The role of exosomes in cancer development has become the focus of much research, due to the many emerging roles possessed by these microscopic messengers. Exosomes are ubiquitously released in to the extracellular milieu, and have been found to regulate immune system function, particularly in tumorigenesis, as well as conditioning future metastatic sites for the attachment and growth of tumor tissue. Through an interaction with a range of host tissues, exosomes are able to generate a pro-tumor environment that is essential for carcinogenesis. Herein, we discuss the contents of exosomes and their contribution to tumorigenesis, as well as their role in chemotherapeutic resistance and the development of novel cancer treatments and the identification of cancer biomarkers.

The tissue or glandular kallikreins (KLK) are members of a highly conserved multigene family encoding serine proteases that are central to many biological processes. The rodent KLK families are large, highly conserved and clustered at one locus. The human KLK gene family is clustered on chromosome 19q13.3–13.4, and until recently consisted of just three members. However, recent studies have identified up to 11 new members of the KLK family that are less conserved than their rodent counterparts. Using a Southern blot and sequence analysis of 10 BACs and cosmids spanning approximately 400 kilobases (kb) either side of the original KLK 60-kb locus, we demonstrated that these genes also lie adjacent to this. We have also clarified the position of several microsatellite markers in relation to the extended KLK locus. Moreover, from Southern blot analysis of the cosmids and BACs with a degenerate oligonucleotide probe to the histidine-encoding region of serine proteases, we have shown that there are no other serine protease genes approximately 400 kb centromeric and 220 kb telomeric of the extended locus. We performed an extensive analysis of the expression patterns of these genes by poly(A)⁺ RNA dot blot and reverse transcriptase-polymerase chain reaction analysis, and demonstrated a diverse pattern of expression. Of interest are clusters of genes with high prostate (KLK2–4) and pancreatic (KLK6–13) expression suggesting evolutionary conservation of elements conferring tissue specificity. From these findings, it is likely that the humanKLK gene family consists of just 14 clustered genes within 300 kb and thus is of a comparable size to the rodent families (13–24 genes within 310 and 480 kb, respectively). In contrast to the rodent families, the newest members of the human KLK family are much less conserved in sequence (23–44% at the protein level) and appear to consist of at least four subfamilies. In addition, like the rat, these genes are expressed at varying levels in a diverse range of tissues although they exhibit quite distinct patterns of expression.

Purpose: Colorectal cancer patients diagnosed with stage I or II disease are not routinely offered adjuvant chemotherapy following resection of the primary tumor. However, up to 10% of stage I and 30% of stage II patients relapse within 5 years of surgery from recurrent or metastatic disease. The aim of this study was to determine if tumor-associated markers could detect disseminated malignant cells and so identify a subgroup of patients with early-stage colorectal cancer that were at risk of relapse. Experimental Design: We recruited consecutive patients undergoing curative resection for early-stage colorectal cancer. Immunobead reverse transcription-PCR of five tumor-associated markers (carcinoembryonic antigen, laminin γ2, ephrin B4, matrilysin, and cytokeratin 20) was used to detect the presence of colon tumor cells in peripheral blood and within the peritoneal cavity of colon cancer patients perioperatively. Clinicopathologic variables were tested for their effect on survival outcomes in univariate analyses using the Kaplan-Meier method. A multivariate Cox proportional hazards regression analysis was done to determine whether detection of tumor cells was an independent prognostic marker for disease relapse. Results: Overall, 41 of 125 (32.8%) early-stage patients were positive for disseminated tumor cells. Patients who were marker positive for disseminated cells in post-resection lavage samples showed a significantly poorer prognosis (hazard ratio, 6.2; 95% confidence interval, 1.9-19.6; P = 0.002), and this was independent of other risk factors. Conclusion: The markers used in this study identified a subgroup of early-stage patients at increased risk of relapse post-resection for primary colorectal cancer. This method may be considered as a new diagnostic tool to improve the staging and management of colorectal cancer.

We have used differential-display PCR (DD-PCR) to compare renal-cell carcinoma (RCC) and normal kidney gene expression with the aim of identifying genes specifically associated with RCC. Using a modified DD-PCR approach, which was non-radioactive, quicker and simpler than the conventional method, 24 cDNA samples were clearly up- or down-regulated in RCC tissue from 4 patients. Fourteen of these showed high similarity to a number of known genes. Eight of these cDNA clones were chosen for further analysis. These were a regulator of G-protein signalling (RGS-5), Notch-3, Na,K-ATPase α subunit, HLA class II antigen, ETS-like protein, transforming growth factor β–stimulated clone (TSC-22), bladder cancer–related protein (BC10) and adipophilin. Semi-quantitative RT-PCR using specific primers to each of these genes confirmed differential expression in 67% to 83% of a further 12 RCC and normal kidney paired samples from 7 of the 8 cDNA clones. Northern analysis further confirmed the up-regulation in expression of RGS-5 and Notch-3 in RCC. Further characterisation of these differentially expressed genes should lead to a better understanding of the changes that occur at the molecular level during RCC development and progression. Int. J. Cancer 88:726–732, 2000.

A gene of Colletotrichum gloeosporioides that is induced by nitrogen starvation in axenic culture and is expressed at the early stages of infection of the host Stylosanthes guianensis has been identified and its role in pathogenicity tested. The sequence of this gene, named CgDN3, indicated that it encodes a protein of 74 amino acids that contains a predicted 18 amino acid signal sequence for secretion of a basic 54 amino acid mature protein with weak homology to an internal region of plant wall-associated receptor kinases. Mutants of C. gloeosporioides were produced by homologous recombination in which part of the coding sequence and promoter region of the CgDN3 gene was replaced with a hygromycin-resistance gene cassette. Mutations in the CgDN3 gene were confirmed in two independent transformants and Northern (RNA) analysis demonstrated the disrupted CgDN3 gene was not expressed. The mutants had faster mycelial growth rates in vitro but produced spores that germinated to form appressoria normally on the leaf surface. However, the CgDN3 mutants were unable to infect and reproduce on intact host leaves. Microscopic analysis revealed small clusters of necrotic host cells at inoculation sites on leaves, suggesting that these mutants elicited a localized, host hypersensitive-like response. The mutants were able to grow necrotrophically and reproduce on leaves when conidia were inoculated directly onto wound sites. The putative promoter region of the CgDN3 gene was fused to a gene encoding a modified jellyfish green fluorescent protein and introduced into the fungus. Following inoculation, strong expression of green fluorescent protein was observed in primary infection vesicles in infected epidermal cells with weaker expression evident in hyphae growing within infected leaf tissue. These findings indicate that CgDN3 encodes a novel pathogenicity determinant associated with the biotrophic phase of primary infection and required to avert a hypersensitive-like response by a compatible host.