Leo L. Stolbach

Ectopic Production of an Alkaline Phosphatase Isoenzyme in Patients with Cancer

Abstract An alkaline phosphatase isoenzyme (Regan isoenzyme) has been identified in the serum of 27 patients with various malignant tumors. This isoenzyme is biochemically and immunologically indistinguishable from placental alkaline phosphatase. In addition to being present in serum, it has been identified in both tumor tissue and malignant effusion fluids. Measurement of this isoenzyme has been clinically useful in monitoring progression or regression of tumor, identifying a source of serum alkaline phosphatase elevation, and identifying malignant effusions.

Serum alkaline phosphatase in hypophosphatasia

It is recognized that serum alkaline phosphatase may reflect enzyme contributions from bone, liver, and intestine. We have investigated serum alkaline phosphatases in two siblings with hypophosphatasia. After administration of long-chain triglycerides, the major alkaline phosphatase component of their sera was shown to be of intestinal origin on the basis of inhibition by l-phenylalanine. Starch block electrophoresis suggested that there were other regions of l-phenylalanine-sensitive alkaline phosphatase in addition to the major slow-moving intestinal band. Medium-chain triglycerides which are absorbed by the portal route did not cause a similar augmentation of intestinal alkaline phosphatase activity. These studies indicate that serum levels of intestinal alkaline phosphatase are increased normally after long-chain fat feeding in hypophosphatasia and may be the major component of total serum alkaline phosphatase activity.

Cooperative groups and community hospitals. Measurement of impact in the community hospitals

The Eastern Cooperative Oncology Group, composed of major cancer treatment centers, has an outreach program which involves community hospitals in ongoing cancer clinical trials. A prevalence survey was carried out in February 1981 among 104 community hospitals and 21 member institutions to determine the characteristics of patients being treated, their staffing, and reasons why patients were not on protocol studies. The survey sampled 25 (50) consecutive patients from community hospitals (member institutions). The purpose of the study was to assess the impact of a community cancer control program. The results of the study demonstrated that 16% of patients surveyed in the affiliated community hospitals were being treated on a research protocol. In addition, a further 35% had their treatment plan influenced by a protocol. Consequently protocols have impacted directly or indirectly on 51% of the patients. The corresponding figures in member institutions were 23% and 38% for a total of 61%. In studying protocol availability, it was found that 66% of all patients were ineligible for any protocol. Of patients eligible for a protocol but not registered on one, 52% were not registered because of physician preference for a specific treatment. The affiliates surveyed were shown to be on average half as large as member institutions in terms of number of beds and staff size. Also, staff/patient ratios are generally smaller in the community hospitals. The median age of patients was considerably lower than SEER incidence data. Also, elderly patients were slightly more prevalent in community hospitals than in member institutions. A clear relationship between disease stage and age in breast cancer patients was noted with the representation of early‐stage disease much higher in young women.

Correlation of Serum l-Phenylalanine-Sensitive Alkaline Phosphatase, Derived From Intestine, with the Abo Blood Group of Cirrhotics

Summary Serum levels of l-phenylalanine-sensitive alkaline phosphatase (LPSAP) were determined in 139 patients with hepatic cirrhosis. A significant correlation exists between the ABO blood type and the ratio of LPSAP to total alkaline phosphatase. Higher mean levels of total alkaline phosphatase were found in patients with blood type O than in patients with type A, and these elevations were due in large part to higher mean levels of the l-phenylalanine-sensitive isoenzyme. The intestinal origin of a major portion of LPSAP is suggested by starch gel electrophoresis of serum which revealed a slowmoving band which was inhibited by l-phenylalanine and which migrated to the location of purified intestinal alkaline phosphatase. The findings support a genetic mechanism controlling the serum levels of l-phenylalanine-sensitive alkaline phosphatase, and in turn total alkaline phosphatase, and contribute to a more precise interpretation of the significance of hyperphosphatasemia.

Comparison of intensive versus moderate chemotherapy of lymphocytic lymphomas. A progress report

In an Eastern Cooperative Oncology Group trial, Cytoxan‐prednisone (CP) Induction was compared to BCNU‐prednisone (BP) in 273 patients with lymphocytic lymphoma. Response rates were comparable, with 21% achieving complete response and 40%, partial response. Patients with a nodular pattern responded better. Maintenance phase comparing cyclic intensive therapy (BCVP) with intermittent chlorambucil revealed the superiority of BCVP as demonstrated by improvement of the quality of response and somewhat longer remissions. The value of the Rappaport classification in the evaluation of lymphoma chemotherapy results is discussed. It is suggested that NHL be separated into “favorable” and “unfavorable” groups, based on the presence or absence of nodularity and treatment schedules devised accordingly.

CLINICAL APPLICATION OF ALKALINE PHOSPHATASE ISOENZYME ANALYSIS

In the past, an elevated alkaline phosphatase has usually been taken as evidence of either bone or liver disease. However, with the development of the isoenzyme techniques described in detail by other participants in this symposium, it has become possible to differentiate with a fair degree of accuracy not only the isoenzymes of bone and liver but also those originating in intestine, placenta and tumor. A combination of isoenzyme analysis techniques have been used in our laboratory to study the serum alkaline phosphatase of patients with various diseases. The use of one or more of these techniques will be described for the following clinical situations: ( 1 ) patients with cirrhosis: (2) patients with tumor in bone or liver; and (3) patients with malignancy whose serum and/or tumor contains the Regan isoenzyme. These studies have made it clear that isoenzyme analysis can result in more accurate interpretations of serum alkaline phosphatase values. TABLE 1 summarizes the characteristics of the alkaline phosphatases originating in the various tissues known to be sources of serum alkaline phosphatase. The alkaline phosphatases of liver and bone are only minimally inhibited by L-phenylalanine, whereas intestinal and placental alkaline phosphatase and Regan isoenzyme are inhibited 77 to 79%. Heat inactivation at 55 C for 16 .minutes causes 90 to 100% inhibition of bone alkaline phosphatase but no inhibition of placental or Regan isoenzyme, whereas liver and intestinal alkaline phosphatase are inhibited 50 to 70% under these conditions. A more drastic condition of heat inactivation at 65 C for five minutes causes complete inhibition of bone, liver, and intestinal alkaline phosphatase but no inhibition of Regan or placental isoenzyme. As has been previously shown, both Regan and placental isoenzyme migrate after starch gel electrophoresis to a position between the bone-liver band and the intestinal band. This band disappears after mixing with antiserum to placental alkaline phosphatase. These isoenzyme characteristics have diagnostic applicability, since the percent inhibition by L-phenylalanine helps to identify patients whose sera contain intestinal alkaline phosphatase, Regan isoenzyme or placental isoenzyme; heat inactivation at 55 C is a useful aid in diagnosing bone or liver disease; and heat inactivation at 65 C identifies patients whose sera contain Regan or placental isoenzyme. At present, all of these determinations are carried out on each serum tested in our laboratory.

Treatment of histiocytic and mixed lymphomas: a comparison of two, three and four drug chemotherapy.

The Eastern Cooperative Oncology Group has studied 187 patients with generalized progressive malignant lymphoma classified as having the histologic sub‐types histiocytic or mixed. Histology review by the Pathology Panel for Lymphoma Clinical Trials demonstrated a 31% disparity with contributing institutions pathologists in regard to cell type, but good agreement with interpretation of nodular or diffuse nodal pattern. Patients were assigned at random to treatment with cyclophosphamide 1 g/m2 on day 1 and prednisone 100 mg/m2 daily for five days (CP); CP plus vincristine 1 mg/m2 on day 1 (CVP); or CVP plus BCNU 60 mg/m2 on day 1 (BCVP). Chemotherapy was given for nine, twenty‐one day cycles. The observed complete remission rates were CP‐21%, CVP 34%, BCVP 34%. Both CVP and BCVP had significantly more complete remissions than CP, but survival following CVP (118 weeks) was significantly longer than that following either BCVP (76 weeks) or CP (74 weeks). Histologic sub‐type, lymph node pattern, response to chemotherapy, performance status and stage of disease were also found to influence survival.

Combination chemotherapy of the malignant lymphomas. A Controlled clinical trial

The Eastern Cooperative Oncology Group has studied 113 patients with generalized progressive malignant lymphomas in a randomized clinical trial. Pathologic diagnosis was subclassified by cell type and nodal pattern by The Pathology Panel for Lymphoma Clinical Trials. Patients were randomly assigned treatment with either cyclophosphamide (C), vincristine (O), and prednisone (P) (COP) or CO without prednisone. Initial treatment was given for 8 weeks and further randomization of responders to observation or additional chemotherapy was carried out. A significant difference in complete remission rate between treatments was shown: with COP, 43%, and with CO, 17%, indicating an important role for prednisone in inducing CR. COP was also associated with longer remission durations and improved survival. Complete remission following initial chemotherapy is also associated with longer duration of disease‐free time and survival. The initial pathologic cell types and nodal pattern also strongly influence survival. Extended “maintainence” CO treatment improved disease‐free remission duration, but not survival.

Pretreatment radiographic evaluation of patients with malignant melanoma.

During a four year period, 53 patients with malignant melanoma underwent extensive pretreatment radiographic evaluation for detection of occult extranodal metastatic disease. This included chest x‐ray with tomography, upper G.I. series with small bowel follow through, barium enama, intravenous pyelogram, and radio‐nuclide scans of the brain, liver and bone. All occult metastatic disease in asymptomatic patients was discovered on routine chest x‐ray examination. There were three false positive examinations, which necessitated further diagnostic tests but there was no change in the final treatment decision. There was no alteration in the management of the remainder of the patients on the basis of the pretreatment radiographic evaluation.

The effect of fatty acid perfusion on intestinal alkaline phosphatase

The purpose of this and the following article is to document experimental evidence of the effect of fatty acid absorption and transport in the small-bowel mucosa on the activity of the intestinal isoenzyme of alkaline phosphatase. The first paper deals with perfusions of fatty acids of increasing carbon length into the duodenum of volunteers, studying changes in luminal concentrations and mucosal location of intestinal alkaline phosphatase. In the second paper, which follows, the preliminary observations in human studies have been investigated in more detail in a rat model. In 15 normal volunteers a segment of proximal small bowel was isolated by a four-lumen, two-balloon tube technique and perfused with micellar solutions of fatty acids (C8∶0, C12∶0, C18∶1) in a taurocholate solution. In control experiments the perfusion fluid lacked fatty acids. The luminal contents were enriched with intestinal alkaline phosphatase over control values as a function of the carbon-chain length of the fatty acids. The extent of sucrase release was less pronounced and that of glycylglycine dipeptidase was negligible. The effect of intraluminal oleic acid on the location of mucosal alkaline phosphatase activity was studied by histochemical techniques in small-bowel biopsies obtained in six additional subjects. Intestinal alkaline phosphatase activity appeared to be in the same location as the dynamic lipid transport, moving from the brush border into the mucosal lymphatics. Our studies indicate bidirectional flow of brush-border intestinal alkaline phosphatase into the bowel lumen and into the intestinal lymphatics during fatty acid absorption.