study warrant further research involving larger numbers of subjects and longitudinal measurements to determine whether GS is associated with decreased BMD, particularly in older GS subjects.     

Continuous infusion of the anti-CD22 immunotoxin IgG-RFB4-SMPT-dgA in patients with B-cell lymphoma: a phase I study

IgG-RFB4-SMPT-dgA consists of deglycosylated ricin A chain (dgA) coupled to the monoclonal antihuman CD22 antibody, RFB4. This study determined the maximally tolerated dose (MTD) of this immunotoxin (IT) administered as a continuous 8-day infusion to 18 patients with B-cell lymphoma (30% CD22+ tumor cells) over 8 days. The MTD was 19.2 mg/m2/192 h (maximum toxicity grade 1), with vascular leak syndrome (VLS) as dose-limiting toxicity (DLT) at 28.8 mg/m2/192 h (grades 3 through 5 in 7 of 11 patients). Predictors of severe VLS included serum IT concentrations greater than 1,000 ng/mL and the absence of circulating tumor cells. Decreased urine sodium excreted in 24 hours provided evidence for mild VLS without notable changes in serum albumin. Four partial responses, 3 minor responses, 6 stable disease, and 3 progression of disease were observed. The mean maximal serum concentration (Cmax) in initial courses at the MTD (19.2 mg/m2) was 443 +/- 144 ng/mL (n = 3; range, 326 to 604). At 28.8 mg/m2/192 h, the Cmax was highly variable (n = 11; mean, 1,102 +/- 702; range, 9.6 to 2,032 ng/mL). Human antimouse or antiricin antibodies developed in 6 of 16 (37.5%) patients after one course of IT. However, 10 eligible patients received multiple courses of IT. Changes in serum cytokines and cytokine receptors did not correlate with toxicity but decreased soluble interleukin-2 receptor concentrations correlated with clinical response. Comparison to a prior study with the same IT administered by intermittent bolus infusions (Amlot et al, Blood 82:2624, 1993) suggests similar clinical response, toxicity, and immunogenicity.     

Ureteral jets in healthy subjects and in patients with unilateral ureteral calculi: comparison with color Doppler US

Color Doppler ultrasound was used to image the ureteral jets in 17 healthy subjects and 26 patients with ureteral calculi proved with intravenous urography. In patients with ureteral calculi, three patterns of ureteral jets were seen: no detectable urine flow from the symptomatic side (12 patients), low-level continuous flow from the symptomatic side (four patients), and periodic ureteral jets on the symptomatic side that were not significantly different from ureteral jets of healthy subjects (10 patients). Of the 12 patients with high-grade obstruction on urograms, 11 had ureteral jets significantly different from those of healthy subjects (either no detectable ureteral jets or continuous low-level jets on the symptomatic side). Only three of the 11 patients with low-grade obstruction or nonobstructing stones had ureteral jets that were different from those of healthy subjects. It is concluded that analysis of ureteral jets with color Doppler can enable detection and qualitative determination of the degree of ureteral obstruction in many patients with unilateral ureteral calculi.     

Perivascular color artifact: pathologic significance and appearance on color Doppler US images

A distinctive type of artifactual extravascular assignment of color was identified in 49 color Doppler ultrasound (US) examinations in 43 patients. This artifact appeared as a random localized mixture of red and blue assigned to perivascular soft tissues; the artifact varied with the cardiac cycle, being most prominent in systole and absent or less prominent in diastole. The artifact was seen in 26 patients (32 examinations) at an anastomotic site or stenotic lesion associated with surgically created arteriovenous fistulas for hemodialysis. It was also observed in ten patients with accidental iatrogenic arteriovenous fistulas (renal transplant [n = 6], femoral artery [n = 3], and iliac artery [n = 1]), five with stenotic arteries not associated with arteriovenous fistulas, and two with arterial aneurysms. The authors believe this artifact reflects perivascular tissue vibration caused by turbulent intravascular blood flow. If properly recognized and accurately interpreted, the artifact can be a valuable color Doppler US sign of underlying vascular abnormality.     

Translumbar inferior vena cava catheters: safety and efficacy in peripheral blood stem cell transplantation

Most patients who need peripheral blood stem cell transplantation do not have peripheral venous access that would allow apheresis for stem cell collection. Subclavian apheresis catheters have an unacceptably high incidence of thrombosis-related access failure. A technique has been developed for translumbar placement of permanent, subcutaneously tunneled, silicone rubber apheresis catheters into the inferior vena cava, and 40 of these catheters have been placed in 36 patients for stem cell collection. Twenty-six catheters have been left in place for venous access during the transplantation procedure. These catheters had a very low rate (2.3%) of apheresis-related related complications. Access failure was attributed to thrombosis in 10 catheters (25%) and to mechanical complications in another 9 (22%), but access was regained in all but 4 of these cases. The catheters functioned well as venous access devices during transplantation, only rarely developing complications during that time. Venograms performed at the time of removal of 16 catheters showed no case of caval occlusion. A residual fibrin sheath was found around 14 catheters. There was no clinical or computed tomographic scan evidence of bleeding after placement or removal of the catheters. Percutaneously placed, translumbar inferior vena cava apheresis catheters provide a safe and effective route for the collection of peripheral blood stem cells for transplantation, and they can be left in place for venous access during transplantation.     

Bone mineral content in early-postmenopausal and postmenopausal osteoporotic women: comparison of measurement methods

To investigate associations among methods for noninvasive measurement of skeletal bone mass, we studied 40 healthy early postmenopausal women and 68 older postmenopausal women with osteoporosis. Methods included single- and dual-energy quantitative computed tomography (QCT) and dual-photon absorptiometry (DPA) of the lumbar spine, single-photon absorptiometry (SPA) of the distal third of the radius, and combined cortical thickness (CCT) of the second metacarpal shaft. Lateral thoracolumbar radiography was performed, and a spinal fracture index was calculated. There was good correlation between QCT and DPA methods in early postmenopausal women and modest correlation in postmenopausal osteoporotic women. Correlations between spinal measurements (QCT or DPA) and appendicular cortical measurements (SPA or CCT) were modest in healthy women and poor in osteoporotic women. Measurements resulting from one method are not predictive of those by another method for the individual patient. The strongest correlation with severity of vertebral fracture is provided by QCT; the weakest, by SPA. There was a high correlation between single- and dual-energy QCT results, indicating that errors due to vertebral fat are not substantial in these postmenopausal women. Single-energy QCT may be adequate and perhaps preferable for assessing postmenopausal women. The measurement of spinal trabecular bone density by QCT discriminates between osteoporotic women and younger healthy women with more sensitivity than measurements of spinal integral bone by DPA or of appendicular cortical bone by SPA or CCT.