National Cancer Institute Protocol Research Articles

Synthesis and antitumor activity of 1,3,4-oxadiazole substituted 2-(5-ylidene-2,4-dioxothiazolidin-3-yl)-acetamides

A series of novel 1,3,4-oxadiazole substituted 2-(5-aryl/heterylidene-2,4-dioxothiazolidine-3-ylidene)-acetamides and their 5-unsubstituted analogues have been synthesized following N-alkylation reaction of 2-chloro-N-(5-aryl-[1,3,4]oxadiazol-2-yl)-acetamides with thiazolidinedione and potassium salts of its arylidene derivatives. The structures of target compounds were confirmed by using 1H NMR spectroscopy and elemental analysis. Evaluation of anticancer activity in vitro for the synthesized compounds was performed in accordance with the National Cancer Institute protocol. A selective influence of some tested compounds against leukaemia MOLT-4 (3e, GP = 76.85%) and K-562 (3e, GP = 79.84%), colon cancer HCT-15 (3d, GP = 76.86%), renal cancer A498 (4a, GP = 74.37%), CAKI-1 (3d, GP = 68.49%) and UO-31 (3b-e, 4a-b, GP = 66.67 ÷ 86.30%) cell lines was established. Grafical abstract:

Analysis of demographic characteristics and disparities of enrollment in cancer clinical research at the U.S. National Cancer Institute (NCI): A 15-year experience.

6529 Background: Racial, gender, and socioeconomic disparities in cancer clinical trials participation have been well documented. The intramural program of the NCI in the National Institutes of Health (NIH) Bethesda campus conducts clinical research where participants are referred by their physician or self-referred, treated without charge and receive aid in travel expenses. We sought to describe the demographics of participants and identify potential disparities. Methods: We used the clinical research data repository of the NIH, Biomedical Translational Research Information System (BTRIS), to extract self-reported demographic data of enrollees in NCI protocols from 2005-2020. Comparisons were made with the U.S. population of patients with cancer as captured in the Surveillance, Epidemiology, and End Results (SEER) Program (2005-2018). The average household income per zip code was derived from the American Community Survey at five-year increments from 2011, 2015, and 2019. Results: We obtained data on 40,007 participants; 38,531 (95%) came from the U.S., of which 18,606 (48%) from DC, Maryland, or Virginia (DMV). Median age at consent was 54 years (range 2 – 89). Proportions of basic demographic attributes and corresponding values from SEER data were presented. Notably, Hispanic male, non-Hispanic female, and white female participants were under-represented in NCI clinical research compared to SEER data. African Americans were under-represented in the non-DMV population (5%) vs. the DMV (19%). Also, for patients from the DMV area, median income was consistently less for African American participants compared to White and Asian participants. Conclusions: The identified disparities may be partially due to the specific research activities available at the NCI, but strategies aiming at increasing the participation and representation of under-represented groups are required. [Table: see text]

Effects of Alvelestat, an Oral Neutrophil Elastase Inhibitor, on Elevated Elastase and Collagen Turnover Biomarkers in Patients with Bronchiolitis Obliterans Syndrome after Hematopoietic Cell Transplantation

IntroductionBronchiolitis Obliterans Syndrome (BOS) is a rare but devastating complication of chronic graft-versus-host disease (GVHD) after allogeneic hematopoietic cell transplantation (HCT) and is associated with a high morbidity and mortality. There is a dearth of treatment options for BOS and new strategies are needed. Airway neutrophilia is a hallmark of BOS, even in the absence of infection, and neutrophil elastase (NE) is an enzyme that has been implicated in the pathogenesis of BOS. We are conducting a phase 1b study of an oral NE inhibitor, alvelestat, in patients with BOS after HCT . Biomarkers, including the elastin breakdown peptides desmosine/isodesmosine (DES/IDES), and stimulated neutrophil elastase assess direct effect on NE activity. Neo-epitope by-products of collagen type 3 and 6 synthesis (PRO-C3 and PRO-C6) and degradation (C3M and C6M) are measured as biomarkers of fibrosis/tissue modellingMethodsPatients age ≥18 years with BOS and chronic GVHD after HCT were recruited to the National Cancer Institute protocol (NCT02669251). This phase 1 study had 2 parts: 8-week intra-patient dose escalation period, followed by a continuation period that allowed for up to 6 months of treatment. Alvelestat was given orally starting at 60mg twice daily, increased every 2 weeks to 120mg twice daily, 180mg twice daily, and finally 240mg twice daily. Peripheral blood samples were collected at baseline and at the end of each dose-escalation stage.Plasma DES/IDES was measured by isotopic dilution liquid chromatography-tandem mass spectrometry (Huang et al Thorax 2012;67:502-508). Ex vivo zymosan stimulated neutrophil elastase activity was measured by ProteaseTag® immunoassay (ProAxsis Ltd, Northern Ireland). PRO-C3, PRO-C6, C3M and C6M were measured by competitive ELISA. (Nordic Biosciences, Denmark). Results are presented as Mean and Standard Error Mean (SEM).ResultsBetween 2016 and 2018, 7 patients were enrolled (3 men and 4 women). Median FEV 1 after bronchodilator at time of enrollment was 44% predicted (range 38-74). All 7 patients were able to tolerate dose escalation of alvelestat up to the maximum dose 240mg twice daily. Preliminary clinical results were previously presented.DES/IDES was elevated at baseline (mean 0.464 (SEM 0.0508) ng/ml, with 6 of 7 subjects above the Upper Limit of Normal (ULN, 0.280 ng/ml)). Levels progressively declined during the dose escalation period to 0.380 (SEM 0.0419) ng/ml by week 8, representing a mean within subject % change from baseline (CFB) of -16.2% (SEM 6.794, Figure 1a)Ex vivo zymosan stimulated elastase activity also showed progressive decrease over the dose escalation period, with some subjects demonstrating 100% suppression (Figure 1b).Collagen synthesis as measured by PRO-C3 and PRO-C6 was increased above ULN at baseline and declined with alvelestat treatment (Figure 1c and 1d). There was no consistent change in collagen degradation biomarkers (C3M, C6M)There was consistency of a suppressive effect on biomarkers of elastase activity and collagen turnover in 6 of 7 treated patients, all of whom had improved or stable lung disease (ranging from change in FEV1 % predicted at end of treatment from +9% to -6%).ConclusionNE can damage lung tissue due to elastin breakdown, pro-inflammatory and pro-fibrotic effects (Sallenave J-M, J Leuk Biol 2015;98:137-139). This is the first evidence of elevated elastase activity as detected by elastin breakdown in patients with BOS and chronic GVHD.Treatment with the selective NE inhibitor, alvelestat was associated with progressive reduction of plasma desmosine levels over 8 weeks of within-subject dose escalation and reduction stimulated neutrophil elastase activity.The consistent suppression of elastase and of collagen synthesis/turnover biomarkers following alvelestat treatment is encouraging for its potential to impact progressive lung fibrosis in BOS and chronic GVHD. DisclosuresParkin: Mereo BioPharma Group: Current Employment. Moore: Mereo BioPharma Group: Current Employment. Pavletic: Center for Cancer Research: Research Funding; National Cancer Institute: Research Funding; National Institutes of Health: Research Funding; Celgene: Research Funding; Actelion: Research Funding; Eli Lilly: Research Funding; Pharmacyclics: Research Funding; Kadmon: Research Funding.

Synthesis and Anti-Cancer Activity Evaluation of Novel 1,3,4-Oxadiazole Substituted 5-Arylidene/Isatinylidene-2-Iminothiazolidin-4-Ones

Following the interaction of 2-chloro-N-(5-aryl-1,3,4-oxadiazole-2-yl) acetamides 1a-b with ammonium thiocyanate in dry acetone, the 5-unsubstituted 2-imino-4-thiazolidinones 4a-b have been synthesized. Compounds 4a-b were subsequently utilized in Knoevenagel condensation with aromatic aldehydes or isatin derivatives to synthesize the series of 5-arylidene/isatinylidene substituted 2-(1,3,4-oxadiazol-2-yl)imino-4-thiazolidinones 5a-h and 6a-d. The structures of target compounds were confirmed by using 1H NMR spectroscopy and elemental analysis. Evaluation of anti-cancer activity in vitro for the synthesized compounds was performed following the National Cancer Institute protocol against leukemia, melanoma, lung, colon, CNS, ovarian, renal, prostate, and breast cancer cell lines. As a result, the most active compound 5a, namely 2-[5-(4-chlorophenyl)-[1,3,4]oxadiazol-2-ylimino]-5-(4-methoxybenzylidene)thiazolidin-4-one was found to be a highly efficient anti-tumor candidate with average logGI50 and logTGI values of -5.19 and -4.09, respectively.

Phase 1 Study of Alvelestat, an Oral Neutrophil Elastase Inhibitor, in Patients with Bronchiolitis Obliterans Syndrome after Hematopoietic Cell Transplantation

Introduction Bronchiolitis Obliterans Syndrome (BOS) is a rare but devastating complication of chronic graft-versus-host disease (GVHD) after allogeneic hematopoietic cell transplantation (HCT) and is associated with a high morbidity and mortality. There is a dearth of treatment options for BOS and new strategies are needed. Airway neutrophilia is a hallmark of BOS, even in the absence of infection, and neutrophil elastase (NE) is an enzyme that has been implicated in the pathogenesis of BOS. We conducted a phase 1 study of an oral NE inhibitor, alvelestat, in patients with BOS after HCT. Methods Patients age ≥18 years with BOS and chronic GVHD after HCT were recruited to the National Cancer Institute protocol (NCT02669251). Patients had stable systemic immunosuppression and FEV1 ≥30% on pulmonary function tests (PFTs). This phase 1 study had 2 parts: 8-week intra-patient dose escalation period, followed by a continuation period that allowed for up to 6 months of treatment. Alvelestat was given orally starting at 60mg twice daily (the dose previously used in patients with chronic lung disease) and increased every 2 weeks as tolerated to 120mg twice daily, 180mg twice daily, and finally 240mg twice daily. Patients continued this dose until completion of the continuation phase, or occurrence of unacceptable toxicity, dose interruption >28 days, or progression of GVHD or BOS. Primary objective was to determine the maximum tolerated dose (MTD) based on dose-limiting toxicities. Secondary objectives included determining pharmacokinetics, markers of NE activity, and markers of inflammation in blood and sputum. PFTs and chronic GVHD evaluations were performed at baseline, 4 weeks and 8 weeks during the dose escalation period, and at 3 months and 6 months during the continuation period. Results Between 2016 and 2018, 7 patients were enrolled (3 men and 4 women). Median FEV1 after bronchodilator at time of enrollment was 44% (range 38-74). All 7 patients were able to tolerate dose escalation of alvelestat up to the maximum dose 240mg twice daily; MTD was not reached. The most common adverse events (AEs) that were possibly related to study treatment were all grade 2, and included increased creatinine (3 patients), ALT or AST elevation (3 patients), and upper respiratory infection (3 patients). The only grade 3 AEs that were possibly related to study drug were gastroenteritis and vomiting requiring hospitalization in 1 patient and pneumonia in 1 patient. Three patients completed the study with 8 weeks + 6 months of treatment. Four patients required dose interruptions, and only 1 of those required dose reduction for grade 3 gastroenteritis (resulting in dehydration and elevated creatinine). Four patients discontinued treatment prior to end of study: 2 patients had dose interruption of >28 days due to adverse events, 1 patient had a decline in FEV1 after pneumonia, and treatment was stopped in 1 patient due to investigator discretion. The median duration of treatment was 6.4 months. Based on NIH chronic GVHD consensus criteria, 6 patients had unchanged disease and 1 patient had progressive disease (decline in FEV1 after pneumonia). Although patients did not achieve the 10% improvement in FEV1 required for an organ response, 2 patients had improvement of 9% in FEV1 and 4 patients had improvement in the Lee chronic GVHD symptom scale lung score. Preliminary pharmacokinetic analyses of the 7 patients showed a linear dose-dependent increase in each exposure metric (steady-state trough and steady-state peak), despite some inter-patient variability. Bronchoalveolar lavage fluid and induced sputum samples are being analyzed for NE activity. Conclusion In this phase 1 study of the oral NE inhibitor, alvelestat, in patients with BOS after HCT, MTD was not reached and the study drug was well tolerated. Six patients had stable disease, while 1 patient had progression in the setting of pneumonia. Two patients notably had improvement in FEV1 of 9%, and 4 patients experienced improvement in symptoms. We have demonstrated that NE inhibition is well tolerated and shows a signal of stabilizing disease in patients with advanced BOS. Further study of the clinical efficacy of alvelestat in BOS after HCT is warranted, especially at an earlier stage of disease and longer duration of drug administration. Disclosures No relevant conflicts of interest to declare.

Synthesis and cytotoxicity evaluation of [(2,4-dichlorophenoxy)methyl]-5-aryl-1,3,4-oxadiazole/4 H -1,2,4-triazole analogues

We report herein the synthesis, characterization, and cytotoxicity evaluations of some newer oxadiazole and triazole analogues (5a-j). The cytotoxicity of all the title compounds were evaluated as per the National Cancer Institute protocol in a one-dose assay (10 $\mu $M) on nine different panels of 59 cancer cell lines. 2-$\{$5-[(2,4-Dichlorophenoxy)methyl]-1,3,4-oxadiazol-2-yl$\}$phenol (5e) showed the maximum cytotoxicity among the series of ten compounds. The cytotoxicity of 5e was comparable to that of the standard anticancer drug, 5-fluorouracil, and better than that of imatinib. The structure activity relationship was also discussed.

How to cure cancer: unbinding entrepreneurs in medicine

PurposeSince the 1960s, experts have predicted that we are on the verge of curing cancer. The purpose of this paper is to explore the obstacles to progress, and to propose policies that will lead more quickly to more success.Design/methodology/approachTo speed future cures, we need to look at the traits, and methods of those innovative medical entrepreneurs who achieved breakthroughs in the past, and learn what institutions and policies enabled, or blocked, their progress.FindingsBreakthrough innovators tend to be less-credentialed outsiders who “see what others do not see,” often by nimble and persistent pursuit of serendipitous discoveries or slow hunches. For example, Freireich and DeVita were less-credentialed outsiders. Freireich cured childhood leukemia and DeVita cured Hodgkin’s lymphoma, by pursuing nimble trial-and-error experimentation in their anti-cancer chemotherapy cocktails. Min Chiu Li pursued his slow hunch that his patients would benefit from longer chemotherapy than the mandated National Cancer Institute protocol allowed. He was fired, but his patients were cured. Today, FDA-mandated regulatory protocols, often defended as applications of the precautionary principle, greatly restrict innovative medical entrepreneurs, thereby delaying cancer cures and costing lives.Originality/valueThe paper proposes a new approach to medical innovation, allowing cancer researchers to engage in trial-and-error experiments that follow up on serendipitous discoveries and plausible hunches. The result will be more cures and longer lives.

Surveillance and expected outcome of acute lymphoblastic leukemia in children and adolescents: An experience from Eastern India

Objective:Research in Eastern India especially among children and adolescents for acute lymphoblastic leukemia (ALL) have not been well documented until recently when it was conducted at a cancer institute of tertiary care with primary objectives of examining and correlating different cell surface markers involved with respect to disease surveillance thereby highlighting it as a strong prognostic marker for future diagnosis and treatment.Materials and Methods:A total of 500 consecutively selected ALL patients were diagnosed and treated according to National Cancer Institute protocol (MCP 841) for a period of 24-88 months during this hospital-based study.Results:Of the total, 50.4% had a higher incidence of T-ALL and 47.6% had pro-B, B-cell precursor ALL. Disease free survival and event free survival were remarkably higher in B-ALL adolescent patients as compared to T-ALL, who had significantly lower overall survival ratio. Prevalence of T-ALL was also observed in relapse cases for adolescent patients.Conclusions:We conclude that there is an increased prevalence of T-ALL among adolescents in Eastern India. Immunophenotypic analysis might help in proper evaluation and prediction of treatment outcomes with an increased thrust on studying age-specific incident rates enabling well planned future treatments for improved and better outcome.

1091P - Impact of Nutrition on Treatment Outcome in Acute Lymphoblastic Leukaemia of Childhood in Developing Country

ABSTRACT Background All conventional modalities of anti-cancer therapy interfere with normal nutrition. Therefore, malnutrition is a major problem in children with cancer. In this study we retrospectively analyzed 500 children suffering from Acute Lymphoblastic Leukemia (ALL), who were being intensively treated by National Cancer Institute protocol (MCP 841) during period beginning from August, 1999 to December, 2011, in a tertiary cancer institute. Our aim was to determine the nutritional status at preliminary diagnosis of ALL children and to study the influence of nutrition on complete remission, disease free survival (DFS) and chemotherapeutic toxicity. Procedure The variables studied were height for age (HFA), weight for age (WFA), mid-arm circumference (MAC), biceps skinfold thickness (BSFT) and serum albumin levels. The HFA, WFA, MAC and BSFT were taken as normal if they were between 3rd and 97th percentile curve of the growth chart as recommended by the Indian Council of Medical Research (ICMR). The albumin level was considered normal if the value was equal to or more than 3g%. Results It was observed that 16.8% children were low weight for age and 10.4% were low height for age during diagnosis. Low WFA (p value 0.001), low HFA (p value 0.0001) and low albumin (p-value 0.0001) were significant in DFS. Conclusion We conclude that malnutrition has high impact on ALL prognoses in developing countries. Major nutritional indicators are HFA, WFA, MAC, BSFT and serum albumin. Patients with malnutrition have less DFS duration, more chances for relapse and more therapeutic toxicity when compared to well-nourished children. Disclosure All authors have declared no conflicts of interest.

Impact of Nutrition on Treatment Outcome in Acute Lymphoblastic Leukaemia of Childhood in Developing Country

Abstract 4744Malnutrition is a major problem in children with cancer. All conventional modalities of anti cancer therapy interfere with normal nutrition. In this study we retrospectively analyzed 331 children of Acute Lymphoblastic Leukaemia (ALL) being intensively treated by National cancer Institute protocol (MCP 841) during period from August, 2000 to December, 2010 in a tertiary cancer institute of the country. Our aim was to determine the nutritional status of children with ALL at diagnosis and to study the influence of nutrition on complete remission, disease free survival (DFS) and toxicity of chemotherapy. The variables studied were height for age, weight for age and serum albumin levels. The height for age and the weight for age were taken as normal if they were between 3rd and 97th percentile curve of the growth chart as recommended by the Indian Council of Medical Research (ICMR). The albumin level was considered normal if the value was equal to or more than 3 gm%. It was seen that 16.9% children were low weight for age and 10.3% were of low height for age at diagnosis. Low weight for age (p value <0.01) and low albumin (p-value <0.005) were significant in DFS.We conclude that malnutrition is having much impact on prognosis of ALL in developing countries like ours. The major nutrition indicators are height for age, weight for age and serum albumin. The patients with malnutrition have less DFS duration, more chances of relapse and more toxicity during therapy as compared to well nourished children.Table 1:Nutritional status of 331 childrenVariablesNormalLowWeight for age275 (83.08%)56 (16.91%)Height for Age297 (89.73%)34 (10.3%)Total Protein274 (82.78%)57 (17.22%)Serum Albumin301 (90.94%)30 (9.06%)Table 2:Effect of Nutritional variables on remission of ALLVariableTotalRemissionNon ResponderInduction DeathTotal failure of Inductionp-valueNormal Weight for age275260 (94.54%)4 (1.45%)11 (4%)15 (5.45%)Low Weight for Age5651 (91.1%)05 (8.9%)5 (8.9%)0.236Normal Height for Age297283 (95.23%)4 (1.35%)10 (3.37%)14 (4.7%)Low Height for Age3428 (82.35%)1 (2.94%)5 (14.7%)6 (17.6%)Normal Albumin301287 (95.35%)2 (0.66%)12 (4%)14 (4.6%)Low Albumin3024 (80%)2 (6.7%)4 (13.3%)6 (20%)Table 3:Effect of Nutritional variables on Disease Free SurvivalVariableTotalDFS (%)Relapse Death (%)Death due to Other Causes (%)Total Deathp-valueNormal Weight for age275198 (72%)59 (21.45%)18 (6.54%)77 (28%)Low Weight for Age5619 (33.9%)25 (44.6%)12 (21.4%)37 (66%).001Normal Height for Age297213 (71.7%)68 (22.9%)16 (5.4%)84 (28.3%)Low Height for Age344 (11.8%)20 (58.8%)10 (29.4%)30 (88.2%)0.0001Normal Albumin301212 (70.4%)62 (20.6%)27 (8.97%)89 (29.6%)Low Albumin305 (16.7%)13 (43.33%)12 (40%)25 (83.3%)0.0001Table 4:Effect of Nutritional variables on Toxicity ProfileVariableMyeloLiverGINeuroMetabolicPancreatitisTotalNormal Weight for age (275)45 (16.36%)15 (5.45%)1 (0.36%)13 (4.72%)2 (0.72%)2 (0.72%)55 (20%)Low Weight for Age (56)21 (37.5%)9 (16.07%)3 (5.36%)5 (8.93%)2 (5.36%)24 (42.8%)Normal Height for Age (297)50 (16.8%)17 (5.72%)2 (0.67%)14 (4.71%)2 (0.67%)2 (0.67%)61 (20.8%)Low Height for Age (34)16 (47%)7 (20.5%)2 (5.8%)4 (11.6%)2 (5.8%)18 (52.9%)Normal Albumin (301)12 (3.98%)3 (0.99%)02 (0.66%)1 (0.33%)64 (21.3%)Low Albumin (30)24 (66.67%)21 (58.33%)4 (11.1%)16 (44.4%)2 (5.55%)3 (8.3%)15 (50%)Table 5:Comparison of the effect of Nutritional variables in Remission vs Relapse Group with Other StudiesVariablesYu LC et al (1994)Present Study (2011)Remission GroupRelapseRemission GroupRelapseWeight %106.4 ± 29.9109.1 ± 19.323.96 ± 13.4421.1 ± 10.1Height %99.3 ± 4.9104.9 ± 16.1121.17 ± 27.02117.85 ± 22.8Protein Mean6.65 ± 0.886.38 ± 1.12Albumin Mean4.23 ± 0.33.59 ± 0.584.01 ± 0.733.63 ± 0.93 Disclosures:No relevant conflicts of interest to declare.

Determinants of Platelet Counts in Patients with Chronic Graft Versus Host Disease.

Abstract 2243Poster Board II-220 Introduction:Although a low platelet count in chronic graft-versus-host disease (cGVHD) patients is a consistent negative survival predictor across many cGVHD studies, this association is poorly understood. Additionally, a subgroup of cGVHD patients has elevated platelets, which is also unexplained. We hypothesized that such platelet abnormalities may reflect the pathophysiology of cGVHD and therefore be associated with specific clinical manifestations. This exploratory study was conducted to investigate determinants and clinical significance of platelet counts in a large cohort of cGVHD patients. Patients and Methods:cGVHD patients were enrolled in the ongoing cross sectional National Cancer Institute protocol (clinicaltrials.gov#NCT00331968). A detailed history, physical and extensive laboratory evaluations were obtained using standardized methods. National Institutes of Health (NIH) severity scores were based on the extent of cGVHD manifestations, degree of functional impairment and the number of affected organs. Wilcoxon rank sum test, Kruskal-Wallis test, and Jonckheere -Terpstra trend test were used in statistical analyses. Results with p-values<0.01 were considered as statistically significant, and those with Spearman rank correlation coefficient 0.3<r <0.5 as moderate to weakly correlated. Multiple regression analysis was performed with potentially predictive variables. Results:The cohort included 120 adult cGVHD patients (median age 48 years, 53% male (M), 47% female (F)). Median time from transplantation to study enrollment was 2.7 years. Median time of cGVHD diagnosis was 0.5 year after transplantation, and median time from cGVHD diagnosis to enrollment was 1.6 years. 75% of subjects received related donor transplant; 84% received peripheral blood stem cells and 16% bone marrow; 63% had a history of acute GVHD (skin 88%, gastrointestinal 45%, liver 22%). Median total NIH organ severity score was 6 (range 2-14), median average total NIH organ severity score was for F 0.750 (range 0.250-1.625), and for M 0.857 (range 0.286-2.000), and median number of organs involved was 4 (range 1-8). Median platelet count was 239,000/μL (range 33,000/μL-648,000/μL). F had significantly higher platelet counts than M (median 275,500 /μL vs. 221,500/μL, p=0.004). Time from transplantation (r=0.35) and time from cGVHD diagnosis (r=0.32) correlated positively with platelet counts (p=0.001). Patients who received TBI based conditioning tended to have higher platelet counts (median 279,000/μL vs. 211,000/μL, p=0.021). Platelet counts varied among the four recipient-donor gender pairings (M recipient/M donor had median platelets 199,500/μL, M recipient/F donor had median platelets 239,500/μL, F recipient/F donor had median platelets 257,000/μL, F recipient/M donor had median platelets 301,000/μL, p=0.010). Among patients with history of acute GVHD, those with liver acute GVHD tended to have higher platelet counts (292,000/μL vs. 226,500 /μL, p=0.027). Absolute lymphocyte count (r=0.35), absolute CD3s (r=0.31), absolute CD4s (r=0.31), absolute NK cells count (r=0.32), and calcium levels (r=0.33) correlated positively with platelet counts (p<0.01). NIH scores for involvement of joints/fascia (p=0.021) and lungs (p=0.039), maximal NIH individual (0-3) organ score (p=0.013), number of involved organs with cGVHD (p=0.0007), total NIH score (p=0.0002), and average total NIH organ severity score (r=0.28; p=0.002) were positively associated with platelet counts. None among the panel of laboratory parameters of acute inflammation correlated with platelet counts. In the linear multiple regression model, female patients (p=0.001), higher absolute lymphocytes count (p<0.0001) and higher average total NIH score (p<0.0001) remained the strongest simultaneous predictors of higher platelet counts. Conclusions:This exploratory analysis demonstrates strong positive association of platelet counts with the lymphocyte counts and with the average total NIH cGVHD organ severity scores. These data implicate abnormalities of platelet counts in cGVHD pathophysiology and warrant further research. Disclosures:No relevant conflicts of interest to declare.

On the need for biopsy confirmation at suspected first recurrence of cancer.

At the time of suspected first recurrence of cancer, it is unclear whether biopsy confirmation is routinely performed, although this is a very common clinical situation. First, 20 oncologists were surveyed to ascertain the pattern of practice in our community. A questionnaire with hypothetical typical cases suspected of having recurrent cancer was distributed. Second, eligibility criteria were reviewed from investigational protocols from the National Cancer Institute (NCI) and the Eastern Cooperative Oncology Group to see whether confirmation of recurrence was specifically required in these research studies. Third, 64 cases from our own practice were reviewed retrospectively to determine our patterns and results in performing biopsies to document suspected recurrence. Finally, criteria were developed that might suggest the need for biopsy confirmation of recurrence and then retrospectively tested against our cases. There was no clear consensus among oncologists regarding the need for tissue confirmation in patients with solid tumor with suspected recurrences, although rebiopsy was routinely requested for recurrent lymphoma. Published Eastern Cooperative Oncology Group and NCI protocols reviewed did not require biopsy proof specifically of recurrence. Retrospective review of our own cases suggested that, in the absence of one of the proposed indicators, the risk of making an erroneous diagnosis without biopsy confirmation is low. It is suggested that biopsy is not routinely necessary for confirmation of recurrence in all cases of suspected recurrent solid tumors, but criteria are proposed that would help to reduce the possibility of misdiagnosis when biopsy of suspected recurrence is not performed.

On the Need for Biopsy Confirmation at Suspected First Recurrence of Cancer

At the time of suspected first recurrence of cancer, it is unclear whether biopsy confirmation is routinely performed, although this is a very common clinical situation. First, 20 oncologists were surveyed to ascertain the pattern of practice in our community. A questionnaire with hypothetical typical cases suspected of having recurrent cancer was distributed. Second, eligibility criteria were reviewed from investigational protocols from the National Cancer Institute (NCI) and the Eastern Cooperative Oncology Group to see whether confirmation of recurrence was specifically required in these research studies. Third, 64 cases from our own practice were reviewed retrospectively to determine our patterns and results in performing biopsies to document suspected recurrence. Finally, criteria were developed that might suggest the need for biopsy confirmation of recurrence and then retrospectively tested against our cases. There was no clear consensus among oncologists regarding the need for tissue confirmation in patients with solid tumor with suspected recurrences, although rebiopsy was routinely requested for recurrent lymphoma. Published Eastern Cooperative Oncology Group and NCI protocols reviewed did not require biopsy proof specifically of recurrence. Retrospective review of our own cases suggested that, in the absence of one of the proposed indicators, the risk of making an erroneous diagnosis without biopsy confirmation is low. It is suggested that biopsy is not routinely necessary for confirmation of recurrence in all cases of suspected recurrent solid tumors, but criteria are proposed that would help to reduce the possibility of misdiagnosis when biopsy of suspected recurrence is not performed.

Antitumor activity of the mixed phosphine gold species chlorotriphenylphosphine-1,3-bis(diphenylphosphino)propanegold(I).

The title compound has been designed for antitumor activity based on structural features of related known antitumor gold agents, that is, gold-monophosphine and gold-diphosphine derivatives. It is a gold complex that contains both types of phosphine ligands, thus suggesting a possible synergistic action. The results of a single crystal X-ray structure determination of this molecule show the metal surrounded by 3 P atoms and one Cl anion in a distorted tetrahedral arrangement. The chloro anion, however, is weakly bound to the metal and so the species shows ionic character. The P NMR study, performed in solution, confirms the structural features observed in the solid and, in addition, indicates partial formation of other known gold(I)-diphosphine antitumor agents. The ionic character and strong Au-P bonds of this novel gold(I) species are similar to those of the most active antitumor gold compounds so far studied. The former feature contributes to solubility in biological fluids, and the latter prevents fast biomolecular attack. In addition, the title compound is less lipophilic, a feature recently correlated to lower liver toxicity. The title compound shows in vitro antitumor activity in the two initial National Cancer Institute protocols against human tumors. In the first screening, a unique dose (0.10 mM) of the title compound reduced cell growth of MCF7 (breast cancer), NCI-H460 (lung cancer), and SF-268 (Central Nervous System cancer-CNS) to 5, 8, and 11%, respectively. In the second protocol a 60-cell line panel was analyzed with the title compound concentration in the 0.1 mM-0.01 microM range. The highest activity was for the breast tumor cell line MCF7 with a LC(50) less than 0.01 microM. LC(50) values in the micromolar range were obtained for 29 cell lines. With the exception of leukemia, these micromolar activities were observed in at least one cell line for each subgroup tumor (non small lung, colon, CNS, melanoma, renal, prostate, breast, and ovarian). The leukemia inactivity was unexpected, as all antitumor gold(I) phosphine compounds in the literature described thus far are active. Melanoma was the most sensitive subgroup screened (five out of seven cell lines).

Reproducibility of measurements of potential doubling time of tumour cells in the multicentre National Cancer Institute protocol T92-0045

We compared the flow cytometric measurement and analysis of the potential doubling time (Tpot) between three centres involved in the National Cancer Institute (NCI) protocol T92-0045. The primary purpose was to understand and minimize the variation within the measurement. A total of 102 specimens were selected at random from patients entered into the trial. Samples were prepared, stained, run and analysed in each centre and a single set of data analysed by all three centres. Analysis of the disc data set revealed that the measurement of labelling index (LI) was robust and reproducible. The estimation of duration of S-phase (Ts) was subject to errors of profile interpretation, particularly DNA ploidy status, and analysis. The LI dominated the variation in Tpot such that the level of final agreement, after removal of outliers and ploidy agreement, reached correlation coefficients of 0.9. The sample data showed poor agreement within each of the components of the measurement. There was some improvement when ploidy was in agreement, but correlation coefficients failed to exceed values of 0.5 for Tpot. The data suggest that observer-associated analysis of Ts and tissue processing and tumour heterogeneity were the major causes of variability in the Tpot measurement. The first two aspects can be standardized and minimized, but heterogeneity will remain a problem with biopsy techniques. © 1999 Cancer Research Campaign

In the Search for New Anticancer Drugs. 27. Synthesis and Comparison of Anticancer Activity in Vivo of Amino Acids, Carbohydrates, and Carbohydrate-Amino Acid Conjugates Containing the [N′‐(2‐chloroethyl)‐N′‐nitrosoamino]carbonyl group

The [N′‐(2‐chloroethyl)‐N′‐nitrosoamino]carbonyl [(2‐chloroethyl)nitrosocarbamoyl, CNC] moiety containing compounds CNC‐glycinamide2d, CNC‐amino acid derivatives7a‐d, and carbohydrate‐CNC‐amino acid conjugates13, 18, 22, 23, 27, and28were synthesized and evaluatedin vivofor their anticancer activities against the murine lymphocytic leukemia P388 using the National Cancer Institute (NCI) protocol. The most active compound was2dwith a 520% increase in life span (% ILS) and 6/6 survivors after 60 days. The CNC‐amino acid analogs7a‐dpossessed high to moderate activities with maximum % ILS values of 270, 174, 141 and 132, respectively. Among the carbohydrate‐CNC‐amino acid derivatives the α‐methyl glycoside derivatives22and23were most active with maximum % ILS values of 277 and 137, respectively, followed by the hemiacetal carbohydrate analogs13and18with % ILS values of 93 and 149, respectively, and the tetra‐O‐acetyl derivatives27and28with % ILS of 110 and 111, respectively. Compounds7b,18,23and28were then testedin vivoagainst the murine lymphoid leukemia L1210 using the NCI protocol. In this case, the hemiacetal type carbohydrate‐CNC‐amino analog18had the highest activity with a maximum % ILS value of 477 and 4/6 survivors on day 60, followed by7b(275% ILS),23(152% ILS) and28(106% ILS). The lipophilicities of all CNC compounds were determined by the partition coefficient using the UV method. A correlation of % ILS values with logPvalues indicated, in general, an increase in cytotoxicity with a decrease in hydrophilicity for the carbohydrate‐CNC‐amino acid conjugates13,18,22,23and the clinical drugs streptozotocin (1e), chlorozotocin (1f), and cymerin (1g).

Sarcomas of the hand and foot: Analysis of local control and functional result with combined modality therapy in extremity preservation

Purpose : The records of 28 patients with sarcomas of the hand and foot treated at the National Cancer Institute (NCI) between 1977 and 1992 were reviewed to assess local control and functional results. Methods and Materials : Histologic types included 15 cases of the Ewing's sarcoma family of tumors, 7 cases of alveolar rhabdomyosarcoma, and 6 cases of nonrhabdomyosarcoma soft tissue sarcomas. Median age of all patients was 18 years (range 4–61), with a median potential follow-up of 114 months following diagnosis. Surgery varied from incisional biopsies for Ewing's Sarcoma and rhabdomyosarcoma lesions to complete excision when possible for nonrhabdomyosarcoma soft tissue sarcoma lesions. Amputation was not primarily performed, except in two patients who underwent ray resections of hand lesions (patients 13 and 24). Radiotherapy generally consisted of 50 Gy/25 fractions (fx)/5 weeks for Ewing's Sarcoma, 54 Gy/30 fx/6 weeks for rhabdomyosarcoma, and 63 Gy/35 fx/7 weeks for nonrhabdomyosarcoma soft tissue sarcomas. Chemotherapy was administered on various NCI protocols. Results : Actuarial local control for Ewing's Sarcoma was 84% at 5 and 10 years. All but one survivor are capable of hand/foot function for routine activities without orthotic requirements. Five of six patients (83%) who died of metastatic disease had functional distal extremities. Actuarial local control for rhabdomyosarcomas was 100% at 5 years, with similar function. Three patients (60%) died of distant disease progression with functional distal extremities. Five-year actuarial local control for nonrhabdomyosarcoma soft tissue sarcomas was 100%, with equivalent function. No patient developed a second malignancy in the treatment field. Conclusions : Although equivalent local control may be achieved in these lesions with either amputation or radiotherapy, a prudent management course would be to defer amputation for management of local recurrences. Many patients with these lesions fail in distant sites only and die without local failure. For these patients and for those who remain long-term survivors, we believe a functional hand and foot provides a better quality of life than a prosthesis.

Antibodies against autologous tumor cell proteins in patients with small-cell lung cancer: association with improved survival.

The frequency and clinical relevance of human antitumor immune responses is not well known, and few target antigens have been identified. This study was designed to determine the frequency of antibodies reactive against extracts of autologous tumor cell lines and to correlate these data with survival. Serum samples were obtained from 40 lung cancer patients treated on National Cancer Institute protocols. These sera were used as probes in immunoblots against protein extracts from tumor cell lines derived from each of these patients. We detected serum antibodies against autologous tumor cell proteins in 21 (58%) of the 36 patients with small-cell lung cancer (SCLC) and three (75%) of the four with non-small-cell lung cancer (NSCLC). Two patients' sera detected the p53 tumor suppressor gene product and two detected the product of the HuD gene (associated with paraneoplastic neurological syndromes) in their autologous tumor cell lysates. SCLC patients with antibodies against autologous tumor cell proteins had improved survivals compared with those in the antibody-negative group (P = .059). All patients who lived longer than 36 weeks were antitumor antibody positive. Sera from six (86%) of seven patients with limited disease were positive for antibodies that reacted against autologous tumor cells, compared with 15 of 29 (52%) of sera from patients with extensive disease. Our results suggest that the sera from patients with SCLC frequently contain antibodies against tumor cell proteins and that these antibodies are associated with improved survival. These data suggest that an antitumor immune response may affect tumor growth, and that the anonymous proteins detected by antitumor antibodies in lung cancer patient sera may represent proteins involved in the development of lung cancer or in its clinical manifestations.

Patterns of disease recurrence following definitive therapy of adenocarcinoma of the pancreas using surgery and adjuvant radiotherapy: Correlations of a clinical trial

Purpose : Between 1980 and 1984, 26 patients with resectable adenocarcinoma of the pancreatic head were enrolled in a National Cancer Institute protocol evaluating intraoperative radiotherapy vs. standard therapy. Methods and Materials : After complete excision of their lesions, patients were observed (Stage I), or randomized to intraoperative radiotherapy vs. external beam radiotherapy (Stages II-IV). The intreoperative dose was 20 Gy in a single fraction using 9–20 MeV electrons. The external beam radiotherapy schema involved daily 150–180 cGy fractions to 45–55 Gy in 5–6 weeks. Chemotherapy was not used for primary disease but was administered off protocol for recurrent disease. Median potential followup for the trial was > 9 years, with a median patient survival of 18 months. Perioperative mortality was 27% (7 patients). Of the remaining 19 patients, one remains alive and without evidence of disease 9 years post-therapy. Twelve patients underwent autopsy and 2 required antemortem laparotomy; histopathologic evidence of disease recurrence was analyzed. Results : Of 15 patients evaluable for intra-abdominal control, 7 (47%) suffered local recurrences and 7 (47%) failed regionally, with 5 patients (35%) failing in both areas. Five patients (35%) developed peritoneal seeding. Of 13 patients evaluable for systemic disease, 8 (62%) suffered distant failure. There were no differences in outcome between intraoperative or external beam radiotherapy or observation in this subset of patients. Conclusion : This report is among the most rigorous descriptions of patterns of failure after resection of pancreatic cancer. It appears clear that advances in local control of this disease are unlikely to translate into increased survival in the absence of improved systemic therapy.

In the Search for New Anticancer Drugs. XXV: Role of M-Nitrosated Amadori Compounds Derived from Glucose-Amino Acid Conjugates in Cancer Promotion or Inhibition

Earlier investigators found that some W-nitrosated Amadori compounds, derived from glucose and amino acid condensation reactions, exhibit mutagenic properties and theorized that these potentially carcinogenic compounds might be formed in the human digestive system. To further investigate these compounds, AAnitrosated Amadori compounds [i.e., N-(1 -deoxy-D-fructos-1 -yl)-L-N-nitroso-glycine (5a), -threonine (5b), -methionine (5c), -valine (5d), -phenylalanine (5e), and -tryptophan (5f)] were synthesized by modifications of known methods. Acute toxicity tests of 5a, 5b, 5c, 5d, 5e, and 5f in male Swiss mice produced the following lowest lethal limits of toxicity: 2000,2000,4000, 3000,2000, and 6000 mg/kg, respectively, whereas the highest tolerated doses were 1750,1500,3000,1500, and 5000 mg/kg, respectively. The 50% lethal dose (intraperitoneally) for 5b in mice was ~1777 mg/kg. This value is at least three times higher than that for the over-the-counter drug ibuprofen (i.e., 495 mg/kg, intraperitoneally, in mice). Compounds 5b, 5c, 5d, and 5f were evaluated in vitro by the National Cancer Institute primary antitumor screen consisting of 60 cell lines. None of the four compounds caused a significant inhibition of cell growth, even at the maximum dosage of 10-4 M. Compounds 5a-f were tested in vivo against the lymphocytic leukemia P388, and 5b and 5f were tested against the lymphoid leukemia L1210 in CDF, male mice following the National Cancer Institute protocol. There were no significant differences in results between the control and drug-treated mice. The percent increase in lifespan ranged from –15 to +15 (T/C = 85–115) for P388 and from –5 to +1 (T/C = 95-101) for L1210, whereas the values for the positive control 5-fluorouracil were 71 and 67 (T/C = 171 and 167), respectively. The combination of very low acute toxicity, low in vitro cytotoxicity in primary tumor screens, and no activity in vivo led to the hypothesis that none of the compounds 5a-f are carcinogenic or cytotoxic. It is possible that these compounds are not metabolicaily activated by either ɑ– or β-oxidations or by retro-aldol cleavage reactions.