Clinical Outcomes of Autologous Hematopoietic Stem Cell Transplant in Multiple Myeloma Patients: A 5-year Experience from a Single Centre in North India.

Swati PabbiIntroduction Multiple myeloma (MM) forms a significant proportion of hematological malignancies. Autologous transplantation continues to be an effective consolidation strategy in resource-restricted settings such as India. Objectives The main objective of the study was to analyze the clinical outcomes of autologous hematopoietic stem cell transplant (HSCT) in MM patients in a single tertiary care center in north India over a period of 5 years. Materials and Methods This retrospective observational study was conducted in a tertiary care center in north India. Data of all MM patients who underwent HSCT between January 2014, and December 2018, were analyzed. The outcome of HSCT was investigated in terms of transplant-related mortality (TRM), progression-free survival (PFS), overall survival (OS), and relapse. PFS and OS were calculated by Kaplan-Meier method and differences between the groups were tested for statistical significance using the two-tailed log-rank test. Life-table method was used for the estimation of survival rate at 1, 3, 5, and 6 years. Results Patient characteristics and survival post-transplant was similar to other published Indian studies. In total, 378 patients were diagnosed with MM in our hospital between 2014 and 2018. One hundred ninety-three patients were found to be eligible for autologous HSCT, out of which 52 ended up having a transplant giving us a high percentage (26.9%) of patients receiving a transplant in our setting. Transplant-related mortality (TRM) was nil in the present study. The mean PFS and OS were 62.8 and 70.1 months, respectively. The mean PFS and OS rates at 5 years were 75.3% and 84.2%, respectively. The average cost estimate of HSCT in our setting was 7.2 lakh Indian national rupees. Conclusion Autologous HSCT is a safe procedure with nil 100-day mortality in present series. Moreover, considering the cost of novel agents, autologous transplant remains a cost-effective way for prolonging remission and time-to-next treatment in India.

Real-world data on renal transplantations from a tertiary-care hospital in North India, in context of Indian regulatory act-transplantation of human organs and tissues act (THOTA): A retrospective analysis.

INTRODUCTION: Renal transplantation is the treatment of choice for patients suffering from end stage renal disease (ESRD). Indian regulations defined under Transplantation of Human Organs and Tissues Act (THOTA), 2014 restricts organ donations to near-related living donors to curb any malpractices like 'paid donors' in living-donor kidney transplantation (LDKT). The aim of our study was to look at real-world data of donor-recipient pairs and to identify relationship of donors (with their respective patients) and the common (or uncommon) DNA profiling methods used for supporting "claimed relationship" in accordance with the regulations. MATERIAL AND METHODS: The donors were categorized and grouped into near-related donor, donors other than near-related donors, swap donors and deceased donors. Claimed relationship was confirmed, commonly by HLA typing, using SSOP method. In few cases, which were uncommon (and infrequent), autosomal DNA analysis, mitochondrial DNA analysis and Y-STR DNA analysis were performed to support the claimed relationship. Data collected included age, gender, relationship, DNA profiling test method. RESULTS: Among the 514 donor-recipient pairs evaluated, numbers of female donors out-numbered male donors. The decreasing order of relationships in near-related donor group were wife>mother>father>sister>son>brother>husband> daughter>grandmother. 11.9% of donors were in the category of donors other than near-related donors. In 97.86% cases, the claimed relationship was supported by HLA typing and in just 2.1% cases autosomal DNA analysis>mitochondrial DNA analysis> Y-STR DNA analysis, in this order, were performed to establish relationship. CONCLUSION: This study brought out gender disparity with women out-numbering men as donors. Among recipients, access to renal transplant was largely restricted to men. As far as relationship of donors to recipients was concerned, mostly near-related family members, like wife, were donors and claimed relationship was almost always (99%) was corroborated by HLA typing.

Implementation of an international barcode labeling standard, International Society of Blood Transfusion 128, and its integration with local regulations at a blood center in India: Step-by-step journey.

The International Society of Blood Transfusion (ISBT) 128 is an internationally endorsed, electronically readable labeling standard that provides a convenient and accurate means of identification, traceability, publication, and storage of information for blood and blood products. The authors' center recently registered with the International Council for Commonality in Blood Banking Automation (ICCBBA) and progressed to ISBT 128 labeling standard. This manuscript was written with the objective of sharing the authors' experience with respect to the implementation of ISBT 128 standards for whole blood donations and integration of ISBT 128 standards with Indian licensing regulations. The authors explore the process of implementation of ISBT 128 standards through a step-by-step journey that included facility registration with International Council for Commonality in Blood Banking Automation (ICCBBA), allotment of facility identification number, development of four-quadrant label for blood components, and integration of local regulatory requirements in the final "composite" label. Acknowledging the lack of any published report from India on ISBT 128 standards implementation, the authors wish to attempt help their peers in understanding and implementation of this global standard at their respective facilities.

Analytical and clinical performance evaluation of enhanced chemiluminescence-based fourth-generation HIV combo assay: Report from tertiary health-care setup in North India.

INTRODUCTION: HIV fourth-generation assay, designed for the detection of HIV p24 antigen along with anti-HIV antibodies of both immunoglobulin M and immunoglobulin G type against HIV 1 and HIV 2 viral antigens, have helped in the early detection of HIV infection and supports in minimizing the transmission risk in the acute phase of infection. The objective of this study was to evaluate the analytical and clinical performance of HIV fourth-generation assay based on enhanced chemiluminescence technology. MATERIALS AND METHODS: The analytical performance of the assay was evaluated in terms of accuracy, precision, limit of detection, type of sample (serum vs. plasma), cross-reactivity (with other transfusion transmissible infections markers), and interference (with endogenous substances). Proficiency control material included kit-controls, archived known positive donor samples, third-party controls, and World Health Organization (WHO)/National Institute for Biological Standards and Controls (NIBSC, MHRA, UK) controls. The clinical performance was evaluated using routine donor and patient samples received during the study period. RESULTS: HIV fourth-generation assay showed reliable and reproducible results measured in terms of coefficient of variation % with kit-controls, archived known positive donor samples, third-party controls, and WHO international standards for anti-HIV 1 and 2 antibodies, HIV1 p24 antigens and HIV2 p26 antigen controls. The analytical sensitivity of the HIV fourth-generation assay was found to be 0.1 IU/mL of HIV1 p24 antigen control and there was no cross-reactivity or interference observed. In the clinical performance of the assay, HIV fourth-generation assay showed reliable performance in both donor and patient samples. CONCLUSION: HIV fourth-generation assay meets the requirements for its use as a screening assay for HIV infection based on the analytical and clinical performance of the assay.

Reference interval of platelet counts and other platelet indices in apparently healthy blood donors in North India according to Clinical and Laboratory Standards Institute guidelines: Need to redefine the platelet count cutoffs for repeat plateletpheresis donation?

BACKGROUND: In clinical practice, laboratory results are of great importance for the diagnosis and treatment. Reference intervals of different parameters aid health-care professionals in the interpretation of results. There are very few studies on reference intervals from India. This prospective study was conducted to determine the reference intervals for platelet count (PLT) and PLT indices; mean PLT volume (MPV), PLT distribution width (PDW), and PLT large cell ratio (P-LCR). These values can be obtained as a part of a routine complete blood count (CBC) and have diagnostic and prognostic significance in certain diseases. PLT count is an important criterion for the selection of donors for repeat plateletpheresis donation. MATERIALS AND METHODS: Sixteen hundred and thirty-four first-time healthy volunteer plateletpheresis donors were enrolled for the study. CBC was done, values of PLT, MPV, PDW, and P-LCR were noted, and the results were analyzed. The 95% of the reference distribution was estimated using the 2.5th and 97.5th percentiles following Clinical and Laboratory Standards Institute guidelines. Adverse donor reactions, if any and quality parameters of single donor PLTs (SDP) were also studied. RESULTS: Reference range values of PLT, MPV, PDW, and P-LCR were 137,825-355,175/µl, 8.1-13.9/fl, 9.1-22.5/fl, and 11.7%-52.9%, respectively, and compared well with other published studies from India. It was observed that reference values of PLT count obtained in the study were lower than reference values that are currently used in most laboratories (150,000-450,000/µl) in India. CONCLUSION: Based on our results, we are of the opinion that the PLT count cutoffs for repeat plateletpheresis donation may need to be revised downwards for our country which would also mitigate the scarcity of apheresis donors.

Comparison of in-vitro and in-vivo parameters of whole blood derived random donor platelets (RDP) after over-night hold with RDP prepared after 2-h hold: Single centre report from India.

BACKGROUND: Random Donor Platelet (RDP) derived from whole blood is the major source of platelets in India. At our centre, we prepare RDPs by buffy coat method after a holding period of 2-hours (THRDP) as per current regulatory guidelines. Overnight hold of buffy coats before RDP preparation (OHRDP) would logistically optimise the manpower usage at our centre. The aim of this study was to compare both in-vitro as well as in-vivo parameters of OHRDPs with THRDPs. METHODOLOGY: Hematological (Platelet, leucocyte counts), physical (pH and Swirling) and biochemical parameters (pO2, pCO2, lactate, bicarbonate and glucose) as well as platelet activation markers were tested in THRDPs and OHRDPs each at Day-1 and Day-5 as in-vitro studies. Separately, in-vivo study was done where Corrected count increment (CCI) and percentage platelet recovery (PPR) were considered. All parameters were expressed as Mean ± Standard deviation and were analysed using paired t-test with level of significance, p < 0.05. RESULTS: OHRDPs had higher platelet counts and lower leucocytes and CD62 P expression than THRDPs. All other markers were well within the quality control range in both groups. No significant differences were seen in the two groups when comparing CCI and PPR. CONCLUSION: OHRDPs were found to be as good or better as compared to the THRDPs in the in-vitro part of our study. Additionally, there were no significant differences between the two groups when they were compared in vivo. This makes us conclude that overnight hold of buffy coats may be implemented at our center.

Evaluation of quality matrix when practice changed from triple bags to quadruple (top and bottom) bags: In vitro analysis of blood components!

BACKGROUND: As a part of continuous quality initiatives, while moving from triple bags to quadruple bags, we undertook a study to compare platelet-rich plasma (PRP) and buffy-coat (BC) methods with respect to all blood components (red blood cells [RBCs], random donor platelet concentrate [RDPC], and fresh frozen plasma [FFP]) prepared by PRP and BC methods. MATERIALS AND METHODS: It was a prospective analysis of different physical and quality parameters of RDPC, RBC, and FFP prepared out of 100 whole blood (WB) donations. Of these, 50 WB units were processed by PRP method using Triple bags and 50 WB units by BC method, using quadruple (top and bottom) bags, with an attached integral filter. RESULTS: RBC prepared by BC method had higher hematocrit (61.3 ± 1.91% vs. 56.03 ± 3.37%; P < 0.05) and lower white blood cell (WBC) contamination (6.3 × 104 ± 6.1 vs. 5.41 × 105 ± 2.5; P < 0.05) in comparison to prepared by PRP method. Higher PLT yield (7.67 × 1010 ± 1.8 vs. 6.47 × 1010 ± 1.5; P < 0.05) and lower WBC count (8.24 × 103 ± 1.1 vs. 1.5 × 104 ± 2.1; P < 0.05) was observed in RDPC prepared by BC method than PRP derived. CD62P expression was lower in RDPC prepared by BC method (31.46 ± 9.7%; P < 0.05) as compared to PRP method (43.35 ± 12.5%; P < 0.05). The BC method also resulted in increased plasma yield (210.56 ± 18.54 ml vs. 187.92 ± 12.93 ml; P < 0.05) in FFP in comparison to PRP method. CONCLUSION: The blood components produced from WB by the BC method have laboratory variables suggestive of superior quality than those produced by the PRP method.

Peripheral blood stem cell mobilization with pegylated granulocyte colony stimulating factor in children.

BACKGROUND AND AIM: We report here our experience of using pegylated granulocyte colony stimulating factor (peg-GCSF) for peripheral blood stem cell (PBSC) mobilization in children. METHODS AND RESULTS: A total of nine children suffering from high-risk/relapsed solid tumors were mobilized with chemotherapy and peg-GCSF (100 microgram/kg single dose). Mean age was 7.7 years (range 2-15 years).The mean time from peg-GCSF administration to PBSC harvest was 9.7 days. Adequate stem cells (median dose 26.9 million/kg) could be harvested in all children by a single apheresis procedure. No major adverse events observed. CONCLUSION: It is feasible and safe to mobilize PBSC with peg-GCSF in children with cancer.

Red blood cell exchange in sickle cell disease patient with multiple alloantibodies.

There are several reports in medical literature about Red Cell Exchange (RCE) being routinely performed pre-operatively in sickle cell disease patients to provide immediate decrease in HbS concentration and prevent post-operative complications. We would like to present one such case of SCD who also had multiple allo-antibodies and had to undergo hemi-arthroplasty for avascular necrosis of head femur. Grouping and antibody screening was performed using column agglutination technique. 3-cell and 11- cell panel were used for antibody screening and identification, respectively. Automated RBC exchange was performed on apheresis machine Com. Tec using the standard PL1 kit (Fresenius Kabi, Germany). Multiple (anti-c, E) allo-antibodies were identified and successful pre-operative RCE was done with corresponding antigen-negative AHG compatible RBC units. Single RCE procedure reduced HbS concentration from 65% to 25%. The patient underwent uneventful hemi-arthroplasty and was discharged on post-operative day-7. Patient is on regular follow-up and continues to do well two months after the day of surgery. This is possibly the first case report from India, which illustrates successful automated RCE in a SCD patient with alloimmunization.

Head-to-head comparison of Enzyme Linked Immunosorbent Assay (ELISA) and Enhanced Chemiluminescence Immunoassay (ECLIA) for the detection of Transfusion Transmitted Disease (TTD) Markers; HIV, HCV and HBV in blood donors, in India.

Safe blood transfusion being the cornerstone of any Blood Transfusion Services requires meticulous testing for Transfusion Transmitted Disease (TTD) markers in donated blood. We performed head-to-head comparison of ELISA and ECLIA for detection of TTD markers for Human Immunodeficiency Virus (HIV), Hepatitis C Virus (HCV) and Hepatitis B Virus (HBV) in 10,164 Indian blood donors. All concordant reactive, discordant reactive and concordant non-reactive samples were re-confirmed using Individual Donor-Nucleic Acid Amplification Test (ID-NAT) as the 'gold standard' test. 223 samples were found reactive; out of which 93 (four HIV, 34 HCV and 55 HBV) samples were concordant reactive and tested positive by both methods while 130 discordant reactive samples were reactive either by ELISA or ECLIA. Out of 130 discordant reactive samples ELISA-reactive and ECLIA-non-reactive samples for HIV, HCV and HBV were 15, eight, and 29 respectively while ECLIA-reactive ELISA-non-reactive samples for HIV, HCV and HBV were 39, 36 and three respectively. Sensitivity of ECLIA and ELISA was 100 % for all three TTD markers, while specificity was 99.62 % and 99.85 % for HIV; 99.64 % and 99.84 % for HCV and 99.97 % and 99.70 % for HBV respectively. Strength of agreement and Kappa Statistics for ECLIA and ELISA compared to the gold standard test was poor and fair for HIV (k = 0.169 and 0.347), moderate and good for HCV (k = 0.539 and 0.763), and very good and good for HBV (k = 0.973 and 0.783). According to this study, it can be concluded both the testing techniques; ELISA and ECLIA have 100 % sensitivity for the detection for HBV, HCV and HIV in blood donors and therefore, either can be used for TTD screening in blood banks in India.

Utility of grey zone testing strategy in transfusion transmissible infection testing in blood bank is of limited value!

Several blood banks use grey zone (GZ) phenomenon (defined as samples with optical density within 10% below the cut off in enzyme immuno-assay [EIA]/chemiluminescence immunoassay [CLIA]) to further augment blood safety. There is paucity of data regarding usefulness of GZ sample and its application in Transfusion Transmissible Infection (TTI) screening procedures in blood transfusion services. We looked at our GZ sample results and their confirmatory test results to verify if it adds to blood safety in our set-up? We performed a prospective analytical study on blood donors' samples over two years. All the donors' samples were screened for TTI using CLIA. Samples with signal/cut-off ratio between ≥0.90 and <1.00 were classified under GZ. They were re-tested in duplicate and submitted to confirmatory testing: Neutralization Test for HBsAg, Immunoblot for HCV, and Western blot for HIV. Among the 50,064 blood donors donating the blood during study period, 573 (1.14%) donors were reactive for HBsAg, HCV, and HIV. Forty-seven (0.1%) TTI samples were GZ, but none was "confirmed positive." The utility of GZ testing seems to be limited. However, this may be continued for sake of "erring on the side of caution" and since this only results in negligible wastage (0.1%) of blood units.

Automated-red cell exchange for methaemoglobinaemia in a G6PD-deficient patient.

Methaemoglobinaemia in G6PD deficiency can be managed by oxidizing agents such as methylene blue and red cell exchange (RCE). We describe a G6PD-deficient patient who presented with oxidative stress with methaemoglobinaemia and was successfully managed with automated-RCE. At presentation, the patient had anaemia, was restless, was tired and had dyspnoea. Co-oximetry showed methaemoglo-binaemia of 10.1 U/g. Further testing revealed the patient had insufficient quantities of G6PD enzyme activity (0.1 U/g Hb). In view of methaemoglobinaemia, severe G6PD deficiency and signs of haemolysis, therapeutic RCE was planned. The patient underwent two automated-RCE procedures on consecutive days, bringing down his methaemoglobin levels from 12.5 to 0.1 U/g. In each procedure, 1.5 volumes of RCE at 100% balance rate was performed using 5 units of red blood cells. The patient responded well to RCE and other supportive treatment and was off medication and doing well at day 100 of follow-up.

Application of sequential serological testing strategy for detection of Hepatitis B surface antigen (HBsAg) for diagnosing HBV infection.

The diagnostic accuracy of any serological test for detection of HBsAg is not 100%. We hypothesized that the sequential testing strategy proven for anti-HIV laboratory diagnosis should also apply to other infectious disease markers like HBsAg. Therefore, we evaluated the diagnostic accuracy of these strategies, I (single test), II (two tests in sequence), III (three tests in sequence) for diagnosis in patients and blood donors and compared it to the confirmatory test for HBsAg (Neutralization Test). Samples were initially tested for HBsAg by A1- Enhanced Chemiluminescent Immuno Assay (ECLIA). Initial reactive (aliquoted donor/patient) samples were reflexly tested by A2- Enzyme Linked Fluorescence Assay (ELFA) and A3- Immuno Chromatography Assay (ICA) assays. Confirmatory neutralization assay was performed on all initial reactive samples. Four strategies (I, II A, II B, and III) that were used in this analysis were; I = A1, IIA = A1 + A2, IIB = A1 + A3, and III = A1 + A2 + A3. The results of all four strategies were compared to Gold Standard (Neutralization Test). A total of, 112, 011 blood samples (75,111 patient samples and 36,900 whole blood donor samples) were initially tested for HBsAg by A-1 (CLIA). Amongst the tested samples, 1,296, 1,188, 1,078, 1,074 samples were found to be reactive by strategy I, IIA, IIB, III respectively. We observed that the PPV (Positive Predictive Value) of Strategy III > Strategy IIB > Strategy IIA > Strategy I. Sequential serological testing strategy comprising of initial sensitive test followed by more specific test increases the diagnostic accuracy of test report as compared to a single test.

Applying newer parameter Ret-He (reticulocyte haemoglobin equivalent) to assess latent iron deficiency (LID) in blood donors-study at a tertiary care hospital in India.

BACKGROUND: It is important to detect Latent Iron Deficiency (LID) to prevent development of an overt iron deficiency anemia. Early detection is difficult by using conventional hematological and biochemical parameters. Soluble transferrin receptor (sTfR) is presently the gold standard for diagnosing LID. We evaluated the utility of Reticulocyte Hemoglobin Equivalent (Ret-He), a newer hematological parameter, to predict LID in blood donors as compared to sTfR. METHODS: This was a randomized prospective study performed on 501 donor samples over a period of three-months. All donors were included after administering medical history questionnaire and a brief physical examination in accordance with national guidelines (Hb ≥12.5). Additional samples were collected during donation according to the institutional standard operating procedure (SOP). All hemograms were performed on the Sysmex XE-2100 analyzer which included Ret-He. sTfR was measured in batch assays by ELISA (Biovendor, Czech Republic). Ret He <28 pg and sTfR≥3µg/ml were used to diagnose LID. Serum Iron, Total Iron Binding Capacity (TIBC) and Serum Ferritin were also measured simultaneously. RESULTS: Of the 501 blood donors, sTfR and Ret-He detected LID in 148 and 135 donors respectively. In comparison to sTfR, Ret-He had sensitivity of 92.7%, a specificity of 97.16%, PPV of 93.1% and NPV of 96.3%. Serum Ferritin, TIBC and serum Iron had comparatively lower sensitivity of 87.16%, 79.7% and 77.7% respectively. CONCLUSION: Ret-He can be used as a routine screening test to detect LID in blood donors. This could provide an opportunity to make appropriate and timely interventions like dietary changes or drug supplementation.