Strategies to improve radiotherapy with targeted drugs

Radiotherapy is a very effective adjuvant treatment for rectal cancer with little side effects. Its killing effect on tumor cells seems to be more profound than the effect on normal tissue. The molecular events caused by irradiation are mainly analyzed in in vitro and animal models; investigations on human material are rare. In the current study, we analyzed the effects of irradiation on gene expression in normal and tumor tissue of rectal cancer patients. Normal and carcinoma tissue of patients from a randomized clinical trial of the benefits of preoperative radiotherapy were analyzed using the Affymetrix Human Cancer Gene Chip. Preoperative radiotherapy was given within 5 days prior to surgery. Results for normal tissue and tumor were compared to investigate the radiation-related differences between normal and tumor cells. We clustered the differentially expressed genes based on their functional annotation. Results were compared with immunohistochemical and literature data. The majority of the investigated cancer-related genes remained unchanged by irradiation (92% in tumor tissue and 93% in normal tissue). The differentially expressed genes varied between tumor and normal tissue except for maspin and IL-8. Both in tumor and normal tissue, differentially expressed genes were present related to cell signaling and cycle control, apoptosis and cell survival and tissue response and repair. However, the spectrum of affected genes was totally different. Pre-existing differences in gene expression between normal tissue and tumor tissue might explain the differences in their responses to radiation. This change in response may explain the clinical beneficial effect of radiotherapy on tumor cells (low local recurrence rate) and the less severe effects on normal tissue (minor side effects).

Background: Disparities in colorectal cancer (CRC) incidence and outcomes are notable between racial and ethnic groups. While these differences may be associated with various factors, established risk factors do not fully explain observed disparities. Increasingly, evidence suggests that the microbiota associated with the tumor microenvironment may play a role in CRC initiation and/or progression. However, few studies have investigated the microbiome of CRC tumor tissue compared to the microbiome in paired normal tissue as a reference. This study explores the microbial landscape of tumor and normal colorectal tissues from diverse racial and ethnic patients with CRC to understand the microbiome’s potential role in CRC disparities. Methods: We conducted bacterial 16S rRNA gene sequencing of colorectal tumor and normal tissue from 283 individuals of varied racial and ethnic backgrounds (African American, Alaska Native, Hispanic, and Non-Hispanic White). We extracted genomic DNA from tumor and normal colon tissue. We used amplicon sequencing of the 16S rRNA gene to classify the relative abundance of different bacterial amplicon sequence variants (ASVs), genera, and phyla in colon tissue. 8 phyla and 48 genera were identified across all samples. We used linear mixed models to identify differences in alpha diversity (assessed as Shannon diversity) between tumor and normal tissue in all participants combined, as well as within each racial and ethnic group. Linear mixed models were also used to identify associations of specific genera with the tumor and normal tissues. All models were adjusted for sex, race, age, ethnicity, tumor stage, tumor site, and year of diagnosis. Results: Our analysis identified significant differences in microbial composition and diversity between normal and tumor tissue. Alpha diversity of the microbiota was significantly lower in tumor tissue compared with normal tissues when all racial and ethnic participants were combined (p < 0.001). There were also significant differences in alpha diversity between normal and tumor tissue within different racial and ethnic populations (p < 0.05) as well as within tumor tissue between different racial and ethnic populations (p < 0.05). After accounting for multiple testing, several genera were found to have a lower relative abundance in tumor tissue compared to normal tissue (p < 0.05). These included Bacteroides, Parabacteroides, Ruminococcus, Faecalibacterium, and other genera. Conclusion: This study enhances our understanding of the CRC-related microbiome. By detailing the diversity and abundance profiles of microbial communities in tumor microenvironments versus normal tissues, we identified differences in the distribution and composition of the microbiome in tumor versus normal tissue, as well as identified specific bacterial genera that were found to have a lower relative abundance in tumor tissues. Citation Format: Scott D. Labrie, Claire E. Thomas, Nicole Loroña, Keith R. Curtis, Hang Yin, Timothy W. Randolph, Ningxin Ma, Jeroen R. Huyghe, Sushma S. Thomas, Li Hsu, Amanda L. Koehne, Sosun Nayemi, Orsalem J. Kahsai, Jane C. Figueiredo, Li Li, Diana G. Redwood, Timothy Thomas, Christopher I. Li, Amanda I. Phipps, Ulrike Peters, Meredith Hullar. Comparative microbiome analysis of tumor and normal colorectal tissue across diverse racial and ethnic groups [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6516.

Oxidative benzo(a)pyrene (BP) metabolism was studied in 12,000 g(S12) supernatant fractions of surgical lung specimens obtained from lung cancer patients undergoing surgical resection. Arylhydrocarbon (BP)hydroxylase (AHH) activity was determined in both normal and tumorous lung tissue specimens from the same patient. A more than 20‐fold inter‐individual variation in AHH activity was found in both the normal and the tumorous lung tissue samples investigated. The apparent KM of the pulmonary enzyme (1–2.5 × 10−4 M) was identical in tumorous and normal tissue. High‐performance liquid chromatography (HPLC) of ethyl acetate‐extractable BP metabolites from the normal lung tissue of six patients identified phenols, dihydrodiols and quinones of BP. The mean yields of these metabolites were very similar to those obtained in assays in the presence of lung S12 from BDVI rats, except that more trans−9, 10‐dihydro‐9, 10‐dihydroxy‐BP (BP‐9, 10‐diol) was formed in the presence of human lung S12. The formation of 3‐hydroxy‐BP (3‐HO‐BP) and BP‐9, 10‐diol in human lung specimens, however showed variations up to 7‐ and 13‐fold, respectively. In normal lung specimens, the total amounts of 3‐HO‐BP and 9‐HO‐BP formed correlated positively with the total amounts of trans‐7, 8‐dihydro‐7, 8‐dihydroxy‐BP (BP‐7, 8‐diol), BP‐9, 10‐diol and trans‐4, 5‐dihydro‐4, 5‐dihydroxy‐BP (BP‐4, 5‐diol) formed (r = 0.83; p < 0.05). A positive correlation was also found between the AHH activity in normal and tumorous tissue from all cancer patients (r = 0.24; p < 0.05). The ratio (R) of AHH activity in tumorous tissue to that in normal tissue from the same lung cancer patient was < 1 in 73 patients, 1 in 2 patients, and > 1 in 11. In 7 of 10 subjects who underwent surgical resection for suspected lung tumours, but in whom histological analysis revealed no malignant tissue, R was > 1 for the AHH activity in the inflammatory tissue to that in normal tissue. The observation of consistently lower AHH activity in malignant lung tissue parallels findings in hyperplastic liver nodules induced in rats by hepatocarcinogens. The frequency distribution of pulmonary AHH activity in normal and tumorous tissue from 105 lung cancer patients, with all types of tumours, was compatible with a unimodal distribution of the enzyme activity. In a subset of 43 patients with squamous‐cell carcinoma, the same distribution was seen; in those patients, AHH activity in tumorous tissue correlated negatively (r = −0.18; p > 0.10) with the number of cigarettes smoked per day prior to surgery. No differences were observed between the mean values for AHH activity in cases of squamous‐cell carcinoma and those of adenocarcinoma. No relationship was found between AHH activity in normal or tumour tissue and the age of patients. Whether differences in pulmonary AHH activity represent a host risk factor in persons who smoke remains to be investigated.

Background:In vitro and preclinical data support the notion that anti-progesterone therapy will have activity against both estrogen and progesterone receptors (ER/PR) positive and negative breast cancer, but biomarkers of efficacy may differ in different types of breast cancer. We have conducted a pre-surgical window trial of oral telapristone acetate (TPA, CDB-4124) treatment in early breast cancer patients. This first ever trial of oral TPA for breast cancer and DCIS patients will provide us pilot biologic data that will help select the right population and the right biomarkers for future trials. Here we report distribution of TPA in plasma and breast normal tissue and tumors collected at surgery. Methods:our trial was a 1:1 randomized, double-blind, and placebo-controlled pre-surgical window trial of oral TPA 12mg (Proellex, CDB-4124, Repros Therapeutics Inc.) treatment for 2-10 weeks. 70 pre and postmenopausal women undergoing surgery for Stage 0-II breast cancer were recruited to the study. The surgical samples of 61 patients were used to determine the concentrations of TPA and its mono-demethylated metabolite (dTPA, CDB-4453) in plasma and matched normal tissue and tumor by Liquid chromatography–tandem mass spectrometry at the Illinois Institute of Technology, while maintaining the blind for the primary endpoint of cell proliferation. Statistical significance and analysis were calculated by Wilcoxon matched-pairs signed rank test and non-parametric Spearman correlation. Results: We found that 32/61 women displayed detectable plasma concentrations of TPA and dTPA (median with IQR) 109ng/mL (71.3, 216) and 46.5 ng/mL (34.2, 73.7), respectively. TPA concentration was 2.3 times higher than dTPA in plasma (p<0.0001). The normal and tumor tissue samples of these 32 women were further analyzed. In normal tissue samples, the concentrations of TPA and dTPA were 283 ng/g (70.7, 326) and 51.0 ng/g (24.4, 122), respectively. TPA concentration was 5.5 fold higher than dTPA in normal tissue (p<0.0001). In tumors, the TPA and dTPA concentrations were 137 ng/g (31.1, 278) and 36.4 ng/g (17.3, 68.7), respectively. TPA concentration was 3.8 fold higher than dTPA in tumors (p<0.0001).Interestingly, TPA and dTPA were more abundant in normal tissue than in tumors (p=0.0005 for TPA, and p=0.0013 for dTPA). We found that TPA and dTPA was highly correlated in plasma (r=0.492, p=0.0042). Plasma TPA concentration was highly correlated with normal tissue concentration (r=0.61, p=0.0003) but non-significantly correlated with tumor concentration (r=0.32, p=0.147). However, the normal and tumor tissue concentrations of TPA and dTPA were highly correlated (r=0.71, p=0.0002 for TPA and r=0.556, p=0.0072 for dTPA). Conclusions: Plasma TPA concentrations reflect concentration in normal breast tissue better than in tumors. However, within the breast, TPA concentration in normal and tumor tissue is correlated. Our trial is to be unblended shortly, and we plan to relate these results to the proliferative rates in tumor and normal tissue. The variability observed in plasma and tissue concentrations also suggests that pharmacogenomics studies may be appropriate in the future. Citation Format: Lee O, Muzzio M, Ivancic D, Rogers C, Allu S, Khan SA. Phase II pre-surgical window trial of telapristone acetate (TPA) in early breast cancer and DCIS patients: Distribution of TPA in plasma, normal breast tissue and tumors [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr OT3-02-09.

In this study, endogenous long chain ceramides were measured in 32 human head and neck squamous cell carcinoma (HNSCC) and 10 nonsquamous head and neck carcinoma tumor tissues, as compared with adjacent noncancerous tissues, by liquid chromatography/mass spectroscopy. Interestingly, only one specific ceramide, C(18:0)-ceramide, was selectively down-regulated in the majority of HNSCC tumor tissues. On the other hand, in nonsquamous tumor tissues, this selectivity for C18-ceramide was not detected. These data suggested the hypotheses that decreased levels of C18-ceramide might impart a growth advantage to HNSCC cells and that increased generation of C18-ceramide may be involved in the inhibition of growth. These roles were examined by reconstitution of C18-ceramide at physiologically relevant concentrations in UM-SCC-22A cells (squamous cell carcinoma of hypopharynx) via overexpression of mammalian upstream regulator of growth and differentiation factor 1 (mUOG1), a mouse homologue of longevity assurance gene 1 (mLAG1), which has been shown to specifically induce the generation of C18-ceramide. Liquid chromatography/mass spectroscopy analysis showed that overexpression of the mLAG1/mUOG1 resulted in increased levels of only C(18:0)-ceramide by approximately 2-fold, i.e. concentrations similar to those of normal head and neck tissues. Importantly, increased generation of C18-ceramide by mLAG1/mUOG1 inhibited cell growth (approximately 70-80%), which mechanistically involved the modulation of telomerase activity and induction of apoptotic cell death by mitochondrial dysfunction. In conclusion, this study demonstrates, for the first time, a biological role for LAG1 and C18-ceramide in the regulation of growth of HNSCC.

Long noncoding RNAs (lncRNAs) are an emerging class of key regulatory RNAs that are greater than 200nt and do not code for protein. They appear to be critical for various biological processes including gene regulation. Increasing evidence has suggested that aberrantly expressed lncRNAs play a key role in the initiation and progression of breast cancer. In most studies of breast cancer, breast tumor tissue was compared to adjacent normal-appearing tissue to identify differentially expressed lncRNAs. However, expression of lncRNAs in normal breast tissue from healthy donors has rarely been studied, and the differences in expression of lncRNAs in breast tumor and normal breast tissue from healthy donors remain largely unknown. Using RNA-sequencing data generated from freshly-frozen breast tissue samples, we examined the expression of lncRNAs in 119 breast tumor, 47 adjacent normal-appearing breast tissue samples, and 23 normal breast tissue samples from healthy donors. Bowtie and RESM algorithms were used to align raw reads and quantify transcript abundance. Differential expression analyses were performed using edgeR software. We assessed the expression of a total of 7,844 mapped lncRNAs using normalized transcript counts. Unsupervised Principal Components Analysis (PCA) demonstrated that the lncRNA expression profile of adjacent normal-appearing tissue was different compared to normal breast tissue from healthy donors, and partially overlapped with the tumor. At various fold change thresholds, more differentially expressed lncRNAs can be detected when the tumor tissue is compared to normal tissue than to adjacent non-tumorous tissue. With a false discovery rate (FDR) < 0.05 and a fold change (FC) of more than ±2, we identified 107 differentially expressed lncRNAs for the tumor vs. normal breast tissue, 53 lncRNAs for tumor vs. adjacent normal-appearing tissue, and 20 lncRNAs for adjacent normal-appearing vs. normal breast tissue. In our analyses of tumor vs. adjacent normal-appearing tissue and tumor vs. normal tissue from healthy donors, we identified differentially expressed lncRNAs including HOTAIR, GAS5 and ANRIL/CDKN2B-AS1 that were previously implicated in breast cancer, with considerably higher fold changes in latter comparison. When using normal breast tissue from healthy donors as the baseline, we identified novel putative breast cancer-associated lncRNAs including EMX2OS, TERC, HOTAIRM1, and WT1-AS that are aberrantly expressed in breast tumor. These lncRNAs were not identifiable in a direct comparison of tumor and adjacent normal-appearing tissue. Our results suggest that adjacent normal-appearing tissue undergoes changes in lncRNAs expression similar to those observed in tumor tissue to some extent, and a number of new and potentially important lncRNAs can only be detected using normal tissue from healthy donors as an optimal baseline tissue. Citation Format: Erin Wagner, Yunlong Liu, Bryan Schneider, Anna Maria Storniolo, Jiali Han, Chunyan He. Differences in expression of lncRNAs in breast tumor, adjacent normal-appearing breast tissue, and normal breast tissue from healthy donors. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3268. doi:10.1158/1538-7445.AM2014-3268

Introduction: Through the remarkable advances in genomics, we have identified many of the genetic alterations causing breast cancer(BC) development. Moreover, recent research revealed that these genetic alterations have been continually occurring and accumulating long before cancer develops. However, the role of normal breast tissue in BC patients has been veiled. In this study, we aimed to determine gene expression affecting cancer development from normal breast tissue using paired breast normal-cancer tissues. Methods: We collected BC-normal paired tissues which harvested during curative BC surgery since August 2008 to February 2013. Using these samples, we performed RNASeq, nCounter customized gene expression analysis and WES. Results: Of 134 BC tissues, 121 tumor tissues and 51 normal tissues were analyzed using nCounter method. Among these samples, 50 were paired BC-normal tissues. In 51 normal breast tissues, there was all normal like intrinsic subtype. Further DEG analysis according to tumor subtype, there was no significant different in normal tissues among BC subtype. We also analyzed the different gene expression between tumor-normal tissues according to BC subtypes. In HR+HER2- BC, MMP11, BRIC5 and KNTC2 were significantly upregulated in tumor tissue compared to normal tissue (all adjusted P value < 0.05 and log2FoldChange > 1). In HR+HER2+BC, only MMP11 was upregulated but SFRP1, ELF5 and FOXC1 expression decreased. TNBC had significantly different gene expression compared to normal breast tissue. Ten genes (ANLN, APOBEC3B, BIRC5, CDCA1,CEP55, EXO1, KIF2C, KNTC2, MYBL2 and ORM2) were overexpressed in tumor tissue whereas seven genes(AR, ESR1, ERBB4, MLPH, PGR, PIP and SCUBE2) expression was decreased. In HR-HER2+BC, ANLN overexpression but ESR1, MIA and KRT14 downregulation were observed. Further DEG analysis according to survival outcome, upregulation of MMP11 and MKI67 were associated to BC recurrence but BIRC5, CEP55M EXO1 and KNTC2 overexpression were associated to BC recurrence free survival regardless of BC subtypes (all adjusted P value < 0.05 and log2FoldChange > 1). Conclusion: Although there were no different gene expression in normal breast tissues according to BC subtype, significant difference of gene expression between tumor and normal tissues were observed according to BC subtype. In particular, TNBC had the highest number of genes with differential expression in tumor and normal tissue. In addition, different expression of some genes was also associated to BC recurrence. Citation Format: J. Kim, K. Park, J. Shin, H. Ahn, J. Ahn, S. Lee, Y. Park. Different gene expression between breast cancer and adjacent normal breast tissue according to breast cancer subtype [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PS4-08-19.

Taking Care with FLASH Radiation Therapy

SUMMARY Concentrations of estrogen (er) and progesterone (pr) receptors were measured by radioreceptor assay in tumor (n = 319) and normal (n = 166) mammary tissue from 248 bitches. Correlations between er and pr and between receptor expression in tumor and normal mammary tissue from the same bitches were evaluated. The influence of tumor, clinical, or hormonal variables on receptor expression also was studied. Approximately 80% of tumor and 95% of normal mammary tissue expressed detectable concentrations of er, pr, or both. Direct correlation was found between er and pr concentrations in normal and tumor tissues. Median er concentrations were significantly higher (46 ± 47 fmol/mg of cytosolic protein vs 27 ± 24 fmol/mg of cytosolic protein; P = 0.0002) in normal than in tumor tissue. On the other hand, pr concentrations were significantly higher (57 ± 52 fmol/mg vs 77 ± 99 fmol/mg; P = 0.03) in tumors (especially benign tumors) than in normal tissue. Poorly differentiated malignant tumors expressed lower concentrations of receptors than did benign or well differentiated malignant tumors. The er and pr concentrations decreased with increasing size of the lesion. Hormonal status of the bitch significantly (P < 0.05) influenced receptor expression in normal tissue: bitches in the luteal phase of the estrous cycle had higher concentrations of er (69 ± 62 fmol/mg) than did ovariectomized bitches (24 ± 19 fmol/mg) or bitches in anestrus (38 ± 45 fmol/mg) or the follicular phase (13 ± 7 fmol/mg). For pr, higher concentrations were observed in normal tissue during anestrus than during pseudopregnancy or in bitches treated with medroxyprogesterone acetate. Similar, but nonsignificant, variations were seen in tumor tissue except in medroxyprogesterone acetate-treated bitches in which pr concentrations were high in tumors and low in normal tissue from the same bitches. Significant relation between age, hormonal history, number or location of lesions, and receptor expression was not observed. However, significantly (P < 0.05) lower receptor concentrations were found in the normal tissue from the 3 cranial pairs of mammary glands (er, 37 ± 45; pr, 43 ± 30 fmol/mg) than from the 2 caudal pairs (er, 51 ± 50; pr, 62 ± 53 fmol/mg). A direct correlation was found for both receptor concentrations between normal tissue adjacent to and distant from the lesions and between normal tissue adjacent to the lesion and the corresponding tumor. In comparison with that in normal tissue, the general mechanism of modulation of receptor expression seems not to be modified in benign lesions and well differentiated malignant tumors. However, hormonal sensitivity is lost in dedifferentiated tumors.

Strategies for vascular targeting in tumors.

Benzo[ a]pyrene diol-epoxide-I-DNA and oxidative DNA adducts associated with gastric adenocarcinoma

Cultured cells can be sensitized to low-temperature hyperthermia (below 43.0 degrees C) by a prior heat shock at a high temperature (above 43.0 degrees C) if the heat above 43.0 degrees C is immediately followed by the heat below 43.0 degrees C. This effect has been termed step-down heating (SDH). We have studied the effect of SDH on the response of murine tumor and normal tissues treated at 45.5 degrees C. Animal tumors were eighth-generation isotransplants of a spontaneous fibrosarcoma in C3Hf/Sed. mice. Tumor response was studied by TG (tumor growth) time assay, i.e., determination of the time required for half the treated tumors to reach 1000 mm3 from the first day of treatment. Normal tissue response was studied in the mouse foot. End point was the time to induction of a score 4.0 (loss of a toe) or greater reaction in half the treated animals, RD50. The SDH at 41.0 degrees C sensitized the response of tumor and normal tissues to 45.5 degrees C. The enhancement ratios were congruent to 1.7 for both tissues, indicating no differential sensitization between tumor and normal tissues. No sensitization was observed if the SDH was given immediately after a second dose of 45.5 degrees C given 6 hr to 5 days following the first dose of 10 min at 45.5 degrees C. The SDH appeared not to inhibit the development of thermal resistance as evidenced by no appreciable changes in the thermal resistance ratio.

Microsatellite alteration in histologically normal lung tissue of patients with non-small cell lung cancer

Objective To investigate the expression and promoter methylation status of p73 gene in children with nephroblastoma and their clinicopathological correlations. Methods The methylation-specific PCR and real-time quantitative PCR were used to detect the mRNA expression and methylation status of p73 gene in 38 cases of nephroblastoma, 15 cases of adjacent tumor and 15 cases of normal renal tissues, then their clinicopathological correlations and how the p73 gene methylation affected its transcription were analyzed. Results Relative quantity(RQ) of p73 mRNA in tumor tissues was 0.79±0.21, 0.74±0.19 in peritumoral tissues, and 0.48±0.17 in normal renal tissues, and differences among the 3 groups were statistically significant, in which tumor and peritumoral tissues were higher than normal tissues(all P 0.05). p73 gene methylation-positive rate in tumor was 26.3%(10/38 cases), adjacent tumor was 33.3%(5/15 cases), normal kidney tissue was 80.0%(12/15 cases), and differences among the 3 groups were statistically significant, in which tumor and peritumoral tissues were both lower than normal tissues(all P 0.05). Difference in RQ values among methylated tumor, adjacent tumor and normal renal tissues was statistically significant(P=0.000), in which tumor and adjacent tumor were both higher than normal renal tissues(P<0.05); But difference of RQ value among unmethylated tumor, adjacent tumor and normal renal tissues was not statistically significant(P=0.075). Conclusions The close correlation between p73 low methylation and high mRNA expression suggests that aberrant promoter methylation is possibly one of the gene expression regulations, and also connected with the development of nephroblastoma.The p73 gene in methylated nephroblastoma may play the role of oncogenes, as there is a negative correlation tendency between the overexpression in transcriptional level and its methylation status. Key words: p73; Nephroblastoma; Tissue; Promoter methylation; mRNA expression

Hyperthermic intraperitoneal chemotherapy (HIPEC) with carboplatin induces distinct transcriptomic changes in ovarian tumor and normal tissues