Autologous immune enhancement therapy in a case of gall bladder cancer stage IV after surgical resection and chemotherapy yielding a stable non-progressive disease.

Advanced gall bladder cancer generally has a poor prognosis and also shows decreased response to conventional therapies like chemotherapy and radiotherapy. Though surgical resection is the most common approach followed, the 1-year survival rate is only 10%. Herein, we report the outcome of administration of autologous natural killer cell and activated T lymphocyte-based autologous immune enhancement therapy (AIET) in a case of gall bladder cancer stage IV which was progressing in spite of surgical resection and several sittings of chemotherapy. There were no adverse reactions after AIET. After three infusions of AIET, an improvement of the quality of life and general condition which is sustaining for more than 6 months and a substantial decrease in the CA 19-9 marker levels from 2938.22 U/mL before AIET to 511 U/mL, 5 months after AIET, in our experience make us recommend AIET along with other conventional treatments in similar cases.

Examining the Gm18 and m(1)G Modification Positions in tRNA Sequences.

The tRNA structure contains conserved modifications that are responsible for its stability and are involved in the initiation and accuracy of the translation process. tRNA modification enzymes are prevalent in bacteria, archaea, and eukaryotes. tRNA Gm18 methyltransferase (TrmH) and tRNA m(1)G37 methyltransferase (TrmD) are prevalent and essential enzymes in bacterial populations. TrmH involves itself in methylation process at the 2'-OH group of ribose at the 18th position of guanosine (G) in tRNAs. TrmD methylates the G residue next to the anticodon in selected tRNA subsets. Initially, m(1)G37 modification was reported to take place on three conserved tRNA subsets (tRNA(Arg), tRNA(Leu), tRNA(Pro)); later on, few archaea and eukaryotes organisms revealed that other tRNAs also have the m(1)G37 modification. The present study reveals Gm18, m(1)G37 modification, and positions of m(1)G that take place next to the anticodon in tRNA sequences. We selected extremophile organisms and attempted to retrieve the m(1)G and Gm18 modification bases in tRNA sequences. Results showed that the Gm18 modification G residue occurs in all tRNA subsets except three tRNAs (tRNA(Met), tRNA(Pro), tRNA(Val)). Whereas the m(1)G37 modification base G is formed only on tRNA(Arg), tRNA(Leu), tRNA(Pro), and tRNA(His), the rest of the tRNAs contain adenine (A) next to the anticodon. Thus, we hypothesize that Gm18 modification and m(1)G modification occur irrespective of a G residue in tRNAs.

Successful transportation of human corneal endothelial tissues without cool preservation in varying Indian tropical climatic conditions and in vitro cell expansion using a novel polymer.

BACKGROUND: Though the transplantation of human corneal endothelial tissue (CET) separated from cadaver cornea is in practice, its transportation has not been reported. We report the successful transportation of CET in varying Indian climatic conditions without cool preservation and the in vitro expansion of Human Corneal Endothelial Precursor Cells (HCEPCs) using a novel Thermo-reversible gelation polymer (TGP). MATERIALS AND METHODS: CET from cadaver corneas (n = 67), unsuitable for transplantation, were used. In phase I, CET was transported in Basal Culture Medium (Group I) and TGP (Group II) and in Phase II, in TGP cocktail alone, from three hospitals 250-2500 km away, to a central laboratory. The transportation time ranged from 6 h to 72 h and the outdoor temperature between 20°C and 41°C. On arrival, CET were processed, cells were expanded upto 30 days in basal culture medium (Group A) and TGP scaffold (Group B). Cell viability and morphology were documented and Reverse transcription polymerase chain reaction (RT-PCR) characterization undertaken. RESULTS: In Phase I, TGP yielded more viable cells (0.11 × 10(6) cells) than Group I (0.04 × 10(6) cells). In Phase II, the average cell count was 5.44 × 10(4) cells. During expansion, viability of HCEPCs spheres in TGP was maintained for a longer duration. The cells from both the groups tested positive for B-3 tubulin and negative for cytokeratins K3 and K12, thereby proving them to be HCEPCs. CONCLUSION: TGP preserves the CET during transportation without cool preservation and supports in vitro expansion, with a higher yield of HCEPCs, similar to that reported in clinical studies.

Smallpox still haunts scientists: results of a questionnaire-based inquiry on the views of health care and life science experts and students on preserving the remaining variola virus stocks.

The World Health Organization (WHO) declared eradication of the dreadful disease "smallpox" in 1980. Though the disease has died down, the causative virus "variola" has not, as it has been well preserved in two high security laboratories-one in USA and another in Russia. The debate on whether the remaining stocks of the smallpox virus should be destroyed or not is ongoing, and the World Health Assembly (WHA) in 2011 has decided to postpone the review on this debate to the 67th WHA in 2014. A short questionnaire-based inquiry was organized during a one-day stem cell meeting to explore the views of various health care and life science specialists especially students on this aspect. Among the 200 participants of the meeting, only 66 had answered the questionnaire. 60.6% of participants who responded to the questionnaire were for preserving the virus for future reference, while 36.4% of the participants were for destroying the virus considering the magnitude with which it killed millions. However, 3% of the respondents were not able to decide on any verdict. Therefore, this inquiry expresses the view that "what we cannot create, we do not have the right to destroy."

Exploring GpG bases next to anticodon in tRNA subsets.

Transfer RNA (tRNA) structure, modifications and functions are evolutionary and established in bacteria, archaea and eukaryotes. Typically the tRNA modifications are indispensable for its stability and are required for decoding the mRNA into amino acids for protein synthesis. A conserved methylation has been located on the anticodon loop specifically at the 37(th) position and it is next to the anticodon bases. This modification is called as m1G37 and it is catalyzed by tRNA (m(1)G37) methyltransferase (TrmD). It is deciphered that G37 positions occur on few additional amino acids specific tRNA subsets in bacteria. Furthermore, Archaea and Eukaryotes have more number of tRNA subsets which contains G37 position next to the anticodon and the G residue are located at different positions such as G36, G37, G38, 39, and G40. In eight bacterial species, G (guanosine) residues are presents at the 37(th) and 38(th) position except three tRNA subsets having G residues at 36(th) and 39(th) positions. Therefore we propose that m1G37 modification may be feasible at 36(th), 37(th), 38(th), 39(th) and 40(th) positions next to the anticodon of tRNAs. Collectively, methylation at G residues close to the anticodon may be possible at different positions and without restriction of anticodon 3(rd) base A, C, U or G.

Cell Based Autologous Immune Enhancement Therapy (AIET) after Radiotherapy in a Locally Advanced Carcinoma of the Cervix.

Radiotherapy is the primary form of treatment in patients with locally advanced cervical carcinoma. However for residual disease in the form of the persistent lymph nodes, surgery or chemotherapy is recommended. As surgery is not acceptable by every patient and chemotherapy has associated side effects, we hereby report the positive outcome of in vitro expanded natural killer cell and activated T lymphocyte based autologous immune enhancement therapy (AIET) for the residual lymphadenopathy in a patient with locally advanced cervical cancer after radiation. After six transfusions of AIET, there was complete resolution of residual lymph nodes and there was no evidence of local lesion. The patient also reported improvement in quality of life. As AIET has been reported as the least toxic among the available therapies for cancer, combining AIET with conventional forms of therapy in similar patients might not only improve the outcome but may also help the patients achieve a good quality of life.

Age-old wisdom concerning cell-based therapies with added knowledge in the stem cell era: our perspectives.

Among the various strategies providing a cure for illness, cell-based therapies have caught the attention of the world with the advent of the "stem cell" era. Our inherent understanding indicates that stem cells have been in existence since the birth of multicellular organisms. However, the formal discovery of stem cells in the last century, followed by their intricate and extensive analysis, has led to clinical and translational efforts with the aim of using them in the treatment of conditions which don't have a definitive therapeutic strategy, has fueled our interest and expectations. Technological advances in our ability to study their cellular components in depth, along with surface markers and other finer constituents, that were unknown until last century, have improved our understanding, leading to several novel applications. This has created a need to establish guidelines, and in that process, there are expressed understandings and views which describe cell therapy along lines similar to that of biologic products, drugs, and devices. However, the age-old wisdom of using cells as tools for curing illness should not be misled by recent knowledge, to make cell therapy using highly complex stem cells equal to factory-synthesized and reproducible chemical compounds, drugs, or devices. This article analyses the differences between these two entities from various perspectives.

In vitro culture and characterization of enteric neural precursor cells from human gut biopsy specimens using polymer scaffold.

In vitro expansion and characterization of neural precursor cells from human gut biopsy specimens with or without Hirschsprung's disease using a novel thermoreversible gelation polymer (TGP) is reported aiming at a possible future treatment. Gut biopsy samples were obtained from five patients undergoing gut resection for Hirschsprung's disease (n = 1) or gastrointestinal disorders (n = 4). Cells isolated from the smooth muscle layer and the myenteric plexus were cultured in two groups for 18 to 28 days; Group I: conventional culture as earlier reported and Group II: using TGP scaffold. Neurosphere like bodies (NLBs) were observed in the cultures between 8th to 12th day and H & E staining was positive for neural cells in both groups including aganglionic gut portion from the Hirschsprung's disease patient. Immunohistochemistry using S-100 and neuron specific enolase (NSE) was positive in both groups but the TGP group (Group II) showed more number of cells with intense cytoplasmic granular positivity for both NSE and S-100 compared to Group I. TGP supports the in vitro expansion of human gut derived neuronal cells with seemingly better quality NLBs. Animal Studies can be tried to validate their functional outcome by transplanting the NLBs with TGP scaffolds to see whether this can enhance the outcome of cell based therapies for Hirschsprung's disease.

Autologous immune enhancement therapy in recurrent ovarian cancer with metastases: a case report.

Current therapeutic modalities for ovarian cancer such as chemotherapy, radiotherapy and surgery have been reported to yield only marginal success in improving survival rates of patients and have associated adverse effects. We report here a case of recurrent stage IV ovarian cancer, treated with cell-based autologous immune enhancement therapy (AIET) along with chemotherapy and followed up for 18 months. A 54-year-old female was diagnosed with a recurrence of ovarian carcinoma 1 year after initial surgical removal followed by chemotherapy for stage IIIC ovarian carcinoma. When diagnosed in 2010 with recurrence, she had liver and spleen metastases with a CA-125 level of 243 U/ml and a stage IV clinical status. Six infusions of AIET using autologous in vitro expanded and activated natural killer (NK) cells (CD3-CD56+) and activated T lymphocytes (CD3+CD56+) were administered in combination with 6 cycles of chemotherapy with carboplatin and doxorubicin. Following this treatment, CA-125 decreased to 4.7 U/ml along with regression of the metastatic lesions and an improved quality of life. No adverse reactions were reported after the AIET transfusions. Eighteen months of follow-up revealed a static nonprogressive disease. Combining AIET with chemotherapy and other conventional treatments has been found to be effective in our experience, as reported earlier, even in patients with advanced ovarian cancer, and we recommend this strategy be considered in treating similar cases.

A comparative analysis of in vitro expansion of natural killer cells of a patient with autoimmune haemolytic anaemia and ovarian cancer with patients with other solid tumours.

The functional profile of natural killer (NK) cells has been reported to be lower in auto-immune haemolytic anaemia (AIHA). In this study, we report a comparative analysis of peripheral blood mononuclear cells (PBMNCs) and the in vitro expansion of NK cells in a patient with AIHA and cancer, with that of other cancer patients without AIHA. PBMNCs and in vitro NK-cell expansion of a 64-year old female patient with ovarian cancer and AIHA was compared with that of four other patients with cancer without AIHA who underwent autologous immune enhancement therapy (AIET). The NK cells were cultured using autologous plasma without feeder layers. The quantities of PBMNCs, NK cells and CD3-CD56+ cells were compared. The average quantity of PBMNCs per ml in Cases I to V were 10.71, 39.2, 49.26, 65.16 and 49.33×10(4), respectively, and the average maximum count of NK cells was 3.9, 1730.03, 1824.16, 1058.61 and 761×10(6), respectively. The average percentage of CD3-CD56+ cells in Cases I to V following in vitro expansion was 1.2, 65.7, 28.63, 65.9 and 40%, respectively. In the present study, probably the first in the literature, the in vitro expansion of NK cells was found to be significantly lower in the AIHA patient. Previously, only a lower NK-cell functional profile was reported. Further studies are required to establish the association between AIHA and NK-cell profile and in vitro expansion, and to find common antibodies between red blood cells (RBCs) and NK cells.