Progressive plasticity during colorectal cancer metastasis.

As cancers progress, they become increasingly aggressive-metastatic tumours are less responsive to first-line therapies than primary tumours, they acquire resistance to successive therapies and eventually cause death1,2. Mutations are largely conserved between primary and metastatic tumours from the same patients, suggesting that non-genetic phenotypic plasticity has a major role in cancer progression and therapy resistance3-5. However, we lack an understanding of metastatic cell states and the mechanisms by which they transition. Here, in a cohort of biospecimen trios from same-patient normal colon, primary and metastatic colorectal cancer, we show that, although primary tumours largely adopt LGR5+ intestinal stem-like states, metastases display progressive plasticity. Cancer cells lose intestinal cell identities and reprogram into a highly conserved fetal progenitor state before undergoing non-canonical differentiation into divergent squamous and neuroendocrine-like states, a process that is exacerbated in metastasis and by chemotherapy and is associated with poor patient survival. Using matched patient-derived organoids, we demonstrate that metastatic cells exhibit greater cell-autonomous multilineage differentiation potential in response to microenvironment cues compared with their intestinal lineage-restricted primary tumour counterparts. We identify PROX1 as a repressor of non-intestinal lineage in the fetal progenitor state, and show that downregulation of PROX1 licenses non-canonical reprogramming.

Organ preservation after neoadjuvant long-course chemoradiotherapy versus short-course radiotherapy.

BACKGROUND: Potential differences in organ preservation between total neoadjuvant therapy (TNT) regimens integrating long-course chemoradiotherapy (LCCRT) and short-course radiotherapy (SCRT) in rectal cancer remain undefined. PATIENTS AND METHODS: This natural experiment arose from a policy change in response to the COVID-19 pandemic during which our institution switched from uniformly treating patients with LCCRT to mandating that all patients be treated with SCRT. Our study includes 323 locally advanced rectal adenocarcinoma patients treated with LCCRT-based or SCRT-based TNT from January 2018 to January 2021. Patients who achieved clinical complete response were offered organ preservation with watch-and-wait (WW) management. The primary outcome was 2-year organ preservation. Additional outcomes included local regrowth, distant recurrence, disease-free survival (DFS), and overall survival (OS). RESULTS: Patient and tumor characteristics were similar between LCCRT (n = 247) and SCRT (n = 76) cohorts. Median follow-up was 31 months. Similar clinical complete response rates were observed following LCCRT and SCRT (44.5% versus 43.4%). Two-year organ preservation was 40% [95% confidence interval (CI) 34% to 46%] and 31% (95% CI 22% to 44%) among all patients treated with LCCRT and SCRT, respectively. In patients managed with WW, LCCRT resulted in higher 2-year organ preservation (89% LCCRT, 95% CI 83% to 95% versus 70% SCRT, 95% CI 55% to 90%; P = 0.005) and lower 2-year local regrowth (19% LCCRT, 95% CI 11% to 26% versus 36% SCRT, 95% CI 16% to 52%; P = 0.072) compared with SCRT. The 2-year distant recurrence (10% versus 6%), DFS (90% versus 90%), and OS (99% versus 100%) were similar between WW patients treated with LCCRT and SCRT, respectively. CONCLUSIONS: While WW eligibility was similar between cohorts, WW patients treated with LCCRT had higher 2-year organ preservation and lower local regrowth than those treated with SCRT, yet similar DFS and OS. These data support induction LCCRT followed by consolidation chemotherapy as the preferred TNT regimen for patients with locally advanced rectal cancer pursuing organ preservation.

Immune checkpoint inhibitors for POLE or POLD1 proofreading-deficient metastatic colorectal cancer.

BACKGROUND: POLE and POLD1 proofreading deficiency (POLE/D1pd) define a rare subtype of ultramutated metastatic colorectal cancer (mCRC; over 100 mut/Mb). Disease-specific data about the activity and efficacy of immune checkpoint inhibitors (ICIs) in POLE/D1pd mCRC are lacking and it is unknown whether outcomes may be different from mismatch repair-deficient (dMMR)/microsatellite instability-high (MSI-H) mCRCs treated with ICIs. PATIENTS AND METHODS: In this global study, we collected 27 patients with mCRC harboring POLE/D1 mutations leading to proofreading deficiency and treated with anti-programmed cell death-ligand 1 alone +/- anti-cytotoxic T-lymphocyte antigen-4 agents. We collected clinicopathological and genomic characteristics, response, and survival outcomes after ICIs of POLE/D1pd mCRC and compared them with a cohort of 610 dMMR/MSI-H mCRC patients treated with ICIs. Further genomic analyses were carried out in an independent cohort of 7241 CRCs to define POLE and POLD1pd molecular profiles and mutational signatures. RESULTS: POLE/D1pd was associated with younger age, male sex, fewer RAS/BRAF driver mutations, and predominance of right-sided colon cancers. Patients with POLE/D1pd mCRC showed a significantly higher overall response rate (ORR) compared to dMMR/MSI-H mCRC (89% versus 54%; P = 0.01). After a median follow-up of 24.9 months (interquartile range: 11.3-43.0 months), patients with POLE/D1pd showed a significantly superior progression-free survival (PFS) compared to dMMR/MSI-H mCRC [hazard ratio (HR) = 0.24, 95% confidence interval (CI) 0.08-0.74, P = 0.01] and superior overall survival (OS) (HR = 0.38, 95% CI 0.12-1.18, P = 0.09). In multivariable analyses including the type of DNA repair defect, POLE/D1pd was associated with significantly improved PFS (HR = 0.17, 95% CI 0.04-0.69, P = 0.013) and OS (HR = 0.24, 95% CI 0.06-0.98, P = 0.047). Molecular profiling showed that POLE/D1pd tumors have higher tumor mutational burden (TMB). Responses were observed in both subtypes and were associated with the intensity of POLE/D1pd signature. CONCLUSIONS: Patients with POLE/D1pd mCRC showed more favorable outcomes compared to dMMR/MSI-H mCRC to treatment with ICIs in terms of tumor response and survival.

Progressive plasticity during colorectal cancer metastasis.

Metastasis is the principal cause of cancer death, yet we lack an understanding of metastatic cell states, their relationship to primary tumor states, and the mechanisms by which they transition. In a cohort of biospecimen trios from same-patient normal colon, primary and metastatic colorectal cancer, we show that while primary tumors largely adopt LGR5 + intestinal stem-like states, metastases display progressive plasticity. Loss of intestinal cell states is accompanied by reprogramming into a highly conserved fetal progenitor state, followed by non-canonical differentiation into divergent squamous and neuroendocrine-like states, which is exacerbated by chemotherapy and associated with poor patient survival. Using matched patient-derived organoids, we demonstrate that metastatic cancer cells exhibit greater cell-autonomous multilineage differentiation potential in response to microenvironment cues than their intestinal lineage-restricted primary tumor counterparts. We identify PROX1 as a stabilizer of intestinal lineage in the fetal progenitor state, whose downregulation licenses non-canonical reprogramming.

Quality of life with encorafenib plus cetuximab with or without binimetinib treatment in patients with BRAF V600E-mutant metastatic colorectal cancer: patient-reported outcomes from BEACON CRC.

BACKGROUND: In the BEACON CRC study (NCT02928224), encorafenib plus cetuximab with binimetinib {9.3 versus 5.9 months; hazard ratio (HR) [95% confidence interval (CI)]: 0.60 [0.47-0.75]} or without binimetinib [9.3 versus 5.9 months; HR (95% CI): 0.61 (0.48-0.77)] significantly improved overall survival (OS) compared with the previous standard of care (control) in patients with BRAF V600E metastatic colorectal cancer (mCRC). Quality of life (QoL) was a secondary endpoint, assessed using validated instruments. PATIENTS AND METHODS: BEACON CRC was a randomized, open-label, phase III study comparing encorafenib plus cetuximab with or without binimetinib and the investigator's choice of irinotecan plus cetuximab or FOLFIRI plus cetuximab (chemotherapy control) in patients with previously treated BRAF V600E mCRC. Patient-reported QoL assessments included the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire Core 30 (EORTC) and Functional Assessment of Cancer Therapy-Colorectal (FACT-C). The primary outcome for these tools was time to definitive 10% deterioration. RESULTS: Encorafenib plus cetuximab, both with and without binimetinib, was associated with longer median times to definitive 10% deterioration versus the control group in the EORTC Global Health Status scale [HR (95% CI): 0.65 (0.52-0.80) versus 0.61 (0.49-0.75), respectively] and the FACT-C functional well-being subscale [HR (95% CI): 0.62 (0.50-0.76) versus 0.58 (0.47-0.72), respectively]. Consistent results were observed across all subscales of the EORTC and FACT-C instruments. QoL was generally maintained during treatment for the global EORTC and FACT-C scales. CONCLUSIONS: In addition to improving OS, encorafenib plus cetuximab with or without binimetinib delays QoL decline in previously treated patients with BRAF V600E-mutant mCRC.

Management of adverse events from the treatment of encorafenib plus cetuximab for patients with BRAF V600E-mutant metastatic colorectal cancer: insights from the BEACON CRC study.

Colorectal cancer is the second leading cause of cancer deaths worldwide, with a 5-year relative survival of 14% in patients with metastatic colorectal cancer (mCRC). Patients with BRAF V600E mutations, which occur in ∼10%-15% of patients with mCRC, have a poorer prognosis compared with those with wild-type BRAF tumours. The combination of the BRAF inhibitor encorafenib with the epidermal growth factor receptor inhibitor cetuximab currently represents the only chemotherapy-free targeted therapy approved in the USA and Europe for previously treated patients with BRAF V600E-mutated mCRC. As a class, BRAF inhibitors are associated with dermatologic, gastrointestinal, and renal events, as well as pyrexia and secondary skin malignancies. Adverse event (AE) profiles of specific BRAF inhibitors vary, however, and are affected by the specific agents given in combination. In patients with mCRC, commonly reported AEs of cetuximab monotherapy include infusion reactions and dermatologic toxicities. Data from the phase III BEACON CRC study indicate that the combination of encorafenib with cetuximab has a distinct safety profile. Here we review the most frequently reported AEs that occurred with this combination in BEACON CRC and best practices for managing and mitigating AEs that require more than standard supportive care.

KRAS mutation effects on the 2-[18F]FDG PET uptake of colorectal adenocarcinoma metastases in the liver.

BACKGROUND: Deriving individual tumor genomic characteristics from patient imaging analysis is desirable. We explore the predictive value of 2-[18F]FDG uptake with regard to the KRAS mutational status of colorectal adenocarcinoma liver metastases (CLM). METHODS: 2-[18F]FDG PET/CT images, surgical pathology and molecular diagnostic reports of 37 patients who underwent PET/CT-guided biopsy of CLM were reviewed under an IRB-approved retrospective research protocol. Sixty CLM in 39 interventional PET scans of the 37 patients were segmented using two different auto-segmentation tools implemented in different commercially available software packages. PET standard uptake values (SUV) were corrected for: (1) partial volume effect (PVE) using cold wall-corrected contrast recovery coefficients derived from phantom spheres with variable diameter and (2) variability of arterial tracer supply and variability of uptake time after injection until start of PET scan derived from the tumor-to-blood standard uptake ratio (SUR) approach. The correlations between the KRAS mutational status and the mean, peak and maximum SUV were investigated using Student's t test, Wilcoxon rank sum test with continuity correction, logistic regression and receiver operation characteristic (ROC) analysis. These correlation analyses were also performed for the ratios of the mean, peak and maximum tumor uptake to the mean blood activity concentration at the time of scan: SURMEAN, SURPEAK and SURMAX, respectively. RESULTS: Fifteen patients harbored KRAS missense mutations (KRAS+), while another 3 harbored KRAS gene amplification. For 31 lesions, the mutational status was derived from the PET/CT-guided biopsy. The Student's t test p values for separating KRAS mutant cases decreased after applying PVE correction to all uptake metrics of each lesion and when applying correction for uptake time variability to the SUR metrics. The observed correlations were strongest when both corrections were applied to SURMAX and when the patients harboring gene amplification were grouped with the wild type: p ≤ 0.001; ROC area under the curve = 0.77 and 0.75 for the two different segmentations, respectively, with a mean specificity of 0.69 and sensitivity of 0.85. CONCLUSION: The correlations observed after applying the described corrections show potential for assigning probabilities for the KRAS missense mutation status in CLM using 2-[18F]FDG PET images.

Changes in the multidisciplinary management of rectal cancer from 2009 to 2015 and associated improvements in short-term outcomes.

AIM: Significant recent changes in management of locally advanced rectal cancer (LARC) include preoperative staging, use of extended neoadjuvant therapies and minimally invasive surgery (MIS). This study was aimed at characterizing these changes and associated short-term outcomes. METHOD: We retrospectively analysed treatment and outcome data from patients with T3/4 or N+ LARC ≤ 15 cm from the anal verge who were evaluated at a comprehensive cancer centre in 2009-2015. RESULTS: In total, 798 patients were identified and grouped into five cohorts based on treatment year: 2009-2010, 2011, 2012, 2013 and 2014-2015. Temporal changes included increased reliance on MRI staging, from 57% in 2009-2010 to 98% in 2014-2015 (P < 0.001); increased use of total neoadjuvant therapy, from 17% to 76% (P < 0.001); and increased use of MIS, from 33% to 70% (P < 0.001). Concurrently, median hospital stay decreased (from 7 to 5 days; P < 0.001), as did the rates of Grade III-V complications (from 13% to 7%; P < 0.05), surgical site infections (from 24% to 8%; P < 0.001), anastomotic leak (from 11% to 3%; P < 0.05) and positive circumferential resection margin (from 9% to 4%; P < 0.05). TNM downstaging increased from 62% to 74% (P = 0.002). CONCLUSION: Shifts toward MRI-based staging, total neoadjuvant therapy and MIS occurred between 2009 and 2015. Over the same period, treatment responses improved, and lengths of stay and the incidence of complications decreased.

Trypanosoma cruzi: a considerable phylogenetic divergence indicates that the agent of Chagas disease is indigenous to the native fauna of the United States.

Thirty U.S. Trypanosoma cruzi stocks isolated mainly from wild mammals were characterized by multilocus enzyme electrophoresis at 22 genetic loci and random amplification of polymorphic DNA for 10 primers. Two main phylogenetic clusters, separated by large genetic distances, were discriminated by both methods, corresponding, respectively, to the formerly described zymodemes I and III. Two stocks isolated from indigenous human cases were identified as zymodeme I. Genetic diversity of the U.S. T. cruzi isolates was considerable, comparable to that scored in similarly sized samples from South America. These results favor the hypothesis that T. cruzi U.S. stocks were not imported at a historical time and are indigenous to the native fauna of the United States. The population structure of these stocks appeared to be basically clonal, as previously reported in South America, and no evidence of hybrid genotypes was found in the United States.

Immunization against Trypanosoma cruzi: adjuvant effect of glucan.

Trypanosoma cruzi, the causative agent of Chagas' disease, infects humans and animals in tropical, subtropical and some temperature regions of the western hemisphere. At present, there is no effective vaccine for T. cruzi infection. Glucan, a beta-1,3 polyglucose biological response modifier, possesses significant adjuvant activity. The present study investigated the adjuvant activity of particulate glucan when combined with a vaccine of glutaraldehyde-killed T. cruzi culture forms. ICR/HSD mice (20 g) were injected s.c. with glutaraldehyde-killed T. cruzi on days 21, 14 and 7 prior to challenge with 50 T. cruzi blood forms. Particulate glucan (1 mg/mouse) was administered s.c. either alone or in conjunction with T. cruzi vaccine. Isovolumetric dextrose served as control. Dextrose, glucan or T. cruzi vaccine as single treatment regimens showed 100% mortality with 20.5, 21.4 and 21.6 day median survival times, respectively. In contrast, glucan administered with T. cruzi vaccine showed an 85% (P less than 0.01) survival at 275 days post-challenge. In addition, the number of T. cruzi observed in the blood of glucan--T. cruzi immunized mice was lower than the appropriate controls. However, immunized mice which survived at 275 days were positive for the presence of T. cruzi by xenodiagnosis. Histopathologic evaluation of glucan--T. cruzi mice revealed no parasites or cardiac pathology, but a mild splenic hyperplasia and inflammation of skeletal muscle were noted. In subsequent studies, mice were immunized with the same regimen of glucan--T. cruzi and challenged with 500 or 5000 T. cruzi. Glucan significantly (P less than 0.05) increased survival as denoted by 60% and 50% survival in the glucan-T. cruzi group vs 0% in controls.(ABSTRACT TRUNCATED AT 250 WORDS)

The prevalence of Trypanosoma cruzi infection in armadillos collected at a site near New Orleans, Louisiana.

A study was made of the prevalence of Trypanosoma cruzi infection in armadillos at a site near New Orleans, Louisiana, where the flagellate was known to occur. Blood cultures, microscopic examination of blood, and direct agglutination tests on sera were employed in 80 armadillos. T. cruzi was isolated in culture from 23 of 80 animals; identity of the parasite was confirmed in mice inoculated with each of the isolates. Only 2 animals were positive by direct examination of blood. Serologic evidence of infection was obtained for the 23 animals that were positive by culture, and for at least 7 of those animals with negative or contaminated cultures. These results suggest that the armadillo is a reservoir of this zoonosis.

Long term cryopreservation of the amastigote stages of hemoflagellates.

Tissue homogenates containing amastigotes of either Leishmania donovani, L. tropica, or Trypanosoma cruzi were rapidly frozen with 10% glycerol as cryoprotectant. Viability and pathogenicity were maintained for at least 23 years with the Khartoum strain of L. donovani, 22 years with the Malakal strain of L. donovani, and 7 years for L. tropica and T. cruzi. Similar results over a shorter period of time were obtained with a slow-freezing technique.

Response to measles vaccine in Haitian infants 6 to 12 months old. Influence of maternal antibodies, malnutrition, and concurrent illnesses.

To study the factors affecting the serologic response to measles vaccination, we evaluated 595 Haitian infants from 6 through 12 months of age, and their mothers, at the beginning of an immunization program. Thirty-four per cent of the infants had preexisting serologic evidence of measles infections by 11 months of age. Among infants more than nine months of age, those who had had measles had a significantly lower nutritional status than those who had not (P less than 0.01). After vaccination, seroconversion rates increased from 45 per cent at 6 months to 100 per cent at 12 months. The lowest rate of vaccine failure compatible with acceptably low rates of natural infections could be achieved by vaccination after eight months of age. Infants born to mothers with low levels of antibody to measles (hemagglutination-inhibition antibody titers less than 1:40) were significantly more likely to have had natural measles (P less than 0.01) or to have seroconversion after vaccination (P less than 0.001) at 6 to 10 months of age than were infants born to mothers with higher of age than were infants born to mothers with higher titers. Malnutrition and acute infections did not affect seroconversion rates. These data support the World Health Organization recommendation to administer measles vaccine in under-developed countries as soon after nine months of age as possible, regardless of nutritional status or the presence of minor illnesses.

An experimental infection of Trypanosoma cruzi in striped skunks (Mephitis mephitis).

Four adult (3 male, 1 female) captive-raised, decented, striped skunks (Mephritis mephitis) were infected experimentaly with a field strain (Texas-Tulane) of Trypanosoma cruzi, originally isolated from a naturally-infected dog. Two skunks were injected intravenously with approximately 4.5 x 10(6) viable T. cruzi trypomastigotes. Two skunks were inoculated per os and per conjunctivum with 10 ml of phosphate buffered saline containing macerated, T. cruzi-infected triatomine intestines and intestinal contents. The skunks had minimal clinical manifestations with no mortalities occurring during 46 days post-exposure. Sera from all skunks were positive at 24 days post-inoculation (PI) by the direct and latex agglutination tests. Blood cultures from the 4 skunksd were positive for T. cruzi at day 24 PI and 3 were positive at day 46 PI. All skunks had mild to moderately severe chronic granulomatous myocarditis of the atria and ventricles. Typical T. cruzi amastigotes were present within myocardial fibers in 3 of 4 skunks.

Naturally occurring trypanosomiasis (Chagas' disease) in dogs.

Nine fatal cases of canine American trypanosomiasis (Chagas' disease) were encountered from November, 1972, through November, 1975. Of the 9 cases, 7 occurred in dogs from 5 central Texas counties, and all but one case was diagnosed during the months of September, October, or November. The source of infection was discovered in only one case--that being a doghouse heavily infested with Triatoma lectularius occulta. Each of 6 bugs collected had infective forms of the protozoan, Trypanosoma cruzi. Trypanosomiasis usually is not considered in the differential clinical diagnosis of cardiac dysfunction in man or other mammals in the United States. An antemortem diagnosis was made in only 1 of the 9 dogs, with diagnosis in the remaining dogs being made at necropsy. The predominant microscopic lesion in all dogs was necrotizing granulomatous myocarditis associated with the amastigotes of T cruzi.