Identification of disease-related aberrantly spliced transcripts in myeloma and strategies to target these alterations by RNA-based therapeutics.

Novel drug discoveries have shifted the treatment paradigms of most hematological malignancies, including multiple myeloma (MM). However, this plasma cell malignancy remains incurable, and novel therapies are therefore urgently needed. Whole-genome transcriptome analyses in a large cohort of MM patients demonstrated that alterations in pre-mRNA splicing (AS) are frequent in MM. This manuscript describes approaches to identify disease-specific alterations in MM and proposes RNA-based therapeutic strategies to eradicate such alterations. As a "proof of concept", we examined the causes of aberrant HMMR (Hyaluronan-mediated motility receptor) splicing in MM. We identified clusters of single nucleotide variations (SNVs) in the HMMR transcript where the altered splicing took place. Using bioinformatics tools, we predicted SNVs and splicing factors that potentially contribute to aberrant HMMR splicing. Based on bioinformatic analyses and validation studies, we provided the rationale for RNA-based therapeutic strategies to selectively inhibit altered HMMR splicing in MM. Since splicing is a hallmark of many cancers, strategies described herein for target identification and the design of RNA-based therapeutics that inhibit gene splicing can be applied not only to other genes in MM but also more broadly to other hematological malignancies and solid tumors as well.

Prevalence of germline TP53 mutations in HER2+ breast cancer patients.

Breast cancer is the most frequent tumor in Li-Fraumeni syndrome (LFS), a rare inherited cancer syndrome associated with germline mutations in the TP53 gene. Recent data show that breast cancer in germline TP53 mutation carriers is commonly HER2+ (63-83 %). We assessed the prevalence of germline TP53 mutations in a cohort of women with HER2+ breast cancer diagnosed age ≤50 years. We identified blood specimens from 213 women with primary invasive HER2+ breast cancer age ≤50 years from a single center. Exon grouping analysis sequencing and multiplex ligation-dependent probe amplification techniques were used to screen for germline TP53 mutations. Among 213 women with HER2+ breast cancer age ≤50 years, 3 (ages at diagnosis 23, 32, 44 years) were found to carry a TP53 mutation (1.4 %, 95 % CI 0.3-4.1 %). ER/PR status was not uniform. Two TP53 carriers met Chompret criteria for LFS; none met classic LFS criteria. Although two-thirds of breast cancers in women with TP53 mutations are HER2+, we observed a low prevalence of germline TP53 mutations among unselected young women with HER2+ breast cancer. Given the potential clinical impact, consideration of germline TP53 testing should be given to young women with HER2+ breast cancer, especially if family cancer history is notable.

Attainment of complete/very good partial response following rituximab-based therapy is an important determinant to progression-free survival, and is impacted by polymorphisms in FCGR3A in Waldenstrom macroglobulinaemia.

The incorporation of rituximab into various regimens has improved depth of response in Waldenstrom macroglobulinaemia (WM), though the impact of achieving better responses remains to be determined. We examined response depth on progression-free survival (PFS) in 159 rituximab-naïve WM patients who received rituximab-based therapy. The median follow-up was 33·5 months, and categorical responses were as follows: complete response (CR, 8·8%); very good partial response (VGPR, 13·2%); partial response (50%); minor response (18·9%); Non-Responders (8·8%). Sequencing for polymorphic variants of FCGR2A, FCGR2B, and FCGR3A was performed, and impact on response depth determined. Achievement of better categorical responses was incrementally associated with improved PFS (P < 0·0001). No separation was observed between CR and VGPR, and attainment of at least a VGPR was associated with improved time-to-progression. Neither age, serum IgM, haematocrit, platelet count, serum ß(2) microglobulin, WM International Prognostic Scoring System score, and treatment group predicted for CR/VGPR. Polymorphisms at FCGR3A-48 and -158 were associated with improved categorical responses, particularly attainment of CR/VGPR (P ≤ 0·03). The attainment of CR/VGPR was associated with significantly longer PFS in rituximab-naïve WM patients undergoing rituximab-based therapy, and was predicted by polymorphisms in FCGR3A.

Gastric cancer in individuals with Li-Fraumeni syndrome.

PURPOSE: Li-Fraumeni syndrome is a rare hereditary cancer syndrome associated with germline mutations in the TP53 gene. Although sarcomas, brain tumors, leukemias, breast and adrenal cortical carcinomas are typically recognized as Li-Fraumeni syndrome-associated tumors, the occurrence of gastrointestinal neoplasms has not been fully evaluated. In this analysis, we investigated the frequency and characteristics of gastric cancer in Li-Fraumeni syndrome. METHODS: Pedigrees and medical records of 62 TP53 mutation-positive families were retrospectively reviewed from the Dana-Farber/National Cancer Institute Li-Fraumeni syndrome registry. We identified subjects with gastric cancer documented either by pathology report or death certificate and performed pathology review of the available specimens. RESULTS: Among 62 TP53 mutation-positive families, there were 429 cancer-affected individuals. Gastric cancer was the diagnosis in the lineages of 21 (4.9%) subjects from 14 families (22.6%). The mean and median ages at gastric cancer diagnosis were 43 and 36 years, respectively (range: 24-74 years), significantly younger compared with the median age at diagnosis in the general population based on Surveillance Epidemiology and End Results data (71 years). Five (8.1%) families reported two or more cases of gastric cancer, and six (9.7%) families had cases of both colorectal and gastric cancers. No association was seen between phenotype and type/location of the TP53 mutations. Pathology review of the available tumors revealed both intestinal and diffuse histologies. CONCLUSIONS: Early-onset gastric cancer seems to be a component of Li-Fraumeni syndrome, suggesting the need for early and regular endoscopic screening in individuals with germline TP53 mutations, particularly among those with a family history of gastric cancer.

Prevalence of early onset colorectal cancer in 397 patients with classic Li-Fraumeni syndrome.

BACKGROUND & AIMS: Hereditary colorectal cancer is associated most commonly with the hereditary nonpolyposis colorectal cancer or familial adenomatous polyposis syndromes. We investigated the prevalence of early onset colorectal cancer and the frequency of p53 germline mutations in 64 families from a Li-Fraumeni syndrome (LFS) registry. METHODS: Patients with documented colorectal cancer and a diagnosis at or before age 50 were included. P53 analyses were performed through germline mutational analyses using standard molecular techniques. RESULTS: Among the 397 patients and 64 families in the classic LFS registry, a total of 11 patients (2.8%) from 10 different families (15.6%) met criteria for classic LFS and had documented colorectal cancer at less than 50 years of age. The mean age at diagnosis in this group was 33 years and of these patients 4 developed colorectal cancer before age 21 (ages, 9, 11, 15, and 20 y). All families that were tested for p53 mutations (8 of 10) had evidence of germline mutations by sequence analysis; therefore, 12.5% of the total number of families in the registry had colorectal cancer at age less than 50 years and a documented germline p53 mutation. Mutations primarily were missense or nonsense and were located between exons 4-10. CONCLUSIONS: LFS patients with germline p53 mutations may have an increased susceptibility to colorectal cancer and present up to several decades earlier than the general population. LFS should be considered when a young patient presents with colorectal cancer.