Lentivirus-mediated gene therapy corrects ribosomal biogenesis and shows promise for Diamond Blackfan anemia.

This study lays the groundwork for future lentivirus-mediated gene therapy in patients with Diamond Blackfan anemia (DBA) caused by mutations in ribosomal protein S19 (RPS19), showing evidence of a new safe and effective therapy. The data show that, unlike patients with Fanconi anemia (FA), the hematopoietic stem cell (HSC) reservoir of patients with DBA was not significantly reduced, suggesting that collection of these cells should not constitute a remarkable restriction for DBA gene therapy. Subsequently, 2 clinically applicable lentiviral vectors were developed. In the former lentiviral vector, PGK.CoRPS19 LV, a codon-optimized version of RPS19 was driven by the phosphoglycerate kinase promoter (PGK) already used in different gene therapy trials, including FA gene therapy. In the latter one, EF1α.CoRPS19 LV, RPS19 expression was driven by the elongation factor alpha short promoter, EF1α(s). Preclinical experiments showed that transduction of DBA patient CD34+ cells with the PGK.CoRPS19 LV restored erythroid differentiation, and demonstrated the long-term repopulating properties of corrected DBA CD34+ cells, providing evidence of improved erythroid maturation. Concomitantly, long-term restoration of ribosomal biogenesis was verified using a potentially novel method applicable to patients' blood cells, based on ribosomal RNA methylation analyses. Finally, in vivo safety studies and proviral insertion site analyses showed that lentivirus-mediated gene therapy was nontoxic.

Early Diagnosis of Oral Cancer and Lesions in Fanconi Anemia Patients: A Prospective and Longitudinal Study Using Saliva and Plasma.

Fanconi anemia (FA) patients display an exacerbated risk of oral squamous cell carcinoma (OSCC) and oral potentially malignant lesions (OPMLs) at early ages. As patients have defects in their DNA repair mechanisms, standard-of-care treatments for OSCC such as radiotherapy and chemotherapy, give rise to severe toxicities. New methods for early diagnosis are urgently needed to allow for treatment in early disease stages and achieve better clinical outcomes. We conducted a prospective, longitudinal study wherein liquid biopsies from sixteen patients with no clinical diagnoses of OPML and/or OSCC were analyzed for the presence of mutations in cancer genes. The DNA from saliva and plasma were sequentially collected and deep-sequenced, and the clinical evaluation followed over a median time of approximately 2 years. In 9/16 FA patients, we detected mutations in cancer genes (mainly TP53) with minor allele frequencies (MAF) of down to 0.07%. Importantly, all patients that had mutations and clinical follow-up data after mutation detection (n = 6) developed oral precursor lesions or OSCC. The lead-time between mutation detection and tumor diagnosis ranged from 23 to 630 days. Strikingly, FA patients without mutations displayed a significantly lower risk of developing precursor lesions or OSCCs. Therefore, our diagnostic approach could help to stratify FA patients into risk groups, which would allow for closer surveillance for OSCCs or precursor lesions.

Development of a mouse model for spontaneous oral squamous cell carcinoma in Fanconi anemia.

Fanconi anemia (FA) patients frequently develop oral squamous cell carcinoma (OSCC). This cancer in FA patients is diagnosed within the first 3-4 decades of life, very often preceded by lesions that suffer a malignant transformation. In addition, they respond poorly to current treatments due to toxicity or multiple recurrences. Translational research on new chemopreventive agents and therapeutic strategies has been unsuccessful partly due to scarcity of disease models or failure to fully reproduce the disease. Here we report that Fanca gene knockout mice (Fanca-/-) frequently display pre-malignant lesions in the oral cavity. Moreover, when these animals were crossed with animals having conditional deletion of Trp53 gene in oral mucosa (K14cre;Trp53F2-10/F2-10), they spontaneously developed OSCC with high penetrance and a median latency of less than ten months. Tumors were well differentiated and expressed markers of squamous differentiation, such as keratins K5 and K10. In conclusion, Fanca and Trp53 genes cooperate to suppress oral cancer in mice, and Fanca-/-;K14cre;Trp53F2-10/F2-10 mice constitute the first animal model of spontaneous OSCC in FA.

Antitumor applications of polyphenol-conjugated turnip mosaic virus-derived nanoparticles.

Background: Filamentous plant virus-derived nanoparticles are biodegradable and noninfectious to humans. Their structure is also amenable to chemical modifications. They constitute an appealing material for biomedical applications including imaging and drug delivery. We had previously used turnip mosaic virus-derived nanoparticles (TuMV-NPs) to increase antibody-sensing in vivo, to prevent biofilm formation and to build biological nanoscaffolds. Materials & methods: We analyzed TuMV-NP biodistribution and tumor homing using in vivo imaging. We studied in vitro the interaction with human cancer cell lines and the antiproliferative effect of epigallocatechin gallate-functionalized TuMV-NPs. Results & conclusion: TuMV-NPs are efficiently internalized by human cells and show good tumor homing. The antiproliferative effect of epigallocatechin gallate-TuMV-NPs suggests that they could offer a potential anticancer therapy.

Cancer is the second leading cause of death worldwide, just behind cardiovascular disease. It accounts for nearly 10 million deaths annually, and new strategies to improve early detection and drug delivery are urgently needed. Nanoparticles are small structures within the nanometer range (1 billionth of a meter) that can be used to deliver either an imaging probe (tracer) to allow the detection of a tumor or drugs to kill tumor cells. There are many types of nanoparticles; those based on plant viruses are especially appealing for biomedical purposes because they are biodegradable and noninfectious to humans. Also, their physicochemical properties, such as symmetry, uniformity and loading capacity, make them excellent nanocarriers. We report here for the first time the ability of nanoparticles derived from the turnip mosaic virus (TuMV), a well-known virus naturally infecting cruciferous plants (e.g., broccoli, turnip, radish, cabbage) but not humans, to deliver a fluorescent imaging probe that allows tumor detection in vivo. Moreover, TuMV nanoparticles were used to deliver a natural chemotherapeutic agent of plant origin to different types of tumor cells (lung, colorectal, breast, and head and neck), showing increased antiproliferative capacity compared to the nonvehiculized drug.

Generating New FANCA-Deficient HNSCC Cell Lines by Genomic Editing Recapitulates the Cellular Phenotypes of Fanconi Anemia.

Fanconi anemia (FA) patients have an exacerbated risk of head and neck squamous cell carcinoma (HNSCC). Treatment is challenging as FA patients display enhanced toxicity to standard treatments, including radio/chemotherapy. Therefore, better therapies as well as new disease models are urgently needed. We have used CRISPR/Cas9 editing tools in order to interrupt the human FANCA gene by the generation of insertions/deletions (indels) in exon 4 in two cancer cell lines from sporadic HNSCC having no mutation in FA-genes: CAL27 and CAL33 cells. Our approach allowed efficient editing, subsequent purification of single-cell clones, and Sanger sequencing validation at the edited locus. Clones having frameshift indels in homozygosis did not express FANCA protein and were selected for further analysis. When compared with parental CAL27 and CAL33, FANCA-mutant cell clones displayed a FA-phenotype as they (i) are highly sensitive to DNA interstrand crosslink (ICL) agents such as mitomycin C (MMC) or cisplatin, (ii) do not monoubiquitinate FANCD2 upon MMC treatment and therefore (iii) do not form FANCD2 nuclear foci, and (iv) they display increased chromosome fragility and G2 arrest after diepoxybutane (DEB) treatment. These FANCA-mutant clones display similar growth rates as their parental cells. Interestingly, mutant cells acquire phenotypes associated with more aggressive disease, such as increased migration in wound healing assays. Therefore, CAL27 and CAL33 cells with FANCA mutations are phenocopies of FA-HNSCC cells.

Registro regional de tumores para el sur de Sonora / Regional record of tumors for the south of Sonora

En el año de 1992 se llevó a cabo un registro regional de tumores malignos para el sur de Sonora, abarcando una población de 1'023,879 habitantes y una superficie de 43,955 km². Se hizo una pesquisa para cada caso nuevo de cáncer informado por centros hospitalarios, consulta externa y laboratorios de anatomía patológica, así como certificados de defunción. Se informan las cifras de cada grupo de tumores por sitio de origen, sexos y decenios de la vida en que se presentaron. Hubo un total de 1063 casos (104 por cada 100,000 habitantes por año); de éstos, 141 correspondieron a fallecimientos para una tasa de mortalidad de 10 por cada 100,000 habitantes por año. Se registraron 479 casos (45 por ciento) en varones y 584 (55 por ciento) en mujeres. La maxíma frecuencia general ocurrió a partir de los 40 años de edad. El tumor más frecuente fue el del cuello uterino con 224 casos (21 por ciento); siguieron los de bronquios y pulmón, piel (no melanoma), glándula mamaria y próstata con 112 (10 por ciento), 111 (10 por ciento), 100 (9 por ciento) y 96 (9 por ciento) casos, respectivamente. El grupo de leucemias y linfomas ocupó el sexto lugar con 54 casos 85 por ciento), también fue el más frecuente en menores de 20 años de edad

Carcinoma cervicouterino localmente avanzado: resultados preliminares con quimioterapia neoadyuvante con fluorouracilo, leucovorin / Locally advanced cervix carcinoma: response at the treatment chemotherapy

Veinte pacientes con carcinoma cervicouterino localmente avanzado fueron tratadas con tres ciclos de quimioterapia neoadyuvante (QT) con fluorouracilo modulado con leucovorin, seguidas de radioterapia convencional (RT). Se evaluó respuesta en 19 de ellas. Hubo buena tolerancia a la QT sola, aunque se observó náuseas y vómitos (47 por ciento), diarrea (21 por ciento) (en todos los casos en grado 1) y toxicidad hematológica en tres pacientes (una con leucopenia y granulocitopenia grado 2, y trombocitopenia grado 1; y las otras dos con leucopenia grado 1). La tolerancia y morbilidad a la RT no se modificaron por el uso previo de la QT. Los resultados a la QT sola fueron: respuestas parcial en cinco casos (26.3 por ciento) y enfermedad estable en ocho (42.1 por ciento). Los resultados a la combinación QT/RT después de 12.5 meses de seguimiento son: respuesta completa (RC) en 14 (73.6 por ciento), otra con RC pero recurrencia pélvica a los nueve meses; lo cual se compara favorablemente con un estudio anterior de pacientes tratados con RT sola con RC de 53 por ciento (p = 0.33, considerada significativa). Estos resultados son alentadores para seguir intentando el uso de la quimioterapia neoadyuvante (o concurrente) en el manejo del cáncer cervicouterino localmenta avanzado en estudios aleatorios, con mayor número de pacientes y más tiempo de seguimiento.

Carcinoma cervicouterino, resultados con radioterapia / Carcinoma of uterine cervix, radiotherapy results

De 1979 a 1981, fueron antendidos 403 casos de carcinoma cervicouterino en el Hospital General , Z-1, IMSS en Cd. Obregón, Sonora. De ellos, 385 correspondieron a carcinoma epidermoide; 14 a adenocarcinoma; 3 a carcinoma adenoescamoso y uno sin biopsia; dos no aceptaron tratamiento. Los 401 casos fueron tratados con Cesio-137 intracavitario con dosificación lenta y radioterapia externa con Cobalto-60. De acuerdo al estado (E), 121 estuvieron en FIGO IB; 117 (30% ) en E-IIB; 78 (19% ) en E-IIA; 54 (13% ) en E-III; 23 en estadiíos cero y IA fue de 100% ; en E-IB, 80.9% ; en E-IIA, 51.2% ; en E-IIB, 35.8% ; en E-III, 27.7% ; en estadio IV, 17.3% en no clasificados, 33.3% ; y 50.% / en el total de casos. La morbilidad fue de ligera a moderada. De los 198 fracasos terapéuticos, el 65% se debió a enfermedad pélvica; el 13% a abandono del tratamiento; el 8% a pérdida del seguimiento; el 6% a metástasis extrapélvicas; el 4% a recurrencia pélvica y metástasis a distancia; y el 4% a fallecimiento por enfermedad intercurrente. El 80% de las recaídas se presentaron alrededor de los 2 años después del inicio de la radioterapia.

Registro regional de tumores malignos para el sur de Sonora / Malignant tumors regional register for south Sonora

Se inició un registro regional de tumores malignos, para el sur de Sonora, México, con una población de 750 000 habitantes. Se hizo una pesquisa de cada caso nuevo de cáncer informado por centros hospitalarios, consulta externa y laboratorios de anatomía patológica, así como en certificados de defunción, durante el periodo de 1982 a 1983 (del 15 de marzo al 14 de marzo siguiente). Se informan cifras de cada grupo de tumores por sitio de origen, sexo y decenios de la vida en que se presentan. Se encontraron en total 1 123 casos, o sea una frecuencia de 1 497.3 por obs: 1 000 000 de habitantes por año, y entre éstos se encontraron 570 defunciones, que constituyen una tasa de mortalidad de 760 por obs: 1 000 000 por año. Fueron 490 los casos (43.6%) en varones y 633 (56.4%) en mujeres. La máxima frecuencia ocurrió a partir de los 40 años. El tumor más frecuente fue el del cuello uterino, 246 casos (21.9%), y lo siguieron los de bronquios y pulmón, glándula mamaria, los linfomas, las leucemias y de piel con 130 (11.6%), 92 (8.2%), 90 (8%) y 86 (7.6%) casos, respectivamente