Raymond P. Warrell

Chromosomal translocation t(15;17) in human acute promyelocytic leukemia fuses RARα with a novel putative transcription factor, PML

A unique mRNA produced in leukemic cells from a t(15;17) acute promyelocytic leukemia (APL) patient encodes a fusion protein between the retinoic acid receptor alpha (RAR alpha) and a myeloid gene product called PML. PML contains a cysteine-rich region present in a new family of apparent DNA-binding proteins that includes a regulator of the interleukin-2 receptor gene (Rpt-1) and the recombination-activating gene product (RAG-1). Accordingly, PML may represent a novel transcription factor or recombinase. The aberrant PML-RAR fusion product, while typically retinoic acid responsive, displays both cell type- and promoter-specific differences from the wild-type RAR alpha. Because patients with APL can be induced into remission with high dose RA therapy, we propose that the nonliganded PML-RAR protein is a new class of dominant negative oncogene product. Treatment with RA would not only relieve this inhibition, but the activated PML-RAR protein may actually promote myelocyte differentiation.

Differentiation therapy of acute promyelocytic leukemia with tretinoin (all-Trans-retinoic acid)

BACKGROUND AND METHODS Patients with acute promyelocytic leukemia have a characteristic (15;17) translocation, with a breakpoint on chromosome 17 in the region of the retinoic acid receptor-alpha (RAR-alpha). Since this receptor has been shown to be involved with growth and differentiation of myeloid cells in vitro, and since recent clinical studies have reported that tretinoin (all-trans-retinoic acid) induces complete remission in patients with acute promyelocytic leukemia we studied the effects of tretinoin on cellular maturation and molecular abnormalities in patients undergoing the induction of remission with this agent. RESULTS Eleven patients with acute promyelocytic leukemia were treated with tretinoin administered orally at a dose of 45 mg per square meter of body-surface area per day. Nine of the 11 patients entered complete remission. In two patients, complete remission was preceded by striking leukocytosis that then resolved despite continued drug treatment. Serial studies of cellular morphologic features, cell-surface immunophenotypic analysis, and fluorescence in situ hybridization with a chromosome 17 probe revealed that clinical response was associated with maturation of the leukemic clone. All patients who responded to treatment who were tested by Northern blot analysis had expression of aberrant RAR-alpha. As patients entered complete remission, the expression of the abnormal RAR-alpha message decreased markedly; however, it was still detectable in several patients after complete morphologic and cytogenetic remission had been achieved. CONCLUSIONS Tretinoin is a safe and highly effective agent for inducing complete remission in patients with acute promyelocytic leukemia. Clinical response to this agent is associated with leukemic-cell differentiation and is linked to the expression of an aberrant RAR-alpha nuclear receptor. Molecular detection of the aberrant receptor may serve as a useful marker for residual leukemia in patients with this disease.

Complete Remission after Treatment of Acute Promyelocytic Leukemia with Arsenic Trioxide

BACKGROUND Two reports from China have suggested that arsenic trioxide can induce complete remissions in patients with acute promyelocytic leukemia (APL). We evaluated this drug in patients with APL in an attempt to elucidate its mechanism of action. METHODS Twelve patients with APL who had relapsed after extensive prior therapy were treated with arsenic trioxide at doses ranging from 0.06 to 0.2 mg per kilogram of body weight per day until visible leukemic cells were eliminated from the bone marrow. Bone marrow mononuclear cells were serially monitored by flow cytometry for immunophenotype, fluorescence in situ hybridization, reverse-transcription-polymerase-chain-reaction (RT-PCR) assay for PML-RAR-alpha fusion transcripts, and Western blot analysis for expression of the apoptosis-associated proteins caspases 1, 2, and 3. RESULTS Of the 12 patients studied, 11 achieved complete remission after treatment that lasted from 12 to 39 days (range of cumulative doses, 160 to 495 mg). Adverse effects were relatively mild and included rash, lightheadedness, fatigue, and musculoskeletal pain. Cells that expressed both CD11b and CD33 (antigens characteristic of mature and immature cells, respectively), and which were found by fluorescence in situ hybridization to carry the t(15;17) translocation, increased progressively in number during treatment and persisted in the early phase of complete remission. Eight of 11 patients who initially tested positive for the PML-RAR-alpha fusion transcript by the RT-PCR assay later tested negative; 3 other patients, who persistently tested positive, relapsed early. Arsenic trioxide induced the expression of the proenzymes of caspase 2 and caspase 3 and activation of both caspase 1 and caspase 3. CONCLUSIONS Low doses of arsenic trioxide can induce complete remissions in patients with APL who have relapsed. The clinical response is associated with incomplete cytodifferentiation and the induction of apoptosis with caspase activation in leukemic cells.

United States Multicenter Study of Arsenic Trioxide in Relapsed Acute Promyelocytic Leukemia

PURPOSE To determine the safety and efficacy of arsenic trioxide (ATO) in patients with relapsed acute promyelocytic leukemia (APL). PATIENTS AND METHODS Forty patients experiencing first (n = 21) or > or = second (n = 19) relapse were treated with daily infusions of ATO to a maximum of 60 doses or until all leukemic cells in bone marrow were eliminated. Patients who achieved a complete remission (CR) were offered one consolidation course of ATO that began 3 to 4 weeks later. Patients who remained in CR were eligible to receive further cycles of ATO therapy on a maintenance study. RESULTS Thirty-four patients (85%) achieved a CR. Thirty-one patients (91%) with CRs had posttreatment cytogenetic tests negative for t(15;17). Eighty-six percent of the patients who were assessable by reverse transcriptase polymerase chain reaction converted from positive to negative for the promyelocytic leukemia/retinoic acid receptor-alpha transcript by the completion of their consolidation therapy. Thirty-two patients received consolidation therapy, and 18 received additional ATO as maintenance. Eleven patients underwent allogeneic (n = 8) or autologous (n = 3) transplant after ATO treatment. The 18-month overall and relapse-free survival (RFS) estimates were 66% and 56%, respectively. Twenty patients (50%) had leukocytosis (> 10,000 WBC/microL) during induction therapy. Ten patients developed signs or symptoms suggestive of the APL syndrome and were effectively treated with dexamethasone. Electrocardiographic QT prolongation was common (63%). One patient had an absolute QT interval of > 500 msec and had an asymptomatic 7-beat run of torsades de pointe. Two patients died during induction, neither from drug-related causes. CONCLUSION This study establishes ATO as a highly effective therapy for patients with relapsed APL.

The Retinoic Acid Syndrome in Acute Promyelocytic Leukemia

OBJECTIVE To describe a novel complication of therapy with all-trans retinoic acid in patients with acute promyelocytic leukemia. DESIGN Case series. SETTING Comprehensive cancer center. PATIENTS Consecutive patients with a morphologic diagnosis of acute promyelocytic leukemia who underwent remission induction treatment with all-trans retinoic acid, 45 mg/m2 body surface area per day. MEASUREMENTS AND RESULTS Nine of 35 patients (26%; 95% CI, 9% to 52%) with acute promyelocytic leukemia who were treated with all-trans retinoic acid developed a syndrome consisting primarily of fever and respiratory distress. Additional prominent signs and symptoms included weight gain, lower-extremity edema, pleural or pericardial effusions, and episodic hypotension. The onset of this symptom complex occurred from 2 to 21 days after starting treatment. Three deaths occurred; post-mortem examinations in two patients showed pulmonary interstitial infiltration with maturing myeloid cells. Six other patients survived, each achieving complete remission (five patients with all-trans retinoic acid only; 1 patient with chemotherapy). In six of the nine cases, the onset of the syndrome was preceded by an increase in peripheral blood leukocytes to a level of at least 20 x 10(9) cells/L. Certain therapeutic interventions, including leukapheresis, temporary cessation of therapy with all-trans retinoic acid, and cytotoxic chemotherapy in moderate doses were not useful after respiratory distress was established. However, the administration of high-dose corticosteroid therapy (dexamethasone, 10 mg IV intravenously every 12 hours for 3 or more days) early in the course of the syndrome resulted in prompt symptomatic improvement and full recovery in three of four patients. CONCLUSIONS The use of all-trans retinoic acid to induce hematologic remission in patients with acute promyelocytic leukemia is associated in some patients with the development of a potentially lethal syndrome that is not uniformly accompanied by peripheral blood leukocytosis. Early recognition of the symptom complex of fever and dyspnea, combined with prompt corticosteroid treatment, may decrease morbidity and mortality associated with this syndrome.

Immunochemical Characterization of Circulating Parathyroid Hormone–Related Protein in Patients with Humoral Hypercalcemia of Cancer

Tumors from patients with humoral hypercalcemia of cancer produce a parathyroid hormone-related protein (PTHRP). We have developed two region-specific immunoassays capable of measuring PTHRP in plasma: an immunoradiometric assay directed toward PTHRP amino acid sequence 1 to 74 and a radioimmunoassay directed toward PTHRP amino acid sequence 109 to 138. Sixty normal subjects had low or undetectable plasma PTHRP (1 to 74) concentrations (mean, 1.9 pmol per liter) and undetectable PTHRP (109 to 138) concentrations (less than 2.0 pmol per liter). Patients with humoral hypercalcemia of cancer (n = 30) had elevated levels of both PTHRP (1 to 74) (mean, 20.9 pmol per liter) and PTHRP (109 to 138) (mean, 23.9 pmol per liter). The plasma concentrations of immunoreactive PTHRP correlated with the levels of urinary cyclic AMP excreted; in some patients, the concentrations decreased after the tumors were resected. Patients with chronic renal failure (n = 15) had plasma PTHRP (1 to 74) concentrations similar to those in the normal subjects, but their plasma PTHRP (109 to 138) concentrations were elevated (mean, 29.6 pmol per liter). The levels of both peptides were normal in patients with hyperparathyroidism and those with hypercalcemia due to various other causes. Breast milk contained high concentrations of PTHRP. An anti-PTHRP (1 to 36) immunoaffinity column failed to extract PTHRP (109 to 138) immunoactivity from plasma, suggesting that the C-terminal region circulates as a separate peptide. We conclude that plasma PTHRP concentrations are high in the majority of patients with cancer-associated hypercalcemia and that the circulating forms of PTHRP in such patients include both a large N-terminal (1 to 74) peptide and a C-terminal (109 to 138) peptide. Measuring the concentrations of PTHRPs may be useful in the differential diagnosis of hypercalcemia.

Leukocytosis and the Retinoic Acid Syndrome in Patients With Acute Promyelocytic Leukemia Treated With Arsenic Trioxide

PURPOSE Arsenic trioxide, like all-trans-retinoic acid (RA), induces differentiation of acute promyelocytic leukemia (APL) cells in vivo. Treatment of APL patients with all-trans RA is commonly associated with leukocytosis, and approximately 50% of patients develop the RA syndrome. We reviewed our clinical experience with arsenic trioxide to determine the incidence of these two phenomena. PATIENTS AND METHODS Twenty-six patients with relapsed or refractory APL were treated with arsenic trioxide for remission induction at daily doses that ranged from 0.06 to 0.17 mg/kg. RESULTS Twenty-three patients (88%) achieved complete remission. Leukocytosis was observed in 15 patients (58%). The median baseline leukocyte count for patients with leukocytosis was 3,900 cells/microL (range, 1,200 to 72,300 cells/microL), which was higher than that for patients who did not develop leukocytosis (2,100 cells/microL; range, 500 to 5,400 cells/microL; P =.01). No other cytotoxic therapy was administered, and the leukocytosis resolved in all cases. The RA syndrome was observed in eight patients (31%). Patients who developed leukocytosis were significantly more likely to develop the RA syndrome (P <.001), and no patient without a peak leukocyte count greater than 10,000 cells/microL developed the syndrome. Among the patients with leukocytosis, there was no observed relation between the leukocyte peak and the probability of developing the syndrome (P =.37). CONCLUSION Induction therapy of APL with all-trans RA and arsenic trioxide is associated with leukocytosis and the RA syndrome. These clinical effects seem to be intrinsically related to the biologic responsiveness and the differentiation process induced by these new agents.

Phase I and II study of fludarabine phosphate in leukemia: therapeutic efficacy with delayed central nervous system toxicity.

Fludarabine phosphate (9-beta-D-arabinofuranosyl-2-fluoroadenine), a novel purine nucleoside, has demonstrated excellent preclinical antitumor activity and little toxicity in phase I clinical trials. We evaluated the clinical use of fludarabine given as a continuous intravenous (IV) infusion for remission induction in patients with relapsed or refractory leukemia. Thirty infusions were administered to 25 patients. At doses less than or equal to 125 mg/m2/d for five days, only three of 17 patients cleared their bone marrow of leukemic cells, and none achieved complete remission (CR). Nine patients received doses of 150 mg/m2/d for five days or 125 mg/m2/d for seven days. Four of these patients achieved CR (three patients with acute nonlymphoblastic leukemia (ANLL), one patient with acute lymphoblastic leukemia (ALL]. However, severe CNS toxicity was encountered in five patients at the two highest dose levels. Initial symptoms of neurotoxicity were delayed from 21 to 43 days after starting treatment and consisted of optic neuritis, cortical blindness, altered mental status, and generalized seizure. Only one patient regained visual and neurologic function; four other patients experienced progressive neurologic deterioration and died. Clinicopathologic evaluation suggested widespread, severe demyelination as the etiology of these reactions. We conclude that fludarabine is an effective drug for remission induction in acute leukemia. However, doses required to achieve CR are associated with unacceptable CNS toxicity. In view of its potent antileukemic activity, further evaluation of fludarabine at lower doses (less than or equal to 75 mg/m2/d for five days) may be warranted in combination with other chemotherapeutic agents for the treatment of patients with acute leukemia.

Gallium Nitrate for Acute Treatment of Cancer-Related Hypercalcemia: A Randomized, Double-Blind Comparison to Calcitonin

STUDY OBJECTIVE To determine whether gallium nitrate therapy is superior to maximally approved doses of calcitonin for acute control of cancer-related hypercalcemia. DESIGN Randomized, double-blind comparison of active treatments. SETTING Comprehensive cancer center. PATIENTS One hundred ninety-eight consecutive hypercalcemic events in 164 patients screened for entry. ELIGIBILITY CRITERIA hospitalization and intravenous hydration for at least 2 days; persistent elevated serum calcium levels of 2.99 mmol/L or greater (adjusted for serum albumin); serum creatinine levels of 221 mumol/L or less; no cytotoxic chemotherapy, radiation, or mithramycin within the preceding 7 days or during study; no concurrent use of aminoglycoside antibiotics; life expectancy greater than 4 weeks; lymphoma and parathyroid carcinoma excluded. Patients were stratified by histologic type of tumor (epidermoid or nonepidermoid). Fifty patients were randomized and treated. INTERVENTIONS Gallium nitrate 200 mg/m2 body surface area for 5 days by continuous intravenous infusion, or salmon calcitonin 8 IU/kg body weight every 6 hours for 5 days by intramuscular injection. Patients randomized to receive gallium nitrate received sham injections of saline to simulate calcitonin; patients randomized to receive calcitonin received 1000 mL 5%-dextrose solution to simulate gallium nitrate. MEASUREMENTS AND MAIN RESULTS All patients were evaluable. Eighteen of twenty-four patients who received gallium nitrate achieved normocalcemia compared with 8 of 26 patients who received calcitonin for an observed difference of 44% (95% confidence interval, 19% to 69%; P = 0.002). Median duration of normocalcemia before other cytotoxic or hypocalcemic therapy was 6 days for patients treated with gallium nitrate compared with 1 day for patients treated with calcitonin (P less than 0.001). Median duration of normocalcemia regardless of intercurrent treatment and without adjustment for serum albumin was 11+ days for patients treated with gallium nitrate and 2 days for patients treated with calcitonin (P less than 0.01). Mean daily fluid intake and mean daily dose of furosemide were similar in both treatment groups. No additional benefit was seen in 9 patients randomized to receive calcitonin who incidentally received corticosteroids. CONCLUSIONS Gallium nitrate therapy is highly effective and superior to maximally approved doses of calcitonin for acute control of cancer-related hypercalcemia.

Treatment of patients with advanced malignant lymphoma using gallium nitrate administered as a seven‐day continuous infusion

Previous trials of gallium nitrate (NSC‐15200) showed that bolus administration produced dose‐limiting nephrotoxicity without substantial antitumor activity. As an effort to increase the therapeutic index of this compound and to establish a satisfactory out‐patient schedule, the authors evaluated the effects of gallium nitrate administered as a continuous infusion in patients with advanced malignant lymphoma. In an initial Phase I trial, four dose levels which ranged from 200 to 400 mg/m2/day in 27 patients were studied. Nausea which impaired oral hydration was found to be dose‐limiting. A dose of 300 mg/ m2/day was chosen for extended Phase II evaluation and 37 additional patients were entered into the study at that dose level. Overall, 16 of 47 patients (34%) who had bi‐dimensionally measurable parameters of disease achieved major antitumor responses (six of 15 with diffuse “histiocytic” lymphoma, five of ten with diffuse poorly‐differentiated lymphocytic lymphoma, two of five with nodular poorly‐differentiated lymphocytic lymphoma, and three of 17 with Hodgkins disease). The median duration of response was 2.5 months. Only 8% of patients who received 300 mg/m2/day developed an increase in serum creatinine concentration >1.1 mg/dl over baseline values. Hypocalcemia occurred in two‐thirds of patients. Other toxic effects, including paresthesiae, diarrhea, and hearing loss, were noted in <5% of patients. There was minimal myelosuppression. The authors conclude that gallium nitrate administered as a continuous infusion for seven days at 300 mg/m2/day is well‐tolerated and effective treatment for patients with advanced malignant lymphoma. Outpatient administration using portable infusion pumps is safe and practical. Further evaluation of the drug administered as a constant infusion is indicated in patients with other neoplastic diseases.