Metabolism of Neoplastic Tissue VII. Effects of Dinitrophenol and Fluoride on Glucose Oxidation in Tumor Homogenates*

CHARLES E. WENNERf AND SIDNEY WEINHOUSE

(Lankenau Horpital Re8earch In8titute and the 1n3t@tuiefor Cancer Re8earch, PhÃ¼adelphia 11, Pa.)

As yet, the high aerobic and anaerobic glycoly his co-workers as a result of an extensive investi sis, an outstanding characteristic of neoplastic gation of phosphate metabolism in normal and cells, remains unexplained. This phenomenon has neoplastic tissues (12, 14â€”16).Potter and Lyle been variously attributed to disturbances of res (15) found that homogenates of such nonneoplas piration, to deficiencies of electron transport en tic tissues as liver, heart, and kidney consumed zymes or co-factors, and to a lack of the citric acid oxygen in the presence of oxalacetate and exhibit cycle (2, 4, 14, 23). However, none of these hy ed relatively low rates of splitting of organic phos potheses has decisive experimental support, and phate. On the other hand, homogenates of the against them can be cited the observations that Walker 256 carcinoma did not absorb oxygen but tumors as a class possess the same electron trans split ATP rapidly. It was reasoned that excessive port components and consume oxygen about as ATP breakdown in the Walker tumor did not al rapidly as do moderately active normal tissues, low maintenance of ATP-dependent oxidative and oxidize fatty acid and carbohydrate interme processes. A key observation in the formulation of diates via the citric acid cycle (10, 24). this theory was the divergent behavior of fluoride, In a search for other explanations for the high which inhibited oxygen consumption in kidney glycolysis of tumor cells, the possibility that this homogenates but stimulated oxygen uptake in type of tissue differs from the non-neoplastic in its homogenates of the Flexner-,Jobling tumor (11). capacities for synthesis and breakdown of phos This finding was explained on the basis of the well phate bonds deserves consideration. The participa known inhibition by fluoride of phosphatase ac tion of phosphorylated intermediates as well as tion; it was reasoned that oxidative activity is lim nucleotides, such as adenosine di- and triphos ited in tumors by excessive dephosphorylation and phate (ADP and ATP), in many steps of glucose is therefore enhanced by fluoride, whereas oxida catabolism implies that the balance between phos tive activity in normal tissues is limited by exces phorylative and dephosphorylative rates might sive phosphorylation and is therefore inhibited by strongly influence the rate of glycolysis. The rapid fluoride. As a further test of this theory, Siekevitz growth of tumors undoubtedly depends on a varie and Potter (19) studied the effects of 2,4-dinitro ty of synthetic processes which require the utiliza phenol (DNP) and fluoride on the oxidation of tion of phosphate bond energy. Thus, the high pyruvate and fumarate in homogenates and mito synthetic activity associated with neoplasia may chondria of various normal and neoplastic tissues. be a primary factor affecting the balance between It was anticipated that DNP, which accelerates phosphate uptake and release, affecting, in turn, ATP breakdown, would act antagonistically to the rate of glucose catabolism. fluoride. No consistent patterns of response to The hypothesis of an altered phosphate metabo these agents were observed, however, and it was lism in neoplasia has been advanced by Potter and concluded that possible differences in phosphoryla tive and dephosphorylative activities may have * Aided by grants from the National Cancer Institute, Public Health Service, the American Cancer Society, and the been largely overshadowed by individual varia United States Atomic Energy Commission, Contract No. tions in enzymatic constitution within each tissue AT(30-1)777. type. t Most of this work was conducted during the tenure, by In view of the importance of this problem to a C. E. W., of a Damon Runyon Memorial Fund Fellowship. proper understanding of carbohydrate metabolism Received for publication January 7, 1955. in neoplasia, a further test of this hypothesis 497

seemed desirable, particularly a test of the effects thickness.â€• They are reported as patoms of labeled glucose of dinitrophenol and fluoride specifically on the carbon converted to CO2. calculated from the formula: oxidative catabolism of glucose. An available tis @iatomsglucose carbon oxidized = sue preparation, suitable for this study, is the c.p.m barium carbonate X @Mbarium carbonate whole, isotonic homogenate. Inasmuch as this preparation will oxidize glucose rapidly, when suit - c.p.m. glucose as barium carbonate ably fortified, and does not require an external MaieriaLt.â€”Diphosphopyridine nucleotide (DPN) was source of high energy phosphate as a driving force obtained from the Pabst Laboratories; it was found to be 85 (27),anymarkeddifferencesbetweentissuesin per cent pure by the enzymatic procedure of Racker (17). their capacities for synthesis and breakdown of fl,4-Dinitrophenol was a recrystallized commercial product. Hexokinase was supplied by the Pabst Laboratories as a dry, phosphate bonds should be manifested in differ lyophilized powder from autolyzed yeast. Cytochrome c, ences in oxidative response to fluoride and dinitro adenosine triphosphate (AlP), fumaric acid, sodium fluoride, phenol. The measurement of the incorporation of and nicotinamide were commercial preparations. Uniformly radioactivity from C'4-labeled glucose into respir labeled glucose-C1' was obtained from the Nuclear Instrument atory carbon dioxide in these experiments pro and Chemical Corporation on allocation by the Isotopes Divi. sion of the United States Atomic Energy Commission. vides a more precise criterion of the oxidation of Tumors.â€”With two exceptions, the tumors used in this this substrate than can be supplied by manometric study have been used in previous studies (26). One exception data alone and thus allows a more critical evalua was the LettrÃ©Ehrlich ascites carcinoma (1), a hyperdiploid tion of the effects of these inhibitors on glucose tumor with a chromosome mode of 45-47 carried in Swiss strain mice. This tumor was used because it is relatively free of blood @ catabolism specifically. cells. The other was the Krebs ascites carcinoma, described by Klein and Klein (7). MATERIALS AND METHODS Incubation of tiwse.â€”The animals, young, adult, well RESULTS nourished rats or mice from our stock colony, were decapitated, Intact cells.â€”Before experiments were conduct and the tissues were rapidly excised. The peripheral tissue of ed with homogenates, a preliminary survey was the tumors was carefully removed, and any grossly necrotic made to ascertain the effect of DNP on glucose portions were discarded. When homogenized tissue prepara tions were used, the experimental procedure was the same as oxidation in tumor slices and in whole ascites cells. @ described previously (%7).The tissue was homogenizedfor As shown in Table 1, there was considerable van minutes by the Potter-Elvehjem procedure (13) in 6 volumes of ation in the consumption of oxygen by the tumor an ice-cold isotonic medium containing KCJ, 0.133 as, and phos cells, ranging from 1 to about 7 @iMinthe DNP phate buffer (pH 7.4), 0.O1@3ii. An aliquot of the suspension, free system. These values correspond to Q, values representing 86 mg. of tissue unless otherwise specified, was added to Chilled Warburg vessels containing the other ions, (,d oxygen/mg dry tissue/hour) of from 1.4 to 10, factors, and substrates constituting the basic medium. This and are thus in the range observed in previous in consisted of the followingsubstances in their final concentra vestigations (4). Despite this variation, the pat tions: MgSO4, S X 10â€”'M; fumarate, 7 X 10' M; cytochrome terns of response to increasing DNP concentration C, 4 X 10' M; phosphate buffer (pH 7.4), 6 X 10-' at; KC1, @ 0.14 M; diphosphopyridine nucleotide, X 10' M; and water were similar. There was a variable stimulation in to make the final volume 1.6 ml. For the experiments with the range, 7 X 10@ to 7 X 10' M, whereas at slices, approximately flOO mg. of tissue was sliced with the 3 X 10@ M inhibitions in oxygen consumption, Stadie-Riggs slicer (@1), and was suspended in @.8ml. of cal ranging from 18 to 56 per cent, were observed. The cium-free Ringer phosphate solution (8) contained in Warburg one exception was the LettrÃ©tumor, in which stim flasks. Substrate and other requirements were added to give a final volume of 3.0 ml. ulation was still evident at the highest DNP level. For carbon dioxide absorption, a filter-paper roll, soaked This pattern of augmentation and inhibition of with 0.1 ml. of 10 N CO,-free sodium hydroxide, was placed respiration in the range of DNP concentrations in the center well. After attachment to the manometers, the herein employed is typical of a wide variety of ani flasks containing tissue slices were filled with oxygen. All mal, plant, and microbial cell types (20). flasks were equilibrated for 10 minutes at 38Â°C.; the manom eter stopcocks were then closed, and the flasks were shaken Data on glucose oxidation, as determined from for 50 minutes. Dilute sulfuric acid was then tipped in from the incorporation of C'4 into respiratory C02, closely side bulb to stop the reaction and liberate the bound CO2. paralleled oxygen consumption. Invariably, glu After being shaken for 10 minutes to allow complete absorption cose oxidation was enhanced at the intermediate of CO2 by the alkali, the flasks were removed, the filter paper and washings were transferred to a stoppered flask, 0.3 m@ of DNP levels and inhibited at the highest level. sodium carbonate was added as carrier, and the carbonate was Thus, it appears that intact tumor cells exhibit precipitated by the addition of excess 1 M barium chloride essentially the same oxidative response to DNP solution. The precipitated barium carbonate was filtered, dried, as do other cell types. and transferred to planchets for counting. Radioactivity Homogenates: effect of dinitrophenol.â€”As with measurements were made with an end-window counter, with the samples spread uniformly over an area of 7.5 sq. cm. slices, considerable variation in oxygen uptake of The values were corrected for self-absorption to â€œinfinite the uninhibited homogenates was observed (Table

2). Of the normal tissues, values were highest for mary carcinoma to 56 per cent for the rhabdomyo heart and kidney, lower for liver, and lowest for sarcoma. Experiments with homogenates of as brain. As in previous studies with homogenates cites tumors were not conducted, owing to difficul (27) or isolated mitochondria (28), the uptake of ties in breaking these cells by methods sufficiently oxygen in tumor homogenates was within the mild to retain oxidative activity. range displayed by normal tissues. Again, a rea Essentially the same pattern of response was sonably consistent pattern of oxidative response observed for glucose oxidation. In each instance, to dinitrophenol was observed, which differed from with the exception of brain, incorporation of radio that in slices in that no stimulation of respiration activity into respiratory CO2 diminished with in occurred with low DNP concentrations. Instead, creasing DNP level. inhibition increased with increased DNP levels, Despite quantitative differences, remarkable the extent ranging from only 25 per cent in brain similarities in the relative effects of DNP on exog to 95 per cent in heart. Stimulation of oxygen con enous and endogenous oxidation were observed. sumption as found in slices also was never ob In heart and kidney, oxygen consumption was served in homogenates of either tissue type. Again, very high (30 LLMoxygen uptake corresponds to a the range of inhibition in tumors was within that Q@of 46). In the experiment with the heart ho displayed for the four non-neoplastic tissue ho mogenate, the 17.8 patoms of glucose carbon oxi mogenates, varying from 20 per cent for the mam dized required the same number of p@tof oxygen,

TABLE 1 EFFECT OF DINITROPIIENOL ON GLUCOSE-C14 OXIDATION BY MOUSE NEO PLASTIC TISSUE SLICES AND WHOLE CELLS Experiments conducted 60 minutes at 88Â°C. with oxygen in the gas phase. Approximately 150 mg. tissue (wet weight) was used per flask. All values are on a uniform basis of 100 mg. tissue (wet weight).

DINITIIOPELNOL cowczaTaanoa (aioz..azzry) 0 7X10 7X10' sx'oâ€”4 Glucose Glucose Glucose Glucose 02 uptake oxidized 0@uptake oxidized Os uptake oxidized 0, uptake oxidized NzopI.AR nc TIS$U@5 (gsM) (patoins C.) (,@is) (1satomsC.) (pM) (guatomsC.) (guI) (patomsC.) Hepatoma 98/15 7.@ 0.9 7.% 1.0 8.0 1.4 4.8 0.4 Sarcoma 87 4.7 0.9 8.9 1.% 4.1 1.8 %.1 0.4 Rhabdomyosarcoma 4.4 1.8 4.% 1.8 8.8 0.9 8.1 0.6 Krebs-@ (ascites) 3.9 1.0 6.7 %.8 4.6 1.2 1.7 0.08 Mammary adenocarcinoma %.9 0.3 %.7 0.8 8.9 0.5 %.4 0.% TAS (ascites) 6.4 1.8 8.1 %.O 5.7 0.7 8.9 0.8 Lettre Ehrlich (ascites) 1.0 0.% %.6 0.9 8.9 1.7 %.8 0.8 %.0 0.5 L6 0.9 4.0 1.6 %.8 0.8

TABLE 2 EFFECT OF DINITROPHENOL ON GLUCOSE OXIDATION IN MOUSE TISSUE HOMOGENATES Experiments conducted 60 minutes at 88Â°C. with air in the gas phase. ATP when used was in a final ooncentration of 0.001 M. Valuesare based on tissue used (86 mg. tissue, wet weight).

DINITROPIIaNOL co@tcmra&nos (uoi.sssrr) 1.QX1Oâ€”' 6X10' L5X104 Glucose Glucose Glucose Glucose 0, uptake oxidized Os uptake oxidized 0, uptake oxidized Os uptake oxidized Movax nssvz (guu) (guatoms C.) (gusu) (jiatoms C.) (guiu) (guatoms C.) (gusu) (juatoms C.) Normal: Heart 27.2 17.8 27.4 17.6 8.1 0.8 1.4 0.8 Kidney 25 8.4 22 1.5 9.6 0.5 11.0 0.5 â€œ ,ATPadded 80 5.8 18 2.7 9.6 0.9 12.0 0.6 Brain 7.5 2.1 5.9 4.6 4.8 5.6 8.7 â€œ ,ATPadded 4.4 1.6 8.5 2.7 4.4 4.2 8.2 8.0 Liver 10.2 0.7 11.2 0.6 7.1 0.28 6.9 0.20 â€œ ,ATPadded 10.2 0.7 9.6 0.7 6.0 0.8% 5.0 0.14 Neoplastic: Hepatoma 98/15 12.4 2.0 10.9 1.8 8.7 0.1 7.8 0.08 Rhabdomyosarcoma 18.9 8.5 12.5 2.6 7.6 1.55 6.1 0.16 Sarcoma 37 7.9 0.59 9.7 0.56 6.8 0.24 6.0 0.19 Mammary adenocarcinoma 5.9 0.87 4.9 0.80 5.4 0.07 4.8 0.07

since glucose oxidation has a respiratory quotient oxidation of the exogenous glucose. In this tissue of 1. Hence 65 per cent (17.8 X 100/27.2) of the it appears that DNP causes an enhanced glucose total oxygen consumed was utilized for oxidation oxidation at the expense of endogenous metabo of the added glucose. At a DNP level of 6 X 10@ lites. These results differ in some respects from M, oxygen consumption was inhibited 89 per cent, those of Tyler (22), who observed considerable in but glucose oxidation was inhibited 98 per cent. creases in oxygen consumption of rat brain ho Thus, it appears that DNP inhibits the oxidation mogenates caused by DNP at levels similar to of glucose more than it does the endogenous sub those employed here; however, his general conclu strates. In kidney, oxidation of the added glucose sion that DNP augments glucose oxidation in brain accounted for a much smaller proportion of the homogenates is in harmony with our isotope data. total respirationâ€”14 and 19 per cent in the two Lack of effect ofA TP on dinitrophenol inhibition. experiments. Again, in this tissue, oxidation of the â€”It was found previously (27) that the whole, for added glucose was more sensitive to DNP than tified homogenate employed in these studies does was that of endogenous substrates. Similar behav not require external ATP for optimal glucose oxi ior was observed in liver, even though oxygen con dation and, therefore, carries out sufficient phos sumption and glucose oxidation are both consider phorylation to activate glucose in the face of all ably lower in this tissue than in kidney and heart. the ATP-splitting activities occurring in this crude system. In testing the possibility that DNP inhib TABLE 3 its glucose oxidation by stimulating ATP break down or by preventing its formation through un EFFECT OF HEXOKINASE AND ATP ON DNP INHIBI coupling of phosphorylation, it was reasoned that TION OF GLUCOSE OXIDATION BY MOUSE under either of these circumstances DNP inhibi KIDNEY WHOLE HOMOGENATE tion should be reversed by the addition of ATP. Experiments conducted for 60 minutes at 38Â°C. with air However, this expectation was not realized. As in the gas phase. Values are expressed in @sM/tissue used (43 mg. fresh weight). demonstrated in Table 2, the addition of ATP to kidney, brain, and liver homogenates had no ef GLUCOSEHexokinaaeADDITIONS OxIDIZED(pg/flask)*ATP DNP 02 UPTAKE fect on the pattern of response to DNP. This lack 0.001 ii 6X10' u (pM) (patoms C.) of effect of external ATP on the oxidative response â€” â€” â€” 14.4 4.7 to dinitrophenol is again clearly shown in the ex â€” â€”4.1â€” â€” 10.2 periment with mouse kidney homogenate in Table + 4.9 1.2 SO + + 6.8 1.6 3. As observed previously (27), the addition of 5.0 + + 5.9 1.2 ATP alone to the fortified homogenate inhibited 1.2â€¢One0.5 Â± + 6.1 somewhat both glucose oxidation and over-all res pg. of hexokinase is equivalent to 0.058 K.M. units (9) at 80Â°C., which is equivalent to the transfer of 0.0084 p@ of phosphate from ATP to piration. The addition of DNP alone brought glucose at 50Â°C.in SOminutes. about a 66 per cent decline in respiration and a 75 per cent decrease in glucose oxidation. Addition of In the tumor homogenates also, the proportion of ATP together with dinitrophenol and several dif the total respiration involved in oxidation of exog ferent concentrations of yeast hexokinase ap enous glucose varied widelyâ€”from about 6 per peared to counteract only slightly the inhibition of cent in the mammary tumor to 25 per cent in the respiration and had little or no effect on glucose rhabdomyosarcoma. Again, however, oxidation of oxidation. Evidently, if dinitrophenol affects the the exogenous glucose was consistently more sen availability of ATP for glucose activation, its ef sitive to DNP than was oxidation of endogenous fect is such that it cannot be reversed by external substrates; maximum DNP inhibitions in oxygen addition of ATP. consumption ranged from 20 to 56 per cent, where That the considerations discussed above apply as inhibition of oxidation of the exogenous glucose as well to the tumor homogenate is disclosed in ranged from about 70 to almost 100 per cent. Table 4, where it is evident that in the mouse hep Oxidative response in brain homogenate.â€”The atoma 98/15, ATP in a concentration of 0.001 M behavior of brain differed notably from that of all failed to alter the typical pattern of dinitrophenol other tissues examined. In the uninhibited system, inhibition in respiration and in glucose oxidation. oxidation of exogenous glucose accounted for 26 It was interesting to note that the inhibition in and 36 per cent of the total respiration. With in oxidative activity resulted in a decreased, rather creasing DNIP level, total oxygen consumption de than an increased, lactic acid production. External creased somewhat, but glucose oxidation increased ATP appeared to increase lactic acid formation greatly; at the two higher DNP concentrations, somewhat but did not appreciably affect the re essentially all the oxygen was being utilized for sponse of glycolysis to dinitrophenol.

Effect of fluoride on glucase OxidatiOn Zfl whole however, the range of inhibition by fluoride was homogenates.â€”In testing the effects of fluoride, it about the same in both the normal and neoplastic was considered advisable to do this both in the ab tissue types. sence and in the presence of DPN, since it seemed Glucose oxidation was more markedly affected likely that any stimulating effects of fluoride ion by fluoride than was oxygen consumption, being might be uncovered more readily in a system in almost completely abolished in every tissue ex hibited by lack of a co-factor rather than in one amined. The Ehrlich ascites cells were responsive maximally stimulated. In Table 5 the effects of neither to DPN nor to fluoride; this is attributed fluoride in a representative group of normal and to their having an intact cell membrane which is neoplastic tissue homogenates are compared. No relatively impermeable to these substances. significant enhancement in either oxygen uptake To see whether fluoride might stimulate oxida or in glucose oxidation was observed. In a few in tion when present in lower concentrations, as does stances, oxygen uptake was increased by fluoride DNP, the experiment shown in Table 6 was car somewhat over the generally low levels observed in ned out. It is evident that fluoride at low concen the absence of DPN. However, these were unac trations does not stimulate either oxygen con companied by increases in glucose oxidation; the sumption or glucose oxidation. It had essentially low levels of glucose oxidation were either un no effect on oxygen consumption or glucose oxida changed or lowered further by fluoride. In the tion in the absence of DPN+, and, in its presence, DPN-stimulated homogenate, fluoride in the con inhibited both of these processes to extents rough centration employed consistently inhibited both ly dependent on the fluoride concentration. glucose oxidation and oxygen uptake. Its effect on oxygen consumption was less marked in brain and DISCUSSION in liver than in the other normal tissues, and this The variability of normal and neoplastic tissues was true also for hepatoma 98/15 as compared in their behavior toward substrates during oxida with the other neoplastic tissues; on the whole, tive metabolism was pointed out by Siekevitz and

TABLE 4 EFFECT OF 2,4-DINITROPHEN0L AND ATP ON GLUCOSE UTILIZATION BY MOUSE HEPATOMA 98/15 HOMOGENATES Experimental conditions are the same as described in Table 2. Values are expressed in pM/tissue used (86 mg. wet weight). ATP, 0.001 M ABSENTPRESENTDINITROPHENOLOxygenLactateGlucoseOxygenLactateGlucoseCONCENTRATIONuptakeformedoxidizeduptakeformedoxidizedMOLAR(pM)(psi)(patoms

C.)[email protected] C.)(pM)(psi)(patoms

X10'8.73.30.106.43.70.382.5 X1O47.32.80.035.63.80.13TABLE

5 EFFECT OF FLUORIDE ON GLUCOSE OXIDATION IN WHOLE HOMOGENATES Experiments conducted as in Table 2. Fluoride when present was in a final concentration of 0.0075 M, and DPN, 0.002 M. Values are based on tissue used (86 mg. wet weight).

ADDITIONS TO BASIC MEDIUM None Fluoride DPN Fluoride+DPN Os Glucose Os Glucose 0, Glucose Os Glucose uptake oxidized uptake oxidized uptake oxidized uptake oxidized Mousx TISSUE (pM) (patomaC.) (psi) (patoms C.) (pM) (patoms C.) (psi) (patomsC.) Normal: Heart 0.4 0.2 1.2 0.1 16.5 11.2 5.8 0.1 Kidney 1.3 0.1 1.4 0.1 19.0 7.2 4.7 0.1 Brain 0.5 0.4 1.3 0.3 5.3 1.6 4.4 0.2 Liver 5.5 0.5 4.1 0.0 7.6 0.6 3.9 0.1 Neoplastic: Hepatoma 98/15 1.6 0.1 2.1 0.0 9.5 1.8 7.2 0.1 Rhabdomyo 1.8 0.1 1.1 0.1 7.7 1.2 3.1 0.2 sarcoma Mammary ade 1.3 0.1 2.8 0.0 6.7 0.8 3.0 0.2 nocarcinoma Sarcoma 87 0 0.1 0 0.0 3.0 0.4 1.2 0.1 Ehrlich ascites 3.5 0.5 3.0 0.4 3.9 0.7 4.2 0.7

Potter (19), and the present study is no exception from non-neoplastic tissues in mechanisms of in this regard. Large differences were observed in phosphorylation and dephosphorylation associat capacity for glucose oxidation, and notable varia ed with glucose metabolism. As pointed out by tions in both oxygen consumption and glucose Reif et at. (18), the possibility of quantitative oxidation were found in response to the addition of aberrations in single steps along the sequences of dinitrophenol and fluoride. Nevertheless, a con reactions involved in the catabolism of glucose, or sistency in the pattern of response to these agents in the electron transport associated with the oxida emerged despite these variabilities. Dinitrophenol tive steps thereof, cannot be denied. As yet, how stimulated the oxidation of glucose in normal tis ever, no such aberrations have been uncovered sue slices at low concentrations, and inhibited at which have been convincingly demonstrated to be higher concentrations; and a similar response was characteristic of the neoplastic state. In view of observed for tumor slices over the same concentra the wide variability in oxidative capability dis tion range of DNP. Both DNP and fluoride inhib played by both normal and neoplastic tissue slices, ited glucose oxidation in homogenates of non-neo it is obvious that the uniformly high glycolysis in plastic tissues, and again their behavior was simi tumors cannot be attributed to quantitative dif lar in tumor homogenates. ferences in respiratory activity. Thus far, no quali Previous isotopic tracer studies (3, 10, 24â€”26) tative differences in the metabolic pathway of nor have emphasized the ability of the intact tumor mal as compared with tumor cells have been dis cell to oxidize glucose, fatty acids, and their inter covered. As stated previously (24), â€œWhatevermay

TABLE 6 EFFECT OF FLUORIDE ON GLUCOSE OXIDATION IN RHABDOMYOSARCOMA HOMOGENATES Experiment conducted as in Table 2 CONCENTRATION OP FLUORIDE. MOLAR 00.00010.0010.01OsGlucoseOsGlucoseOsGlucoseOsGlucoseuptakeoxidizeduptakeoxidizeduptakeoxidizeduptakeoxidizedDPN(psi)(patoms

C.)Absent2.20.151.50.101.60.151.50.09Present8.00.957.20.464.00.174.00.15C.)(psi)(patoms C.)(psi)(patoms C.)(jiM)(patoms

mediates at rates similar to those of non-neoplastic be the cause of the high aerobic glycolysis in tu.. tissues. The complete oxidation of these sub mors . . . it is not due to quantitative or qualita stances by oxygen to CO2 and water occurs via tive peculiarities of oxidative metabolism.â€• The highly integrated processes, in which phosphoryla cause of this phenomenon remains as obscure now tions and dephosphorylations occur at many as it was to its investigators 30 years ago. stages. Glucose, whose catabolism via fructose-1,6- diphosphate requires two phosphorylations initial SUMMARY ly for activation, can be readily oxidized to com The effects of 2,4-dinitrophenol and fluoride pletion in whole tumor homogenates, without the ions on the oxidation of glucose in normal and tu external addition of ATP, just as it can in normal mor tissue slices and homogenates of the mouse tissue homogenates (26). This indicates that the were investigated by the use of uniformly labeled generation of ATP in tumor is sufficiently rapid to glucose-Câ€•. maintain its level in the face of all the ATP-de Dinitrophenol stimulated glucose oxidation in stroying processes occurring in this crude system. neoplastic and non-neoplastic tissue slices at low Moreover, glucose oxidation is affected in the same concentrations and inhibited at higher concentra manner in both tissue types by other factors which tions. Both dinitrophenol and fluoride were inhibi limit oxidative activity, such as the lack of ions or tory toward glucose oxidation in homogenates of co-enzymes or hypotonicity (27). Several recent both tissue types. investigations have found that phosphorylation associated with specific oxidations occurs in en REFERENCES zyme preparations of neoplastic tissues just as it 1. BAYREUTRER,v. K. Der Chromosomenbestand des Ehr does in similar preparations of normal tissues (5, lich-Ascites Tumors der Maus. Ztschr. Naturforsch., 6, 9, 30). 7(b): 544, 1952. With all these findings taken into consideration, 2. BuRr, D. A Colloquial Consideration of the Pasteur and Neo-Pasteur Effects. Cold Spring Harbor Symp. Quant. the opinion seems inescapable that the neoplastic Biol., 7:420â€”59, 1939. cell does not differ in any fundamental respect 8. CHAPMAN, D. D.; BROWN, D. W., JR.; CHAIxos'@, I. L;

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Downloaded from cancerres.aacrjournals.org on September 26, 2021. © 1955 American Association for Cancer Research. Metabolism of Neoplastic Tissue: VII. Effects of Dinitrophenol and Fluoride on Glucose Oxidation in Tumor Homogenates

Charles E. Wenner and Sidney Weinhouse

Cancer Res 1955;15:497-503.

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